| [Background] Acute pancreatitis(AP) is a clinical common disease, the main cause of AP is biliary disease in China. However, in recent years, the incidence of AP increased year by year because the living standards and lifestyle of people changed, about 25% of AP could develop into severe acute pancreatitis(SAP). While the pathogenesis of SAP is still unclear, and no specific medicine in clinical treatment of SAP at present. Chinese medicine QingYi Grain(QYG) can improve the clinical symptoms, shorten the disease course, and reduce the complications for patients, which showes a good using prospect. QYG is a new formulation of QingYi Decoction. Therefore, to explore the therapeutical mechanism of QYG in SAP is necessary, which will contribute to study the pathogenesis of SAP.[Objective]: To investigate the protective mechanism and the influence of QYG on aquaporin 4(AQP-4) expressed in pancreas and kidney of rats with SAP. The rats were treated with Chinese medicine QYG by gavage, and the study will provide a theoretical basis for QYG father research in the future.[Methods] Eighty Sprague-Dawley(SD) rats were randomly divided into sham-operation(SO) groups, SAP groups and QYG groups, and 24 rats in each group. Eight rats in normal control group. The SAP model of rats were induced by injecting 5% sodium taurocholate(STC)(0.1ml/100g) retrogradely though pancreatic duct. Experimental methods:①The pathological changes of pancreas and kidney were observed by HE staining,②The ratio of dry weight /wet weight(D/W) of kidney and the renal index of rats were tested by electronic scale,③The level of amylase(AMS) in serum were tested by artificial iodine colorimetry, total superoxide dismutase(SOD) in serum were examined by hydroxylamine colorimetric, malondialdehyde(MDA) in serum were examined by thiobarbituric acid(TBA) assay.④The level of interleukin-1(IL-1), interleukin-6(IL-6) and tumor necrosis factor-α(TNF-α) in serum were tested by ELISA kits,⑤The expression of AQP-4 protein in pancreas and kidney of rats were detected by immunohistochemistry and western blot, and the AQP-4-mRNA were tested by Reverse Transcription Polymerase Chain Reaction(RT-PCR).[Results]①The ratio of renal D/W weight in SAP groups were smaller than that in SO groups(P<0.05), which was improved after QYG treatment, there was not statistically significant in comparing with that in SO groups at 72h.②Renal index of rats in SAP groups were smaller than that in SO groups and QYG groups, but there were no statistics significant difference among three groups.③The levels of AMS and MDA in serum were significantly higher in SAP groups and QYG groups than that in SO groups, however, AMS was reduced significantly after QYG intervention for 24h and 48h(P<0.05 ), but there was no obviously difference compared with that in SO groups at 72h(P>0.05). MDA was lower significantly than that in SAP groups(P<0.05), and shew a fall-off trend. Total content of SOD in SAP groups and QYG groups were less than that in SO groups, which was increased after QYG treatment for 48h (P<0.05).④The levels of IL-1, IL-6 and TNF-αin serum in SAP groups and QYG groups were higher than that in SO groups at each time point, which reached to the peak amplitude at 48h in SAP groups. The levels of IL-1, IL-6 and TNF-αin serum were dropped significantly after QYG treatment for 72h(P<0.05). There were positive correlation among IL-1, IL-6 and TNF-α(r=0.984, r=0.972, r=0.966), and the effect of QYG exhibited a time-dependence relationship.⑤No pathological changes was observed in pancreas and kidney of rats in SO groups. However, different degrees of edema and exudation, the inflammatory cell infiltration have happened in pancreatic and renal tissues of rats in SAP groups, partial pancreatic or renal tissues had happened hemorrhage and necrosis, the structure of which had changed into blur, and even into a slice. The pathological damages of pancreatic and renal tissues were improved after QYG treament, the pathological scores in QYG groups were lower-down(P<0.05).⑥AQP-4 protein stainning cell haven't been seen in pancreas of rats in all groups, while different extents of AQP-4 protein positive staining cells have been observed in kidney of rats in all groups, and the content of AQP-4 protein expressiones were correlated to the severity of rats with SAP. AQP-4 protein expression was decreased generally in kidney after QYG intervention, the same change tendency of AQP-4 protein and its mRNA expression in kidney happened both in western blot and RT-PCR.⑦The statistical correlation analysis showed a positive correlation between AQP-4 protein expression and inflammatory mediators.⑧QYG displayed a time dependence and positive correlation effect in treatment of rats with SAP.[Conclusions]①AMS rised obviously in serum of rats with SAP, the result indicated STC induced SAP model of rats were succesful, and QYG can reduce AMS production in serum.②QingYi Grains(QYG) can effectively reduce the pathological damages of pancreas and kidney.③The levels of AMS and MDA and SOD in serum of rats with SAP were improved by QYG, which evaluated the antioxidant factors and the ability of scavenging oxygen free radicals. Therefore, we supposed that antioxidation maybe one of the treatment mechanisms of QYG in SAP.④AQP-4 protein expressed in pancreas and kidney of rats was different, almost none AQP-4 protein was discovered in pancreas, but different levels of AQP-4 protein expressed in kidney of rats.⑤The contents of IL-1, IL-6 and TNF-αin serum of rats with SAP was increased gradually, which shew a positive correlation when analyzed with AQP-4 protein products expressed in kidney. QYG decreased the levels of IL-1, IL-6 and TNF-αin serum, and downregulated AQP-4 prorein expression in kidney, we presumed that QYG protected renal injury of rats with SAP though inhibiting the inflammatory response, and downregulating aquaporin 4 expression in kidney. QYG mitigated renal edema and secondary pathological changes, reduced complications and shortened the course of SAP. |
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