MD Degree in Plastic Surgery Thesis

Study of the Effect of Two Different Techniques of Early Burn Wound Excision on the Alteration of Interleukin-6 and Tumor
Necrosis Factor-Alpha Levels in Severe Burns

A Thesis Submitted for Partial Fulfillment of M.D. degree in Plastic and Reconstructive Surgery
Doctor Mohamed Ahmed El Rouby Consultant of Plastic & Reconstructive Surgery Ain Shams University Cairo Egypt +2 0101556023 +2 0126531265 http://www.elroubyegypt.com http://tajmeel.ohost.de elroubyegypt@elroubyegypt.com elroubyegypt@gmail.com elroubyegypt@hotmail.com dr_mohamed_a@yahoo.com . - -
To my Sons and family!
Keywords - Burn. - Tangential excision. - Down-to-fascia excision. - Fascial excision. - IL-6. - TNF-.
Index Index List of Figures List of Tables List of Charts List of Abbreviations Introduction Aim of the work Review of Literature
- Immune-Inflammatory Response to Burn
The Mediators of Inflammation The Immune-Competent and Effector Cells The Burn Toxin (Lipid Protein Complex) (LPC) Paradoxes in Burn Immune Failure: The Activation-Induced Cell Death Theory (AICD) (Apoptosis) The Role of LPC in Paradoxes in Burn Immune Failure The Role of Burn Wound Sepsis in the Post-burn Inflammatory Response The Pathogenesis of Multiple Organ Failure (MOF) Following Severe Burns
A
A C D E F 1 4 5 7 9 26 38 43 44 49 52 54 54 59 62 63 64 65 67 70 71
- Evaluation of the Burn Wound Management Decisions

I- Estimation of burn wound depth II- Surgical Burn Wound Management A- Surgical Methods of Burn Wound Closure B- Timing of Burn Wound Excision C- Extent of Burn Wound Excision D- Depth of Burn Wound Excision E- Techniques of Burn Wound Excision and Closure F- Coverage of Excised Burn Wound III- Impact of burn wound excision on the ImmuneInflammatory Response to Burn
Index Patients and Methods

- Study design - Patient population - Management Protocol - Monitoring of the patients - Statistical Methodology
B
75 75 75 77 83 87 89 89 91 91 93 95 98 98 103 108 109 110 111 112 125 130
Results
- Demography of Patients Population - Analysis of Clinical Outcomes
Clinical outcomes of first group Clinical outcomes of second group Analysis of the clinical outcomes in both groups
- Analysis of Laboratory Investigations

Laboratory results of first group Laboratory results of second group Comparison between IL-6 assay levels of survivors in both groups Comparison between TNF- assay levels of survivors in both groups Comparison between IL-6 assay levels of non-survivors in both groups Comparison between TNF- assay levels of non-survivors in both groups
Discussion Summary and Conclusion References Arabic Summary
List of Figures
No. Description 1 2 3 Complement activation pathways The arachidonic acid cascade Cytokines as communication links within the immune system, and between the immune system and other organs Biological functions of TNF- case no. 7 in first group (tangential excision) case no. 12 in second group (down-to-fascia excision)
C
Page 12 16 19
4 5 6
22 80 82
List of Tables
No. Description 1 2 3 4 5 6 7 8 9 10 11 The major inflammatory mediators The major cytokines released following thermal injury Paradoxes in Burn Immune Failure Patient Population of first group Patient Population of second group Clinical outcomes of first group Clinical outcomes of second group IL-6 assay results in pg/ml of first group TNF- assay results in pg/ml of first group IL-6 assay results in pg/ml of second group TNF- assay results in pg/ml of second group
D
Page 9 20 44 89 90 92 94 99 100 104 105
List of Charts
No. Description 1 2 3 Comparison between Average Hospital stay (AHS) of survivors and non-survivors in both groups Comparison between clinical outcomes of both groups IL-6 assay levels of survivors and non-survivors of first group at preoperative, 3rd, 7th and 14th postoperative (PO) days TNF - assay levels of survivors and non-survivors of first group at preoperative, 3rd, 7th and 14th postoperative (PO) days IL-6 assay levels of survivors and non-survivors of second group at preoperative, 3rd, 7th and 14th postoperative (PO) days TNF- assay levels of survivors and non-survivors of second group at preoperative, 3rd, 7th and 14th postoperative (PO) days IL-6 assay levels of survivors in both groups at preoperative, 3rd, 7th and 14th postoperative (PO) days TNF- assay levels of survivors in both groups at preoperative, 3rd, 7th and 14th postoperative (PO) days IL-6 assay levels of non-survivors in both groups at preoperative, 3rd, 7th and 14th postoperative (PO) days TNF- assay levels of non-survivors in both groups at preoperative, 3rd, 7th and 14th postoperative (PO) days
E
Page 96 97 101
102
106
107
7 8 9 10
108 109 110 111
List of Abbreviations
5-HT ABG AHS AICD AIDS Alb anti-PGPS IgM APCs b.p.m B - cells BDI bFGF BSA BUN C1 C3a, C3bBbP C3dg C4, C4b, C4a C5a CBC CD-system CD-14 CGRP CPK C-RP DIC EASIA EGF ET-1 Fn GIT GM-CSF
5-Hydroxytryptamine ( Serotonin) Arterial Blood Gases Average Hospital Stay The Activation-Induced Cell Death Theory (Apoptosis) Auto-Immunodeficiency Syndrome Serum albumin Antibody of Bacterial Cell Wall Antigen-PGPS Antigen-Presenting Cells beat per minute B-lymphocytes (The antibody-producing cells). Burn Depth Indicator Basic Fibroblast Growth Factor Burned Surface Area Blood Urea Nitrogen Complement factor 1 Complement factor 3 (Activated, blocked) Degraded product after activation of C3 Complement factor 4 (Activated, blocked) Activated Complement factor 5 Complete Blood Count Cluster Of Differentiation-System Lipopolysaccharide Receptor Calcitonin Gene-Related Peptide Creatine Phosphokinase C-Reactive Protein Disseminated Intravascular Coagulation Immunoenzymometric Quantitative Assay of Human Serum Cytokines Epidermal Growth Factor Endothelin-1 Fibronectin Gastro-intestinal tract Granulocyte-Macrophage Colony Stimulating Factor
gram+ve gram-ve Hb Hct HSR IFN-, and IGF-1 IgG IgM IL IL-1, IL-1 IL-2 IL-2R IL-6 IL-8 LG LPC LPS LTB4, LTD4 MBL MILP MOD MOF MRI N. NK OFRs -values PAF PaO2 PBD PBMC PDGF Bacteria positive to stain Bacteria negative to stain Hemoglobin Concentration Hematocrite Value Ratio of Helper To Suppressor T-lymphocytes Interferon , and Insulin-Like Growth Factor-1 Immunoglobulin G Immunoglobulin M Interleukins Interleukin-1 , Interleukin-2 IL-2 Receptor Interleukin-6 Interleukin-8 Large Granular Lymphocytes Heat Induced Toxin Lipoprotein Complex Gram-ve Bacterial Endotoxin Lipopolysaccharide Leukotrienes B4, D4 Mannose-binding lectin Mitogen-Induced Lymphocyte Proliferation Multiple Organ Dysfunction Multiple Organ Failure Proton Magnetic Resonance Imaging Non-Survivor Natural Killer (type of T-lymphocytes) Oxygen Free Radicals Probability value Platelet Activating Factor Oxygen pressure in arterial blood gases test Post-Burn Day Peripheral Blood Mononuclear Cells Platelet Derived Growth Factor
PGD2 PGE2 PGF2 PGI2 PGPS PMNLs PT PTT S. SaO2 SD SGOT SGPT SIRS SR STF TBSA TC T- cells TGF, TGF1, TGF2 TH TLC TM TNF- TS TSS TSST-1 TX A2 Prostaglandins D2 Prostaglandin E2 Prostaglandin F2 Prostacyclin Bacterial Cell Wall Antigen Polymorphonuclear Leukocytes Prothrombin Time Partial Thromboplastin Time Survivor Oxygen saturation in arterial blood gases test Standard Deviation Serum Glutamic Oxaloacetic Transaminase Serum Glutamic Pyruvic Transaminase Systemic Inflammatory Response Syndrome Survival Rate Subeschar Tissue Fluid Total Body Surface Area Burned T-Cytotoxic (type of T-lymphocytes)
T-lymphocytes (The Thymus-Dependent Lymphocytes)
Transforming Growth Factors , 1 and 2 T-Helper (type of T-lymphocytes) Total Leucocytic Count Thrombomodulin Tumor Necrosis Factor-Alpha T-Suppressor (type of T-lymphocytes) Toxic Shock Syndrome Toxic Shock Syndrome Toxin-1 Thromboxane A2
Introduction
Introduction
Although burn injuries are frequent in our society, many surgeons feel uncomfortable in managing patients with major thermal trauma. The burn wound is the source of virtually all ill effects, local and systemic, seen in a burned patient. Burn eschar exerts a systemic immune response that cascades through cytokine pathways leading to Systemic Inflammatory Response Syndrome (SIRS), which may
progress to Multiple Organ Failure (MOF) (Monafo et al., 1992). In addition, eschar acts as a nidus for infection that is aggravated by immune suppression state. This may progress to sepsis or sepsis-induced SIRS (Monafo et al., 1992). Munster in 1996, suggested that high serum level of tumor necrosis factor-alpha (TNF-) and low serum
level of Interleukin-6 (IL-6) may be considered to be the most important poor prognostic factors related to
Introduction
systemic
inflammation
and
multiple
organ
failure
following thermal injury. In addition, Yamamoto et al in 1996 stated that patients who are subjected to early escharectomy
showed a significant increase in blood platelets, decrease in fibronectin, white proteins and no blood cells, albumin and total variations in C-reactive
significant
protein level. Therefore, surgical removal of the burn wound in resuscitated patients especially when done early, results in improvement in survival rates and morbidity (Hart et al., 2003). There are two main techniques for early burn
wound excision, namely tangential and down-to-fascia excision (Herndon et al., 1999). It is hypothesized of that the accumulation fluid (STF) and may
reabsorption
subeschar
tissue
increase the morbidity and mortality rates in severely burned patients. Therefore, the unburned tissues at the
Introduction
m argin and the depth of the burn may be affected and may exaggerate the systemic inflammation (Chen et al., 2000). The study of the alteration of the immunological profile in relation to timing and extend of early excision has been established. However, the impact of technique of early burn wound excision on the immunological profile changes has not yet been studied.
Aim of The Work
Aim of The Work
The aim of this thesis is to compare the effect of two different techniques of early burn wound excision (tangential alteration excision of and down-to-fascia (IL-6) as and excision) tumor for on the
interleukin-6 (TNF-)
necrosis
factor-alpha
levels
indicators
immunological profile alterations. This would enable the burn surgeons to decide the proper technique and proper depth of early burn wound excision that would decrease the systemic inflammatory response in order to decrease hospital stay, decline the morbidity and improve clinical outcome and survival rate in extensively burned patients.
Review of Literature
The integument is the principle site of interaction with the surrounding world. It serves as a protective barrier, immunologic surveillance and thermoregulation. It consists
of two mutually dependent layers, the epidermis and dermis, which rest on hypodermis, which is a fatty subcutaneous layer, the panniculus adiposus (Van De Graff et al., 1986). Epidermal thickness is variable in different anatomic locations, sexes, and ages of man and ranges between 0.5 to 1.5 mm. This varying thickness primarily represents a
difference in dermal thickness, as epidermal thickness is rather constant throughout life and from one anatomic
location to another. The thickest epidermis is found in the palms and soles, while the thinnest epidermis is found on the eyelids and in the post-auricular region. Male skin is thicker than female skin in all anatomic locations. Children have relatively thin skin, which progressively thickens until the fourth or fifth decade of life when it begins to thin. This thinning is also primarily a dermal change, with loss of elastic fibers, epithelial appendages, and ground substance (Moore et al., 1998). Following a major burn injury a myriad of physiologic
changes occur that together comprise the clinical scenario of the burn patient (Herndon et al., 1999). Fluid and electrolyte imbalance, which results in
systemic intravascular losses of water, sodium, albumin and red blood cells and unless intravascular volume is rapidly restored, evidenced shock by develops. increased Metabolic resting disturbances are
oxygen
consumption
(hypermetabolism), an excessive nitrogen loss (catabolism), and a pronounced weight loss (malnutrition). Bacterial
contamination of tissues is another complication of major burns where patients are unable to mount an adequate
immunologic defense, increasing the risks for septic shock (Lowry, 1993). Later on, vital organs dysfunctions occur which
include, renal insufficiency can result from hypoperfusion or from nephron obstruction with myoglobulin; pulmonary
dysfunction may be caused
from initial respiratory tract
damage or from progressive respiratory insufficiency due to pulmonary edema, bronchopneumonia adult and respiratory distress gastrointestinal syndrome or
complications
which
include paralytic ileus and gastrointestinal ulcerations. Small
bowel ischemia and stasis promote bacterial translocation as a mechanism for endogenous infection (Beal et al., 1994).
Immune-Inflammatory Response to Burn

The burn wound is the source of virtually all ill effects, local and systemic, seen in a burned patient. Locally, the burn wound is characteristically made up of several
concentric three-dimensional zones of tissue damage due to different heat transfer. The zone of coagulative necrosis, where irreversible skin death occurs, is surrounded by an intermediate zone of stasis and the zone of hyperaemia, which is the outermost zone. Tissue damage in the zone of hyperaemia and the zone of necrosis, which do not show an inflammatory response, is primarily attributed to the direct effect of heat on blood vessels and tissue respectively. In contrast, in the zone of stasis, thermal injury triggers a pronounced inflammatory reaction (Knabl et al., 1999). Burn eschar through heat induced toxin lipoprotein complex (LPC) exerts a systemic immune response leading to Systemic Inflammatory Response Syndrome (SIRS),
which may progress to Multiple Organ
8
Dysfunction
Syndrome (MOD) or Multiple Organ Failure (MOF). In addition, eschar acts as a nidus for infection that is
aggravated by immune suppression state. This may progress to sepsis or sepsis-induced SIRS through lipopolysaccharides (LPS) which present in microorganisms (Ikeda et al., 2000). The systemic inflammation has a cellular component (consisting of different kinds of immune competent effector cells including leukocytes, platelets, and
vascular
endothelial cells, mast cells, and fibroblasts), and a vascular component permeability). active (related Both to blood flow of and the microvascular inflammatory Inflammatory
components collectively
response are governed by a wide variety of biologically products, termed Mediators forming networks and cascades which involved in the process of wound healing and tissue repair. These mediators are produced by the circulating immune-
inflammatory cells (e.g. the leukocytes), the plasma, and the tissue (Table 1). The inflammatory mediators fall into groups depending upon their origin and specific regulatory function (Cioffi et al., 1993).
Table 1: The major inflammatory mediators, (Arturson, 1996).
Mediator
Bradykinin
9
Action(s)
Pain, Vasodilatation Increased microvascular permeability Smooth muscle contraction Increased microvascular permeability PMNL and macrophage chemotaxis Mast cell degranulation Smooth muscle contraction Mast cell degranulation PMNL activation PMNL and macrophage chemotaxis Smooth muscle contraction Vasodilatation Increased microvascular permeability Increased microvascular permeability Smooth muscle contraction Chemokinesis Increased microvascular permeability Smooth muscle contraction Increased microvascular permeability Smooth muscle contraction PMNL activation Vasodilatation Vasoconstriction , PMNL chemotaxis Increased microvascular permeability Smooth muscle contraction
Origin(s)
Kinin system (kininogen) Coagulation system Complement C3 Complement C5
Fibrinopeptides Fibrin split products C3a C5a
Substance P Histamine
Sensory nerve endings Mast cells, Basophils
5-Hydroxytryptamine (5HT = serotonin) Platelet activating factor (PAF) PGE2 PGF2 LTB4 LTD4
Platelets , Mast cells PMNL, Macrophages, Basophils

Cyclooxygenase pathway Cyclooxygenase pathway
Lipoxygenase pathway Lipoxygenase pathway
I- The Mediators of Inflammation: 1- Oxygen Free Radicals (OFRs): The phagocytotic cells (polymorphonuclear leukocytes,
monocytes, and macrophages) are known to be a potent source for oxygen free radicals (OFRs), which are produced
10
in connection with their oxidative metabolism. OFRs are very short lived and their existence and pathophysiological role are extremely difficult to demonstrate (Haglund et al., 1991). Although they have no direct effect, yet they can markedly microvascular influence integrity effector in target cell movement through and the
organs
stimulation of lipid peroxidation, with subsequent production of biologically active arachidonic acid metabolites (Lipid
peroxides) (Kumar et al., 1995). 2- The Kinin System: The kinin system is activated in two different pathways, thereby and generating the bradykinin. inflammatory mediators, Activated Hageman bradykinin of the
lysyl
factor
clotting system acts on prekallikrein to generate kallikrein, which in turn releases bradykinin from low molecular weight kininogen. Activation of the plasmin system as well as enzymes release from generate damaged tissue tissue cells which act on
prekallikrein to Lysyl bradykinin
Kallikrein, low
releases weight
(Kallidin)
from
molecular
kininogen. Bradykinin is a very powerful vasoactive mediator,
11
which causes venular dilatation and increased microvascular permeability. Furthermore, it has an indirect effect on cell movement and microvascular integrity through its ability to activate phospholipase of the cell membrane, thus resulting in stimulation of the arachidonic acid cascade (Arturson, 1996). 3- The Coagulation and Fibrinolytic (Plasmin) Systems: The hypercoagulable state following thermal injury
results from activation of Hageman factor with subsequent activation of the coagulation cascade and ends by fibrin formation. This state is paralleled with increased activity of the plasma proteolytic enzyme plasmin, which serves to slowly degrade fibrin to fibrin degradation products. This can explain the high incidence of disseminated intravascular coagulation (DIC) in association with severe burns (GarciaAvello et al., 1998). 4- The Complement System Cascade (Opsonins): The complement system is a group of around 20
different serum proteins whose overall action is the control of inflammation. Complement occurs through two activation following thermal independent pathways, the
injury
classical 1996). and alternative pathway (Figure 1)
12
(Arturson,
Fig.1: Complement activation pathways (Kaneko et al., 2001)
The process of activation occurs in a cascade fashion that is closely similar to the blood-clotting mechanism.
13
Activation of either pathway ends up by a central common event, which is the activation of complement factor 3 (C3). The active fragments of C3 start to regulate the function of the phagocytic leukocytes (macrophages and neutrophils) via specific receptors on their surface (Kaneko et al., 2001). There are three major biological activities of the
complement system, the chemotaxis (attraction of phagocytic leukocytes towards their target), the opsonization (coating of the target so that it can be easily recognized by the phagocytic cells) and phagocytosis and lysis of target cells (Ono et al., 1993). Dibirdik et al. in 1995 conducted a clinical study to evaluate the changes in serum complement levels following thermal injury. They demonstrated an initial rise of serum complement C3 level, followed by a sustained decrease
during the next two to three weeks. Their explanation of the phenomenon was that, the initial rise was due to stimulation of the complement cascade by thermal injury, and the
subsequent decrease in levels was secondary to the initial increased consumption, thus resulting in depletion of the various complement components.
5- Neurotransmitters: Lofgren Substance were the P and and Lundberg Calcitonin in 1994 demonstrated peptide the
14
that,
gene-related sharing the of in
(CGRP)
neurotransmitters They modulate
inflammatory element of
process.
vascular vascular
inflammation effects.
through
alteration
permeability
6- Vasoactive Amines: They play an important role in the modulation of blood flow at the site of inflammation increased through vasodilatation, permeability. serotonin originate (5from amines, are
vasoconstriction, The most
and
microvascular include which
important and
components histamine, and
hydroxytryptamine) mast cells, their
basophils
platelets. effect on
Vasoactive
through
modulatory
microcirculation,
believed to share in post burn oedema formation, both locally at the site of injury and systemically in distant organs (Sanchez, 2002). 7- Platelet Activating Factor (PAF): It is produced by platelets, neutrophils (PMNL),
eosinophils, monocytes and vascular endothelial cells. It is
15
considered to induce marked enhancement of microvascular permeability following thermal injury, thus resulting in
oedema formation in local and systemic organ tissue. PAF seems to act synergistically with vasoactive amines to
produce marked alteration of microcirculation within target tissue (Schenfeld et al., 1990). 8- The Arachidonic Acid Cascade: Arachidonic acid results from the action of the enzyme phospholipase A2 on phospholipids of the cell membrane. The stimuli to this a process, variety which of is termed lipid
peroxidation, thrombin,
include
hormones, complex,
collagen, bacterial
bradykinin,
antigen-antibody
peptides, and oxygen free radicals (OFRs). Arachidonic acid is the main precursor for the biosynthesis different of many
biologically pathways
active involving
mediators,
through
enzymatic and
Cyclooxygenase,
5-Lipoxygenase,
15-Lipoxygenase enzymes. The mediators derived from the effect of the enzymatic cascade on arachidonic (PGI2), acid include thromboxane PGE2 A2, and
prostacyclin
prostaglandins
(PGD2,
PGF2), Lipoxins, and the leukotrienes (A4, C4, B4, D4 and
PE4). These mediators play an important role in
16
the both
thermally induced
inflammatory response,
acting on
components of inflammation (Figure 2). Those mainly acting on the cellular component can promote adhesiveness,
chemotaxis of leukocytes (e.g. Lipoxins, Leukotriene B4), platelet aggregation (Thromboxane A2), and antiaggregation (PGI2).
Phospholipides Lipase Arachidonic acid
Cyclo-oxygenase PGG2 TX synthetase isomerase TXA2 PGD2 PGE2 PGF2 TXB2 6-Keto PGF1 PGH2 PGI2 synthetase PGI2 hydrolase LTC4 synthetase 6-lipoxygenase LTC4
LTB4
LTC4 LTD4 LTE4
Fig. 2: The arachidonic acid cascade (Arturson, 1996)
The mediators acting on the vascular component (e.g. Prostaglandins, Leukotriene C4) are capable of modulating the microcirculatory and status increasing through vasoconstriction, permeability.
vasodilatation,
microvascular
Some mediators exert a dual effect (both cellular
17
and
vascular), such as thromboxane A2, PGI2, and Lipoxins. Therefore, the increased lipid peroxidation might be partially responsible for the indirect (ischaemia induced) local tissue damage following thermal injury (Arturson, 1990). 9- Cytokine Cascade: Cytokines are intercellular signal proteins or peptides that modulate the inflammatory response following trauma. They were previously termed lymphokines, as they were thought However, to have other the cells lymphocytes including as their only source. cells (PMNL, and
phagocytic
macrophages),
platelets,
fibroblasts,
endothelial
cells,
even keratinocytes, were all found to share in the production of such mediators. Cytokines act primarily on the cellular component of inflammation from which they originate, and serve to regulate leukocytes fall into the and a complex other interaction effector of cells groups, between (Figure of the 3).
different Cytokines
number
which
Interleukins consist the largest group. There is about 12 interleukins (IL-1 to IL-12), produced by the leukocytes and other effector cells, but not all of them
18
are of clinical significance following thermal injuries. The other cytokines that play an important role in thermal injury include the tumour necrosis factor-alpha (TNF-), interferon (IFN), neopterin, factor and granulocyte-macrophage (Table 2). colony are
stimulating
(GM-CSF)
Cytokines
known to exert their effect by binding to specific receptors present on the surface of immunecompetent and other
effector cells (Lowry, 1993). Interleukin-1 macrophages and (IL-1) acts to is produced mainly by and
stimulate
T-Lymphocytes
neutrophils. It can also act on the hypothalamus to induce fever, and liver cells to induce the production of acute phase proteins (e.g. C-reactive protein) (Dinarello et al., 1993). Interleukin-2 (IL-2) is produced by T-Lymphocytes and acts chiefly on all types of T-cells where it is the most powerful activator and growth factor. Besides T-cell, it can also activate macrophages (Teodorczyk-Injeyan et al., 1992). Interleukin-6 (IL-6) is produced by T and B cells, macrophages, fibroblasts, and endothelial cells and acts on many cells. In the liver, it stimulates the production of acute
phase proteins. Furthermore, it induces B cells
19
to
differentiate into antibody forming cells. Increasing levels have been observed in severely burned patients, denoting strong activation of cellular and humoral immune
mechanisms. IL-6 is also increased in burn blister fluid, suggesting a possible correlation with wound healing
(Bellomo, 1992).
Fig. 3: Cytokines as communication links within the immune system, and between the immune system and other organs (Arturson, 1996)
20
Table 2: The major cytokines released following thermal injury (Arturson, 1996).
Cytokine IL-1 Immune system Macrophages LGLs, B cells Other cells Endothelial cells Fibroblasts Main targets T cells, B cells Macrophages Endothelial cells Tissue cells T cells Main functions Activation of lymphocytes and macrophages. Leucocyte/endothelial adhesion. Acute-phase proteins. Activation of lymphocytes and macrophages. T-cell proliferation and differentiation. Induce acute-phase proteins. B-cell differentiation. Chemotaxis.
IL-2
T cells
IL-6 IL-8
T cells, B cells Monocytes
Fibroblasts
B cells Hepatocytes PMNLs Basophils Macrophages Granulocytes Tissue cells
TNF
Macrophages Lymphocytes Mast cells
Activation of macrophages, granulocytes and cytotoxic cells. Leucocyte/endothelial adhesion. Stimulation of acute-phase proteins and angiogenesis. Cachexia and pyrexia. Activation of macrophages. Leucocyte/endothelial adhesion.
IFN
T cells, NK cells
Epithelial cells Fibroblasts
Leukocytes Tissue cells
Interleukin-8 (IL-8) is a proinflammatory cytokine with a chemoattractant activity. It is produced by monocytes, endothelial stimulates cells, keratinocytes, and neutrophils. phagocytosis IL-8 and
neutrophil
IgG-mediated
oxidative burst. Furthermore, patients with total body surface area burn covering more than 40% have significantly higher
21
concentrations of IL-8 in their plasma than patients with small burns i.e. IL-8 is a major contributor of the systemic inflammatory response that follows major burns (Schroder et al., 1992). Tumour activated necrosis factor- and is (TNF-) is produced together by with
macrophages
considered,
interleukin-1, to be an alarm cytokine, that can stimulate TLymphocytes. TNF- Besides the the activation of of other T-Lymphocytes, cytokines, promoting and their regulates production and
stimulates
neutrophils
monocytes,
endothelial adhesiveness, phagocytosis, oxidative burst, and degranulation. In addition to its regulatory function on the cellular component of the inflammatory response, it can stimulate liver cells to produce acute phase proteins (e.g. Creactive protein), and the hypothalamus to induce fever
(Figure 4) (Tracey et al., 1993). Interferon (IFN) can be divided into three groups: IFN, and . IFN- is the only interferon that is known to play an important immunoregulatory function in thermal injury. It is produced by lymphocytes. It is considered to be the most important inducer of macrophage activation (Suzuki et al.,
1982).
22
TNF
Low quantities (plasma conc. <10-9 M)
Moderate quantities
High quantities (plasma conc. >10-7 M)
Local inflammation
Systemic effects
Septic shock
Leukocyte
activation
Brain
fever
Heart
Low output
Liver
Acute phase protein
Blood vessels
Endothelial cell
Adhesion molecule
IL-1, chemokines
Bone marrow
Thrombus
Low Resistance
Liver
Leucocytes Hypoglycemia
Fig. 4: Biological functions of TNF- (Lowry, 1993).
Neopterin works in close correlation with IFN-, being primarily released by macrophages upon stimulation by the latter. Neopterin is considered to be a marked activator of cellular mechanisms (Grabosch et al., 1992). The granulocyte-macrophage colony-stimulating factor
(GM-CSF) is a cytokine that plays an important role in cellular immune mechanisms. It stimulates the proliferation and differentiation of granulocyte and macrophage
23
progenitor cells in the bone marrow. GM-CSF is not only effective on immature cells but can also stimulate various functions of mature granulocytes (PMNL) and macrophages. It is evident that GM-CSF enhances the oxidative
metabolism, phagocytosis, and cytotoxic capacity for both granulocytes and macrophages (Kaufman et al., 1989). Munster in 1996, suggested that high serum level of TNF- and low serum level of IL-6 may be considered to be the most important poor prognostic factors related to
systemic inflammation and multiple organ failure following thermal injury. Deveci and colleagues in 2000 stated that high levels of IL-6 decrease the levels of TNF-. Therefore, it may be postulated that IL-6 inhibits the severity of the inflammatory response in the early period of thermal injury. On contrary, there is evidence that cytokines are
paradoxically involved in wound healing and tissue repair mechanisms. Ono et al. in 1995 demonstrated the presence of numerous cytokines and growth factors in the burn blister fluid. These included IL-1, IL-1, IL-6, IL-8, epidermal
24
growth factor (EGF), basic fibroblast growth factor (bFGF), platelet derived growth factor (PDGF), transforming growth factors (TGF, TGF1 and TGF2). Therefore, although
cytokines were primarily described as mediators of systemic inflammation with its related morbidity, they could
simultaneously promote wound healing procedures. 10- Fibronectin (Fn): Fibronectin is a glycoprotein secreted by many cells, e.g. the hepatocytes. It exerts a wide range of biological functions, which are primarily related to the cellular element of the post-burn immune-inflammatory response. Its function closely resembles that of the complement cascade, which acts as an opsonic system, but for the polymorphonuclear neutrophils phagocytic fibronectin leukocytes function activates of (PMNL). the Besides enhancing the
monocyte-macrophage and
system, the
thrombocytes,
regulates
activities of many other effector cells contributing in the acute inflammatory response. In addition, Fn is known to participate through in the sites organization of thrombus on formation, thus
special
binding fibrin
fibronectin,
enhancing the removal of soluble fibrin by macrophages.
25
Furthermore, Fn can play a beneficial role in the acceleration of wound healing, by promoting the phagocytosis of cellular debris by macrophages (Mosher et al., 1981). 11- Acute Phase Proteins (Acute Phase Reactants): The hepatocytes respond to trauma by liberating a
series of proteins into the circulation, collectively known as the acute phase proteins. The rise in plasma level of such proteins, does not only reflect the change in hepatic protein synthesis in response to trauma, but also reflects their
integral role in the host-protective mechanisms and tissue restoration procedures. The release of acute phase proteins by liver cells in response to thermal injury is triggered by the circulating cytokines, namely IL-1, IL-6 and TNF-
(Dowton et al., 1988). The acute phase proteins known to play a crucial role in the post-burn inflammatory response include C-reactive 2which protein, 1-antitrypsin, and 2-antichymotrypsin, Liver proteins,
macroglobulin,
haptoglobin.
exhibit a rise in the course of the acute phase response, are termed positive acute phase proteins. Other proteins, derived
26
from the liver, exhibit a marked drop in their plasma concentration, and hence are referred to as negative acutephase proteins. These include albumin and transferrin
(Castell et al., 1990). C-reactive protein is one of the sensitive indicators of the status of the post-burn inflammatory response as it is correlated well with the extent of the burn. Its exact
physiological role is not definitely understood (Latha et al., 1997). II- The Immune-Competent and Effector Cells: 1- Polymorphonuclear Neutrophil Leukocytes (PMNL): PMNLs constitute cells. over Their 90% main is of the circulating within the They
polymorphonuclear
function
immune-inflammatory
response
phagocytosis.
constitute one of the two major arms of phagocytosis, the second being mediated by the monocytic-macrophage
system. Neutrophils are derived from the bone marrow. The initial event that immediately follows stimulation to each is the other
phenomenon of
aggregation of leukocytes
(Rolling). This is followed by adhesion of the leukocytes to
27
the vascular endothelium in the target tissue (Adherence). The phenomenon of rolling and adherence, which result in segregation of leukocytes from the circulation, are followed by extravasation, then migration of the effector cells towards their target (Chemotaxis). Leukocytes eventually bind, and then phagocytose their target cell. Phagocytosis is followed by a process of lysis of the ingested microorganism
(Arturson, 1996). The specific phenomena receptor-ligand of rolling interactions and adherence by involve three
modulated
families of cell surface proteins. These are the selectins (which mediate the phase of rolling and aggregation), the integrins (which mediate the phase of adhesion) and the platelet-endothelial adhesion molecule 1 (which mediates
the phase of endothelial transmigration) (Muller et al., 1993). The chemotaxis is modulated by chemotactic molecules acting on leukocytes such as activated complement 5 (C5a), leukotriene B4 (LTB4), and interleukin-8 (IL-8). The
phenomenon of opsonization (phagocytosis) is facilitated by the opsonic proteins which include immunoglobulin G (IgG), complement 3 (C3), fibronectin, and C-reactive protein
(Solomkin et al., 1984).
28
The process of lysis is mediated through two main pathways. The first pathway is the oxygen-dependent killing, in which the leukocyte by utilizes energy derived enzyme from with
oxidative
metabolism
myeloperoxidase
production of Oxygen free radicals (OFRs), as metabolic wastes. The second pathway of intracellular killing is known as the oxygen-independent killing, under the direct effect of the lysozymes manufactured by the leukocyte secretory
granules (Arturson, 1990). The cytoplasm of PMNLs contains many secretory
granules with specific enzymatic protein contents. The most important secretory granules are those responsible for the production of myeloperoxidase enzyme, and the lysozymes. Other secretory granules are responsible for the production of various enzymes including collagenase, elastase, neutral proteases, and lactoferrin. Lactoferrin can affect stem-cell proliferation into mature neutrophils and regulate all types of neutrophils activities. Proteolytic enzymes (e.g. collagenase and elastase) are believed to exert a damaging effect in target tissues, and hence can also share in the pathogenesis of
functional organ failure (Arturson, 1996).
29
The initial early post-burn phase of strong activation of PMNLs function is eventually followed by phase
characterized by marked impairment of their activities. This serious dampening of neutrophil functions eventually complications
predisposes to the occurrence (El-Falaky et al., 1985). Thus, both OFRs and
of infectious
proteolytic
enzymes
are
considered to be of great pathological importance, as they can mediate much of the tissue damage and organ
dysfunction attributed to systemic inflammation after major thermal injuries (Cioffi et al., 1993). Vindenes and Bjerknes in 1997 reported that, the
second stage where there occurs a marked failure of PMNLs function might be attributed to abnormalities in the
contractile protein actin. Actin is a key component of the neutrophil cytoskeleton. Continuous polymerization and
depolymerization of actin is responsible for generating the force for several forms of motile responses, principally
locomotion, but also shape change, pseudopod formation,
30
phagocytosis and secretion. Furthermore, a strong correlation exists between the neutrophil microfilament apparatus and the surface adhesion molecules, which mediate the
leukocyte-endothelial serve as the link
interaction. between
Thus the
surface
molecules and the
cytoskeleton
extracellular matrix. This link depends upon a similarity between actin filaments, and the adhesion molecules on the surface of leukocytes and endothelial cells. 2- The Monocytic-Macrophage System: Monocytes are always circulating in the blood stream. Upon stimulation, they enter into target tissues where they undergo maturation into macrophages. Besides its major
function in phagocytosis, the monocytic-macrophage system is known to work in intimate lymphocytes. alarm This occurs by correlation with the the production effect on Tof T-
through
interleukin-1 and tumour necrosis factor (TNF), which act as cytokines their triggering
lymphocytes. Neopterin is another cytokine originating from the monocytic-macrophage system, and can trigger many of the cellular immune-inflammatory mechanisms (Arturson,
1996).
Therefore, macrophages can modulate
31
the
inflammatory response
through two major
pathways. The
first pathway is related to cytokine production (cytokine cascade) and results in marked activation of many effector cell lines, mainly the T-lymphocytes. The second pathway is related to OFRs production, and results in activation of the arachidonic acid cascade, which in turn plays an integral role in both vascular and cellular elements of inflammation
(Sparkes, 1997). Macrophages can also participate in tissue restoration mechanisms by phagocytosis of cellular debris and
production of proteolytic enzymes (e.g. collagenase), thus promoting the separation of necrotic tissue (e.g. eschar). Furthermore, many of the cytokines produced by
macrophage activation can promote healing by stimulating the proliferation of fibroblasts and keratinocytes (Leibovich, 1984). 3- The lymphoid System: Lymphocytes express a large number of different
molecules (markers) on their surfaces. These molecules are peptide (protein) in nature. Some of these molecules function
32
as receptors for cytokines. The peptide nature of these molecules (markers) enables their identification by specific monoclonal antibodies, constituting the CD-system (cluster of differentiation-system). Thus, one can easily know the ratio between the different lymphocytes and the status of activity of the various cytokine receptors (Arturson, 1996). a- The B-lymphocytes: (The antibody-producing cells). They differentiate into plasma cells, which produce
antibodies. Antibodies exert many functions including the neutralization of toxins, and the enhancement of
phagocytosis by PMNLs. Furthermore it may play a role in triggering the arachidonic acid cascade, which plays an
important immune-regulatory function (Roitt et al., 1993). The B-lymphocytes are the first to recognize the
antigen, and then present it to the T-lymphocytes. Thus, they act as antigen-presenting cells (APCs) for the T-
lymphocytes, which start to produce cytokines, those are essential Therefore, for it is antibody clear production that, although by B-lymphocytes. are
B-lymphocytes
responsible for the initial event (antigen presentation), yet their further participation in the immune-inflammatory
response is through a cytokine-mediated
33
(T-lymphocyte-
derived) triggering of proliferation and differentiation into active plasma cells (Arturson, 1996). Teodorczyk-Injeyan demonstrated immunoglobulin denoting the an M initial (IgM) and rise in an the coworkers in plasma in level 1989 of
early
post-burn
phase, A
triggering of
inflammatory response.
parallel rise in interleukin-2 (IL-2) level in plasma was observed. This indicated that, immunoglobulin production by B-lymphocytes is a cytokine (IL-2 dependent) event. b- T-lymphocytes: (The thymus-dependent lymphocytes). The component T-lymphocytes of the constitute the other major system.
immune-regulatory
lymphoid
They are stimulated by the antigen-presenting cells (APCs). Being stimulated, into a T-lymphocytes wide include variety the proliferate of T-helper and
differentiate subpopulations.
T-lymphocyte (TH), T-
These
suppressor (TS), T-cytotoxic (TC), natural killer (NK), and large granular lymphocytes (LG) (Bach et al., 1979). According to the CD-system, T-Helper cell (TH cell)
equals equals CD4+ CD8+. and T-suppressor/T-cytotoxic the CD-system (TS/TC) can
34
ratio
Similarly,
identify
interleukin-2 (IL-2) receptors as CD25, activation inducer molecules as CD69, and transferrin receptors as CD71.
Natural killer cells (NK) are recognized as CD16 and Blymphocytes as CD20 (Arturson, 1996). Each T-lymphocyte subpopulation is concerned with a specific immune-regulatory function within the
inflammatory response. The various components of the Tlymphocyte system are complexly interrelated via the
cytokine network. Cytokines also help in integrating the Tlymphocyte functions. affect the functions Thus with other immune-competent through cells vascular of cytokines, the cell can
T-lymphocytes, of other
functions system,
immunecells,
inflammatory
including
endothelial
fibroblasts, mast cells, and phagocytic cells (PMNLs and macrophages) (Lowry, 1993). Thermal injury results in enhancement of T-lymphocyte functions, which is indicated by an increase in the ratio of helper to suppressor cells (HSR). This initial stimulation of the T-lymphocytes is followed by a phase of marked
35
exhaustion of their functions. Suppression of T-lymphocyte functions eventually creates a status of marked increase in the susceptibility of extensively burned patients to infectious complications (Rioja et al., 1993). 4- The Mast Cells: Mast cells lies close to the blood vessels in all tissues. They constitute an important Upon content cellular stimulation, of element they of start The the to most
inflammatory liberate their
response. granule
mediators.
important of these is histamine with its potent effect on the vascular element of inflammation. Other mediators include arachidonic cytokines. acid Thus, metabolites mediators (lipid derived IL-8, peroxidases) from mast and cell
degranulation have three main physiological effects. They are chemoattractants (TNF-, PAF, LTB4, PAF), and
vasoactivators
(Histamine,
kininogenase),
spasmogens (Histamine, PGD2, LTC4, LTD4). Therefore, it is clear that mast cells, through their rich content of
mediators, can markedly influence effector cell movement, blood flow, and microvascular integrity (permeability) in
target tissue (Gordon et al., 1990).
5- Platelets:
36
Platelets are derived from megakaryocytes in the bone marrow. They have receptors for coagulation factors,
enabling them to share actively in the process of blood clotting. In addition, platelets response, are and also is involved rich in in the
immune-inflammatory
secretory
granules which contribute in both inflammation and tissue restoration, Those through many biologically active products.
concerned
with
inflammation
include
Thromboxane
A2 (vasoconstrictor and proaggregator), platelet factor IV (chemotactic), platelet activating factor (PAF) (enhancement of microvascular permeability). On the other hand, plateletderived cytokines that are known to mediate healing (IGF-1),
mechanisms involve
insulin-like growth factor 1
platelet derived growth factor (PDGF), epidermal growth factor (EGF), and transforming growth factor (TGF-). These mediators can promote the proliferation of both
fibroblasts and keratinocytes (McGrath, 1990). 6- Vascular Endothelium: The endothelium of small vessels (arterioles, capillaries and postcapillary venules) plays important regulatory
37
functions throughout the course of inflammation. This occurs through affecting the production vascular The chief of biologically and cellular active mediators of
both
elements
inflammation.
endothelium-derived
mediators
include prostacyclin (PGI2) which is lipid peroxide with vasodilator effect, and various cytokines (Arturson, 1996). The vascular endothelial cells have surface receptors (protein in nature), which play an important integral function in the locomotion of phagocytic cells (PMNLs and
macrophages) (Bevilacqua et al., 1993). In other addition, biologically (ET-1), be vascular endothelial products. cells produce are ET-1 two the is
significant and
These (TM). of
endothelin-1 believed to
thrombomodulin in the onset
involved
disseminated
intravascular coagulation (DIC) when produced in excess by vascular endothelial cells. TM is a membrane protein with antithrombus activity and is also involved in DIC (Ishibashi et al., 1991). Nakae and colleagues in 1996 found a significant
increase in the plasma levels of ET-1, TM, and TNF- in
38
patients who developed MOF (multiple organ failure) and died in comparison to those who survived. Their findings strongly suggested that the three mediators are intimately related and can reflect the severity of the inflammatory response in systemic organs and subsequently the degree of multiple organ dysfunctions. III- The Burn Toxin (Lipid Protein Complex) (LPC): Careful biochemical analysis of the eschar enabled the isolation of a heat-induced toxic material. The isolated toxic material from the burned skin proved to be a polymerized aggregate of lipids and proteins of the cell membranes in the burned skin. The burn toxin (LPC) is elaborated in the eschar under the polymerization effect of heat, and then, is
continuously absorbed into the circulation at the escharsubeschar (viable-nonviable) interface. Thus, LPC can also be isolated by careful biochemical assay of the sera of burn victims. In the circulation, the eschar (LPC), by acting as an antigen, results in triggering of a system inflammatory
response (Schoenenberger et al., 1971). When a patch of skin is separated from an animal,
39
burned, and then applied back to the area of the mouse from where it has been removed, it became highly toxic, and resulted in death of the mice from severe inflammation in systemic organ and eventually multiple organ failure (MOF). If a barrier is placed under the burned skin patch, mortality does not develop. These findings strongly suggest the
absorption of toxic material from the burned skin (LPC), with subsequent strong triggering of a systemic
inflammatory response (Schoenenberger et al., 1971). Animal experiments showed that, a sterile homogenate of burned skin, injected into the mouse peritoneal cavity, killed the mice whereas sterile homogenates of unburned normal skin were not toxic. These findings confirmed the capacity of burned skin to initiate a sustained systemic inflammatory response, by being a constant source for burn toxin (LPC) (Schoenenberger et al., 1975). Demling and Lalonde in 1990 showed that in more than half of the patients dying from the systemic inflammatory response with its deleterious effects on vital organ function, bacteria are not demonstrated. Their suggestion was that, the uncontaminated eschar, by being a constant source for
40
(LPC), could strongly enhance the systemic inflammatory response. Following severe burns, the gut, due to altered bacterial endotoxin and the The
permeability may become a constant source of toxins, including of the cell gram-ve bacterial (LPS),
(lipopolysaccharide gram+ve exotoxins
membrane)
(superantigens)
(enterotoxins).
translocating toxins may result in triggering of a systemic inflammatory response and subsequent vital organ
dysfunction. Thus, in the absence of burn wound infection, there is much controversy about the triggering antigen of the systemic inflammatory response, whether it is the escharderived LPC or the gut-derived LPS and enterotoxins (Yao et al., 1995). Yao and colleagues in 1995 showed a significant
lowering of plasma endotoxin level that was associated with marked improvement of survival rate after selective
intestinal decontamination. On the other hand, Likewise, Carsin et al. in 1997 failed to show a significant correlation between plasma levels of
41
IL-6, and TNF-, being significant members of the cytokine cascade, and high They plasma concluded levels that of the gut-derived post-burn
endotoxaemia.
inflammatory response is mainly LPC-dependent, and thus can occur irrespective of gut-derived toxaemia. The fact that the sterile burn eschar is a continuous source for a specific burn toxin (LPC), which can trigger a vigorous systemic inflammatory response with multiple
organ dysfunctions, provided enough evidence that the burn eschar is a dangerous thing to preserve. Early excision of the burn eschar and wound closure eventually eliminates the pernicious burn toxin before gaining access into the
circulation at the interface between the eschar and subeschar viable tissue. Thus, the improvement in survival rate
following extensive burns, by prompt excision and wound closure, is primarily attributed to marked dampening of the post-burn LPC-triggered systemic inflammatory response
and the related vital organ dysfunction (Allgower et al., 1995). Kistler et al. 1990, conducted an experimental study on rats to evaluate the effect of cerium nitrate on the liberation
42
of the burn toxin (LPC) from the eschar. Rats covered with cerium-treated control group burned covered skin survived, in contrast skin. to the They
saline-soaked
burned
concluded that cerium nitrate, by having a high binding affinity for LPC, is able to fix to LPC within the eschar, thereby preventing and it from gaining in its access into the and
circulation,
resulting
denaturation
neutralization. Thus, cerium nitrate exerts a simple chemical excision of the eschar. Kistler et al. also suggested the possibility of using cerium nitrate in human burns, to affect a non-traumatic (LPC), thus fixation avoiding of the burned loss, skin-derived trauma, toxin and
blood
surgical
removal of second degree burn skin, which all accompany the practice of early surgical excision of the eschar. Scheidegger and coworkers in 1992 and Lamaie and colleagues in 1999, achieved dramatic improvement in
survival of burned patients by bathing them, at the time of hospital admission, in cerium nitrate on a 30-minutes basis. They suggested that such non-traumatic excision of the burn eschar should be a reliable alternative to early surgical excision practice.
43
IV- Paradoxes in Burn Immune Failure: The ActivationInduced Cell Death Theory (AICD) (Apoptosis) (Table 3): When T-lymphocytes, taken from burned patients, are stimulated in vitro, they show marked reduction in their main product, interleukin-2. They also show a markedly deficient expression of IL-2 receptor (IL-2R) on their surface. These findings suggest a definite failure in T-cell function.
However, paradoxically, the serum of burn patients is found to contain high levels of IL-2 and IL-2R. This clearly indicates that immune cell activation event takes place in vivo, in response to the burn, and that the in vitro results represent the activity of cells, which became exhausted or refractory. Therefore, it is evident that T-lymphocyte failure is not an initial event, but follows a state of vigorous initial activation in vivo (Teodorczyk-Injeyan et al., 1991). In addition, In vitro culture of macrophages withdrawn from burn patients show marked reduction in interleukin-1 production, despite a high level of the same cytokine in vivo. Thus, macrophages exhibit failure, secondary to a strong in vivo activation (Liu et al., 1994).
* Immune paradox of burns Tissue ischemia and destruction. Loss of barrier functions. Foreign body introduction. Inoculation of microorganisms. Helper-cell dysfunction. Neutrophil dysfunction. Complement depletion. Abnormal opsonic activity. Stress-associated hormones. Immunosuppressive substances. Increased PGE2 level.
44
Table 3: Paradoxes in Burn Immune Failure (Atiyeh et al., 2005).
Vindenes and coworkers in 1994 described the pattern of behaviour of PMNLs following thermal injury in the form of an initial systemic activation of all functions including the expression of surface adhesion molecules, surface receptors for opsonins (immunoglobulins and complement),
phagocytosis, oxidative burst and intracellular degradation of ingested microorganisms. The following impairment of
PMNL functions included the whole pattern of previously enhanced activities. The Role of LPC in Paradoxes in Burn Immune Failure: Schoenenberger et al. in 1975 concluded that LPC is a significant mediator of weakened host defenses following a
thermal insult.
45
Consistent with the previous finding, Echinard et al. in 1982 demonstrated that, immediate excision of the burn eschar in Guinea pigs could significantly enhance the host defense mechanisms. They stressed that the eschar-derived LPC should be the main cause for the observed post-burn status of immunesuppression. They also stated that, their results are perhaps convincing in the way that prompt
excision of the burn eschar should similarly enhance host defenses in human burns. Heberer et al. in 1982 studied the behaviour of
peripheral blood phagocytotic cells, when incubated in vitro, in the presence of LPC. They found that LPC initially enhanced phagocytosis but further contact with the
granulocytes and monocytes proved an exhaustive effect on their function. They concluded that the burned skin-derived LPC has a dual effect on immune cells, an initial triggering effect and a later exhaustive effect, leading to suppression of all functions. Munster and colleagues in 1986 suggested that gut-
46
derived bacterial toxins, due to bacterial translocation might play a role in the pathogenesis of the phase of post-burn immunesuppression. circulating They found was that markedly although reduced the by
endotoxaemia
administration of polymixin B, yet it failed to show any enhancement in T-lymphocyte function. Munster et al.
concluded that, although gut-derived toxaemia is a common finding following thermal insult, yet it could not be
considered as the main trigger for the observed immuneinflammatory response, nor for the subsequent
immunefailure. Instead, it is suggested that the burned skinderived LPC is the primary aeteological factor for both aspects of the post-burn immune dysfunction, i.e. the initial stimulation, and the later on suppression. Sparkes in 1991 studied the behaviour of T-
lymphocytes when cultured in vitro, in the presence of LPC. He found that LPC was able to stimulate IL-2 production in resting competent cells, therefore acting as an antigen.
However, paradoxically (LPC) inhibited the cells that are already IL-2-dependent. This latter ability of (LPC) to arrest the growth of cells that are already IL-2-dependent is termed
the activation induced cell death (AICD) (Apoptosis).
47
Consequently, the LPC can be considered as the chief mediator of both the post-burn inflammatory response and the following status of immune-failure with increased
susceptibility to infection. Cioffi et al. in 1993 stated that, thermal injury, like any other major trauma, triggers the release of corticosteroids and catecholamines from the suprarenal a glands. Such
endogenous
hormones
have
well-known
immune-
suppressive effect. Therefore, they are suspected to be the cause of post-burn immune failure. However, Cioffi and coworkers concentrated upon the fact that the post-burn
immune failure is far more serious than in any other major trauma (e.g. blunt trauma). Thus, it is suggested that postburn immune failure is a specific (LPC)-mediated
phenomenon, while endogenous hormones are believed to be secondary dysfunction. Allgower peripheral et al. in 1995 studied cells the behaviour i.e. of Tnon-specific contributors in immune cell
blood
mononuclear
(PBMC),
48
lymphocytes, withdrawn from burn patients and cultured in vitro. They compared the in vitro capacity of lymphocytes to produce IL-2 and and to express IL-2R by in patients once treated in
conventionally,
patients
treated
bathing
cerium nitrate. They found that, in the first group, the Tlymphocyte function was markedly dampened in vitro. In contrast, lymphocyte withdrawn from cerium nitrate-bathed patients showed a sustained capacity to produce IL-2 and to express IL-2R. The explanation was that, cerium nitrate by fixing LPC can prevent much of its paradoxical suppressive effect on T-lymphocyte function, thereby achieving an
increased tolerance to infection. Sparkes in 1997 stated that the burn-induced, escharderived toxin (LPC) lies behind all the observed post-burn immune cell dysfunctions. Initially, the LPC, released into the circulation at the viable-nonviable interface, acts as an antigen, thus resulting in a systemic immune-inflammatory response (SIR). This LPC-triggered systemic inflammation
includes various cascades (e.g. the cytokine cascade), and takes place in all vital organs, hence the name internal inflammation. The magnitude of the LPC-triggered SIR is
49
directly proportionate to the total body surface area burned (percentage of TBSA), and can eventually end up by vital organ dysfunction and failure (MOF). Thus, the first issue is that the pernicious LPC can directly kill the burn victim, without the presence of bacteria. Measures like prompt
excision and bathing in cerium nitrate proved efficiency in improving survival by eliminating the burn toxin, and thus dampening the systemic inflammatory response.
V- The Role of Burn Wound Sepsis in the Post-burn Inflammatory Response: It is strongly is not evident, a nowadays, that burn in wound burn burn infection, primary even a contributor
pathophysiology.
Thus,
non-contaminated
eschar can threaten a burn patients life, by the continuous leaching of the pernicious burn toxin into the circulation. It is therefore not surprising to have records of 50 % mortality from extensive burns without evidences of burn wound
infection. It is only when the initial LPC-mediated (SIR) is not strong enough to deteriorate vital organ functions, that burn wound infection comes to the scene. In such situation,
50
bacterial toxins start to menace the immune-competent cells. Due to the immune failure, paradoxically induced by LPC, the bacterial toxin-mediated inflammatory response becomes enhanced and sustained, thus resulting in vital organ
dysfunction. Therefore, it is clear that, the LPC, by inducing immunesuppression, creates a status of persistent bacterial toxin-mediated 1990). Bacterial toxins, which can menace the immunesystemic inflammation (Demling et al.,
inflammatory system, fall into two major groups. They are either derived from the cell membrane of gram-ve bacteria, hence the name (endotoxins), or produced by gram+ve
organisms, hence the name (exotoxins). Endotoxins of gram -ve bacteria are lipopolysaccharide in nature (LPS), while the exotoxins of gram+ve species are alternatively termed bacterial superantigens (enterotoxins). Both groups are
known to drive the infection-mediated post-burn systemic inflammatory response (SIR). Thus they can induce vital organ dysfunction and shock, i.e. an endotoxin or enterotoxic shock syndrome, comparable to the burn toxic shock
mediated by the (LPC) (Allgower et al., 1995).
51
Childs et al. 1999, studied the pattern of illness in gram+ve sepsis of burn wound, in relation to the type of exotoxin produced. They demonstrated that all types of
gram+ve exotoxins could trigger a systemic inflammatory response, with subsequent development of toxic shock
syndrome (TSS). TSS is specific to a special strain of staph. aureus producing a special exotoxin, which was termed the toxic shock syndrome toxin-1 (TSST-1). Tanaka comparative and clinical colleagues study to in 1995 the conducted a
evaluate
hemodynamic
changes resulting from toxic shock syndrome toxin-1 staph aureus sepsis, and the endotoxin-producing gram-ve rod
sepsis in patients with severe burns. They stated that, in spite of the widely spread data concerning gram-ve rod sepsis, i.e. the endotoxic shock syndrome, little information is known about the severity of the toxic shock syndrome toxin-1 gram+ve coccal sepsis. The findings of the study reported that toxic shock syndrome toxin-1 gram+ve coccal sepsis induces hyperdynamic hypermetabolic responses that are
equal or even more profound than does the endotoxinproducing gram-ve rod sepsis. The results of Tanaka and co-
52
workers are considered to be the first clinical report that TSST-1 sepsis may result in more profound responses than does endotoxin sepsis. This may be explained based on a stronger stimulation of the inflammatory cascades (e.g.
cytokine cascade), by the TSST-1 than by the endotoxin.
VI- The Pathogenesis of Multiple Organ Failure (MOF) Following Severe Burns: Cioffi and coworkers in 1993 stated that, following thermal injury, there occurs a sustained triggering of a systemic inflammatory response (SIR). The initial chief
stimulation is mediated by the burn-induced eschar-derived toxin (LPC). Subsequent stimulation of immune-competent
cells is mediated by wound bacterial toxins, resulting from an LPC-mediated immune failure. Both groups of bacterial toxins, the gram-ve derived endotoxins (LPS), have and an the equal
gram+ve-derived
exotoxins
(Superantigens),
capacity to trigger systemic inflammation. Thus, systemic inflammation is initially triggered by the LPC, then sustained and enhanced by the LPS and bacterial superantigens. The systemic inflammatory response (SIR) entails the activation
53
of various cascades of mediators that coordinate the function of immunecompetent cells. Eventually, the persistent
inflammation in systemic organs (e.g. heart, liver, kidney, and GIT) results in dysfunction (MOD), then complete
failure of function (MOF) and death of the burn victim. Arturson, 1996, stated that, although the pathogenesis of MOF is primarily attributed to the burn toxin (LPC) and wound bacterial toxins (LPS and superantigens); yet other factors may also contribute to the systemic inflammatory response. Surgical procedures and gut-derived toxins (due to translocation) may add to the initial LPC-driven systemic inflammatory response (SIR). Surgical trauma and gut-
derived sepsis may also result in leukocyte exhaustion, thus enhancing the specific LPC-mediated immunesuppression. It is thus concluded that surgical trauma and gut-derived toxins are only contributors to immunestimulation and exhaustive immune failure, while the chief mediator of both aspects of immune-dysfunction is the burn toxin (LPC).
54
Evaluation of Burn Wound and Management Decisions

The hydrophilic human skin possesses a high specific heat and a low thermal conductivity. Therefore, skin
becomes overheated slowly, but also cools slowly. As a result, thermal damage continues after the burning agent is extinguished or removed (Carvajal et al., 1979). In addition to the appearance (i.e. extent and depth) of the wound, other factors can determine burn wound
management decisions. These factors include the type of burn, the age of the patient, and the circumstances
surrounding the injury (Herndon, 2001). For many years, However, deep modern burns were treated strategies
conservatively.
treatment
involve early aggressive surgical removal of the deep burns. Therefore, an accurate estimation of burn depth becomes crucial (Herndon et al., 1986). I- Estimation of burn wound depth: The standard technique for determining burn depth has long been clinical observation of the wound. Unfortunately, the difference in burn depth between a deep dermal burn and
55
a full-thickness burn may be only a matter of only a few tenths of a millimeter. Further, a burn is a dynamic process; therefore, what appears shallow on first day may appear deep by third post-burn day. Finally, the kind of topical wound care used can dramatically change the appearance of the burn (Hlava et al., 1983). Because increased of these in limitations, planning and because burn of its
importance
definitive
wound
care, interest has been stirred and technology has brought numerous devices and techniques to determine burn depth more precisely than clinical observation. These techniques have been used based on the physiology of the skin and alterations produced by burn. These techniques take
advantage of, the ability to detect dead cells or denatured collagen (e.g. biopsy, ultrasound, vital dyes), altered blood flow (e.g. fluorescein, laser Doppler, and thermograph), the color of the wound (e.g. light reflectance), and physical changes, such as edema (e.g. magnetic resonance imaging) (Wachtel, 1989 and Herndon, 2001). The burn depth indicator (BDI) should be 100%
accurate for shallow and full-thickness wounds for any of
these techniques (Herndon, 2001).
56
Histological wound biopsy would seem to be the most precise diagnostic tool. However, biopsies leave permanent scars in partial-thickness wounds, they are expensive and they require an experienced pathologist to tell live from denatured collagen and cells. Further, there is no guarantee that areas adjacent to the biopsy are the same depth (Jackson, 1953). Ultrasound technique has a problem that collagen
denatures at 65C while the epidermal cells, from which the burn must heal, are destroyed at about 47C. As a result apparent depth is likely to be underestimated by ultrasound technique (Brink et al., 1986). Vital would be Dyes useful application in directly to the burn and wound also in
detecting dead
tissue
determining the depth of excision. Davies and coworker in 1980, described important characteristics of such a dye. It should stain only dead tissue, not be removable with wound treatment, be nontoxic, provide a sharp demarcation between living and dead tissue, penetrate all dead tissue, and be
compatible with topical treatments usually used in burn care.
57
Methylene blue, which is metabolized to a colorless compound sulfadiazine discoloration by and living cells, when mixed a with silver blue
applied
topically, 48
significant which
appeared
within
hours,
remained
after vigorous washing (Zawacki et al., 1970). Fluorescein Fluorometry is another technique, where
fluorescein injected systemically. It is delivered through the patients circulation and fluoresces under ultraviolet light. Partial-thickness burns uniformly will exhibit fluorescence
within few minutes, but full-thickness burns will show nil (Grossman et al., 1984). Laser Doppler Flowmetry technique depends on the
electrical signal of blood flow in normal versus burned areas of skin. The laser Doppler has the advantage of being easy to use and non-invasive (Park, 1998). Thermography is based on the concept that the
diminished blood flow to deep dermal and full-thickness burns makes them cooler. However it is highly dependent on room and patient temperature, the patients anxiety and
stress level (Henane et al., 1981). Light Reflectance (Spectroscope) could estimate
58
the
depth as the skin is relatively transparent to short wavelength infrared light, and reduced hemoglobin absorbs more of the light than oxygenated hemoglobin (Anselmo et al., 1973). Proton Magnetic Resonance Imaging (MRI) correlate
with tissue water content, where full-thickness burns result in slower resorption of wound edema than partial-thickness burns (Koruda et al., 1986). Based on these widely differing techniques, it becomes apparent that precise determination of burn depth awaits further assessment 2001). Age of the patient is another determining factor for burn wound management decisions. There are many factors which make burn mortality in the geriatric patient is higher than the rest of the population with similar burns (Heimbach, 1987). As man ages, the skin atrophies with thinning of the refinement remains of the instrumentation, not-so-golden and clinical (Herndon,
standard
59
dermis and disappearance of skin appendages. The thin skin makes the diagnosis of burn depth difficult and the grafts on fat often do not survive (Deitch, 1985). The current plan is to excise to fascia full-thickness burns, which will not heal, from the periphery or by
contraction, and to graft them with meshed grafts taken at 0.008 inch from the back. Indeterminate burns are generally to be treated conservatively, as outpatients if possible
(Herndon, 2001).
II- Surgical Burn Wound Management

During the past thirty years, the treatment of deep burns by experienced burn surgeons has changed dramatically!
(Heimbach, 1987). Previously, nearly all large, deep burns were treated expectantly, eschar was permitted to slough spontaneously and the wounds were left to granulate before they were skin grafted. Split-thickness skin grafts were procured and
applied in many instances, not in sheets, but using a variety of free-hand techniques. Small patches or stamps of graft,
60
or even smaller punch grafts were applied to maximize epithelial perimeter and, hopefully to minimize graft loss from the heavily contaminated wounds. Weeks or,
frequently, months passed before wound closure could be achieved with these methods. Survival rates were very low especially if the burn surface area exceeded more than 40% of the body surface area. Perceptive surgeons were of course fully aware of the many disadvantages associated with such passive, expectant therapy (Haynes, 1987). During two World Wars, the idea of the prompt
excision of all devitalized tissue was elicited. This axiom clearly seemed applicable to burn treatment, but numerous practical clinical constraints prevented its general application (Haynes, 1987). In more extensive burns, effective were measures lacking, for dense
controlling
wound
microbial
growth
wound colonization almost occurred within the first few days and commonly. This was the source of invasive infection of normal tissue at the burn margins, systemic sepsis and death.
61
There were few safe, effective antibiotics and the importance of nutritional support was ill-appreciated (Gray et al., 1982). Then, effective topical therapy was introduced. This one advance, more than any other, re-awakened widespread interests by the surgical community in burn care (Jackson, 1969). Other important advances particularly in the area of critical potent care and medicine, relatively effective safe mechanical ventilation, power
antibiotics,
precision
dermatomes, mesh-expansion techniques of the grafts, and interested personnel in burn management contributed to the dramatic overall improvement in burn care (Herndon et al., 1987). Nowadays, aggressive, earlier and more frequent use of definitive surgical therapy for deep burns has become the norm in the western world. The question remain open, however, as to the timing, extent and depth to which surgical wound closure should be utilized in patients with burns so extensive that survival is problematic (Hart et al., 2003).
62
Surgical Methods of Burn Wound Closure

The excision of necrotic tissue and the covering of excised areas give local advantages, by eliminating lacticacid-rich necrotic tissues. These tissues influence collagen synthesis, by reducing the soluble mediators that act on the microcirculation and the impermeability of the blood vessels, increasing antibacterial defense from external contamination (Gray et al., 1982). General advantages are also obtained by removing toxic materials with protease activity, reducing hydro-electrolytic, protein and heat loss, decreasing the risk of sepsis and shortening the length of hospital stay (Heimbach, 1987). Excision therapy may also reduce protein catabolism, immunosuppression, and evaporative water losses. In some cases, early excision can improve hypertrophic scarring (Gray et al., 1982). Therefore, the ultimate solution of burn management is closure of the burn wound through surgical intervention. The alternative extent, burn-care and philosophies technique of differ the in the timing, procedure cosmosis by reducing
depth
surgical
(Herndon et al., 1986)
63
A- Timing of Burn Wound Excision

Timing of excision therapy is debatable. The patient should undergo excision therapy when surgical risks do not increase risk of mortality nor compromise anticipated
functional and cosmetic results. It may be immediate (within first 24 hours post-burn), early (within 3rd -5th PBD) or late (within 4th to 14th PBD) when the acute resuscitation period is well over (Burke et al., 1976). Experimentally, immediate complete excision of the
wound within 24 hours of injury prevented hypermetabolism and immune suppression in the post-burn period but
theoretically, it may aggravate the hemodynamic instability in the major burns. Clinically, in children with greater than 60% TBSA burns, immediate excision therapy resulted in improved survival (Desai et al., 1990). Many surgeons prefer early excision of the burn wound within 3rd and 5th PBD of the injury as soon as hemodynamic stability, physiological tolerance, reliable determination of
burn depth are ascertained and prior to bacterial colonization
of the wound (Heimbach, 1987). For most flame burns, excision therapy can
64
be
completed within 48 hours of admission unless delayed by serious inhalation injury, concomitant injuries, frailty from extremes of age, or pre-existing medical conditions.
However, in scald burns, delay of excision for one week reduces blood loss and areas of skin grafting (Rose et al., 1996). Partial-thickness flame burns that will spontaneously
heal within 14-21 days may be not excised and if treated conservatively, deep partial-thickness burns produce poorer scars, more complications and prolonged hospitalization.
Therefore, deep partial-thickness wounds are often treated similar to full-thickness injuries by surgical excision as soon as possible (Heimbach, 1987).
B- Extent of Burn Wound Excision

The extent of excision is determined by the stability of the patient, the burn size, the speed of the surgical team, the adequacy of anesthesia, the rate of blood loss and the availability of skin graft or its substitute (Engrav et al.,
1983). Blood losses pressure, topical are minimized and by use topical of of or
65
tourniquets, subcutaneous producing with
thrombin
epinephrine. hypertension
However, overdoses or paroxysmal
epinephrine do
tachycardia
occur
injudicious topical use, especially in children. In burns less than 40% TBSA, excision can be completed in a single procedure (Rose et al., 1996). However, the practice of early excision and grafting is ideally performed for 10 to 15% of TBSA per session, giving priority as always to the face and hands. All full-thickness burns can be excised first, so that deep dermal and
indeterminate depth wounds are addressed later, preventing excision of potentially viable tissue (Herndon, 2001).
C- Depth of Burn Wound Excision

For instance, the ideal depth of burn wound excision has not yet been established (Hart et al., 2003). However, theoretically, injured tissue following thermal trauma
presents a central area of necrosis surrounded by a stasis zone in which cell metabolism is slowed down, creating
favorable 1986). The bacterial colonization is responsible for conditions for wound sepsis (Herndon et
66
al.,
the
deepening of the wounds by lysis of the surrounding healthy tissue, the most feared and serious complication in the burn patient (Cope et al., 1947). Episodes of sepsis also lead to ischemic necrosis of subcutaneous fat subsequent to poor peripheral perfusion and microvascular stasis, that leads to late graft loss and these ischemic areas become portals for invasive wound sepsis (Herndon et al., 1984). Deeper excisions is also indicated for life-threatening invasive wound sepsis with fungi and yeast such as
aspergillus and candida and also for large areas with failed graft take (Engrav et al., 1983). It is hypothesized that the accumulation and
reabsorption of subeschar tissue fluid (STF) may increase the morbidity and mortality rates in severely burned patients. Therefore, the unburned tissues at the argin and the depth m of the burn may be affected and may exaggerate the systemic
inflammation (Chen et al., 2000).
67
D- Techniques of Burn Wound Excision

The aim of surgical treatment is to transform the avascular burn wound into a uniformly bleeding non-
contaminated surgical wound, at the same time avoiding the process of eschar demarcation due to granulation tissue, which is the cause of hypertrophic scars. The areas prepared in this way must then be covered to obtain definitive healing (Herndon et al., 1989). a- Primary closure of excised burns or donor sites Small deep burns on lax skin areas such as the buttock, female breast or the limbs or torso in the elderly can on occasion be excised and closed primarily, particularly if cosmosis is not an issue. In the elderly also, donor sites harvested from lax skin areas can at times primarily (Herndon et al., 1990). b- Amputation When a major portion of a limb, hand, or digit has been destroyed, or its potential for functional recovery is judged to be closed
68
be nil, amputation may be appropriate or even life saving, especially when doing so eliminates the deep burn and the stump can be closed primarily (Brown et al., 1994). c- Tangential excision The tangential or so-called laminar method of
sequentially shaving the eschar from the wound surface until a viable-tissue in plane 1970s. is It reached was is particularly first described for by the
Janzekovic
suitable
excision of deep dermal or shallow, superficial full-thickness burns. The procedure is usually performed using a hand-held blade equipped with a calibrated depth guard (0.010-0.025 inches), such as the small Goulian knife or the larger, more cumbersome Humby knife (Gray et al., 1982). For broad, relatively flat burn deep burns, a power dermatome such as the Brown, with its depth gauge set appropriately, is convenient for rapidly performing
tangential excision, but it is not suitable for use on the hand, foot or face (Heimbach, 1987). In experienced hands, tangential excision is rapid and permits salvage of viable reticular dermis, although
69
hemorrhage may be considerable. On limbs, a tourniquet probably helps to reduce blood loss, although it makes the differentiation of viable from non-viable tissue more
difficult. An acceptable wound bed is identified by active punctuate bleeding. By using this technique, a maximum of viable tissue is preserved and optimal functional and
cosmetic results are achieved (Herndon et al., 1989). d- Down to fascia excision (degloving or avulsion) Fascial excision is recommended if the subcutaneous fat is burned, and in selected large burns with more than 60% TBSA full-thickness who have high risks for infection, blood loss, or skin graft slough. Excision to this plane minimizes bleeding and provides a reliable, clean, vascular bed. Linear escharotomies placed 180 apart on a limb, and/or at the wound margins otherwise, which is best limited at the level of wrist or ankle (Herndon et al., 1989). Fascial excision results in damage to lymphatics and cutaneous nerves and loss of subcutaneous fat (Rode, 2001). The excised fat never regenerates and can give a spindly appearance to the areas excised while any increase in body
70
fat is deposited in the remaining beds of adipose tissue. Moon faces and thick necks can result (Heimbach, 1987).
E- Coverage of Excised Burn Wound

Preferably, coverage of excised burn wound is
performed with permanent skin autograft, but closure can also be achieved with skin allograft, other biological
dressings or skin substitutes. Without immediate coverage, desiccation or infection can increase tissue loss and negate the benefits of early excision (Heimbach, 1987). In burns less than 40% TBSA, wide availability of donor sites permits wound coverage with autograft. In burns more than 40% TBSA, when immediate permanent coverage is not possible after surgical excision, the area may be temporarily covered with cadaver skin, which may later be replaced by skin amniotic autograft. Alternatively, may be even used if for less this
satisfactory,
membrane
function (Herndon, 2001). The improved outcome only of burns optimal treatment operating will times be and further clear
when
technical criteria (extent, depth and technique of excision)
will have been established (Hart et al., 2003).
71
III- Impact of burn wound excision on the ImmuneInflammatory Response to Burn

Studies have shown that cell mediated immunity is suppressed markedly following thermal injury. Macrophages and the activation of an inflammatory cascade that includes interleukins IL-1, IL-6, tumor necrosis factor-alpha (TNF-) and PGE2 have been implicated as causative factors
(Schwacha et al., 2000). Burn wound excision and grafting restores cellular
immunity. It normalized TNF- production to sham levels, independent of when post-burn the procedure was other PGE2) and is
conducted. In contrast, the elevated
production of
inflammatory mediators (IL-1, IL-6, nitric oxide, post-burn grafting. was unaffected splenic by burn wound
excision proliferation
Moreover,
T-lymphocyte
also suppressed at 7th day post-burn and is not improved by burn wound excision and grafting (Schwacha et al., 2000). Therefore, excision and the grafting beneficial are effects to of be burn related wound to the
likely
72
normalization of macrophage TNF- production as well as the maintenance of skin barrier function (Schwacha et al., 2000). In addition, burn injury inhibits viral-specific cytotoxic T-lymphocyte augments activity. Early, complete wound excision Improved
cytotoxic
T-lymphocyte
function.
cytotoxic T-lymphocyte activity after burn may reduce the risk of infection, providing an immunologic rationale for expeditious wound excision (Hultman et al., 1997). Yamamoto et al. in 1996 stated that all B-cell functions are significantly suppressed by burn injury. Immediate
excision and grafting restores anti-PGPS IgM synthesis to normal, while nonspecific B-cell functions are not changed significantly. However, early excision and grafting fails to improve significantly any B-cell functions. Immediate but not early excision restores antibody synthesis to the bacterial cell wall antigen (PGPS). Immediate excision may therefore lead to a decrease in bacterial infection after burn injury. Huang et al., 1999 stated that eschar excision en masse at one operation is feasible and effective in preventing and
73
treating early post-burn organ dysfunction and multiorgan failure, mainly by alleviating systemic inflammatory
response syndrome and endothelial cell injury. Recent reports have suggested that very early excision (less than 24 hours post-burn) and primary closure of burn wounds might circumvent the immunosuppression, which
follows severe thermal trauma. The trauma of excision and grafting alone results in depression of cell-mediated
immunity. This deterioration is due to the systemic cytokine response, which is predominantly that of IL-6. It is also related to the release of TNF- (Herndon, 2001). The inhibit subeschar tissue fluid possesses the ability to
mitogen-induced
lymphocyte
proliferation
(MILP);
this suppressive nature is stable, persisting for prolonged periods. The gradual absorption of STF likely contributes to the serologic evidence of cell-mediated immune suppression documented in victims of severe thermal injury (Dyess et al., 1991). Therefore, excision may removing better than dead tissue, down-to-fascia because it
tangential
excision
removes large amounts of immunosuppressive
74
subeschar
tissue fluid (Schwacha et al., 2000). These data call into question the ability of very early excision and grafting to alter the immunosuppression, which follows severe thermal trauma (Carsin et al., 2002). Therefore, aggressive, earlier and more frequent use of definitive surgical therapy for deep burns has become the standard management of severe burns. However, the
outcome of burns treatment will be further improved when optimal operating times and clear technical criteria (extent, depth) will have been established (Hart et al., 2003).
Patients a nd Methods
Patients and Methods
75
A- Study Design: The study was concerned with the comparison between the technique of early burn wound excision (tangential excision and down-to-fascia excision) and the immunological profile changes, using alteration of interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-) levels as indicators.
B - Patient Population: This prospective comparative study was conducted in the Burn Unit of Ain Shams University Hospitals, in the period from January 2004 until March 2006. The study included 30 acutely burned adult patients admitted to the Burn Unit. All patients had a combination of superficial and deep dermal burns. Patients were chosen irrespective to age or sex (15 females, 15 males). The ages ranged between 20 - 50 years, (29.2 + 9.8 years). The burned surface area (BSA) ranged between 21 and 70% according to the Lund and Browder chart, (42.8 + 13.4 %). The deep areas, (i.e. deep dermal) were ranging between 15 and 25% of TBSA, (21.9 + 2.9%). Out of
76
the 30 patients, 25 had flame burns (83 %), 4 had scalds (13 %) and 1 had flash burns (3 %). The shortest hospital stay was 8 days, and the longest stay was 39 days, (20.9 + 10 days).
All patients were admitted within 8 hours from the injury. Patients with preexisting medical diseases (e.g. renal or liver impairment, diabetes mellitus, immunodeficiency syndrome as AIDS, leukemia, lymphoma, lymphocytopenia) were excluded. All patients received parental antibiotics (according to our burn unit protocol, it was ciprofloxacin) in the first week. Escharotomy was performed in the emergency room under general anesthesia for circumferential deep burns of the upper limb, lower limb, neck, and trunk. Careful haemostasis was performed to minimize blood loss to exclude immune response to stress and blood loss especially with tangential excision,
(Basill, 2004).
77
The 30 patients were divided into two groups according to the technique of the excision: First group (15 patients: 9 females and 6 males): This group included the patients who were candidates for tangential early burn wound excision and application of allograft. Second group (15 patients: 6 females and 9 males): This group included the patients who were candidates for down-to fascia early burn wound excision and application of allograft. C- Management Protocol: All patients were weighed in the admission room, prior to resuscitation and the following protocols were applied: I- Resuscitation Protocol: Resuscitation was performed using the Modified Parkland formula (Baxter, 1974):
Total amount of fluid = 3ml / kg body weight / % BSA.
78
The success of resuscitation was assessed by monitoring the pulse rate, blood pressure, urine output per hour, and central venous pressure. Throughout resuscitation, alterations could be made on the amount of fluids given, depending upon the hemodynamic status of the burned patient. In the first 24 hours, the only solution given was Ringers Lactate. In the following days, a combination of crystalloids and colloids was given, depending upon the hemodynamic status of the patient. Blood transfusion, fresh frozen plasma, and albumin, were given according to the laboratory data. II- Nutritional Protocol: The caloric and protein requirements were calculated according to the Curreri formula, 1974:
Total caloric requirements /day = 25 x weight in kg + 40 x % BSA.
Feeding was started within 24 hours from the time of admission, if the intestinal sounds were audible. The requirements were given either orally or by tube feedings. Multivitamins and trace elements were also supplemented on daily basis.
79
III- Post-resuscitation Management: 1- Wound management: Burn wounds were cleansed on admission using aqueous povidone iodine (10 %). Superficial and deep dermal burns were dressed with tulle grass. The change of dressing was performed on a daily or twice daily basis. In most cases, the dressing was performed in the hydrotherapy room, preferably under general anesthesia. 2- Surgical Procedures (early excision and grafting): - Timing of excision: Early excision and grafting was attempted once the resuscitation has been accomplished, and the patients general condition has become stable. The burn wound excision was started within first five PBDs and it was completed by the eleventh PBD. -Extent of excision: Extent of excised area ranged between 5% and 16% of TBSA per session, (9.8 + 2.6%). A tourniquet or subeschar adrenaline 1/200,000 infiltration was applied and the
80
electrosurgical unit was utilized especially in down to fascia excision, in order to minimize blood loss. In extensive burns, provided that the general condition of the patient permitted, multiple sessions of excision were performed. - Techniques and depth of excision: a- Tangential excision: The tangential method - sequentially shaving the eschar from the wound surface until a viable-tissue plane - was applied for first patient group. An acceptable wound bed was identified by active punctuate bleeding.
Fig. 5: case no. 7 in first group (a) preoperative, (b) bed after tangential excision, (c) after application of meshed autograft
81
The procedure was usually performed using a hand-held blade equipped with a calibrated depth guard (0.010-0.025 inches), such as skin graft knife. For broad, relatively flat burn deep burns, a power Brown dermatome, with its depth gauge set appropriately ( 0.15 inch), was convenient for rapidly performing tangential excision. b- Down to fascia excision: Excision to this plane minimized blood loss and provided a reliable, clean, vascular bed. Linear escharotomies were placed 180 apart on a limb, and/or at the wound margins otherwise, which was limited at the level of wrist or ankle. The procedure was usually performed using a scalpel or electrocautery unit (cutting or coagulation settings).
82
Fig. 6: case no. 12 in second group (a) preoperative, (b) bed after down-to-fascia excision, (c) excised eschar, (d) after application of amniotic membranes
- Coverage of Excised Burn Wound The total areas excised were covered by allograft (homograft or amniotic membranes) except case no.7 in first group and cases no. 1, 9 and 15 in second group, where autografts were used.
83
D - Monitoring of the patients: Organ dysfunction was based on the following set of clinical and/or laboratory criteria: a- Clinical Monitoring: i- Systemic monitoring: a) Central nervous system: Alteration in the level of consciousness and Changes in temperature (hyper or hypothermia). A low-grade fever, i.e. up to 38oC was considered to reflect a hypermetabolic status and it thus not indicative of sepsis. b) Cardiovascular system: Severe arrhythmias, Heart rate > 160 b.p.m and lasting for more than 48 hours or Decrease in blood pressure needing pharmacological support. c) Renal system: Anuria, oliguria or renal replacement therapy, Tachypnea, orthopnea and/or cyanosis. Assisted ventilation for more than 5 days, d) Respiratory system:
84
ii- Local (burn wound) monitoring:

Local signs suggestive of burn wound infection. Progression of partial-thickness to full-thickness injury. Change in wound color (focal areas of red, brown, or black discoloration). Green discoloration of the subcutaneous fat. Discoloration and edema of wound margins.
b- Laboratory Monitoring: All laboratory tests were assessed at one day before operation and 3rd, 7th and 14th days after escharectomy. Some of the tests were performed on a daily basis to ensure careful monitoring. i. Routine laboratory investigations:
1) Complete blood count (CBC) Hemoglobin concentration (Hb) (male: 13.8 ~ 17.2 gm/dL, female: 12.1 ~ 15.1 gm/dL). Hematocrite value (Hct) (male: 40.7~50.3%, female: 36.1~44.3 %). Total leucocytic count (TLC) (4000~10.000 cells/ mm3):
85
o Any change from the base line was considered to be alarming of sepsis. A decrease in count was to be taken more serious than leukocytosis in the evaluation of the severity of sepsis Platelet count (150,000 ~ 300,000 / mm3). 2) Coagulation profile Prothrombin time (PT) (11 ~ 13.5 seconds) and the partial thromboplastin time (PTT) (25 ~ 35 seconds). o These tests were performed to monitor the possible occurrence of disseminated intravascular coagulopathy. 3) Random blood sugar 4) Serum albumin (Alb) (3.5 ~ 5.5 gm/L). o Persistently low levels of serum albumin in spite of adequate replacement reflected a sepsis-mediated hypercatabolic state. 5) C-reactive protein (less than 0.6 mg/dL)
ii. Specific Laboratory investigations were done for detection of organ dysfunction (Yang et al., 1992):
Pulmonary function tests: arterial blood gases (ABG). o PaO2 < 50 mm Hg or SaO2 < 90%, Renal function tests: Serum creatinine (0.6 ~ 1 mg/d) and/or Blood urea nitrogen (BUN) (10 ~ 15 mg/100 ml).
86
Hepatic function tests: elevated SGOT (male: 8-46 u/L, female: 4-35 u/L), SGPT (male: 7-46 u/L, female: 4-35 u/L). Cardiac function tests: SGPT and creatine phosphokinase (CPK) (30 ~ 200 U/L).
iii. Assay of serum levels of cytokines: This was done in Ain Shams University Hospital Laboratories (Immunity Lab) Interleukin-6 (IL-6) assay: assay IL-6 was for in done the by an
immunoenzymometric measurement of
quantitative (EASIA)
human
serum
(Biosource Europe S.A., Belgium). The detection limits were 80 ~ 2024 pg/ml for IL-6 assay Tumor necrosis factor- alpha (TNF-) assay: was done by an immunoenzymometric assay for the quantitative measurement of human TNF- in serum (EASIA) (Biosource Europe S.A., Belgium). The detection limits were 50 ~ 1800 pg/ml for TNF- assay.
87
E - Statistical Methodology:
i. To compare the effect of technique of burn wound excision on the clinical outcome, these tests were performed: 1. Survival rate (SR) for each group, (percentage). 2. Average hospital stay (AHS) for each group, (Mean+SD). 3. Comparison between SR of both groups, (Fishers exact test). 4. Comparison between AHS of both groups, (Fishers exact test). ii. To compare the effect of technique of burn wound excision on the immunological profile changes, these tests were performed: According to Mann-Whitney test (independent samples), probability indices (-values) were calculated for preoperative, 3rd, 7th and 14th postoperative days IL-6 assay and TNF- assay levels of each group. (A -value less than 0.05 was considered statistically significant otherwise, it was insignificant). a- For first group: The comparison between: (Mean+SD) 1) Serum IL-6 assay levels in survivors and non-survivors.
88
2) Serum TNF- assay levels in survivors and non-survivors. b- For second group: The comparison between: (Mean+SD) 1) Serum IL-6 assay levels in survivors and non-survivors. 2) Serum TNF- assay levels in survivors and non-survivors. c- For both groups: The comparison between: (Mean+SD) 1) Serum IL-6 assay levels in survivors of both groups. 2) Serum IL-6 assay levels in non-survivors of both groups. 3) Serum TNF- assay levels in survivors of both groups. 4) Serum TNF- assay levels in non-survivors of both groups.
Results
Results A- Demography of Patients Population I- Patient Population of first group:
89
This group included 6 males and 9 females. Their ages ranged between 50-20 years, (29.9 + 11 years). The total burned surface area (BSA) ranged between 23-60% of total body surface area (TBSA), (42 + 9%). The deep areas, (i.e. deep dermal) were ranging between 15 and 25% of TBSA, (22 + 2%). Out of these 15 cases, 13 had flame burns (87 %) and 2 had scalds (13%).
Table (4): Patient Population of first group:
Patients 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15
* Female
Gender F* M** M M F F M F M F F M F F F
** Male
Age 50 y*** 20 y 27 y 26 y 35 y 20 y 50 y 26 y 20 y 20 y 20 y 44 y 37 y 20 y 34 y
% BSA**** 50 % 50 % 48 % 42 % 50 % 30 % 23 % 30 % 42 % 60 % 40 % 40 % 46 % 40 % 40 %
% Deep Burn
Type of burn
25 22 23 20 25 22 16 22 22 25 23 20 24 20 23
Flame Flame Flame Flame Flame Flame Scald Scald Flame Flame Flame Flame Flame Flame Flame
***years
**** Total Burned Surface Area
Results II- Patient Population of second group:
90
This group included 9 males and 6 females. Their ages ranged between 45-20 years, (28.5 + 7.3 years). The total burned surface area (BSA) ranged between 21-70% of total body surface area (TBSA), (44 + 16%). The deep dermal areas were ranging between 15 and 25% of TBSA area, (21.7 + 3.5%). Out of these 15 cases, 12 had flame burns (80 %) 2 had scalds (13 %) and 1 had flash burns (7 %).
Table (5): Patient Population of second group:
Patients 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15
* Female
Gender F* M** M M F M M M F M M M F F F
** Male
Age 30 y*** 30 y 44 y 20 y 29 y 30 y 45 y 27 y 22 y 26 y 23 y 21 y 25 y 25 y 30 y
% BSA**** 25 % 35 % 40 % 35 % 45 % 65 % 30 % 64 % 70 % 55 % 30 % 70 % 21 % 30 % 45 %
% Deep Burn
Type of burn
15 22 25 24 25 22 16 23 24 25 23 24 16 19 23
Scald Flame Flame Flame Flame Flame Flame Flame Flame Flame Flash Flame Scald Flame Flame
***years
**** Total Burned Surface Area
Results B- Analysis of Clinical Outcomes I- Clinical outcomes of first group:
91
Hospital stay ranged between 20-39 days in survivors, (31.8 + 7 days) and between 8-16 days in non-survivor, (12.5 + 3 days). The average hospital stay (AHS) for all cases was 24.1 + 11.4 days. The mortalities were 6 of 15 (40%) (2 cases with single organ failure and 4 cases with multiple organs failure) (Table 6). Timing of tangential excision was within first five postburn days. Extent of excised area ranged between 5% and 16% of TBSA per session, (9.8 + 2.6%). All the excised areas were covered by allograft except in one case, where autograft was used.
Results
Table (6): Clinical outcomes of first group:
Patient
92
Time of excision session
extent of excision per session
Hospital Stay
Organ Dysfunction
Outcome
1 2nd PBD* 2 4th,7th PBDs 3 2nd PBD 4 3rd,6th PBDs 5 3rd,6th,11th PBDs 6 4th,7th PBDs 7 5th PBD 8 4th,6th,11th PBDs 9 4th,7th PBDs 10 3rd, 7th PBDs 11 2nd,6th PBDs 12 4th,6th PBDs 13 3rd PBD 14 2nd PBD 15 3rd PBD * = Post-burn day S = Survivor N = Non-survivor A = autograft
10% 10% , 12% 10% 10%,10% 10%,10%,5% 10%,12% 16% A 10%,5%,7% 10%, 12% 10%,10%,5% 10%,13% 10%,10% 10% 5% 14%
8 days 39 days 12 days 35 days 38 days 14 days 20 days 35 days 32 days 39 days 27 days 22 days 15 days 10 days 16 days
Multiple Single Multiple Multiple Single Multiple
N S N S S N S S S S S S N N N
Results II- Clinical outcomes of second group:
93
Hospital stay ranged between 17-39 days in survivors, (26 + 8.1 days) and between 9-18 days in non-survivors, (12.8 + 3.2 days). The average hospital stay (AHS) for all cases was 17.2 + 8.2 days. The mortalities were 10 of 15 (66.7 %) (3 cases with single organ failure and 7 cases with multiple organs failure) (Table 7). Timing of down-to-fascia excision was within first five post-burn days. Extent of excised area ranged between 5% and 14% of TBSA per session, (9.3 + 2.7%). All the excised areas were covered by allograft except in three cases, where autograft was used.
Results
Table (7): Clinical outcomes of second group:
Patient
94
Time of excision session
extent of excision per session
Hospital Stay
Organ Dysfunction
Outcome
1 2nd, 5th PBD* 2 4th PBDs 3 2nd PBD 4 3rd PBDs 5 3rd PBDs 6 4th PBDs 7 5th PBD 8 4th PBD 9 4th,6th,11th PBDs 10 3rd PBDs 11 2nd,6th PBDs 12 4th,6th PBDs 13 3rd, 5th PBD 14 2nd,6th PBD 15 3rd,5th PBD * = Post-burn day S = Survivor N = Non-survivor A = autograft
10%,5% A 8% 10% 12% 11% 10% 6% 11% A 7% , 10%,6% 12% 10%,13% 11%,6% 10%,6% 14%,5% 9% A, 14%
25 days 17 days 12 days 11 days 9 days 14 days 14 days 14 days 18 days 11 days 27 days 8 days 17 days 22 days 39 days
Multiple Single Multiple Multiple Multiple Single Single Multiple Multiple Multiple -
S N N N N N N N N N S N S S S
Results III- Analysis of the clinical outcomes in both groups:
95
The Average Hospital stay (AHS) of all cases of first group was 24.1 + 11.4 days and it was 17.2 + 8.2 days for all cases of second group. By comparing the AHS, it was significantly higher in first group than in second group (-value = 0.0674). The AHS for survivors in first group was 31.8 + 7 days and it was 26 + 8.1 days in second group. Wherever, The AHS for nonsurvivors in first group was 12.5 + 3 days and it was 12.8 + 3.2 days in second group. The AHS for survivors of both groups were not significantly different (-value = 0.1845), as well as, the AHS for nonsurvivors in both groups were not significantly different (-value = 0.8554). (Chart 1).
Results
96
Days
40 35 30 25 20 15 10 5 0 Group I
Survivors Non-survivor
31.8 26 12.5 12.8

Group II
Error bars represent SD
Survivors AHS Non-Survivors AHS
Group I Group II -value
31.8 + 7 days 26 + 8.1 days 0.1845
12.5 + 3 days 12.8 + 3.2 days 0.8554
Chart (1): Comparison between Average Hospital stay (AHS) of survivors and non-survivors in both groups.
The first group mortalities were 6 cases (2 cases with single organ failure and 4 cases with multiple organs failure). The second group mortalities were 10 cases (3 cases with single organ failure and 7 cases with multiple organs failure).
Results
97
The incidence of MOD in non-survivors in first group (66.7%) was not statistically significant than that of second group (70%) (-value = 0.6761). (Chart 2). By comparing the Survival Rates (SR) of both groups, the SR of first group (60 %) was not statistically significant than the SR of second group (33.3%) (-value = 0.2714).
15
Number of cases
12 9 6 3 0 Group I 9 4 2
Survivors Single OD (non-survivors) Multiple OD (non-survivors)
7 5 3 Group II
Survivors (Non-Survivors) Organ Dysfunction Single Multiple
Group I (15 cases) 9 cases Group II (15 cases) 5 cases -value = 0.2714
2 cases 3 cases
4 cases 7 cases
-value = 0.6761
Chart (2): Comparison between clinical outcomes of both groups
Results C- Analysis of Laboratory Investigations I- Laboratory results of first group:
98
All laboratory tests were assessed at one day before operation and 3rd, 7th and 14th days after tangential escharectomy. All patients had high levels of all blood elements in the preoperative samples (with first five PBDs) (due to
haemoconcentration) that began to resolve after proper fluid therapy. Total leucocytic count was elevated in all patients without clinical manifestations of infection. No patient
experienced leucopenia. Thrombocytosis occurred in 4 patients who had MOD. Elevated liver transaminases were noticed in 5 patients (4 patients had MOD and 1 patient had single organ dysfunction). C-reactive protein and creatine phosphokinase (CPK) values were high in 4 patients who had MOD. Albumin, fasting blood sugar, serum creatinine and blood urea/nitrogen (BUN) levels were within the normal values throughout the study. Assessment of serum levels of cytokines interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-) were done by EASIA
Results
99
(Biosource Europe S.A., Belgium). Detection limits were of 80~2024 pg/ml for IL-6 assay and 50~1800 pg/ml for TNF- assay. IL-6 assay results are listed in table (8) and TNF- assay results are listed in table (9).
Table (8): IL-6 assay results in pg/ml of first group:
Patients Preoperative 3rd Postoperative 7th Postoperative 14th Postoperative Day day day day 1** 700 650 2024 2* 200 500 2024 1000 3** 570 200 850 4* 500 2024 700 450 5* 600 1000 1500 1000 6** 500 450 1300 2024 7* 600 650 200 150 8* 400 2024 430 400 9* 600 1000 700 450 10* 800 600 510 200 11* 700 510 1000 300 12* 600 450 1000 100 13** 1000 2024 1700 2024 14** 1250 650 2024 15** 620 700 2024 * Survivor ** Non-survivor
Results
Table (9): TNF- assay results in pg/ml of first group:
Patients
100
Preoperative 3 Postoperative 7 Postoperative 14 Postoperative Day day day day 1400 700 1600 500 700 1200 300 240 320 300 400 200 1400 1800 1800 200 100 200 1800 150 160 220 130 100 80 1200 -
rd
th
th
1** 490 400 2* 300 450 3** 340 300 4* 350 500 5* 480 550 6** 350 400 7* 360 500 8* 240 300 9* 480 400 10* 640 500 11* 420 400 12* 360 450 13** 700 900 14** 1000 1200 15** 500 700 * Survivor ** Non-survivor
Results
101
Chart 3; illustrates serum IL-6 assay levels of survivors and non-survivors of first group at preoperative, 3rd, 7th and 14th postoperative days. Analysis of data revealed significant higher levels of IL-6 assay of non-survivors compared to survivors at 7th (896 + 568 pg/ml for survivors, 1654 + 486 pg/ml for non survivors) and 14th (450 + 336 pg/ml for survivors, 2024 + 0 pg/ml for non survivors) postoperative days (-value = 0.0360, 0.0004 respectively).
2000
Survivors Non-survivors
IL-6 assay (pg/ml)
1600 1200 800 400 0
Preop.
3rd P.O.
7th P.O.
14th P.O.
Post-operative Days Error bars represent range Highest value Lowest value
Preop. day(1)
800 200
3rd PO day(2)
2024 450
7th PO day(3)
2024 200
14th PO day(4)
1000 100
Survivors
Mean + SD
Highest value Lowest value
555+147
1250 500
973+629
2024 200
896+568
2024 850
450+336
2024 2024
Nonsurvivors
Mean + SD
773+291
779+637
1654+486
2024+0
(1) -value = 0.2238 (3) -value = 0.0360
(2) -value = 0.5287 (4) -value = 0.0004
Results
102
Chart (3): IL-6 assay levels of survivors and non-survivors of first group at preoperative, 3rd, 7th and 14th postoperative (PO) days. Chart 4; illustrates serum TNF- assay levels of survivors and non-survivors of first group at preoperative, 3 rd, 7th and 14th postoperative days. Analysis of data revealed significant higher levels of TNF- assay of non-survivors compared to survivors at 7th (406 + 187 pg/ml for survivors, 1533+242pg/ml for non survivors) and 14th (148+50pg/ml for survivors, 1500+424pg/ml for non survivors) postoperative days (-value = 0.0004, 0.0004 respectively).
TNF-alpha assay (pg/ml)
Survivors
1600
Non-survivors
1200 800
400 0
Preop.
3rd P.O. 7th P.O. Postoperative days
14th P.O.
Error bars represent range Highest value Lowest value
Preop. day(1)
640 240
3rd PO day(2)
550 300
7th PO day(3)
700 200
14th PO day(4)
220 80
Survivors
Mean + SD
403+118
1000 340
450+75
1200 300
406+187
1800 1200
148+50
1800 1200
Nonsurvivors
Mean + SD
563+250
650+350
1533+242 1500+424
(1) -value = 0.2238 (3) -value = 0.0004
(2) -value = 0.6889 (4) -value = 0.0004
Results
103
Chart (4): TNF- assay levels of survivors and non-survivors of first group at preoperative, 3rd, 7th and 14th postoperative (PO) days.
II- Laboratory results of second group: All laboratory tests were assessed at one day before operation and 3rd, 7th and 14th days after down-to-fascia burn wound excision. All patients had high levels of all blood elements in the preoperative samples (with first five PBDs) (due to
haemoconcentration) that began to resolve after proper fluid therapy. Total leucocytic count was elevated in all patients without documented source of infection. No patient experienced leucopenia. Thrombocytosis occurred in 7 patients who had MOD. Elevated liver transaminases were noticed in 8 patients (7 patients had MOD and 1 patient had single organ dysfunction). C-reactive protein and creatine phosphokinase (CPK) values were high in 7 patients who had MOD.
Results
104
Albumin, fasting blood sugar, creatinine phosphokinase (CPK), serum creatinine and blood urea/nitrogen (BUN) levels were within the normal values throughout the study. Assessment of serum levels of cytokines interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-) were done by EASIA (Biosource Europe S.A., Belgium). Detection limits were of 80~2024 pg/ml for IL-6 assay and 50~1800 pg/ml for TNF- assay. IL-6 assay results are listed in table (10) and TNF- assay results are listed in table (11).
Table (10): IL-6 assay results in pg/ml of second group:
Patients 1* 2** 3** 4** 5** 6** 7** 8** 9** 10** 11* 12** 13* 14* Preoperative 3rd Postoperative 7th Postoperative 14th Postoperative Day day day day 300 600 1200 1000 500 800 500 300 700 1250 570 620 600 570 510 600 2024 1100 510 350 200 500 600 2024 510 1300 2024 700 1000 350 1900 700 800 800 250 2024 900 2024 1000 2024 400 400 510 1450 2024 2024 510 300 2024 800 200 100
Results
15* 650 * Survivor ** Non-survivor 350 600 200
105
Table (11): TNF- assay results in pg/ml of second group:

Patients Preoperative 3rd Postoperative 7th Postoperative 14th Postoperative Day day day day 460 400 400 500 520 720 500 500 504 600 400 500 600 350 300 360 900 1200 800 900 1200 1600 1800 1600 900 640 1600 260 380 370 250 1400 1400 1200 1400 1600 1800 100 200 200 100
1* 180 2** 360 3** 840 4** 600 5** 400 6** 560 7** 300 8** 180 9** 420 10** 1000 11* 400 12** 370 13* 360 14* 340 15* 520 * Survivor ** Non-survivor
Results
106
Chart 5; illustrates serum IL-6 assay levels of survivors and nonsurvivors of second group at preoperative, 3 rd, 7th and 14th postoperative days. Analysis of data revealed significant higher levels of IL-6 assay of non-survivors compared to survivors at 7th (680+303 pg/ml for survivors, 1177+730 pg/ml for non survivors) and 14th (362+289 pg/ml for survivors, 1643+796 pg/ml for non survivors) postoperative days (-value = 0.0710, 0.0027 respectively).
2000
Survivors Non-survivors
IL-6 assay (pg/ml)
1600 1200 800 400 0 Preop. 3rd P.O. 7th P.O. Postoperative days 14th P.O.
Preop. day(1)
650 300
3rd PO day(2)
2024 350
7th PO day(3)
1000 400
14th PO day(4)
800 100
Survivors
Mean + SD
538+137
1250 300
819+685
2024 200
680+303
2024 250
362+289
2024 300
Nonsurvivors
Mean + SD
747+313
921+667
1177+730
1643+796
(1) -value = 0.2065 (3) -value = 0.0710
(2) -value = 0.7679 (4) -value = 0.0027
Chart (5): IL-6 assay levels of survivors and non-survivors of second group at preoperative, 3rd, 7th and 14th postoperative (PO) days.
Results
107
Chart 6; illustrates serum TNF- assay levels of survivors and non-survivors of second group at preoperative, 3rd, 7th and 14th postoperative days. Analysis of data revealed significant higher levels of TNF- assay of non-survivors compared to survivors at 7th (402 + 141 pg/ml for survivors, 1250 + 371pg/ml for non survivors) and 14th (170 + 67 pg/ml for survivors, 1466 + 206 pg/ml for non survivors) postoperative days (-value < 0.0001, < 0.0001 respectively).
Survivors
1600
Non-survivors
1200
800
400
0 Preop. 3rd P.O. 7th P.O. Postoperative days 14th P.O.
Preop. day(1)
520 180
3rd PO day(2)
600 300
7th PO day(3)
640 260
14th PO day(4)
250 100
Survivors
Mean + SD
360+122
1000 180
422+115
720 400
402+141
1800 800
170+67
1800 1200
Nonsurvivors
Mean + SD
503+252
514+92
1250+371
1466+206
(1) -value = 0.2065 (3) -value < 0.0001
(2) -value = 0.7753 (4) -value < 0.0001
Chart (6): TNF- assay levels of survivors and non-survivors of second group at preoperative, 3rd, 7th and 14th postoperative (PO) days.
Results
108
III- Comparison between IL-6 assay levels of survivors in both groups: Chart 7; illustrates serum IL-6 assay levels of survivors in both groups at preoperative, 3rd, 7th and 14th postoperative days. Analysis of data revealed no significant variations in levels of IL-6 assay of first group survivors compared to second group survivors (-value > 0.4376).
2000
Group I Group II
IL-6 assay (pg/ml)
1600 1200 800 400 0
Preop.
14th P.O.
Preop. day(1)
800 200
3rd PO day(2)
2024 450
7th PO day(3)
2024 200
14th PO day(4)
1000 100
Group I
Mean + SD
555+147
650 300
973+629
2024 350
896+568
1000 400
450+336
800 100
Group II
Mean + SD
538+137
819+685
680+303
362+289
(1) -value = 0.6064 (3) -value = 0.4376
(2) -value = 0.5185 (4) -value = 0.6064
Chart (7): IL-6 assay levels of survivors in both groups at preoperative, 3rd, 7th and 14th postoperative (PO) days.
Results
109
IV- Comparison between TNF- assay levels of survivors in both groups: Chart 8; illustrates serum TNF- assay levels of survivors in both groups at preoperative, 3rd, 7th and 14th postoperative days. Analysis of data revealed no significant variations in levels of TNF- assay of first group survivors compared to second group survivors (-value > 0.4376).
Group I
1600
Group II
1200
800
400
Preop.
14th P.O.
Error bars represent range
Preop. day(1)
640 240
3rd PO day(2)
550 300
7th PO day(3)
700 200
14th PO day(4)
220 80
Group I
Mean + SD
403+118
520 180
450+75
600 300
406+187
640 260
148+50
250 100
Group II
Mean + SD
360+122
422+115
402+141
170+67
(1) -value = 0.6064 (3) -value = 0.8981
(2) -value = 0.4376 (4) -value = 0.6064
Chart (8): TNF- assay levels of survivors in both groups at preoperative, 3rd, 7th and 14th postoperative (PO) days.
Results
110
V- Comparison between IL-6 assay levels of non-survivors in both groups: Chart 9; illustrates serum IL-6 assay levels of non-survivors in both groups at preoperative, 3rd, 7th and 14th postoperative days. Analysis of data revealed no significant variations in levels of IL-6 assay of first group non-survivors compared to second group survivors (-value > 0.4198).
2000
Group I Group II
IL-6 assay (pg/ml)
1600 1200 800 400 0
Preop.
14th P.O.
Preop. day(1)
1250 500
3rd PO day(2)
2024 200
7th PO day(3)
2024 850
14th PO day(4)
2024 2024
Group I
Mean + SD
773+219
1250 300
779+637
2024 200
1654+486
2024 250
2024+0
2024 300
Group II
Mean + SD
747+313
921+667
1177+730
1643+796
(1) -value = 0.8749 (3) -value = 0.4198
(2) -value = 0.8749 (4) -value = 0.5676
Chart (9): IL-6 assay levels of non-survivors in both groups at preoperative, 3rd, 7th and 14th postoperative (PO) days.
Results
111
VI- Comparison between TNF- assay levels of non-survivors in both groups: Chart 10; illustrates serum TNF- assay levels of non-survivors in both groups at preoperative, 3rd, 7th and 14th postoperative days. Analysis of data revealed no significant variations in levels of TNF- assay of first group non-survivors compared to second group survivors (-value > 0.5806).
1600 1200 800
Group I Group II
400 0
Preop.
3rd P.O. 7th P.O. 14th P.O. Postoperative days
Preop. day(1)
1000 340
3rd PO day(2)
1200 300
7th PO day(3)
1800 1200
14th PO day(4)
1800 1200
Group I
Mean + SD
563+250
1000 180
650+350
720 400
1533+242
1800 800
1500+424
1800 1200
Group II
Mean + SD
503+252
514+92
1250+371
1466+206
(1) -value = 0.7925 (3) -value = 0.5806
(2) -value = 0.9578 (4) -value = 0.6132
Chart (10): TNF- assay levels of non-survivors in both groups at preoperative, 3rd, 7th and 14th postoperative (PO) days.
Discussion
Discussion
112
The Second World War was an important time stone for management of severe burns. The idea of down-to-fascia excision technique was elicited to
remove all devitalized tissues. In the early eighties, with the advances in immunological studies of burn, another technique that is tangential excision was introduced and became the mainstay method of escharectomy. Since this time, the technique and depth of burn wound excision has been always controversial especially in severe burn. However, the early escharectomy was applied, by any of these two techniques, to restore cellular and humoral immunity and modulates the stress response in burned patients that leads to reduce the incidence of SIRS and MOF. Many previous studies compared the merits and drawbacks of the tangential excision versus the down-tofascia excision in different points of view. Since, the tangential excision technique has shown the advantages of preserving subdermal fat over bony prominences for
Discussion
113
cosmetic reasons, as well as maintaining the integrity of lymphatic drainage and cutaneous nerves. On the other hand, the grafting onto subdermal fat and exaggerated blood loss are eminent disadvantages of this technique (Herndon et al., 1999). On the contrary, down-to-fascia excision assures a viable bed for skin grafting with minimal blood loss, but it destroys the lymphatics, cutaneous nerves and
subcutaneous fat that compromise the cosmetic outcome (Rode, 2001). Recently, Chen and colleagues in 2000 suggested that the accumulation and reabsorption of subeschar
tissue fluid (STF) likely contributes to the serologic evidence of cell mediated immunological abnormalities documented in severe thermal injury. They stated that down-to-fascia excision may be better than tangential excision because it removes larger amount of subeschar tissue fluid.
Discussion
114
After
revising literature,
no
study compared
the
impact of each technique on alteration of immunological profile of severely burned patients. Therefore, this work aimed at comparing the effect of these two different techniques of early burn wound excision on alteration of immunological profile of severely burned patients. Moreover, it could interpret the results to suggest the impact of elimination of larger amount of subeschar tissue fluids (STF) on altering the immunological profile in burned patients. This would enable the burn surgeons to decide the proper technique (i.e. proper depth) of early burn wound excision that would decrease the
incidence of SIRS in order to decrease the morbidity, improve clinical outcome and increase survival rate in extensively burned patients. For these aims, the study was designed to divide 30 acutely burned patients into two groups (each group included 15 patients) according to technique of escharectomy. We unified the patients inclusion criteria
Discussion
115
for both groups to make the technique of excision as the only variable that would have a possible effect on the changes in serum levels of the most important prognostic factors of SIRS, which are TNF- and IL-6 cytokines. Statistical analysis of both groups revealed no
significant difference in all parameters of the study. Il-6 assay showed no difference between survivors of both groups (-value > 0.4376). Moreover, there was also no significant difference in non-survivors of both groups (-value > 0.4198). In addition, neither there was insignificant variation in levels of TNF- assay of survivors in both groups nor non-survivors in both groups at all time points (-value > 0.4376, -value > 0.5806 respectively). These tangential findings excision conclude and the equal effect of
down-to-fascia
excision
techniques on altering of immune response of severely burned patients.
Discussion
116
Therefore, in deep dermal burns, there does not appear to be any advantage to routinely performing a down-to-fascia excision, since the immunological profile changes are similar in both techniques of excision. Conversely, many previous studies aimed at the
evaluation of the role of subeschar tissue fluids (STF) in development of multiorgan failure (MOF). Ferrara in 1988 and Dyess in 1991 suggested that: STF may act as both an immunologic barrier to microbial clearance in otherwise viable subcutaneous tissue and a reservoir for systemically reabsorbed immuno-suppressive factors.
Similar findings were encountered in the study of Chen and colleagues in 2000 who suggested that STF might be one of the inducing factors involved in the genesis of SIRS and the development of MOF in the early postburn stage. However, Rong and colleagues in 2003 stated that the cells and large molecules seems to be more difficult to enter subeschar tissue fluid compared with small
Discussion
117
molecules and no marked local inflammatory response occurs in subeschar tissue fluid during early stage of severe burn, and subeschar tissue fluid has no lethal effect. Consequently, the results of this study disagreed with the concept of down-to-fascia excision may better than tangential excision because it removes larger amount of subeschar tissue fluid. Additionally, it could state that subeschar tissue fluid removal may has no role in suppression of immune response in thermal induced SIRS and MOF. Moreover, there has been always a controversy in the correlation between the IL-6 and TNF- serum levels and mortality rate in severely burned patients. Munster in 1996 studied the correlation between serum levels of IL-6 and MOD in severely burned patients. He
suggested that low serum level of IL-6 and high serum level of TNF- might be considered the most important
Discussion
118
poor
prognostic
factors
related
to
SIRS
and
MOF
following thermal injury. A similar conclusion was reached out in the study of Deveci and colleagues in 2000 who stated that IL-6 inhibits the severity of the inflammatory response in the early period of thermal injury by decreasing serum
levels of TNF-. However, Hack and co-workers in 1989, and Drost and colleagues in 1993 found a positive correlation between high IL-6 and TNF- serum levels and
incidence of multiorgan failure (MOF) and consequently mortality rates in severe thermal injuries. Conversely, Rodriguez and colleagues in 1993
reported no association between mortality and IL-6. By analysis of mean values of IL-6 assay and TNF assay in another point of view, comparisons between survivors and non-survivors of the same group were established. The results showed significantly higher levels of IL-6 assay and TNF- assay of non-survivors
Discussion
119
compared to survivors in both group (-value <0.0710 and <0.0004 respectively) especially at 7 th and 14th
postoperative days. The authors results who confirmed the that findings there of previous significant
emphasized
were
higher levels of TNF- assay of non-survivors compared to survivors. However, it was found significant higher levels of IL-6 assay also in non-survivors compared to survivors, which disagreed with the suggestion of
previous authors. An in-depth analysis of the results, the curves of mean values of IL-6 assay and TNF- assay in survivors in both groups were compared. It was found that burn wound excision normalized TNF- serum levels, in both groups. In contrast, the elevated serum levels of IL-6 were decreased by burn wound excision, but they did not reach the normal levels. These results were similar to the findings of
Schwacha and colleagues in 2000, who studied the
Discussion
120
impact of wound excision on the immunological profile. Their study revealed TNF- was that escharectomy and of grafting which elevated production, In independent contrast, the
normalized procedure
applied.
production of IL-6 post-burn was decreased by burn wound excision and grafting but did not reach the normal levels. These results could be explained by the fact that the half-life of some cytokines is different due to difference in their renal clearance. Therefore, decline in levels of cytokines in severe burn after escharectomy is not the same. Concerning the clinical outcomes, three parameters were used to explore the possible impact of technique of burn wound excision on the clinical outcome in severely burned patients, namely, the Average Hospital Stay
(AHS), the incidence of multiorgan dysfunction (MOD) in non-survivors of both groups and the Survival rate (SR).
Discussion
121
The
correlation
between
the
technique
of
burn
wound excision and the Average Hospital stay (AHS) revealed that the AHS of patients of group of tangential excision was significantly higher than group of down-tofascia excision (-value = 0.0674). This could be explained by that in tangential
excision that the exaggerated blood loss needs more time to resuscitate the operated patients even with careful haemostasis before undergoing another session of
excision. Additionally, the accuracy of escharectomy is surgeons dependant factor so the bed after excision may be unhealthy and need further debridement or the
grafting onto subdermal fat results in a lower success. Moreover, infection rates are higher in tangential
excisions than those of down-to-fascia excisions due to abundant vascularity of the bed in the second type of escharectomy. Taking the depth of excision into consideration, the incidence of multiorgan dysfunction (MOD) in non-
Discussion
122
survivors in group of tangential excision (66.7%) was statistically insignificant compared to that of group of down-to-fascia excision (70%) (-value = 0.6761). Thus, it can clearly conclude that incidence MOD is not related to depth of burn wound excision. The Survival Rates (SR) of group of tangential excision was not statistically significant compared to the SR of group of down-to-fascia excision (-value =
0.2714). Many studies elaborated on the correlation between the results of burn wound excision techniques on one hand and the the local issue outcomes of the on the of other the hand.
However,
value
different
techniques of burn wound excision in improving the general condition of severely burned patients, and hence in the proper prediction of the average hospital stay and survival rate has not been investigated by any burn center. Therefore, to explain these results, further
detailed investigations are needed.
Discussion
123
On light of all these results, it was concluded that both methods of excision, namely down-to-fascia and tangential excision, affect immunological profile
similarly, STF is an inflammatory response to burn and it may has no immunosuppressive effect, escharectomy decline serum levels of both IL-6 and TNF- and normalizes TNF- serum levels and
escharectomy
decreases serum levels of IL-6 but not to normal levels. Additionally, clinical outcomes of both techniques of excision survival are similar, since, incidence application of of MOD and
rates.
However,
tangential
excision technique needs more hospital stay. These conclusions recommend down-to-fascia
excision for full-thickness injury and tangential excision through or below the dermis for deep dermal injury. However, it is recommended that following initial
evaluation, wound excision could be carried beyond the deepest level of injured tissue, according to surgeons preference, facilities and location of burn.
Discussion
124
Additionally,
this
piece
of
work
recommend
changing protocol of Ain Shams University Burn Unit in management of deep dermal burn to start with
escharectomy and avoid conservative management. Finally, further study to backbone this study results are needed.
Summary a nd Conclusion
Summary and Conclusion
125
This prospective comparative study was conducted in the Burn Unit of Ain Shams University Hospitals, in the period from January 2004 until March 2006. It aimed to compare the effect of two different techniques of early burn wound excision (tangential excision and down-tofascia excision) on alteration of interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-) levels as indicators for the immunological profile alterations. The study included 30 acutely burned adult patients. All patients were chosen irrespective to sex. The ages ranged between 50 - 20 years, (29.2 + 9.8 years). The burned surface area (BSA) ranged between 21 and 70%, (42.8 + 13.4 %). The deep areas, (i.e. deep dermal) were ranging between 15 and 25% of TBSA, (21.9 + 2.9%). Out of the 30 patients, 25 had flame burns (83 %), 4 had scalds (13 %) and 1 had flash burns (3 %). The 30 patients were divided prospectively into two groups according to the technique of the excision: First group (15 patients), included the patients who were candidates for tangential early burn wound excision. This group
126
included 6 males and 9 females. The average hospital stay (AHS) was 24.1 + 11.4 days. The survival rate was 60%; (the mortalities were 6 of 15 [2 cases with single organ failure and 4 cases with multiple organs failure]). Second group (15 patients), included the patients who were candidates for down-to fascia early burn wound excision. This group included 9 males and 6 females. The average hospital stay (AHS) was 17.2 + 8.2 days. The survival rate was 33.3%; (the mortalities were 10 of 15 [3 cases with single organ failure and 7 cases with multiple organs failure]). For each patient in both groups the Resuscitation Protocols were applied according to Ain Shams University Burn Unit protocols. Burn wound excision was started within 5th post-burn days and was completed within 11th PBDs. Extent of excised area ranged between 5% and 16% of TBSA per session. All the excised areas were covered by allograft except in three cases, where autograft was used. Monitoring of the patients was done by Clinical Monitoring (Systemic, Local) and Laboratory Monitoring; which included routine laboratory investigations (e.g. CBC, PT, PTT, Random
127
blood sugar, Alb. and C-RP), Laboratory criteria of multiorgan dysfunction (MOD) (e.g. ABG, Renal function [serum creatinine and blood urea/nitrogen (BUN)], Hepatic function [SGOT, SGPT] and cardiac functions as SGOT) and assay of serum levels of cytokines IL-6 and TNF- by EASIA. By comparing the AHS, it was significantly higher in first group than in second group (-value = 0.0674). The incidence of MOD in non-survivors in first group (66.7%) was not statistically significant than that of second group (70%) (-value = 0.6761). By comparing the Survival Rates (SR) of both groups, the SR of first group (60 %) was not statistically significant than the SR of second group (33.3%) (-value = 0.2714). As regards, IL-6 assay for comparison between survivors and non-survivors of the same group, the results showed significantly higher levels of IL-6 assay of non-survivors compared to survivors in both group at 7th and 14th postoperative days (for 1st group, -value = 0.0360 and 0.0004 respectively, and for 2nd group, -value = 0.0710 and 0.0027 respectively). In addition, TNF- assay for comparison between survivors and non-survivors of the same group emphasized that there were
128
significant higher levels of TNF- assay of non-survivors compared to survivors in both group at 7th and 14th postoperative days, (for 1st group, -value = 0.0004, 0.0004 respectively, and for 2nd group, -value < 0.0001, < 0.0001 respectively). On the other hand, by comparing the curves of mean values of IL-6 assay and TNF- assay in survivors in both groups, it was found that burn wound excision normalized TNF- serum levels, in both groups. In contrast, the elevated serum levels of IL-6 were decreased by burn wound excision, but they did not reach the normal levels. Concerning impact of depth of excision on serum levels of IL6 and TNF-, analysis of data revealed insignificant variation in levels of IL-6 assay of neither survivors in both groups nor nonsurvivors in both groups at all time points (-value > 0.4376, value > 0.4198 respectively). In addition, neither there was insignificant variation in levels of TNF- assay of survivors in both groups nor non-survivors in both groups at all time points (-value > 0.4376, -value > 0.5806 respectively).
129
Consequently, these findings disagreed with the concept of fascial excision may better than tangential excision because it removes large amounts of subeschar tissue fluid. On light of these results, it is concluded that in deep dermal burns, there does not appear to be any advantage to routinely performing a fascial excision, since the immunological profile changes and clinical outcomes are similar in tangential and fascial excision. These finding clearly indicated that following initial evaluation, wound excision is carried beyond the deepest level of injured tissue, where fascial excision is used for full-thickness injury and tangential excision is used in or below the dermis for deep dermal injury.
References
References
130
Allgower M, Schoenenberger GA and Sparkes BG (1995): Burning the Largest Immune Organ. Burns, vol. 21 (Suppl. 1). Anselmo VJ, Zawacki BE (1973): Infrared Photography as a Diagnostic Tool for the Burn Ward. Proc. Soc. Photo-optical Instr. Engrs., 8:181. Arturson G (1990): Possible Involvement of Arachidonic Acid Metabolites in Thermal Trauma. In: Dolecek R, Brizio-Molteni L and Traber D. (eds.). Endocrinology of Thermal Trauma: Pathophysiologic Mechanisms and Clinical Interpretation. Philadelphia: Lea & Febiger, 238. Arturson G (1996): Pathophysiology of the Burn Wound and Pharmacological Treatment. The Rudi Hermans Lecture; 1995. Burns, 22(4): 255-274. Atiyeh BS, Hayek SN and Gunn SW (2005): New Technologies for Burn Wound Closure and Healing. Burns, 31(8):944-56. Bach FH, Bonavida B, Vitetta ES, et al. (eds.) (1979): T and B Lymphocytes: Recognition and Function. New York, Academic Press. Basill Bruite (2004): Personal communication in Joint meeting of The Middle East Burn and Fire Disaster Socity and the Plastic and Reconstructive Surgery Department of Ain Shams University, Cairo, Egypt. Baxter CR, Curreri PW and Marvin JA (1973): The Control of Burn Wound Sepsis by the Use of Quantitative Bacteriologic
References
131
Studies and Subeschar Clysis with Antibiotics. Surg. Clin. North Am., 53: 1509-18. Beal A and Cerra FB (1994): Multiple Organ Failure Syndrome in the 1990s. Systemic Inflammatory Response and Organ Dysfunction. JAMA, 271: 226. Bellomo R (1992): The Cytokine Network In the Critically Ill. Anaesth. Intens. Care., 20: 288. Bevilacqua MP and Nelson RM (1993): Selectins. J. Clin. Invest., 91: 379. Brink JA, Shhets PW and Dines KA (1986): Quantitative Assessment of Burn Injury in Porcine Skin with High Frequency Ultrasonic Imaging. Invest. Radiol., 21: 645-51. Brown RL, Greenhalgh DG, Kagan RJ and Warden GD (1994): The Adequacy of Limb Escharotomies-Fasciotomies after Referral to a Major Burn Center. J. Trauma, 37(6): 916920. Burke JF, Quinby WC and Bondoc CC (1976): Primary Excision and Prompt Grafting As Routine Therapy for the Treatment of Thermal Burns in Children. Surg. Clin. North America, 56: 477-494. Carsin H, Assicot M, Feger F, Rox O, Pennacino H, Le Bever H, Ainaud P and Bohuon C (1997): Evolution and Significance of Circulating Procalcitonin Levels Compared with IL-6, TNF-, and Endotoxin Levels Early after Thermal Injury. Burns, 23(3): 218-224.
References
132
Carsin H, Bargues L, Stephanazzi J, Paris A, Aubert P and Le Bever H (2002): Inflammatory Reaction and Infection in Severe Burns. Pathol. Biol. (Paris), 50(2):93-101. Carvajal HF, Linares HA and Brouhard BH (1979): Relationship of Burn Size to Vascular Permeability Changes in Rats. J. Surg. Gyn. & Obs., 149:193-202. Castell JV, Gomez-Lechon M, David M, Fabra R and Heinrich PC (1990): Acute Phase Response of Human Hepatocytes: Regulation of Acute Phase Protein Synthesis by IL-6. Hepatology, 12: 1179-1186. Chen J, Zhou YP and Rong XZ (2000): An Experimental Study on Systemic Inflammatory Response Syndrome Induced By Subeschar Tissue Fluid. Burns, 26 (2): 149-155. Childs CN, Edwards JV, Healthcote DM, DawsonM and Davenport PJ (1999): Toxic Shock Syndrome Toxin-1 (TSST1) Antibody Levels in Burned Children. Burns, 25(6):473-6. Cioffi WG, Burleson DG and Pruitt BA (1993): Leucocyte Response to Injury. Arch. Surg., 128: 1260. Cope O, Laugohr H, Moore FD and Webster R (1947): Expeditious Care of Full-Thickness Burn Wounds by Surgical Excision and Grafting. Ann. Surg., 125: 1-22. Curreri PW, Richmond D, Masrvin J, et al. (1974): Dietary requirements of patients with major burns. J. American Diet Association, 65:415.
References
133
Davies M, Adendorff D, Rode H and Van De Reit IS (1980): Coloring the Damaged Tissues on the Burn Wound Surface. Burns, 6: 156. Deitch E (1985): A Policy of Early Excision and Grafting in Elderly Burn Patients Shortens the Hospital Stay and Improves Survival. Burns Incl. Therm. Inj., 12: 109-114. Demling RH and Lalonde C (1990): Identification and Modifications of the Pulmonary and Systemic Inflammatory and Biochemical Changes Caused by a Skin Burn. J. Trauma, 30: 557. Desai MH, Herndon DN, Broemeling L, Barrow RE, Nichols RJ and Rutan RL (1990): Early Burn Wound Excision Significantly Reduces Blood Loss. Ann. Surg., 211(6): 753-762. Deveci M, Eski M, Sengezer M and Kisa U (2000): Effects of Cerium Nitrate Bathing and Prompt Burn Wound Excision on IL-6 and -TNF Levels in Burned Rats. Burns, 26:41-45. Dibirdik I, Durak N, Kislaoglu E, Kuluay T and Aytemiz C (1995): Effects of Prophylactic Intravenous Immunoglobulin-G Therapy on Humoral and Cellular Immune Components and Their Functions in Burned Patients. Burns, 21(2): 130-135. Dinarello CA and Wolff SM (1993): The Role of Interleukin1 in Disease. N. Engl. J. Med., 328: 106. Dowton SB and Colten HR (1988): Acute Phase Reactants in Inflammation and Infection. Serntn. Hematol., 25: 84-90.
References
134
Drost A and Pruitt B (1993): Plasma Cytokines Following Thermal Injury and Their Relationship with Patient Mortality, Burn Size and Time Post Burn. J. Trauma, 35:335-339. Dyess DL, Ferrara JJ, Luterman A and Curreri PW (1991): Subeschar Tissue Fluid: A Source of Cell-Mediated Immune Suppression in Victims of Severe Thermal Injury. J. Burn Care Rehabil., 12(2):101-105. Echinard CE, Sajdel SE, Burke PA and Burke JF (1982): The Beneficial Effect of Early Excision on Clinical Response and Thymic Activity after Burn Injury. J. Trauma, 22: 560-565. EL-Falaky MH, Abdel-Hafez A and Houtah AH (1985): Phagocytotic Activity of Polymorphonuclear Leukocytes in Burns. Burns, 11: 185. Engrav LH, Heimbach DM, Reus JL, Harnar TJ and Marvin JA (1983): A Randomized Prospective Study of Early Excision and Grafting of Indeterminate Burns Less Than 20% TBSA. J. Trauma, 23: 1001-1004. Ferrara JJ, Dyess DL, Luterman A and Curreri PW (1988): The Suppressive Effect of Subeschar Tissue Fluid upon in Vitro Cell-Mediated Immunologic Function. J. Burn Care Rehabil., 9(6):584-8. Garcia-Avello A, Lorente JA, Cesar-Perez J, Garcia-Frade LJ, Alvarado R, Arevalo JM, Navarro JL and Esteban A (1998): Degree of Hypercoagulability and Hyperfibrinolysis is Related To Organ Failure and Prognosis after Burn Trauma. Thromb Res., 15, 89(2):59-64.
References
135
Gordon JR, Burd PR and Galli SJ (1990): Mast Cells as a Source of Multifunctional Cytokines. Immunol. Today, 11: 458. Grabosch A and Rokos H (1992): Neopterin as Parameter of Cell-Mediated Immunity Response in Thermally Injured Patients. Burns, 18: 113. Gray D, Pine R and Harner T (1982): Early Excision versus Conventional Therapy in Patients with 20% to 40% Burns. Am. J. Surg., 149: 76. Grossman A and Zuckerman A (1984): Intravenous Fluorescein Photography in Burns. J. Burn Care Rehabil., 5: 66. Hack CE, De Groot ER, Felt-Bersma RJ, et al. (1989): Increased Plasma Levels of Interleukin-6 in Sepsis. Blood, 74: 1704. Haglund U and Gerdin B (1991): Trends in Shock Research. Oxygen Free Radicals and Circulatory Shock. Circ. Shock, 34: 405. Hart DW, Wolf SE, Chinkes DL, Beauford RB, Mlcak RP, Heggers JP, Wolfe RR and Herndon DN (2003): Effects of Early Excision and Aggressive Enteral Feeding on Hypermetabolism, Catabolism and Sepsis after Burn. J. Trauma, 54:755-764. Haynes BW (1987): The History of Burn Care. In: Boswick JAJ (ed.). The Art and Science of Burn Care, 3-9.
References
136
Heberer M, Allgower M, Durig M, Beuther P, Hasler P and Schoenenberger GA (1982): Chemiluminescence of Phagocytotic Cells Following Incubation of Whole Blood With Human Burn Toxin. Eur. Surg. Res., 14: 107-108. Heimbach DM (1987): Early burn excision and grafting. Surg. Clin. North America, 67: 93-107. Henane R, Bittel J and Banssilon V (1981): Partitional Calorimetry Measurements of Energy Exchanges in Severely Burned Patients. Burns, 7:180. Herndon DN, Abston S and Stein MD (1984): Increased Thromboxane B2 Levels in The Plasma of Burned and Septic Burned Patients. J. Surg. Gyn. & Obst., 159:210-213. Herndon DN and Parks DH (1986): Comparison of Serial Debridement and Autografting and Early Massive Excision with Cadaver Skin Overlay in the Treatment of Large Burns in Children. J. Trauma, 26(2): 149-152. Herndon DN, Stein MD, Rutan TC, Abston S and Linares H (1987): Failure of TPN Supplementation to Improve Liver Function, Immunity and Mortality In Thermally Injured Patients. J. Trauma, 27: 195-204. Herndon DN, Barrow RE, Rutan RL, et al (1989): A comparison of Conservative Versus Early Excision Therapies in Severely Burned Patients. Ann. Surg., 209:547-553. Herndon DN, Barrow RE, Kunkel KR, Broemeling L and Rutan RL (1990): Effects of Recombinant Human Growth
References
137
Hormone on Donor-Site Healing in Severely Burned Children. Ann. Surg., 212:424-429. Herndon DN, Ramzy PI, Barret JP (1999): Thermal Injury. Crit. care Clin., 15:333-52. Herndon DN (2001): Total Burn Care (2nd Edition), W.B. Saunders, London, England. Hlava P, Moserov AJ and Konigov AR (1983): Validity of Clinical Assessment of the Depth of a Thermal Injury. Acta Chir. Plast., 25: 202-208. Huang Y, Zongcheng Y, Faming C, Roger SC and Ao Li (1999): Effects of Early Eschar Excision En Masse at One Operation for Prevention and Treatment of Organ Dysfunction in Severely Burned Patients. World J. Surg., 23:1272-1278. Hultman CS, Yamamoto H, De Serres S, Frelinger JA and Meyer AA (1997): Early But Not Late Burn Wound Excision Partially Restores Viral-Specific T-Lymphocyte Cytotoxicity. J. Trauma., 43(3):441-7. Ikeda H and Kobayashi K (2000): Immunological Response to Severe Burn. Ryoi. Shok. Shirizu. (Japanese), (32):487-90. Ishibashi M, Saito K, Watanabe K, Uesugi S and Furue H (1991): Plasma Endothelin-1 Levels in Patients with Disseminated Intravascular Coagulation. N. Engl. J. Med., 324: 1516. Jackson D (1953): The Diagnosis of the Depth of Burning. Br. J. Surg., 40: 588 -96.
References
138
Jackson D (1969): Second Thoughts on the Burn Wound. J. Trauma, 9: 839. Kaneko H, Orii KE, Yamada M, Inoue R and Kondo N (2001): Long-Term Duration of Reduced Serum Complement Level Following Burn Injury. J. Investig. Allergol. Clin. Immunol.,11(4):303-4. Kaufman SE, Di Persio JF and Gasson JC (1989): Effects of Human GM-CSF on Neutrophil Degranulation in Vitro. Exp. Hematol., 17: 800. Kistler D, Hafermann B, Schoenenberger GA and Hettich R (1990): Increased Survival Rates by Topical Treatment of Burn with Cerium Nitrate. Eur. Surg. Res., 22: 283. Knabl JS, Bauer W, Andel H, Romer W, Choi MS, Horvat R, Meissl G, et al. (1999): Progression of Burn Wound Depth by Systemically Application of a Vasoconstrictor: An Experimental Study with a New Rabbit Model. Burns, 25(8):715-21. Koruda MJ, Zimbler A. Settle PG (1986): Assessing Burn Wound Depth Using in Vitro Nuclear Magnetic Resonance (NMR). J. Surg. Res., 40: 475-81. Kumar R, Seth RK, Sekhon MS, et al. (1995): Serum Lipid Peroxide and Other Enzyme Levels of Patients Suffering From Thermal Injury. Burns, 21: 96.
References
139
Lamaie K, Hakem I. Hemeda M. and Awad M (1999): The role of Cerium Nitrate in Major burn. Master degree thesis, Ain Shams University. Latha B, Ramakrishnan KM, Jayaraman V and Babu M (1997): Action of Trypsin Chymotrypsin (Chymoral Forte DS) Preparation on Acute-Phase Proteins Following Burn Injury in Humans. Burns, 23(1): 3-7. Leibovich SJ (1984): Mesenchymal Cell Proliferation in Wound Repair: The Role of Macrophages. In: Hunt TK, Heppenstall RB, Pines E, et al. (eds.). Soft and Hard Tissue Repair. New York: Praeger., 36. Liu XS, Yang ZC, Luo ZH, et al. (1994): Clinical Significance of the Changes of Blood Monocytic Interleukin-1 Production in Vitro in Severely Burned Patients. Burns, 20: 302. Lofgren O and Lundeberg T (1994): Calcitonin-Gene Related Peptide Reduces Thermally Induced Oedema in the Rat Paw. Neurosci. Lett., 1: 1. Lowry SF (1993): Cytokines Mediators of Immunity and Inflammation. Arch. Surg., 128: 1235. Marano MA, Fong Y, Moldawer LL, et al. (1990): Serum Cachectin/Tumor Necrosis Factor in Critically Ill Patients with Burns Correlates with Infection and Mortality. J. Surg. Gyn. & Obst., 170: 32.
References
140
McGrath MH (1990): Peptide Growth Factors and Wound Healing. Clin. Surg., 17: 421. Moore KL and Persuad TVN (1998): The Integumentary System. Before We Are Born: Essentials of Embryology and Birth Defects, 5th ed., 481-496. Mosher DF, Proctor RA and Grossman JE (1981): Fibronectin: Role in Inflammation. Adv. Inflam. Res., 2: 187207. Muller WA, Weigl SA, Deng X, et al. (1993): Platelet / Endothelial Adhesion Molecule-1 [PECAM-1] is Required For Transendothelial Migration of Leukocytes. J. Exp. Med., 178: 449. Munster AM, Winchurch RA, Thupari JN and Ernst CB (1986): Reversal of Postburn Immunosuppression with LowDose Polymixin-B. J. Trauma, 26: 995. Munster AM (1996): The Immunological Response and Strategies For Intervention. In: Herndon D, editor. Total burn care, 2nd ed. W.B. Saunders, 279-292. Nakae H, Endo S, Indada K, Yamada Y, Takakuma T and Yoshida M (1996): Plasma Levels of Endothelin-1 and Thrombomodulin in Burn Patients. Burns, 22(8): 594-597. Ono Y, Kunii O, Kobayashi K and Kanegasaki S (1993): Evaluation of Opsonophagocytic Dysfunctions in Severely Burned Patients by Luminol-Dependent Chemiluminescence. Microbiol. Immunol., 37(7):563-71.
References
141
Ono I, Gunji H, Zhang JZ, Marvyama K and Kaneko F (1995): A Study of Cytokines in Burn Blister Fluid Related to Wound Healing. Burns, 21(5): 352-355. Park DH (1998): Use of Laser Doppler Flowmetry for Estimation of the Depth of Burns. Plast. Recon. Surg., 101(6): 1516-1523. Rioja LF, Alonso P, De Haro J, et al. (1993): Prognostic Value of the CD4/CD8 Lymphocyte Ratio in Moderately Burned Patients. Burns, 19: 198. Rode H. (2001): Burn Injuries in Pediatric Patients. Crit. care Clin., 26:133-152. Rodriguez JL, Miller CG, Garner WL, et al. (1993): Correlation of the Local and Systemic Cytokine Response with Clinical Outcome Following Thermal Injury. J. Trauma, 34: 684-95. Roitt I, Brostoff J and Male D (1993): Immunology, 3rd ed. London. Moshy. Rong XZ, Li QH, Huang XH, Bei CH, Liu ZQ and Su YM (2003): Changes in Proinflammatory Cytokines and Immune Function of Subeschar Tissue Fluid During Early Stage After Severe Burn. Zhongguo Wei Zhong Bing Ji Jiu Yi Xue,15(10):612-4. Rose JK, Barrow RE, Desai MH and Herndon DN (1996): Advances in Burn Care. Adv. Surg., 30: 71-95.
References
142
Sanchez R (2002): Initial Shock from Burns. Physiopathology: Therapeutic Principles. (Paris) Pathol. Biol.,50(2):82-92. Scheidegger D, Sparkes BJ, Lu N, Schoenenberger G and Allgower M (1992): Survival in Major Burn Injuries Treated by One Bathing in Cerium Nitrate. Burns, 18: 296. Schenfeld W, Kasimir S, Koeller M, et al. (1990): Metabolism of Platelet Activating Factor (PAF) and Lyso-PAF in Polymorphonuclear Granulocytes from Severely Burned Patients. J. Trauma, 30: 1554. Schoenenberger GA, Bauer UR, Cueni LB, Eppenberger U and Allgower M (1971): Isolation and Characterization of a Cutaneous Lipoprotein with Lethal Effects by Thermal Injury in Mouse Skin. Biochem. Biophys. Res. Commun., 42: 975. Schoenenberger GA, Burkhardt F, Kalberer F, et al. (1975): Experimental Evidence for a Significant Impairment of Host Defense for Gram-ve Organisms by a Specific Cutaneous Toxin Produced by Severe Burn Injuries. J. Surg. Gyn. & Obst., 141: 555. Schroder JM and Christophers E (1992): The Biology of NAP-1/IL-8, a Neutrophil-Activating Cytokine. Immunol. Series, 57: 387. Schwacha MG, Knoferl MW and Chaudry IH (2000): Does Burn Wound Excision after Thermal Injury Attenuate Subsequent Macrophage Hyperactivity and Immunosuppression? Shock, 14(6):623-8.
References
143
Solomkin JS, Nelson RD, Chenoweth DE, et al. (1984): Regulation of Neutrophil Migratory Function by Complement Activation Products. Am. Surg., 200: 742. Sparkes BG (1991): Influence of Burn-Induced Lipid-ProteinComplex on IL-2 Secretion by PBMC in Vitro. Burns, 17: 129. Sparkes BG (1997): Immunological Responses to Thermal Injury. Burns, 23(2): 106-113. Suzuki F and Pollard RB (1982): Alteration of Interferon Production in a Mouse Model of Thermal Injury. J. Immunol., 129: 1806. Tanaka H, Mituo T, Yukioka T, Matsuda H, Shimazaki S and Igarashi H (1995): Comparison of Hemodynamic Changes Resulting from Toxic Shock Syndrome Toxin-1Producing Staphylococcus Aureus Sepsis and EndotoxinProducing Gram-ve Rod Sepsis in Patients with Severe Burns. J. Burn Care and Rehabil., 16(6): 616-621. Teodorczyk-Injeyan JA, Sparke BG and Peters WJ (1989): Regulation of IgM Production in Thermally Injured Patients. Burns, 15: 241. Teodorczyk-Injeyan JA, Sparkes BG, Mills G and Peters W (1991): Immunesuppression Follows Systemic T-lymphocyte Activation in the Burn Patient. Clin. Exp. Immunol., 85: 515. Teodorczyk-Injeyan JA, Sparkes BG, Lalani S, et al. (1992): Regulation of Soluble IL-2 Receptor Levels Following Thermal Injury. Clin. Exp. Immunol., 90: 36.
References
144
Tracey KJ and Cerami A (1993): Tumor Necrosis Factor: An Update Review of Its Biology. Crit. Care Med., 21: 415. Van De Graaff KM and Fox SI (1986): Concepts of Human Anatomy and Physiology. Dubuque: Wm. C. Brown. Vindenes HA and Bjerknes R (1994): Activation of Polymorphonuclear Neutrophil Granulocytes Following Burn Injury: Alteration of Fc-receptor and Complement-receptor Expression and of Opsonophagocytosis. J. Trauma, 36: 161. Vindenes HA and Bjerknes R (1997): Impaired Actin Polymerization and Depolymerization in Neutrophils from Patients with Thermal Injury. Burns, 23(2): 131-136. Wachtel TL (1989): Initial Care of Major Burns. Postgrad. Med., 85(1): 178-196. Yamamoto H, Siltharm S, De Serres S, Hultman CS and Meyer AA (1996): Immediate Burn Wound Excision Restores Antibody Synthesis to Bacterial Antigen. J. Surg. Res., 63:15762. Yang HM, Guo ZR and Gao WY (1995): The Association of Circulating Endotoxaemia with the Development of Multiple Organ Failure in Burned Patients. Burns, 21(4): 255-258. Yang ZC, Zhao ZD and Huang YS (1992): Clinical Study of the Pathogenesis of Multiple Organ Failure after Burns. Chin. J. Plast. Surg. Burns, 8: 8-16.
References
145
Zawacki BE and Walker HI (1970): An Evaluation of Patent Blue-V, Brophenol Blue and Tetracycline for the Diagnosis of Burn Depth. Plast. Recon. Surg., 54: 459-66.
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Doctor Mohamed Ahmed El Rouby Consultant of Plastic & Reconstructive Surgery Ain Shams University Cairo Egypt +2 0101556023 +2 0126531265 http://www.elroubyegypt.com http://tajmeel.ohost.de elroubyegypt@elroubyegypt.com elroubyegypt@gmail.com elroubyegypt@hotmail.com dr_mohamed_a@yahoo.com . - -

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Study of the Effect of Two Different Techniques of Early Burn Wound Excision on the Alteration of Interleukin-6 and Tumor

Necrosis Factor-Alpha Levels in Severe Burns

To my Sons and family!

- Evaluation of the Burn Wound Management Decisions

Index Patients and Methods

- Analysis of Laboratory Investigations

Discussion Summary and Conclusion References Arabic Summary

108 109 110 111

Aim of The Work

Aim of The Work

dysfunction may be caused

from initial respiratory tract

include paralytic ileus and gastrointestinal ulcerations. Small

Immune-Inflammatory Response to Burn

Fibrinopeptides Fibrin split products C3a C5a

Sensory nerve endings Mast cells, Basophils

Platelets , Mast cells PMNL, Macrophages, Basophils

Lipoxygenase pathway Lipoxygenase pathway

prekallikrein to Lysyl bradykinin

kininogen. Bradykinin is a very powerful vasoactive mediator,

Fig.1: Complement activation pathways (Kaneko et al., 2001)

gene-related sharing the of in

neurotransmitters They modulate

vasoconstriction, The most

microvascular include which

components histamine, and

hydroxytryptamine) mast cells, their

eosinophils, monocytes and vascular endothelial cells. It is

PGF2), Lipoxins, and the leukotrienes (A4, C4, B4, D4 and

LTC4 LTD4 LTE4

Fig. 2: The arachidonic acid cascade (Arturson, 1996)

T cells, B cells Monocytes

B cells Hepatocytes PMNLs Basophils Macrophages Granulocytes Tissue cells

Macrophages Lymphocytes Mast cells

Epithelial cells Fibroblasts

Leukocytes Tissue cells

High quantities (plasma conc. >10-7 M)

Fig. 4: Biological functions of TNF- (Lowry, 1993).

enhancing the removal of soluble fibrin by macrophages.

(Rolling). This is followed by adhesion of the leukocytes to

granules (Arturson, 1990). The cytoplasm of PMNLs contains many secretory

locomotion, but also shape change, pseudopod formation,

leukocyte-endothelial serve as the link

molecules and the

through two major

inflammatory liberate their

target tissue (Gordon et al., 1990).

insulin-like growth factor 1

thrombomodulin in the onset

increase in the plasma levels of ET-1, TM, and TNF- in

(lipopolysaccharide gram+ve exotoxins

Table 3: Paradoxes in Burn Immune Failure (Atiyeh et al., 2005).

mediated by the (LPC) (Allgower et al., 1995).

cytokine cascade), by the TSST-1 than by the endotoxin.

Evaluation of Burn Wound and Management Decisions

accurate for shallow and full-thickness wounds for any of

II- Surgical Burn Wound Management

Surgical Methods of Burn Wound Closure

A- Timing of Burn Wound Excision

burn depth are ascertained and prior to bacterial colonization

B- Extent of Burn Wound Excision

tourniquets, subcutaneous producing with

However, overdoses or paroxysmal