Study On The Mechanism Of Gli2 Mediated Inflammatory Cytokines In Acute Pancreatitis In Mice

objective: Hedgehog signaling pathway plays an important role in acute pancreatitis,but its downstream cytokines and specific mechanisms are not very clear.The aim of experiment was to select cytokines which changed obviously from the cell array in the model of caerulein-induced acute pancreatitis and verified these cytokines in vivo and in vitro.Methods: Thirty C57 mice were randomly divided into three groups: control pancreas group,acute pancreatitis group and inhibited pancreas group.The model of acute pancreatitis in mice was induced by caerulein,then uesd HE stain and serum amylase test to verify the model of success.RT-qPCR,Western blot and immunohistochemistry were used to detect the expression of Shh and Gli2 in the Hedgehog signaling pathway in acute pancreatitis.we started experiment in vitro,the mouse pancreatic acinar cells 266-6 were cultured in vitro.Afer overexpres Gli2,cytokine array was used for screen out cytokines which changed obviously and then these cytokines were tested by RT-qPCR in vitro.Then validation in vivo,next used Gant61(Gli family specific inhibitor)to inhibit Gli2,then took the mouse serum for Elisa detection to test the selected cytokines.Results:(1)compared with the control pancreas group,the levels of amylase in the acute pancreatitis group and the inhibited pancreas group were significantly higher(P<0.05)and the HE staining showed pancreatic acinar cells edema and volume increased.(2)compared with the control pancreas group,the expression of Shh and Gli2 in each tissue(pancreas,lung,liver,intestine and kidney)in acute pancreatitis group was significantly increased(P < 0.05),and Gli2 was mainly expressed in the cytoplasm of cells.(3)After overexpress Gli2 in vitro,IFN-?,Fasl,IL-6,G-csf,Cxcl9 and Tnfr1 a were selected by cell array.RT-qPCR test found that in overexpress Gli2 group IFN-?,Fasl and IL-6 decreased,G-csf,Cxcl9 and Tnfr1 a increased compared with vector group.(4)In reverse validation,after inhibition of Gli2 by Gant61,these selected cytokines were detected with Elisa.The expirement found that the expression of IFN-?,Fasl and IL-6 in inhibited pancreas group increased significantly compared with the control pancreas group,acute pancreatitis group(P<0.05)and the expression of G-csf,Cxcl9 and Tnfr1 a in inhibited pancreas group decreased significantly compared with the control pancreas group,acute pancreatitis group(P<0.05).Conclusin:(1)The Shh-Gli2 signal axis participates in the damage of the pancreas,lung,liver,intestine and kidney in acute pancreatitis induced by caerulein in mice;(2)Gli2 is involved in the regulatory process of acute pancreatitis by mediated cytokines IFN-?,Fasl,IL-6,G-csf,Cxcl9 and Tnfr1a.