Study Of The Role And Underlying Mechanisms Of Rho/Rho-kinase Signaling Pathway On Acute Necrosis Pancreatitis In Rats

The pathogenesis of acute necrosis pancreatitis (ANP) remains unclear.Premature activation of pancreatic enzymes, dysfunction of pancreatic microcirc-ulation, excessive activation of inflammatory cytokines, and the noxious role ofoxygen free radicals,etc.,seem to be among its pathogenic mechanisms. Recentstudies indicate that Rho/Rho kinase signaling pathway as an important molecularswitche involves in various biological behavior in vivo, such as the regulation ofpancreatic enzyme secretion and the formation of oxygen free radicals. Inhibit Rhokinase by Y-27662has been widely used to study and clarify the biological role ofRho/Rho kinase signaling pathway in vivo.There is no report regarding the functionof the Rho/Rho kinase signaling pathway in ANP domainly.Objective: To investigate the effect and mechanism of inhibiting the Rho/Rhokinase signaling pathway on ANP in rats.Methods:24healthy male Sprague-Dawley rats(240-280g weight) wererandomly divided into three groups: ANP (group A), pretreatmant with Rho/Rhokinase inhibitor Y-27632group (group B), saline control group (group C).In group A,the model of ANP induced by the retrograde injection of3.5%sodiumtaurocholate(1ml/kg) in the pancreatic duct. In group B, pretreatment withintra-abdominal injection of Y-27632(5mg/kg) in30minutes preoperative,otherprocessing the same to group A. In group C saline retrograde into the pancreatic duct.Blood was collected from the tail vein at postoperative4h, the animals weresacrificed at24h.Observations:(1)The macroscopic change of pancreaticinflammation;(2)The change of the serum amylase;(3)The change of the serumsuperoxide dismutase (SOD) and maleic dialdehyde (MDA);(4)The change ofpancreatic tissue myeloperoxidase (MPO);(5)The histopathological change ofpancreas；(6)The immunohistochemical detection of Rho-associated protein kinase I(ROCK I). Result:(1)Pancreatic macroscopic change: In group A,pancreas was glay, matt,hyperemia edema, with large areas of necrosis, and pancreatic surface gastroepiploicvisible saponification spots, intraperitoneal with bloody ascites.In group B, pancreasbecame dark red, matt, hyperemia edema, with little areas of necrosis, and littlesaponification spots, clear intraperitoneal ascites.In group C, pancreas appeareduniformly of pink, good gloss, no edema, no or little clear intraperitoneal ascites, nosaponification spots.(2)The changes of the serum amylase: At4h,24h observation point, comparedwith the group C(902.25±92.02,646.75±102.81),serum amylase was significantlyincreased in group A (3747.38±256.90,2933.75±181.78) and group B(1694.25±326.58,1323.25±313.53),the difference has statistics significance (P <0.01).The difference of group A and group B also has statistics significance (P <0.01).(3)Compared with C group,the levels of serum SOD was significantlyincreased and serum MDA was significantly decreased in group A(P <0.01). Thedifference of group A and group B has statistics significance (P <0.01).(4)The MPO activity of pancreatic tissue was significantly elevated in ANP rat,Y-27632inhibition of Rho/Rho kinase signaling pathway can reduce the elevationof MPO.Compared with erch group,the MPO activity of pancreatic tissue hasstatistics significance (P <0.01).(5)Pancreatic histopathological injury score: Pancreatic histopathological injuryscore in group A was increased significantly as compared with in group C(P <0.01).The difference between group A and group B was significant (P <0.05)(6)Expression of pancreatic ROCK I protein: Group A ROCK I showed strongpositive expression of brown, group B ROCK I showed positive expression of paleyellow, group C ROCK I showed weakly positive expression of pale yellow. ROCKI semi-quantitative analysis, each group difference has statistics significance (P <0.05)Conclusion:(1)Y-27632could inhibits Rho/Rho kinase signaling pathway,reduces the expression of ROCK1protein in pancreatic tissue in the ANP rats.(2)Inhibiton of Rho/Rho kinase signaling pathway can reduce the degree ofpathological damage and inflammatory cell infiltration in pancreatictissue,suggesting that it has a protective effect to ANP.(3) Its mechanisms may beassociated with inhibition of pancreatic enzymes activation, reduction of oxygen freeradical formation, reduction of the outcom of inflammory factor and inflammatorycascade effect.