Effect of Compound 21, a Selective Angiotensin II Type 2 Receptor Agonist, in an Abdominal Adhesion Murine Model
Colton Boudreau, MSc1, Courtney Jones, BSc1, Alison Gareau, PhD2, Terry Levatte, BSc1, Stephanie Legere, MSc1, Michael Bezuhly, MD, MSc, SM, FRCSC, FAAP1.
1Dalhousie University, Halifax, NS, Canada, 2Calgary Lab Services, Calgary, AB, Canada.
Purpose: Abdominal adhesions are fibrous bands that form in response to surgical trauma, and connect visceral and/or peritoneal surfaces. Adhesions occur in over 90% of patients post-laparotomy and can lead to serious long-term complications, including infertility, small bowel obstruction and perforation, and chronic pain. This study uses a murine model of induced abdominal adhesions to study the anti-fibrotic effect of a novel selective angiotensin II type 2 receptor agonist, compound 21 (C21), in reducing abdominal adhesion formation.
Methods: The effects of C21 in vivo were assessed using a cecal abrasion model in female BALB/c mice. A laparotomy was performed and the cecum and overlying parietal peritoneum was abraded with fine grit sandpaper. Mice were divided into systemic (oral gavage) or local (intraperitoneal injection) treatment groups and were treated with C21 (10 μg/kg) or saline (vehicle control) daily for 7 days. Mice were sacrificed 8 days post-surgery and adhesions were graded by a blinded observer. Peritoneal fluid was obtained at time of sacrifice and ELISA used to quantify TGFβ levels. Laparotomy incisions were excised and CD31, CD68, and αSMA immunostaining, and picrosirius red staining were performed to assess surgical laparotomy incisional wound properties. To study the in vitro effects of C21, parietal peritoneal fibroblasts and visceral mesothelial cells were isolated and scratch wound assays performed using C21 (10 μM), angiotensin II (AngII, 1 μM), or both.
Results: Post-operative C21 administration, both systemically and locally, greatly reduced the formation of abdominal adhesions in vivo. Additionally, TGFβ in peritoneal fluid was reduced in mice treated with C21. Histological analysis of surgical incisions revealed no statistical difference in the number of CD31+ vessels or CD68+ cells. Expression of αSMA was reduced in C21-treated animals. Picrosirius red staining revealed no difference in collagen I and collagen III distribution in laparotomy scars between control and C21-treated animals. Laparotomy scar collagen density and surrounding dermis thickness was not significantly different between treatment groups. Cell migration of isolated parietal peritoneal fibroblasts and visceral mesothelial cells in vitro was markedly reduced in the presence of C21 compared to control or AngII.
Conclusions: C21 markedly reduced or completely prevented adhesion formation both with local and systemic adminstration. These findings may be attributed to decreased levels of pro-fibrotic TGFβ in vivo and decreased cell migration of parietal peritoneal fibroblasts and visceral mesothelial cell migration in the presence of C21. Importantly, C21 did not appear to have histologically quantifiable effects on laparotomy wounds compared to controls. This study suggests that C21 could reduce abdominal adhesions without impeding laparotomy wound healing.
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