Effects of Hyaluronic
Acid–Carboxymethylcellulose
Antiadhesion Barrier on Ischemic
Colonic Anastomosis
An Experimental Study
Suphan Erturk, M.D.,* Serdar Yuceyar, M.D.,* Muhyittin Temiz, M.D.,*
Baki Ekci, M.D.,* Nevin Sakoglu, M.D.,* Huriye Balci, Ph.D.,† Ahmet Dirican, M.D.,‡
Ali Cengiz, M.D.,* Haluk Saner, M.D.*
From the *Department of General Surgery, †Central Research Laboratory, and ‡Department of Biostatistics,
Istanbul University Cerrahpasa Medical Faculty, Istanbul, Turkey
PURPOSE: Intraperitoneal adhesions may help the healing
of marginally viable bowel ends. If adhesion formation is
prevented by various methods, the integrity of ischemic
bowel anastomosis may be compromised. Thus, we decided
to study the effects of hyaluronic acid–carboxymethylcellu-
lose, an antiadhesion barrier, on ischemic bowel anastomo-
sis. METHODS: Thirty Wistar-Albino rats were divided into
three groups. In Group A (control), a well-perfused distal
colonic segment was transected, and free ends were anas-
tomosed. In Group B, an ischemic colonic segment was
prepared, then divided and anastomosed. In Group C, after
completion of ischemic colonic anastomosis, hyaluronic
acid–carboxymethylcellulose film was wrapped around the
anastomosis. In all groups, rats were killed on the seventh
day. Intraperitoneal adhesions were graded by adhesion
score, and healing of the anastomosis was assessed by mea-
surement of bursting pressure and hydroxyproline levels in
the anastomotic tissue. RESULTS: A statistically significant
difference was found between hydroxyproline levels of the
control group and the ischemic group (P ⫽ 0.02). HP level
was also significantly higher in the hyaluronic acid–car-
boxymethylcellulose group than in the ischemic group (P ⫽
0.01). There was no difference in hydroxyproline levels
between the control and hyaluronic acid–carboxymethyl-
cellulose groups. Compared with the control group, burst-
ing pressure was lower in the ischemic group (P ⫽ 0.02).
Hyaluronic acid–carboxymethylcellulose wrapping in-
creased the bursting pressure significantly (P ⬍ 0.001).
However, there was no difference in bursting pressure
between the control group and the hyaluronic acid–car-
boxymethylcellulose group (P ⫽ 0.13). A marked increase
in the adhesion score was observed in the ischemic group
(P ⫽ 0.01). The difference between adhesion scores of the
hyaluronic acid–carboxymethylcellulose and ischemic
groups was not found to be significant, although the adhe-
sion score in the hyaluronic acid–carboxymethylcellulose
Presented at the joint meeting of the Mediterranean Society of
Coloproctology and the Turkish Association of Colorectal Surgeons
and the Ninth Turkish National Congress of Colorectal Surgery,
Kemer-Antalya, Turkey, September 9 to 13, 2001.
No reprints are available.
529
group was lower (P ⫽ 0.16). There was no difference in
adhesion score between the control and hyaluronic acid–
carboxymethylcellulose groups. CONCLUSIONS: Applica-
tion of hyaluronic acid–carboxymethylcellulose in ischemic
colonic anastomosis did not compromise anastomotic integ-
rity. The adverse effect of ischemia on healing of colonic
anastomosis was counteracted by hyaluronic acid–car-
boxymethylcellulose. [Key words: Intraperitoneal adhe-
sions; Prevention; Ischemia; Intestinal anastomosis; Hyal-
uronic acid–carboxymethylcellulose]
Erturk S, Yuceyar S, Temiz M, Ekci B, Sakoglu N, Balci H,
Dirican A, Cengiz A, Saner H. Effects of hyaluronic acid–
carboxymethylcellulose antiadhesion barrier on ischemic
colonic anastomosis: an experimental study. Dis Colon Rec-
tum 2003;46:529–534.
A fter abdominal operations, intraperitoneal adhe-
sions are seen in approximately 60 to 90 percent
of cases and may lead to considerable complications,
i.e., bowel obstruction, fistula, chronic abdominal
pain, sterility in females, and difficulties experienced
during reoperative interventions.1–4 Various methods
have been tried to prevent postoperative adhesion
formation, including intraperitoneal application of
mechanical barriers.5 Hyaluronic acid–carboxymeth-
ylcellulose (HA-CMC) has been used for this purpose
in experimental and clinical conditions and has been
shown to be effective in decreasing postoperative
adhesions.2–4
Postoperative adhesions develop during the pro-
cess of repairing the damaged peritoneal surfaces.
Peritoneal injury associated with ischemia interferes
with fibrinolysis, and the tendency to adhesion for-
mation increases.6 It has been proposed that intraperi-
toneal adhesions develop in response to ischemic
conditions, and they function as a vascular graft to
protect marginally viable tissues.7 From this point of
530 ERTURK ET AL
view, especially after anastomosis of partially isch-
emic bowel ends, adhesion formation can be protec-
tive.6
The effect of HA-CMC on healing of bowel anasto-
mosis is controversial.3,4,8 However, we could not
find any study concerning the effect of HA-CMC on
ischemic bowel anastomosis. Therefore, we studied
the effects of an antiadhesive agent, HA-CMC, on
ischemic bowel anastomosis in rats.
MATERIALS AND METHODS
In this study, 30 female Wistar-Albino rats weighing
between 190 and 230 g were used. They were housed
in the Production and Experimental Research Center
of the Cerrahpasa Medical Faculty. All rats were
healthy and were kept in the same physical and en-
vironmental conditions. Throughout the period of the
study, they were kept in plastic cages and were fed
with standard laboratory diet and water ad libitum.
The institution’s guide for the care and use of labora-
tory animals was followed. The rats were divided into
three groups of ten animals each. Nonischemic co-
lonic anastomoses, ischemic colonic anastomoses,
and ischemic colonic anastomoses with HA-CMC ap-
plication were performed in Group A (control),
Group B, and Group C, respectively.
Technique
Rats were allowed to drink water until 12 hours
before the procedure. Operations were performed
under general anesthesia with 40 mg/kg intramuscu-
lar ketamine (Ketalar®, Parke Davis-Eczacibasi/Istan-
bul, Turkey). After shaving and cleansing of the an-
terior abdominal wall with 10 percent povidone-
iodine solution, the peritoneal cavity was opened by
a vertical midline incision.
In Group A, during preparation of the sigmoid
colon and mesentery, the vascular structures were
preserved. The sigmoid colon was transected hori-
zontally 3 cm proximal to the peritoneal reflection.
End-to-end anastomosis was performed with sepa-
rated atraumatic 5-0 silk sutures by a single-layer
technique. The abdominal wall was closed continu-
ously by use of 3-0 polyglactin for fascia and 3-0 silk
for skin.
In Group B, an ischemic colonic segment 4 cm in
length was prepared initially.9 For this purpose, mar-
ginal arteries and the vasa recti of the sigmoid colonic
segment were ligated separately with 4-0 atraumatic
Dis Colon Rectum, April 2003
silk sutures. The ischemic segment was cut horizon-
tally from the mid point, and then bowel continuity
was reestablished by the technique described in
Group A.
In Group C, immediately after transection and anas-
tomosis of the ischemic colonic segment, HA-CMC
film (Seprafilm®, Genzyme Corp., Cambridge, MA) 1
cm ⫻ 2 cm in size was wrapped around the anasto-
mosis. The abdominal wall was closed with the same
technique. This chosen size is sufficient to cover the
whole anastomotic line together with neighboring
ischemic bowel segments in rats. In clinical studies on
human subjects, a maximum of four Seprafilm® mem-
branes per patient were used intraperitoneally as an
antiadhesive barrier.10 Seprafilm® is commercially
available as a 12.7-cm ⫻ 15.2-cm (5-inch ⫻ 6-inch)
single-packaged membrane. When large-bowel anas-
tomosis is performed in humans, one Seprafilm mem-
brane is adequate to cover and protect the anasto-
motic region from adhesion formation.
Evaluation of Anastomotic Healing
Anastomotic Bursting Pressure. On the seventh
postoperative day, all rats were killed with a high
dose of ether anesthesia. HA-CMC film remains in
vivo as a mechanical barrier for approximately one
week. This period allows sufficient time for peritoneal
surfaces to heal without adhesion formation. During
this period, Seprafilm® exerts its functions almost
completely. After Day 7, it disappears from the peri-
toneal cavity and essentially has no additional effect
on adhesion formation. Thus, we killed rats on the
seventh postoperative day for evaluation. The ante-
rior abdominal wall containing the previous incisional
scar on the midline was everted like an inferiorly
based U-shaped flap, and the peritoneal cavity was
opened. The peritoneal cavity and site of the anasto-
mosis were observed macroscopically for complica-
tions, and adhesion scoring was done. Then, a co-
lonic segment 4 cm in length that contained the line of
anastomosis at the mid point was resected carefully
without destruction of any visceral adhesion. Colonic
content was gently cleaned manually. One end of the
colonic segment was ligated with silk. The other end
was connected to the system to measure the bursting
pressure by feeding tube. For this purpose, 1 tip of a
3-way stopcock was connected to the feeding tube,
the second tip was connected to a 50-ml syringe that
continuously insufflated the system at a constant rate,
and the third tip was attached to the manometer. The
Vol. 46, No. 4 HA-CMC IN ISCHEMIC COLONIC ANASTOMOSIS 531

resected colonic segment was totally submerged in RESULTS

water. The bursting pressure was recorded in milli-
grams of mercury when the first gas bubble appeared. Two rats from Group B and one rat from Group C
Hydroxyproline Level. After bursting pressure had died on the second and third postoperative days,
been measured, the bowel wall that contained the respectively. There were no abnormal macroscopic
line of anastomosis was excised 1 cm in length to findings at the autopsy of these rats. New rats were
determine the level of hydroxyproline (HP). The added to the groups to fulfill the original numbers.
specimens were preserved in physiologic saline at Although there were two perianastomotic abscesses
⫺80°C. After the samples had been thawed, dried, in groups B and C, anastomotic dehiscence was not
weighed, and homogenized separately, the HP con- observed in any rat.
tents were determined according to the method of HP values (mean ⫾ SD) were found to be 9.94 ⫾
Prochop and Kivirikko11 as mg/100 g of tissue. 1.79, 7.36 ⫾ 1.31, and 12.60 ⫾ 6.20 mg/100 g of tissue
in Group A, Group B, and Group C, respectively.
There was a statistically significant difference among
Adhesion Scores
HP levels of all groups by Kruskal-Wallis test (P ⫽
Adhesions were graded on a scale from zero to four 0.01). When the groups were compared with a post
as described by Nair et al12 (Table 1). The degree of hoc test, the HP level of Group B was lower than that
adhesion observed in each subject was recorded. On of Group A. This difference was found to be signifi-
Day 7, all rats in the 3 groups were killed, and adhe- cant (P ⫽ 0.02). The difference between Group B and
sion scores were studied in a blinded fashion to avoid Group C was also significant (P ⫽ 0.01). In Group C,
bias and to evaluate data objectively. We did not the HP level was higher than in Group A. However,
measure HP and bursting pressure values in a blinded this difference was not significant (P ⫽ 0.97). HP
fashion, because instrumental measurement was levels of the groups are shown in Table 2.
done objectively. Bursting pressures (mean ⫾ SD) were 220 ⫾ 15.63,
147.5 ⫾ 37.43, and 266 ⫾ 46.23 mmHg in Group A,
Group B, and Group C, respectively. There was a
Statistical Analysis highly significant difference among bursting pressure
levels of all groups by Kruskal-Wallis test (P ⬍ 0.001).
Nonparametric Kruskal-Wallis one-way analysis of
variance was used for statistical evaluation. Post hoc
Table 2.
tests used were Dunn (multiple comparisons) and HP Levels, Anastomotic Bursting Pressures, and
Dunnett (comparisons against a control group) for Adhesion Scores of Groups
Kruskal-Wallis. The UNISTAT威 5.0 statistical package
95% Confidence
for Windows™ (UNISTA Ltd., London, United King- Interval for Median
dom) was used for statistical analyses. Values of P ⬍ Parameters Median
0.05 and P ⬍ 0.01 were considered significant and Lower Upper
Bound Bound
highly significant, respectively.
HP level (mg/100 gr tissue)
Group A 9.5 8.3 11.5
Table 1. Group B 7.1 6.2 8.4
Adhesion Scoring System Group C 12.2 6.5 16.3
Anastomotic bursting pressure (mmHg)
Degree of
Findings Group A 218 210 240
Adhesion
Group B 163 110 175
0 No adhesion Group C 280 250 300
1 One adhesion band between the organs or Adhesion
one organ and peritoneum score
2 Two adhesion bands among the organs or Group A 0 0 1
one organ and peritoneum Group B 3 0 4
3 More than two adhesion bands among the Group C 1 0 2
organs or a mass of adhesion formed by HP ⫽ hydroxyproline; Group A ⫽ control group; Group
intestines not adhered to peritoneum B ⫽ ischemia group; Group C ⫽ hyaluronic acid-car-
4 Organs are adhered to peritoneum or boxymethyl cellulose group.
massive adhesions n ⫽ 10 in each group.
532 ERTURK ET AL
Bursting pressure of Group B was lower than that of
Group A, and in post hoc tests, the difference was
highly significant (P ⫽ 0.02). The value for Group C
was higher than that for Group A, but this difference
was not found to be significant (P ⫽ 0.13). A highly
significant difference was present between Groups B
and C (P ⬍ 0.001). Bursting pressure values of the
groups are shown in Table 2.
Adhesion scores (mean ⫾ SD) were 0.5 ⫾ 0.7071 in
Group A, 2.5 ⫾ 1.5092 in Group B, and 1.1 ⫾ 0.8756
in Group C. There was a highly significant difference
among the adhesion scores of all groups by Kruskal-
Wallis test (P ⫽ 0.01). When the groups were com-
pared with a post hoc test, the score for Group B was
higher than that for Group A, and there was a highly
significant difference between the two groups (P ⫽
0.01). The score for Group C was lower than that for
Group B, but the difference between these groups
was not significant (P ⫽ 0.16). Although the adhesion
score in Group C was higher than in Group A, the
difference between them was statistically not signifi-
cant (P ⫽ 0.41). Adhesion scores of the groups are
shown in Table 2.
DISCUSSION
Intraperitoneal adhesions occur in as many as 60 to
90 percent of patients undergoing abdominal sur-
gery.1,2,6 Although only a few of them lead to symp-
toms or clinical sequelae, they can cause important
postoperative complications, such as intestinal ob-
struction, fistula, chronic abdominal pain, infertility
in women, and difficulty in reoperative proce-
dures.1–4,6,13,14
Postoperative adhesions result from the healing
process of injured peritoneal surfaces. Adhesion for-
mation begins with development of fibrin matrix dur-
ing coagulation. Within a few days, this matrix is
gradually replaced by vascular granulation tissue. At
Day 5, most of the existing fibrin has been completely
removed from the environment.1 Fibrinolysis plays an
important role in the resolution of the inflammatory
exudates, thereby minimizing the risk of adhesion
formation. Adequate blood supply is essential for nor-
mal fibrinolysis. Peritoneal injury associated with isch-
emia interferes with fibrinolysis and leads to organi-
zation rather than resolution of the inflammatory
exudates and increases the tendency to adhesion for-
mation.6 Similarly, in the present study, peritoneal
adhesion in the ischemic group was significantly
higher than that in the control group, and the differ-
Dis Colon Rectum, April 2003
ence was significant (P ⫽ 0.01). It has been proposed
that intraperitoneal adhesions develop to function as
a vascular graft for ischemic tissues.7 Therefore, espe-
cially after anastomosis of ischemic bowels, adhe-
sions may play a protective role.6,15 In the present
study, however, despite the increased adhesions in
the ischemic group, they had no positive effect in the
anastomotic healing process. The differences in both
HP levels and bursting pressures between the control
and ischemic groups were found to be significant (P
⫽ 0.02). Therefore, there is a contradiction between
the present results and previous reports claiming peri-
toneal adhesions can be protective in ischemic bowel
anastomosis.
Various methods and agents have been tested to
prevent postoperative adhesion formation. One such
method is the intraperitoneal application of mechan-
ical barriers.4,5 Hyaluronic acid (HA) and carboxym-
ethylcellulose (CMC) are two such agents. HA is a
glucosaminoglycan that is found normally in vitreous
and synovial fluid, umbilical cord, skin, cartilage, and
ligamentous tissues.1,2 CMC, a nontoxic polysaccha-
ride made by monochloroacetate and cellulose, is
used in the cosmetic, food, and drug industries.3 HA
and CMC were combined into a single agent (HA-
CMC film) in an effort to create a biologically safe and
degradable substance that would prevent adhesions.1
It remains in vivo as a mechanical barrier for approx-
imately one week. This allows sufficient time for peri-
toneal surfaces to heal without adhesion formation.1,3
There are several experimental and clinical studies
with different results regarding the effects of HA-CMC
on prevention of adhesion formation.2–4,16–18 It has
been suggested that the effective mechanism of HA-
CMC is related to the separation of peritoneal surfaces
during epithelial regeneration.15,19,20 Another expla-
nation is that it may decrease the activity or prolifer-
ation of fibroblasts, prevent fibrin deposition on the
injured serosal surfaces, and inhibit movement of in-
flammatory cells and cellular elements during perito-
neal repair.20 In the present study, peritoneal adhe-
sions were decreased in the ischemic group by use of
HA-CMC, although the difference was not statistically
significant (P ⫽ 0.16). There was also no significant
difference in adhesion scores of the control and HA-
CMC groups (P ⫽ 0.41). The present results are in
accordance with relevant literature.
Peritoneal adhesions are seen during healing of
peritoneal surfaces, and the cascade of events is sim-
ilar to the stages seen in the early periods of wound
healing.1,14 Likewise, there is a similarity between
Vol. 46, No. 4 HA-CMC IN ISCHEMIC COLONIC ANASTOMOSIS
healing of bowel anastomosis and wound healing
seen in other organs. Agents that prevent adhesion
formation during the inflammatory period can also
adversely affect healing of bowel anastomosis. Effects
of HA-CMC on bowel anastomosis have been studied,
and different results have been reported. Bowers et
al.4 observed that among previously irradiated rats
undergoing small-bowel resection and anastomosis,
HA-CMC film was associated with a greatly increased
rate of abscess formation at the operative site. Felton
et al.15 and Talbert et al.21 observed an increased rate
of leakage from the anastomotic line in rats to which
CMC had been applied. Similarly, Uzunkoy et al.8
indicated that CMC had adverse effects on intestinal
anastomosis in rats. However, in an experimental
study by Medina et al.,3 HA-CMC did not prevent
anastomotic healing of the large bowel in a rabbit
model. The results of Buckenmaier et al.22 and van
Oosterom et al.23 with HA-CMC application were sim-
ilar to those of Medina et al.3 In another experimental
study, it was demonstrated that application of N,O-
carboxymethyl chitosan (NOCC), which has structural
similarities to HA, did not decrease the strength of a
large-bowel anastomosis in the rat.14 In the present
study, decreased levels of HP and bursting pressure in
the ischemic group were significantly increased after
application of HA-CMC (P ⫽ 0.01 and P ⬍ 0.001,
respectively).
CONCLUSIONS
Potential beneficial effects of intraperitoneal adhe-
sions on ischemic bowel anastomosis were not ob-
served in the present study. In contrast, adverse ef-
fects of ischemia dominated. Furthermore, adverse
effects of ischemia on colonic anastomosis were
counteracted by HA-CMC application.
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