Targeting Stromal Cells in Chronic Inflammation

Andrew Filer, M.D., Ph.D., Karim Raza, M.D., Ph.D., Mike Salmon, Ph.D., and Christopher D.
Buckley, M.D., Ph.D.
Rheumatology Research Group, MRC Center for Immune Regulation, Division of Immunity and
Infection, University of Birmingham, Birmingham B15 2TT, United Kingdom
Summary
Why chronic inflammatory reactions persist in specific sites, such as rheumatoid arthritis in the
joints, remains a mystery. Current models of inflammation have concentrated upon the responses
of lymphocytes such as B and T cells to specific antigens, and have attempted, often
unsuccessfully, to address the causative agent. However recent studies have shown that stromal
cells such as macrophages, endothelial cells, and fibroblasts play important roles in the switch that
turns a spontaneously resolving acute inflammatory response within a tissue into chronic and
persistent disease. Therapeutic manipulation of the stromal microenvironment has been
particularly effective in treating cancer and is likely to provide novel therapies to achieve
improved control of chronic inflammatory disease.
A characteristic feature of chronic inflammatory reactions is their persistence and
predilection for certain sites. Why for example does rheumatoid arthritis not get better
spontaneously? Why does it localize predominantly to the joints, while the inflammation of
psoriasis localizes predominantly to the skin? It has been assumed that inflammation is a
generic response that follows a common recipe, resulting in endothelial cell activation,
immune cell infiltration, and tissue repair. However the advent of biological therapies, such
as anti-TNF blockade, has taught us that there are features of the inflammatory response that
are not generic (public), but instead remain unique or private to the tissue where
inflammation occurs.
Current models of inflammation have concentrated upon the role of antigen-specific
lymphocyte responses (production of antibodies by B lymphocytes and pro-inflammatory
activity by T lymphocytes) and have attempted, unsuccessfully in many cases, to address the
causative agent. However, recent studies have begun to challenge the primacy of the
lymphocyte and have instead begun to focus on an extended immune system in which
stromal cells such as macrophages, endothelial cells, and fibroblasts play a role in the
persistence and the anatomic location of inflammation. It turns out that tissue-resident
stromal cells are much more important than initially thought in determining the site at which
inflammation occurs. Furthermore, stromal cells play an important part in that crucial switch
which turns a spontaneously resolving acute inflammatory response into chronic and
persistent disease such as that occurs in rheumatoid arthritis. Inflammation is therefore not
generic but contextual, and differences in the response of different inflammatory diseases to
therapy are likely to be due to intrinsic differences in the behavior of stromal cells within
different tissue microenvironments.
Corresponding author: Dr. Christopher D. Buckley (c.d.buckley@bham.ac.uk)..
UKPMC Funders Group
Author Manuscript
Discov Med. Author manuscript; available in PMC 2011 August 24.
Published in final edited form as:
Discov Med. 2007 February ; 7(37): 2026.
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What Are Stromal Cells and How Do They Contribute to Chronic
Inflammation?
In this review we define stromal cells as those cells responsible for defining tissue
architecture through their elaboration of extracellular matrix and cytokines including tissue
resident macrophage-like cells. This broad definition includes fibroblasts, vascular
endothelial cells, and tissue specific macrophages such as the Kupffer cell of the liver. Each
set of tissue specific stromal cells appears to produce a distinctive set of matrix and soluble
proteins that not only define the specialized architecture of organs and tissues, but also
characterize the chemical scent of that tissue. For example we now know that tissue-
specific stromal cells are able to determine the nature and number of immune cells that
accumulate in tissues during inflammatory responses. At the resolution of such responses,
infiltrating immune cells which are no longer useful either undergo cell death or leave via
draining lymphatics under the direction of gradients of specialized signals called
chemokines. Stromal cells contribute to the withdrawal of survival signals and normalization
of chemokine gradients that allow the resolution of immune responses to occur. Subversion
of these pathways, which normally result in a well-choreographed influx and departure of
immune cells, leads to a switch to persistent inflammation which then appears to remain
remarkably stable (Buckley et al., 2001).
Furthermore, stromal cells play an important part in that crucial switch which
turns a spontaneously resolving acute inflammatory response into chronic and
persistent disease such as that occurs in rheumatoid arthritis. Inflammation is
therefore not generic but contextual, and differences in the response of different
inflammatory diseases to therapy are likely to be due to intrinsic differences in the
behavior of stromal cells within different tissue microenvironments.
Why Target Stromal Cells?
It is clear that current anti-inflammatory therapies, while effective, do not cure immune
mediated inflammatory diseases. This may be because current therapies do not target the
right type of cells, at the right time and in the right place. Recent studies suggest that
ignoring the contribution of interactions between immune cells and stromal cells may
account for the failure of current therapies to effect a permanent cure. These interactions
offer a new family of potential organ-specific targets in order to treat such disease. In
addition, stromal cells are a source of a new family of naturally occurring anti-inflammatory
agents, termed resolvins, whose active production during inflammatory reactions helps bring
about the resolution of inflammation (Serhan et al., 2007). Rather than negatively targeting
pro-inflammatory signals, part of future therapies may involve the identification and
exploitation of these endogenous anti-inflammatory signals. In targeting the stromal
microenvironment we are finally starting to address the issue of the switch that underlies
persistence of chronic inflammatory diseases rather than their immediate control.
Stromal Cells as Targets in Inflammation and Cancer
Fibroblasts
The most abundant cells of the stroma are fibroblasts, which are responsible for the
synthesis and remodeling of extracellular matrix components and therefore tissue
remodeling and repair. Rheumatoid synovial fibroblasts provide a convincing example of
how stromal cells contribute to the persistence of inflammation. These cells are critical to
the destruction that occurs in the rheumatoid joint as they directly invade and destroy
cartilage and bone. Furthermore this behavior is imprinted: when rheumatoid fibroblasts are
grown in culture for many weeks then reimplanted into mice, they continue to invade
Filer et al. Page 2
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cartilage. In this way, fibroblasts directly contribute to disease outcome (Ritchlin, 2000).
Rheumatoid fibroblasts also play a role in maintaining the presence of immune cell
populations in the joint (Buckley et al., 2001). Not only do fibroblasts keep activated, pro-
inflammatory T lymphocytes alive, they also produce cytokine signals such as transforming
growth factor (TGF-) and interleukin 15 (IL-15), which instruct T lymphocytes to
express receptors for a specific chemokine, CXCL12. CXCL12 is a chemokine with very
specific roles in the developing immune system and normally plays a role in guiding the
movement and development of bone marrow cells. However high levels were unexpectedly
seen in the rheumatoid joint, resulting in the capture and inappropriate retention of activated
T lymphocytes in this microenvironment (Parsonage et al., 2005). Specialized immune
signalling molecules such as CD40 have also been found on fibroblasts (Brouty-Boye et al.,
2000). These molecules, previously thought to be restricted to immune cells within
lymphoid tissues, allow communication with T lymphocytes, which in the rheumatoid joint
leads to further fibroblast production of CXCL12. Therefore an emerging theme is that
chronically inflamed tissues, such as the rheumatoid joint, begin to take on characteristics of
lymphoid tissues such as the bone marrow. Furthermore fibroblasts, as key determinants of
tissue landscape, appear to contribute to this false regional identity. How fibroblasts do
this is an area of active research but intriguing evidence suggests that fibroblasts can
originate from a number of unexpected sources including their recruitment from the
circulation as monocyte-derived fibrocytes or circulating stromal progenitor cells. Therefore
understanding how fibroblast numbers and subsets change during the course of
inflammatory disease will be an important avenue for further research.
Like the field of inflammation, cancer medicine has also been experiencing a renaissance in
interest in the biology of fibroblasts and stromal cells (Bhowmick et al., 2004). A number of
important cytokines have been described which contribute to cancerous transformation of
healthy cells by so-called tumor associated fibroblasts. These include hepatocyte growth
factor (HGF) and TGF-. Crucially, tumor associated fibroblasts appear able to help
transform normal cells, in addition to pre-malignant cells, into tumor cells. The importance
of tumor associated fibroblasts has been demonstrated in breast cancer, where human cancer
cells were unable to grow successfully when implanted into mice unless human tumor-
derived fibroblasts were implanted with them. Intriguing parallels have been shown with
inflammatory disease such as rheumatoid arthritis in the predilection for cancer cells to
metastasize to certain sites. In particular the presence of the chemokine CXCL12 and its
receptor CXCR4 has been implicated in the persistence and pattern of tissue metastases in
breast cancer.
How can we start to target fibroblasts in inflammation and cancer (Figure 1)? Direct
targeting and depletion is as yet a distant possibility, as we do not have specific markers for
these cells. However, cytokines and chemokines are excellent targets for therapy, and
blockade of CXCL12 using small chemical entities in animal models of arthritis has yielded
encouraging results. Blockade of other specialized molecules such as CD40 may also be
beneficial. In cancer research, blockade of TGF- and HGF have been effective in inhibiting
cancer development and metastasis in animal models. Novel future approaches may include
targeting of circulating fibroblast precursors by depletion or blockade of transdifferentiation
from other cells such as fibrocytes.
Recent studies suggest that ignoring the contribution of interactions between
immune cells and stromal cells may account for the failure of current therapies to
effect a permanent cure. These interactions offer a new family of potential organ-
specific targets in order to treat such disease.
Filer et al. Page 3
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Endothelial Cells
Endothelial cells are responsible for the regulation of vascular tone and thrombosis, as well
as orchestrating the movement of cells from the blood into tissues. Immune cells migrate
across a diverse spectrum of blood vessels from large arteries to capillaries and specialized
lymphatic endothelium. In chronic inflammation and during the growth of cancerous tissue,
angiogenesis (new blood vessel formation) is required to support the increased metabolic
demand. Angiogenesis requires the presence of new organ-specific vascular cells in addition
to a network of associated supporting stromal cells known as pericytes. Angiogenesis is
normally tightly controlled via signals such as vascular endothelial growth factor (VEGF)
and fibroblast growth factor. It is now clear that angiogenesis is essential for cancer
progression and metastasis. Tumor invasion into the extracellular matrix results in release of
angiogenic factors such as VEGF. In fact studies of skin cancer have indicated that
expression of VEGF is wide-spread in both benign and malignant tumors; however, the
presence of corresponding VEGF receptors by stromal endothelial cells is the key signal
which triggers certain tumors to invade and spread (Bergers and Benjamin, 2003). So not
only are angiogenic signals important, but the cancer cells rely on their surrounding stromal
cells to metastasize. There is good evidence that both endothelial cells and pericytes are, like
some fibroblasts, recruited as precursors from the circulation. These precursor cells have
been shown to be attracted in some cancers by the chemokine CXCL12, yet again providing
a further shared molecular link between the processes that go wrong in cancer and chronic
inflammation.
Finding an effective means to block angiogenesis has become the focus of much research,
some of which has produced encouraging results (Figure 2): VEGF inhibitors suppress new
vessel growth in inflamed tissue, and in animal models reduce arthritis incidence and
severity. Studies of angiogenesis inhibitors and antibodies against extracellular matrix
components that promote angiogenesis such as tenascin have been favorable. Furthermore,
inhibiting conversations between endothelial cells and their closely related pericytes by
blocking a specific signal, platelet derived growth factor, has been highly effective, helping
even late stage tumors to regress when VEGF inhibitors alone had failed. This illustrates the
importance of targeting the stroma as a whole, rather than individual cell types.
Tissue Resident Macrophages
Within tissues, monocytes differentiate into a wide variety of cells including macrophages,
dendritic cells (which find and present invading antigens to the immune system), and
osteoclasts (which degrade bone during remodeling). In tissues, macrophages perform a
sentinel role, identifying dangerous invaders and removing dying cells by phagocytosis.
However they also have important regulatory effects upon other stromal cells such as
fibroblasts, and are potentially amenable to direct targeting as excellent cellular markers that
are relatively specific for macrophages have been identified.
An intriguing model of liver inflammation developed by Duffield et al. (2005) has shown
that depletion of macrophages at key stages of liver fibrosis leads to an alteration of disease
pattern, suggesting that there is potential for just such an approach. During fibrosis of the
liver, specialized liver fibroblasts called stellate cells transform under the influence of the
cytokine TGF- into myofibroblasts, which cause tissue damage and lay down fibrotic tissue
leading to scarring. Macrophages promote this step by secretion of TGF-. In contrast,
during recovery from fibrosis in animal models, myofibroblasts undergo cell death, leading
to regression of fibrosis. This time the signal involved is TNF-related apoptosis-inducing
ligand (TRAIL), which once again is produced by macrophages. What was remarkable in
Duffields study was that, using a mouse model, macrophages could be rapidly depleted
when required. Removal of macrophages during the damaging, fibrotic phase of disease
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resulted in fewer myofibroblasts and decreased scarring. However when macrophages were
depleted during the recovery phase removing the source of TRAIL, sustained accumulation
of fibrotic, scarred tissue occurred as death of myofibroblasts was blocked (Duffield et al.,
2005) (Figure 3). Thus macrophages are able to switch the roles and the survival of other
stromal populations. As we learn more about the relationships between stromal cells in
disease, it is likely that tissue resident macrophage cells will become a viable target for
depletion allowing us to modify the outcome of chronic inflammatory diseases.
Conclusion
Stromal cells such as endothelial cells, fibroblasts, and tissue-resident macrophages have an
emerging potential as therapeutic targets. They are responsible for orchestrating and
maintaining the presence of immune cells in persistent inflammatory conditions and for
supporting the spread and tissue-specific metastasis of tumor cells in cancer. The success of
anti-TNF blockers to treat conditions such as rheumatoid arthritis illustrates the
effectiveness of a classical approach to blocking interactions between immune and stromal
cells. Harnessing our improving knowledge of the stromal microenvironment is likely to
lead to further successful therapies. Lastly, naturally occurring, stromally derived factors
that actively drive the resolution of inflammation are exciting new replacement therapies
for the future and are likely to be important targets for further research.
Acknowledgments
The University of BirminghamRheumatology Research Group is part of the Medical Research Council (MRC)
Centre for Immune Regulation. It is supported by grants fromthe MRC and arc (Arthritis Research Campaign).
References and Further Readings
Bergers G, Benjamin LE. Tumorigenesis and the angiogenic switch. Nature Reviews Cancer. 2003;
3(6):401410.
Bhowmick NA, Neilson EG, Moses HL. Stromal fibroblasts in cancer initiation and progression.
Nature. 2004; 432(7015):332337. [PubMed: 15549095]
Brouty-Boye D, Pottin-Clemenceau C, Doucet C, J asmin C, Azzarone B. Chemokines and CD40
expression in human fibroblasts. European J ournal of Immunology. 2000; 30(3):914919.
[PubMed: 10741409]
Buckley CD, Pilling D, Lord J M, Akbar AN, Scheel-Toellner D, Salmon M. Fibroblasts regulate the
switch from acute resolving to chronic persistent inflammation. Trends in Immunology. 2001;
22(4):199204. [PubMed: 11274925]
Duffield J S, Forbes SJ , Constandinou CM, Clay S, Partolina M, Vuthoori S, Wu S, Lang R, Iredale J P.
Selective depletion of macrophages reveals distinct, opposing roles during liver injury and repair.
J ournal of Clinical Investigation. 2005; 115(1):5665. [PubMed: 15630444]
Parsonage G, Filer AD, Haworth O, Nash GB, Rainger GE, Salmon M, Buckley CD. A stromal
address code defined by fibroblasts. Trends in Immunology. 2005; 26(3):150156. [PubMed:
15745857]
Ritchlin C. Fibroblast biology. Effector signals released by the synovial fibroblast in arthritis. Arthritis
Research. 2000; 2(5):356360. [PubMed: 11094448]
Serhan CN, Brain SD, Buckley CD, Gilroy DW, Haslett C, ONeill LA, Perretti M, Rossi AG, Wallace
J L. Resolution of inflammation: state of the art, definitions and terms. FASEB J ournal. 2007;
21:325332. [PubMed: 17267386]
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Figure 1.
Fibroblasts are responsible for maintaining the persistence of inflammation in diseases such
as rheumatoid arthritis by secreting cytokines that keep activated lymphocytes alive, and
chemokines that prevent lymphocytes from leaving the joint. They also contribute to
increased inflammatory signal generation via direct communication using molecules such as
CD40. Potential therapeutic approaches include blockade of specific cytokines, chemokines,
and CD40. Some of the Proinflammatory fibroblasts in tissues appear to originate from
distinct circulating precursors, which are potential targets for depletion within the blood. An
alternative, novel approach would be to block their differentiation from precursor monocyte
populations.
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Figure 2.
Endothelial cells are responsible for posting of inflammatory cytokines and chemokines
on their cell surface and thereby provide an area-post code for infiltrating leucocytes.
Blockade of cytokines or chemokine presentation pathways are potential targets for therapy.
The production of new blood vessels (angiogenesis) is essential to support increased
inflammatory or cancerous progression. Blockade of angiogenesis and/or recruitment of
circulating endothelial cell precursors are therapeutic tools already in clinical trials for
certain diseases.
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Figure 3.
Tissue macrophages modulate the differentiation and function of fibroblasts, and are an
emerging therapeutic target. In a model of liver fibrosis (top left), macrophages control the
differentiation of stellate cells to myofibroblasts that produce fibrotic tissue. Depletion of
macrophages during this stage results in less fibrosis (bottom left). During recovery (top
right), macrophages promote the programmed cell death (apoptosis) of myofibroblasts
leading to decreased fibrosis. Macrophage depletion at this stage (bottom right) prevents
death of myofibroblasts and results in worsening of disease.
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