Page iii

Atlas of Gynecologic Oncology

Second Edition
Edited by
J Richard Smith, MB ChB, MD, FRCOG
Consultant and Honorary Senior Lecturer in Gynaecology
West London Gynaecological Cancer Centre,
Hammersmith Hospitals NHS Trust
London, UK
Adjunct Associate Professor of Gynecology
New York University School of Medicine
New York, USA
Giuseppe Del Priore, MD, MPH, FACOG
Vice President
New York University Downtown Hospital,
New York, USA
Associate Professor, Albert Einstein College of Medicine,
New York, USA
John P Curtin, MBA, FACOG, MD
Stanley Kaplan Professor
Chairman Ob. Gyn
New York University School of Medicine
New York, USA
John M Monaghan, MB ChB, FRCS(Ed), FRCOG
Whitton Grange
Whitton
Northumberland, UK
With artwork by

Dee McLean
Joanna Cameron
LONDON AND NEW YORK

Page 66

impossible. Antibiotic prophylaxis is given if there is any evidence or suspicion

of a urinary tract
infection.
Flexible cystoscopy is usually carried out in the endoscopy suite under local
anesthesia. Lignocaine
(lidocaine) gel inserted into the urethra acts as both lubricant and local
anesthetic agent. If possible the
patient should void prior to examination to ensure the bladder is empty.

Instrumentation
Rigid cystoscope
The rigid cystoscope (Figure 4) is composed of a sheath, a bridge and a
telescope: it is 30 cm long. The
sheath has both an inlet and an outlet port for irrigation and is attached to the
bridge with a watertight
lock. The endoscope is introduced into the sheath through the bridge, and is also
fitted with a watertight
lock. The telescope comprises a hollow metal cylinder containing a series of
solid rod lenses and a
magnifying eye-piece. In front of the eye-piece is a pillar connected to a
fiberoptic light source which
transmits light to the visual field. The bridge has one or two other ports for the
introduction of biopsy
forceps and electrodes, and a director which allows the passage of a ureteric
catheter and its
advancement into the ureteric orifice. Endoscopes with viewing angles of 0,
30, 70 and 90 are
available.

Flexible cystoscope
The flexible cystoscope (Figure 5) is 3540 cm long and consists of a control
head with eye-piece and
conFigure 4

Rigid cystoscope
Figure 5

Flexible cystoscope
trols, a multichannel flexible shaft and a controllable tip. The flexible shaft
contains fiberoptic channels

carrying the optics and light source to the visual field, an irrigation channel and
a biopsy channel.
Movement of the tip occurs in one plane and ranges from 145 to 180,
controlled by a deflecting level
adjacent to the eye-piece.

Operative procedure
Rigid cystoscope
The patient is placed on the operating table in the lithotomy position. The
cystoscope sheath is lubricated
and introduced into the urethra. The female urethra is about 4 cm long and has a
relatively uniform
caliber from the meatus to the bladder outlet. Upon entering the bladder the
telescope is removed to
allow the residual urine and irrigant to drain from the bladder: this may be sent
for cytological and
bacteriological analysis. Approximately 50 mL of saline is inserted and the
fundus of the bladder is
identified by finding the air bubble. With incomplete distension the bladder
mucosa appears rugated, but
as the irrigant fluid distends the bladder the mucosa becomes smooth. The
ureteric orifices are visualized
on the interureteric ridge at the superolateral corners of the trigone (Figure 6).
By regular sweeping of
the cystoscope backwards and forwards and rotation of the endoscope the entire
bladder mucosa can be
visualized. Views of the anteroinferior bladder are obtained by suprapubic
compression with the hand.
At the completion of the examination, the irrigating fluid is evacuated from the
bladder by removing the
telescope and
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67

Figure 6

Ureteric orifices
the instrument is slowly withdrawn. A bimanual examination of the pelvis is
performed after the
procedure.
Bladder biopsy
Bladder biopsy (Figures 7, 8) is the procedure most commonly performed
during cystoscopy. Biopsy
forceps are introduced down the cystoscope sheath via a port in the bridge,
sometimes together with a

diathermy wire. This allows cup biopsies of the mucosa to be taken. If required,
the biopsy sites are
Figure 7

Bladder biopsy
Figure 8

Bladder biopsy
then cauterized with diathermy to prevent excessive bleeding.
Ureteric catheterization and stenting
The instrumentation and stenting of ureters should only be performed by
clinicians such as gynecologic
oncologists trained in this procedure since it is easy to damage the ureteric
orifices and ureters. Ureteric
catheterization and the placement of double J stents is achieved with the 30
telescope. There is a special
port for the introduction of the stents which can be directed towards the ureteric
orifices. A floppytipped,
Teflon-coated guide wire is first placed into the ureteric orifice and advanced
under fluoroscopic
control into the renal pelvis. The double J stent is slid over the guide wire
through the channel of the
cystoscope and into the ureter (Figure 9). The stent is radio-opaque and its
position is monitored by
fluoroscopic control. Excessive force used in insertion of the guide wire or stent
should be avoided. The
proximal and distal ends curl to form a J shape when they are correctly placed
in the renal pelvis and
bladder respectively.

Flexible cystoscope
The patient is placed on the operating table or bed in the frog-leg position. The
cystoscope is lubricated
and introduced into the urethra. The end of the cystoscope is passed into the
bladder and deflected
upwards. The midline of the anterior bladder is examined by
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68

withdrawing the instrument until the bladder outlet is encountered. The

cystoscope is then pushed back
into the bladder, rotated 30 and withdrawn again. This process is continued
until the entire bladder has
been
Figure 9

Introduction of the double J stent

inspected. Biopsy of the bladder mucosa can also be achieved by the passage of
biopsy forceps down the
instrumental channel of the cystoscope.

Postoperative care
No special postoperative measures are needed.

Bibliography
Calne RY, Pollard SG (1992) Operative surgery. London: Gower.
Carter DC, Russell RCG, Pitt HA (1996) Atlas of general surgery, 3rd edn. London:
Chapman & Hall.
Cotton PB, Williams CB (1990) Practical gastrointestinal endoscopy, 3rd edn. Oxford:
Blackwell.
Jackson E, Fowler JR (1992) Urological surgery. Mastery of Surgery series. New York:
Little, Brown.
Reuter HJ (1987) Atlas of urological endoscopy: diagnosis and treatment. New York:
Thieme. Page 69

6
Ovarian tissue cryopreservation and
transplantation
techniques
Erkan Buyuk
Kutluk H Oktay

Introduction
Modern improvements in cancer treatment regimens using aggressive
chemotherapy radiotherapy, as
well as bone marrow transplantation, can result in cure rates exceeding 90% for
many cancers (Baird et
al. 1999). However, this success has been accompanied by loss of fertility and
premature menopause in
many women cured of their disease. Ovarian cryopreservation and
transplantation is one of the options
aimed to preserve fertility in women who face a threat to their fertility.
Discovery of modern
cryoprotectants and progress in cryopreservation techniques led to successful
cryopreservation of
gametes, embryos and ovarian tissue. However, there is significant room for
improvement in
revascularization of tissues after auto-transplantation, as nearly two thirds of the
ovarian reserve is lost
during the initial ischemic state after grafting (Morales et al. 1995, Imthurn et
al. 2000, Demirci et al.

2001).

Ovarian tissue cryopreservation

Ovarian tissue cryopreservation for future transplantation can be done in cancer
patients as well as for
other benign conditions where chemotherapy, radiotherapy, or surgically
induced ovarian failure is
anticipated. Table 1 summarizes the indications for ovarian tissue banking.

Tissue harvesting
As long as there is no contraindication, ovarian tissue is collected via
laparoscopy In adult patients, we
generally remove one ovary to obtain a large reserve of primordial follicles.
However, in pediatric age
groups, a large cortical biopsy may be enough since their ovaries harbor a larger
number of follicles than
thePage 70
Table 1 Indications for ovarian cryopreservation and transplantation
1. Cancer patients
Breast cancer (stage 0III)
Cervical cancer
Childhood cancers
Hodgkins lymphoma
non-Hodgkins lymphoma (except Burkitt lymphoma)
Osteosarcoma
Ewings sarcoma
Wilms tumor
2. Bone marrow transplant patients
Aplastic anemia
Sickle-cell anemia
Autoimmune and immunodeficiency diseases (e.g. rheumatoid arthritis)
3. Autoimmune diseases
Collagen vascular diseases (e.g. SLE)
Acute glomerulonephritis
Behets disease
4. Adjunctive oophorectomy
Recurrent breast cancer
Endometriosis
5. Benign ovarian tumors
Recurrent cysts
Endometriosis
6. Prophylactic oopherectomy
BRCA-1 or -2 mutation carriers
SLE, systemic lupus erythematosus.

adult ovary (Newton et al. 1998). The whole ovary or ovarian cortical pieces are
removed by a
laparoscopic approach using a 5 mm scope inserted in the umbilicus and 5 mm
and 12 mm trochars in

the lower quadrants. Use of electrocautery is not recommended in order to avoid

damage to ovarian
cortex containing the follicles. The ipsilateral fallopian tube is left intact to
allow a spontaneous
pregnancy to occur in case an orthotopic transplantation is performed in the
future. An endoscopic
specimen bag is used to remove the ovary through the 12 mm trocar; the trocar
is pulled out and the
specimen is delivered through the 12 mm incision. This incision may need to be
widened to extract a
large ovary.
Figure 1

Preparation of ovarian tissue for cryopreservation. The ovary is bivalved

through its hilum.
Note that the primordial follicles are located predominantly in the cortex

Processing of the ovarian tissue

The aim of processing ovarian tissue before cryo-preservation is to obtain
ovarian pieces small and thin
enough for the cryoprotectants to easily permeate .The sample is transported to
the laboratory on ice in
Leibovitz L-15 medium. In the case of a whole ovary, it is bivalved through its
hilum (Figure 1) and the
cortex
Figure 2

The cortex is dissected from the stromaPage 71

is separated from the medullary portion (stroma) using a number 10 blade
(Figure 2). This step is
undertaken because the primordial follicles are contained in the cortical portion
and the medullary
portion may decrease tissue permeation of cryoprotectants. The cortex is then
divided into 1051 mm
strips using a no. 10 or 11 blade (Figure 3). The preparation is performed under
a laminar flow hood and
the tissue is kept in the medium throughout the process. The cortical pieces are
then put in cryovials
containing 1.5 ml of ovarian freeze solution (1.5 M 1,2 propanediol, 20%
patients own serum and 0.1
M sucrose in Leibovitz L-15 medium).

Cryopreservation and thawing

The cryovials are kept in ice for 30 minutes for the equilibration of the
cryoprotectants.
Cryopreservation is performed using a slow freeze protocol in a programmable
freezer. The pieces are

cooled to 7C and seeded at this temperature. They are then cooled to 140C
and plunged into liquid
nitrogen (Oktay 2001).
Thawing is done by a rapid thaw protocol in a 30C water bath, followed by
washing the tissues in
decreasing gradients of cryoprotectant (Oktay 2001).
Figure 3

The cortex is divided into 1051 mm strips

Ovarian transplantation techniques

Before performing ovarian transplantation, the risk of reseeding occult cancer
cells should be kept in
mind and the decision to perform ovarian transplantation should be made
accordingly. Table 2
summarizes the risk of ovarian involvement in different cancers. For instance,
the risk of ovarian
involvement is higher in leukemia and neuroblastoma patients, compared to
lymphoma or Wilms tumor
patients.
Transplantation can be done using an orthotopic or heterotopic approach. In
orthotopic transplantation,
the tissue is placed in the ovarian fossa (Oktay and Buyuk 2002). Although
spontaneous pregnancy can
in theory be achieved using this technique, the procedure is technically more
challenging, and when
there is a higher likelihood of ovarian involvement with cancer, it may be less
desirable to graft ovarian
tissue retroperitoneally. In heterotopic transplantation, the tissue is grafted into a
place other than the
ovarian fossa. The operation is done under local anesthesia and follow-up is
easier with heterotopic
transplantation.
Table 2 Risk of metastases to ovaries in various cancers
High risk
Leukemia
Neuroblastoma
Burkitt lymphoma
Moderate risk
Breast cancer
Stage IV
Infiltrative lobular histological subtype
Adenocarcinoma/adenosquamous carcinoma of the cervix
Colon cancer
Low risk
Breast cancer

Stages IIII
Infiltrative ductal histological subtype
Squamous cell carcinoma of the cervix
Non-Hodgkins lymphoma
Hodgkins lymphoma
Wilms tumor
Ewings sarcoma
Nongenital rhabdomyosarcoma
Osteogenic sarcoma
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Page 72

However, patients will always need an in vitro fertilization (IVF) procedure in

order to conceive (Oktay
et al. 2001a, b). In case of cancer recurrence, tissue sampling and removal will
also be easily
accomplished. The risk and significance of recurrent cancer at the transplant site
are unknown.

Pelvic orthotopic transplantation

Following thawing and washing, the ovarian pieces are placed in a petri dish
containing transport
medium, and transported on ice to the operating room. In the operating room, 6
0 vicryl is used to string
the strips by passing the needle between the cortex and the stroma of each strip
under a microsurgical
microscope (Figure 4). Several strings are formed depending on the number of
the strips, and they are
then anchored to a Surgicel frame (Ethicon, Somerville, NJ). 10 vicryl is used
on the sides to further
strengthen this frame. 0 vicryl sutures are then tagged to the apex and the base.
Synchronously, the
patient is anesthetized, and three trocars are inserted: an 11 mm one in the
umbilicus, a 5 mm one in the
right lower quadrant, and a 13 mm one (with fascia anchor) suprapubically.
Using sharp and blunt
dissection, a pocket is created in the ovarian fossa, posterior to the broad
ligament, superior to the
ureters, and inferior to iliac vessels in
Figure 4

The strips are strung on to 60 vicryl

Figure 5

The graft is loaded retrogradely into the 13 mm trocar

the supine position. The graft is then loaded retrogradely, into a 13 mm trocar
(Figure 5), which is

reinserted in the fascia anchor suprapubically. Pulling on the leading suture, the
graft is dropped in the
pelvis. The leading suture is then placed in the most dependent portion of the
pocket, approximately 1
cm above the ureter, and the needle is passed through the peritoneum into the
pelvic cavity (Figure 6).
By pulling on this suture, the graft is wedged in the pelvic pocket. Next, the
base suture is passed
through the upper
Figure 6

The leading suture is placed in the most dependent portion of the pelvic pocket
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73

Figure 7

The graft is flattened and stretched against the pelvic side-wall

Figure 8

The pelvic peritoneum is closed with interrupted sutures

edge of the peritoneal pocket. The graft is stretched and flattened against the
pelvic side-wall by pulling
this suture from the intraperitoneal site (Figure 7). With an abundance of pieces,
a second graft may be
prepared and placed superior and caudal to the first one. Then, using an
extracorporeal knot placement
technique, the peritoneum is approximated with interrupted sutures (Figure 8).
The base of the Surgicel
frame is also included into the suture while closing the peritoneum to further
secure the graft in place.
The patient is given 150 IU/day of follicle-stimulating hormone (FSH)
systemically for 7 days (Oktay et
al. 2003). As beneficial effects on graft survival have been observed in animal
studies (Richardson et al.
1987). The patient is also given 80 mg per day aspirin for 7 days, and started on
hormone replacement
therapy within 48 hours of the transplant, as it is thought that these treatments
may enhance angiogenesis
(Ries et al. 1999).

Forearm heterotopic transplantation

After thawing and washing as described previously, ovarian cortical strips are
placed in phenol red-free
Minimum Essential Medium Alpha Medium (with L-glutamine, ribonucleosides
and
deoxyribonucleosides, Invitrogen, cat. no. 41061029), supplemented with 20%
of the patients own

serum and 10 g/mL cefotetan, and kept on ice. Then, each strip is tagged with
40 vicryl as described
previously (Figure 9). The needle is cut, and the cortical pieces are left in the
medium until the surgical
site is ready for transplantation. To create a pocket for the graft under the skin of
the forearm, a 1 cm
transverse incision is made over the brachioradialis muscle, 5 cm below the
antecubital fossa. If there is
a cosmetic concern, the incision and the transplantation may be made more
medially. A pocket is created
between the fascia and the subcutaneous tissues using blunt dissection (Figure
10). Since this area is
relatively vascular, attention must be given to avoid major bleeding. As the
ovarian tissue will acquire
its blood supply from these vessels, extensive cauterization should be avoided.
Figure 9

Each ovarian strip is tagged using 40 vicryl