Effect of bevacizumab on postoperative adhesion formation in a rat uterine horn adhesion model and the correlation with vascular endothelial growth factor and Ki-67 immunopositivity

We investigated whether adhesion formation is prevented by the inhibition of angiogenesis in a rat uterine horn adhesion model. A single-dose of bevacizumab was effective in preventing postoperative intraperitoneal adhesion formation among 30 Wistar albino rats that underwent serosal injury by use of a standard technique after laparotomy with intraperitoneal bevacizumab. We investigated whether adhesion formation is prevented by the inhibition of angiogenesis in a rat uterine horn adhesion model. A single-dose of bevacizumab was effective in preventing postoperative intraperitoneal adhesion formation among 30 Wistar albino rats that underwent serosal injury by use of a standard technique after laparotomy with intraperitoneal bevacizumab. The development of peritoneal adhesions after abdominal and pelvic surgery leads to clinical problems, including intestinal obstruction, chronic abdominal pain, infertility, and chronic pelvic pain (1Aritaş Y. Akcan A. Erdogan A.R. Akgün H. Saraymen R. Akyildiz H. Effects of melatonin and phospholipid on adhesion formation and correlation with vascular endothelial growth factor expression in rats.Ulus Travma Acil Cerrahi Derg. 2009; 15: 416-422PubMed Google Scholar). Studies have reported postoperative intra-abdominal adhesions in 50% to 95% of women who undergo gynecologic surgery (2Hellebrekers B.W.J. Trimbos-Kemper G.C.M. Blitterswijk V.C.A. Bakkum E.A. Trimbos J.B.M.Z. Effects of five different barrier materials on postsurgical adhesion formation in the rat.Hum Reprod. 2000; 15: 1358-1363Crossref PubMed Scopus (117) Google Scholar). The injuries have been attributed to various causes, including mechanical trauma, ischemia at suture sites, ischemia caused by electrocautery, foreign bodies, tissue desiccation, and infection (3Ferland R. Mulani D. Campbell P.K. Evaluation of a sprayable polyethylene glycol adhesion barrier in a porcine efficacy model.Hum Reprod. 2001; 16: 2718-2723Crossref PubMed Scopus (75) Google Scholar). Postoperative inflammation develops after tissue injury. When the fibrin gel structure that is produced during wound healing is not broken down by fibrinolytic activity, permanent fibrous tissue forms and adhesion develops (4Barbul A. Wound healing.in: Brunicardi F.C. Andersen D.K. Billiar T.R. Dunn D.L. Hunter J.G. Pollock R.E. Schwartz’s principles of surgery. 8th ed. McGraw-Hill, Philadelphia2005: 223-248Google Scholar). Vascular endothelial growth factor (VEGF) expression increases during wound healing and angiogenesis, and is necessary for adhesion formation (5Mitchell R.N. Wound healing.in: Kumar V. Cotran R.S. Robbins S.L. Robbins basic pathology. 7th ed. W.B. Saunders, Philadelphia2003: 61-78Google Scholar, 6Beddy D. Watson R.W.G. Fitzpatrick J.M. O’Connell P.R. Increased vascular endothelial growth factor production in fibroblasts isolated from strictures in patients with Crohn’s disease.Br J Surg. 2004; 91: 72-77Crossref PubMed Scopus (61) Google Scholar). Adhesion formation may, therefore, be prevented by inhibiting angiogenic activity (7Chiang S.C. Cheng C.H. Moulton K.S. Kasznica J.M. Moulton S.L. TNP-470 inhibits intraabdominal adhesion formation.J Pediatr Surg. 2000; 35: 189-196Abstract Full Text Full Text PDF PubMed Scopus (33) Google Scholar, 8Greene A.K. Alwayn I.P. Nose V. Flynn E. Sampson D. Zurakowsky D. et al.Prevention of intra-abdominal adhesions using the antiangiogenic COX-2 inhibitor celecoxib.Ann Surg. 2005; 242: 140-146Crossref PubMed Scopus (76) Google Scholar, 9Ferrara N. Henzel W.J. Pituitary follicular cells secrete a novel heparin-binding growth factor specific for vascular endothelial cells.Biochem Biophys Res Commun. 1989; 161: 851-858Crossref PubMed Scopus (2000) Google Scholar).Bevacizumab (Avastin; Genentech/Roche, San Francisco, CA) is a full-length recombinant humanized monoclonal antibody that inhibits VEGF (10Hurwitz H. Fehrenbacher L. Novotny W. Cartwright T. Hainsworth J. Heim W. et al.Bevacizumab plus irinotecan, fluorouracil, and leucovorin for metastatic colorectal cancer.N Engl J Med. 2004; 350: 2335-2342Crossref PubMed Scopus (9100) Google Scholar, 11Moshfeghi A.A. Rosenfeld P.J. Puliafito C.A. Michels S. Marcus E.N. Lenchus J.D. Venkatraman A.S. Systemic bevacizumab (Avastin) therapy for neovascular age-related macular degeneration: twenty-four week results of an uncontrolled open-label clinical study.Ophthalmology. 2006; 113 (2002.e1–12)Abstract Full Text Full Text PDF PubMed Scopus (198) Google Scholar, 12Spaide R.F. Laud K. Fine H.F. Klancnik Jr., J.M. Meyerle C.B. Yannuzzi L.A. et al.Intravitreal bevacizumab treatment of choroidal neovascularization secondary to age-related macular degeneration.Retina. 2006; 26: 383-390Crossref PubMed Scopus (595) Google Scholar). Given the apparent close association between VEGF and inflammation, bevacizumab inhibition of VEGF might influence or possibly inhibit the inflammation and wound healing involved in postoperative adhesion formation.The antigen Ki-67 is a nuclear protein used as a marker of cell proliferation (13Karak A.K. Sarkar C. Chumber S. Tandon N. MIB-1 proliferative index in parathyroid adenoma and hyperplasia.Indian J Med Res. 1997; 105: 235-238PubMed Google Scholar). Recent work has shown good correlation between the expression of Ki-67 and the number of mitotic cells (14Ueda T. Aozasa K. Tsujimoto M. Ohsawa M. Uchida A. Aoki Y. et al.Prognostic significance of ki-67 immunoreactivity in soft tissue sarcomas.Cancer. 1989; 63: 1607-1611Crossref PubMed Scopus (181) Google Scholar).Our study was approved by the institutional review board of the Ankara Educating and Research Hospital in Ankara, Turkey, and was performed at the hospital’s Animal Research Center (IP: 0378, tarih: 14.07.2010). The institutional review board’s guidelines for animal care and use were followed. Thirty mature, nonpregnant female Wistar albino rats (aged 10 to 12 weeks) were used as a model for experimental induction of postoperative intra-abdominal and uterine horn adhesions. The rats were randomly divided into three groups before surgery, with 10 animals in each group.The animals were anesthetized using ketamine hydrochloride (40 mg/kg intramuscularly) and xylazine (2 mg/kg intramuscularly). The adhesion model of Basbug et al. (15Basbug M. Aygen E. Tayyar M. Kaya E. Narin F. Oktem O. Hyaluronic acid plus heparin for improved efficacy in prevention of adhesion formation in rat uterine horn model.Eur J Obstet Gynecol Reprod. 1998; 78: 109-112Abstract Full Text Full Text PDF PubMed Scopus (27) Google Scholar) was followed. A syringe of 1 mL of Ringer’s lactate (RL) containing 5 IU and 7.5 IU of bevacizumab was injected intraperitoneally into the uterine horns (0.5 mL into each uterine horn) of groups 1 and 2, respectively. Only 1 mL of RL was injected intraperitoneally into the uterine horn of the control group (16Kim T.I. Chung J.L. Hong J.P. Min K. Seo K.Y. Kim E.K. Bevacizumab application delays epithelial healing in rabbit cornea.Invest Ophthalmol Visual Sci. 2009; 50: 4653-4659Crossref PubMed Scopus (77) Google Scholar).The midline incision was closed within 10 minutes of the completion of the procedure. One rat in the control group died during the study. After a 3-week recovery period, the rats were killed, and the adhesion areas were evaluated and graded. Tissue samples were taken from all serosal surfaces where adhesion had developed.The investigators scoring the adhesions were blinded to which group the rats belonged. The extent and severity of the adhesions were evaluated using an established scoring system (17Linsky C.B. Diamond M.P. Cunnigham T. Constantine B. DeCherney A.H. Di Zerega G.S. Adhesion reduction in a rabbit uterine horn model using an absorbable barrier TC-7.J Reprod Med. 1987; 32: 17-20PubMed Google Scholar). The severity of the adhesions was measured as follows: 0 = no resistance to separation, 0.5 = some resistance, 1 = sharp dissection required. The total grade was additive, yielding a range of adhesion scores from 0 to 4, which represented both extent and severity.Samples from the adhesion areas of the formaldehyde-fixed specimens and the paraffin-embedded tissue blocks were removed after staining with dye. Sections 4 μm thick were cut and stained with hematoxylin and eosin according to standard laboratory procedures. The histopathologic adhesion classification system of I to IV of Zühlke et al. (18Zühlke H.V. Lorenz E.M. Straub E.M. Sawas V. Pathophysiology and classification of adhesions [article in German].Langenbecks Arch Chir Suppl II Verh Dtsch Ges Chir. 1990; : 1009-1016PubMed Google Scholar) was employed.For Ki-67 (cat. no. RM-9106-S; Labvision/NeoMarkers Corp., Fremont CA) and VEGF (cat. no. RB-222-P; Labvision/NeoMarkers), staining procedures were performed according to the manufacturers’ instructions. The Ki-67 index was expressed as a percentage value of the number of labeled cells in a region containing at least 100 cells. All slides were evaluated with a Leica DMI 4000 B light microscope (Leica, Wetzlar, Germany). Among the nuclei cells positive for Ki-67, 0 to 5 cells were negative and 5 to 25 cells were positive.To prevent interindividual bias, all tissues were evaluated by the same histologist (M.C.), who was blinded to the origin of the samples. The relative intensity of immunoreactivity staining was assessed quantitatively using the process previously described by McCarty et al. (19McCarty Jr., K.S. Miller L.S. Cox E.B. Konrath J. KS Sr, McCarty Estrogen receptor analyses: correlation of biochemical and immunohistochemical methods using monoclonal antireceptor antibodies.Arch Pathol Lab Med. 1985; 109: 716-721PubMed Google Scholar), which takes into account both the intensity and the distribution of specific staining (0 = no expression, 1 = mild, 2 = moderate, and 3 = intense) (20Budwit-Novotny D.A. McCarty K.S. Cox E.B. Soper J.T. Mutch D.G. Creasman W.T. et al.Immunohistochemical analyses of estrogen receptor in endometrial adenocarcinoma using a monoclonal antibody.Cancer Res. 1986; 46: 5419-5425PubMed Google Scholar).Statistical data analysis was performed using SPSS for Windows release 11.5 (SPSS, Inc., Chicago, IL). Kruskal-Wallis, one-way analysis of variance (ANOVA), Mann-Whitney U, the chi-square test, and Bonferroni-corrected were applied. When correction was necessary, alpha was set at 0.0167.Adhesions were not present in eight of the rats in group 2 (80%). In the two rats in which adhesions developed, the adhesion was between the omentum and the incision line. All of the animals in the control group exhibited adhesions between the omentum and the uterine horns or the incision line. Adhesion formation was statistically significantly lower in the bevacizumab-treated groups than in the control group (P<.05). The total adhesion score in group 2 was statistically significantly lower compared with that of the control group (P=.000). However, there was no statistically significant difference in the total adhesion scores of group 1 and group 2. The adhesion severity scores were compared separately between the groups. The severity scores of the treatment groups were statistically significantly less than the control group (P<.05). In group 2, the severity score (median = 0.0, min-max = 0–0) was much lower compared with the control group (median = 0.7, min-max = 0.5–1.0) and group 1 (median = 0.5, min-max = 0–1) (Table 1).Table 1Macroscopic scoring of adhesion formation in the rat uterine horn model comparing two different-dose bevacizumab-treated rats and control group 17Linsky C.B. Diamond M.P. Cunnigham T. Constantine B. DeCherney A.H. Di Zerega G.S. Adhesion reduction in a rabbit uterine horn model using an absorbable barrier TC-7.J Reprod Med. 1987; 32: 17-20PubMed Google Scholar, 22Rose B.I. MacNeill C. Larrain R. Kopreski M.M. Abdominal instillation of high-molecular-weight dextran or lactated Ringer’s solution after laparoscopic surgery: a randomized comparison of the effect on weight change.J Reprod Med. 1991; 36: 537-539PubMed Google Scholar, 26di Zerega G.S. Verco S.J.S. Young P. Kettel M. Kobak W. Martin D. et al.A randomized, controlled pilot study of the safety and efficacy of 4% icodextrin solution in the reduction of adhesions following laparoscopic gynecological surgery.Hum Reprod. 2002; 17: 1031-1038Crossref PubMed Scopus (174) Google Scholar.GroupNo.Mean ± standard deviationMedianP valueFor all groupsaKruskal-Wallis one-way analysis of variance (ANOVA) test (α = 0.05).Group 1 and group 2bMann-Whitney U test corrected with Bonferroni (α = 0.0167).Group 1 and group 3bMann-Whitney U test corrected with Bonferroni (α = 0.0167).Group 2 and group 3bMann-Whitney U test corrected with Bonferroni (α = 0.0167).Group 1: Bevacizumab, 5 IU (1.25 mg/1 mL of Ringer’s lactate)101.75 ± 1.042.12< .001.011.04< .001Group 2: Bevacizumab, 7.5 IU (1.875 mg/1 mL of Ringer’s lactate)100.65 ± 0.330.50Group 3: Control, 1 mL of Ringer’s lactate9cOne rat died.2.83 ± 1.093.0a Kruskal-Wallis one-way analysis of variance (ANOVA) test (α = 0.05).b Mann-Whitney U test corrected with Bonferroni (α = 0.0167).c One rat died. Open table in a new tab The median histologic adhesion score was 3 (2 to 4), 1 (0 to 3), and 1 (0 to 2) in the control group, group 1, and group 2, respectively. The difference between the control and the treatment groups was statistically significant (P<.05). In the treatment groups, there were no statistically significant differences in the histopathologic adhesion scores (P>.05). The adhesion areas contained dense collagen fibers and were infiltrated with lymphocytes and mast cells. The adhesion areas also contained foreign body granulomas with giant cells containing highly birefringent material in polarized light. In cases of mild fibrosis, only subtle changes were observed, with interposition of abdominal fat tissue between the serosal surfaces and small areas of fibrosis with minimal or no granulation tissue.The Ki-67 staining values of the adhesion areas were statistically significantly higher in the control group compared with those in the treatment groups (P<.05).The difference between the Kİ-67 staining values in group 1 and group 2 was not statistically significant (P>.05).Staining for VEGF was statistically significantly higher in adhesion areas of the control group compared with the other groups (P<.05). There was no difference between the VEGF staining values of group 1 and group 2.The adhesions occur as a result of damage to the mesothelial cells on the serosal surface; the cell damage is followed by inflammation, the release of exudates, and the formation of a soft fibrin gel matrix. If this soft fibrin is not fractioned and removed by fibrinolytic activity, dense fibrinous and vascular changes develop within 2 weeks (4Barbul A. Wound healing.in: Brunicardi F.C. Andersen D.K. Billiar T.R. Dunn D.L. Hunter J.G. Pollock R.E. Schwartz’s principles of surgery. 8th ed. McGraw-Hill, Philadelphia2005: 223-248Google Scholar). Our follow-up period of 3 weeks was sufficient to identify adhesion maturation. As bevacizumab remains in the circulation for up to 6 weeks, it also allowed enough time for adhesion maturation to be affected by the reabsorbed drug (21Ignjatovic D. Aasland K. Pettersen M. Sund S. Chen Y. Spasojevic M. Nesgaard J.M. Intra-abdominal administration of bevacizumab diminishes intra-peritoneal adhesions.Am J Surg. 2010; 200: 270-275Abstract Full Text Full Text PDF PubMed Scopus (31) Google Scholar). Various approaches have been adopted to treat adhesion formation, such as intra-abdominal fluid injection, which has been used to ensure the separation of the deperitonealized areas (22Rose B.I. MacNeill C. Larrain R. Kopreski M.M. Abdominal instillation of high-molecular-weight dextran or lactated Ringer’s solution after laparoscopic surgery: a randomized comparison of the effect on weight change.J Reprod Med. 1991; 36: 537-539PubMed Google Scholar, 23Verco S.J. Peers E.M. Brown C.B. Rodgers K.E. Roda N. di Zerega G. Development of a novel glucose polymer solution (icodextrin) for adhesion prevention: pre-clinical studies.Hum Reprod. 2000; 15: 1764-1772Crossref PubMed Scopus (108) Google Scholar, 24Wiseman D.M. Trout J.R. Diamond M.P. The rates of adhesion development and the effects of crystalloid solutions on adhesion development in pelvic surgery.Fertil Steril. 1998; 70: 702-711Abstract Full Text Full Text PDF PubMed Scopus (122) Google Scholar, 25Metwally M. Watson A. Lilford R. 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In recent studies of fibrinolytics, including streptokinase and recombinant human tissue plasminogen-activating factor, statins and cyclooxygenase-2 inhibitors prevented postoperative adhesion (3Ferland R. Mulani D. Campbell P.K. Evaluation of a sprayable polyethylene glycol adhesion barrier in a porcine efficacy model.Hum Reprod. 2001; 16: 2718-2723Crossref PubMed Scopus (75) Google Scholar, 32Hellebrekers B.W. Trimbos-Kemper T.C. Trimbos J.B. Emeis J.J. Kooistra T. Use of fibrinolytic agents in the prevention of postoperative adhesion formation.Fertil Steril. 2000; 74: 203-212Abstract Full Text Full Text PDF PubMed Scopus (184) Google Scholar, 33Aarons C.B. Cohen P.A. Gower A. Reed K.L. Leeman S.E. Stucchi A.F. Becker J.M. 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Vascular endothelial growth factor is a heparin-binding glycoprotein, which plays an important role in angiogenesis; and VEGFR-2 is the primary mediator of growth and permeability in endothelial cells (36Shalaby F. Rossant J. Yamaguchi T.P. Gertsenstein M. Wu X.F. Breitman M.L. Schuh A.C. Failure of blood-island formation and vasculogenesis in Flk-1-deficient mice.Nature. 1995; 376: 62-66Crossref PubMed Scopus (3332) Google Scholar, 37Carmeliet P. Ferreira V. Breier G. Pollefeyt S. Kieckens L. Gertsenstein M. et al.Abnormal blood vessel development and lethality in embryos lacking a single VEGF allele.Nature. 1996; 380: 435-439Crossref PubMed Scopus (3427) Google Scholar, 38Gerber H.P. Hillan K.J. Ryan A.M. Kowalski J. Keller G.A. Rangell L. et al.VEGF is required for growth and survival in neonatal mice.Development. 1999; 126: 1149-1159Crossref PubMed Google Scholar). Also, VEGF is released from injured tissues and aids tissue growth and vascular remodeling. The recent studies demonstrated the expression of VEGF in the endothelium, fibroblasts of pelvic adhesions in females (39Wiczyk H.P. Grow D.R. Adams L.A. O’Shea D.L. Reece M.T. Pelvic adhesions contain sex steroid receptors and produce angiogenesis growth factors.Fertil Steril. 1998; 69: 511-516Abstract Full Text Full Text PDF PubMed Google Scholar, 40Rout U.K. Oommen K. Diamond M.P. Altered expressions of VEGF mRNA splice variants during progression of uterine peritoneal adhesions in the rat.Am J Reprod Immunol. 2000; 43: 299-304Crossref PubMed Scopus (45) Google Scholar, 41Diamond M.P. El-Hammady E. Munkarah A. Bieber E.J. Saed G. Modulation of the expression of vascular endothelial growth factor in human fibroblasts.Fertil Steril. 2005; 83: 405-409Abstract Full Text Full Text PDF PubMed Scopus (54) Google Scholar). In another study, fewer adhesions developed in mice injected with an anti-VEGF antibody (42Saltzman A.K. Olson T.A. Mohanraj D. Carson L.F. Ramakrishnan S. Prevention of postoperative adhesions by an antibody to vascular permeability factor/vascular endothelial growth factor in a murine model.Am J Obstet Gynecol. 1996; 174: 1502-1506Abstract Full Text Full Text PDF PubMed Scopus (42) Google Scholar, 43Kim S. Lee S. Greene A.K. Arsenault D.A. Le H. Meisel J. et al.Inhibition of intra-abdominal adhesion formation with the angiogenesis inhibitor sunitinib.J Surg Res. 2008; 149: 115-119Abstract Full Text Full Text PDF PubMed Scopus (25) Google Scholar). Ignjatovic et al. (21Ignjatovic D. Aasland K. Pettersen M. Sund S. Chen Y. Spasojevic M. Nesgaard J.M. Intra-abdominal administration of bevacizumab diminishes intra-peritoneal adhesions.Am J Surg. 2010; 200: 270-275Abstract Full Text Full Text PDF PubMed Scopus (31) Google Scholar) reported that a single dose of bevacizumab administered intraperitoneally statistically significantly reduced the grade and severity of intra-abdominal adhesions in a rat model.In common with other studies, we clearly demonstrated the preventive effect of bevacizumab against adhesion formation (16Kim T.I. Chung J.L. Hong J.P. Min K. Seo K.Y. Kim E.K. Bevacizumab application delays epithelial healing in rabbit cornea.Invest Ophthalmol Visual Sci. 2009; 50: 4653-4659Crossref PubMed Scopus (77) Google Scholar, 44Nomoto H. Shiraga F. Kuno N. Kimura E. Fujii S. Shinomiya K. et al.Pharmacokinetics of bevacizumab after topical, subconjunctival, and intravitreal administration in rabbits.Invest Ophthalmol Vis Sci. 2009; 50: 4807-4813Crossref PubMed Scopus (209) Google Scholar, 45Fernando N.H. Hurwitz H.I. Targeted therapy of colorectal cancer: clinical experience with bevacizumab.Oncologist. 2004; 9: 11-18Crossref PubMed Scopus (86) Google Scholar). We showed that bevacizumab prevents adhesion formation via the inhibition of VEGF. The use of a VEGF receptor inhibitor would make up-regulation ineffective and prevent adhesion formation. Our finding that the VEGF staining scores of the treatment group were much lower than in the control group correlates well the findings of previous studies (21Ignjatovic D. Aasland K. Pettersen M. Sund S. Chen Y. Spasojevic M. Nesgaard J.M. Intra-abdominal administration of bevacizumab diminishes intra-peritoneal adhesions.Am J Surg. 2010; 200: 270-275Abstract Full Text Full Text PDF PubMed Scopus (31) Google Scholar, 39Wiczyk H.P. Grow D.R. Adams L.A. O’Shea D.L. Reece M.T. Pelvic adhesions contain sex steroid receptors and produce angiogenesis growth factors.Fertil Steril. 1998; 69: 511-516Abstract Full Text Full Text PDF PubMed Google Scholar, 40Rout U.K. Oommen K. Diamond M.P. Altered expressions of VEGF mRNA splice variants during progression of uterine peritoneal adhesions in the rat.Am J Reprod Immunol. 2000; 43: 299-304Crossref PubMed Scopus (45) Google Scholar, 41Diamond M.P. El-Hammady E. Munkarah A. Bieber E.J. Saed G. Modulation of the expression of vascular endothelial growth factor in human fibroblasts.Fertil Steril. 2005; 83: 405-409Abstract Full Text Full Text PDF PubMed Scopus (54) Google Scholar, 42Saltzman A.K. Olson T.A. Mohanraj D. Carson L.F. Ramakrishnan S. Prevention of postoperative adhesions by an antibody to vascular permeability factor/vascular endothelial growth factor in a murine model.Am J Obstet Gynecol. 1996; 174: 1502-1506Abstract Full Text Full Text PDF PubMed Scopus (42) Google Scholar, 43Kim S. Lee S. Greene A.K. Arsenault D.A. Le H. Meisel J. et al.Inhibition of intra-abdominal adhesion formation with the angiogenesis inhibitor sunitinib.J Surg Res. 2008; 149: 115-119Abstract Full Text Full Text PDF PubMed Scopus (25) Google Scholar). We found that bevacizumab decreased the number and percentage of cells positive for Ki-67 in a concentration-dependent manner and that the KI-67 staining scores of the treatment groups were statistically significantly lower than the control group. These data correlate with the findings of a similar experimental study (16Kim T.I. Chung J.L. Hong J.P. Min K. Seo K.Y. Kim E.K. Bevacizumab application delays epithelial healing in rabbit cornea.Invest Ophthalmol Visual Sci. 2009; 50: 4653-4659Crossref PubMed Scopus (77) Google Scholar). The development of peritoneal adhesions after abdominal and pelvic surgery leads to clinical problems, including intestinal obstruction, chronic abdominal pain, infertility, and chronic pelvic pain (1Aritaş Y. Akcan A. Erdogan A.R. Akgün H. Saraymen R. Akyildiz H. Effects of melatonin and phospholipid on adhesion formation and correlation with vascular endothelial growth factor expression in rats.Ulus Travma Acil Cerrahi Derg. 2009; 15: 416-422PubMed Google Scholar). Studies have reported postoperative intra-abdominal adhesions in 50% to 95% of women who undergo gynecologic surgery (2Hellebrekers B.W.J. Trimbos-Kemper G.C.M. Blitterswijk V.C.A. Bakkum E.A. Trimbos J.B.M.Z. Effects of five different barrier materials on postsurgical adhesion formation in the rat.Hum Reprod. 2000; 15: 1358-1363Crossref PubMed Scopus (117) Google Scholar). The injuries have been attributed to various causes, including mechanical trauma, ischemia at suture sites, ischemia caused by electrocautery, foreign bodies, tissue desiccation, and infection (3Ferland R. Mulani D. Campbell P.K. Evaluation of a sprayable polyethylene glycol adhesion barrier in a porcine efficacy model.Hum Reprod. 2001; 16: 2718-2723Crossref PubMed Scopus (75) Google Scholar). Postoperative inflammation develops after tissue injury. When the fibrin gel structure that is produced during wound healing is not broken down by fibrinolytic activity, permanent fibrous tissue forms and adhesion develops (4Barbul A. Wound healing.in: Brunicardi F.C. Andersen D.K. Billiar T.R. Dunn D.L. Hunter J.G. Pollock R.E. Schwartz’s principles of surgery. 8th ed. McGraw-Hill, Philadelphia2005: 223-248Google Scholar). Vascular endothelial growth factor (VEGF) expression increases during wound healing and angiogenesis, and is necessary for adhesion formation (5Mitchell R.N. Wound healing.in: Kumar V. Cotran R.S. Robbins S.L. Robbins basic pathology. 7th ed. W.B. Saunders, Philadelphia2003: 61-78Google Scholar, 6Beddy D. Watson R.W.G. Fitzpatrick J.M. O’Connell P.R. Increased vascular endothelial growth factor production in fibroblasts isolated from strictures in patients with Crohn’s disease.Br J Surg. 2004; 91: 72-77Crossref PubMed Scopus (61) Google Scholar). Adhesion formation may, therefore, be prevented by inhibiting angiogenic activity (7Chiang S.C. Cheng C.H. Moulton K.S. Kasznica J.M. Moulton S.L. TNP-470 inhibits intraabdominal adhesion formation.J Pediatr Surg. 2000; 35: 189-196Abstract Full Text Full Text PDF PubMed Scopus (33) Google Scholar, 8Greene A.K. Alwayn I.P. Nose V. Flynn E. Sampson D. Zurakowsky D. et al.Prevention of intra-abdominal adhesions using the antiangiogenic COX-2 inhibitor celecoxib.Ann Surg. 2005; 242: 140-146Crossref PubMed Scopus (76) Google Scholar, 9Ferrara N. Henzel W.J. Pituitary follicular cells secrete a novel heparin-binding growth factor specific for vascular endothelial cells.Biochem Biophys Res Commun. 1989; 161: 851-858Crossref PubMed Scopus (2000) Google Scholar). Bevacizumab (Avastin; Genentech/Roche, San Francisco, CA) is a full-length recombinant humanized monoclonal antibody that inhibits VEGF (10Hurwitz H. Fehrenbacher L. Novotny W. Cartwright T. Hainsworth J. Heim W. et al.Bevacizumab plus irinotecan, fluorouracil, and leucovorin for metastatic colorectal cancer.N Engl J Med. 2004; 350: 2335-2342Crossref PubMed Scopus (9100) Google Scholar, 11Moshfeghi A.A. Rosenfeld P.J. Puliafito C.A. Michels S. Marcus E.N. Lenchus J.D. Venkatraman A.S. Systemic bevacizumab (Avastin) therapy for neovascular age-related macular degeneration: twenty-four week results of an uncontrolled open-label clinical study.Ophthalmology. 2006; 113 (2002.e1–12)Abstract Full Text Full Text PDF PubMed Scopus (198) Google Scholar, 12Spaide R.F. Laud K. Fine H.F. Klancnik Jr., J.M. Meyerle C.B. Yannuzzi L.A. et al.Intravitreal bevacizumab treatment of choroidal neovascularization seco