The Study On The Function And Molecular Mechanism Of Down-regulation Of Aquaporin 10 In Cholestasis

Background&PurposeCholestasis(cholestasis)is a complex clinical syndrome caused by abnormal bile secretion and excretion,including abnormal bile acid synthesis and abnormal expression of bile acid transporter.Hepatic fibrosis and cirrhosis may occur along with the development of the disease.Even the phenomenon of liver failure.In the past research,people have made great progress in the molecular mechanism of cholestasis,but the research subjects mostly focus on the bile acid transporter and bile acid synthesis key protein,and the water with the highest proportion in bile and transport.There are few studies on the function and mechanism of channel proteins in water in cholestasis.Aquaporin 10(AQP10)is a member of the water channel family,and AQP3,AQP7,AQP8,and AQP9 belong to the water-glycerol channel subfamily.These aquaporins transport glycerol in addition to transporting water molecules.Urea,also plays an important role in the metabolism of fat.In previous studies,AQP10 expression was found to be associated with abnormalities in fat metabolism and some herpes-like skin diseases and abnormal expression of AQP10 in the duodenum and jejunum[1],adipose tissue and skin[2].However,the role of AQP10 in cholestasis has not been studied.In this paper,we aimed to study the expression changes of aquaporin 10(AQP10)in the liver of patients with cholestasis,the mechanism of action of this effect and its related functional studies.Method1.we determined the expression of AQP10 in hepatocytes of patients with cholestasis by Western Blot(WB),reverse transcription-PCR(RT-qPCR),and immunofluorescence(IF).Secondly,according to the patient’s serum biochemical index data,it is analyzed whether the indicators are related to the expression level of AQP10.2.A cell model of in vitro cholestasis PLC/PRF/5-ASBT stable cell line(ASBT)was constructed using dual luciferase gene reporter and chromatin immunoprecipitation(Chromatin Immunoprecipitation,ChIP)studies the mechanism by which this effect occurs.3.The AQP10 was overexpressed in PLC/PRF/5-ASBT cells and the intracellular bile acid content was measured after treatment with conjuated bile acids.Result1.The expression of AQP10 in liver cells of patients with cholestasis is reduced by one time compared with the control group.Total bile acid content in serum and liver tissue of patients with cholestasis was negatively correlated with the expression of AQP10 in liver tissue of patients.2.Taurocholic acid(TCA)activates signaling pathway.The dual luciferase gene report demonstrates that the binding site of nuclear factor Kappa B(NF-κB)to the AQP10 promoter is located at-434 bp/-228 bp of the AQP10 promoter.ChIP results showed that NF-κB directly regulate the expression of AQP.3.After treated with the same concentration of bile acid,the total bile acids content of PLC/PRF/5-ASBT&AQP10 cells were significantly lower than that of PLC/PRF/5－ASBT cells.ConclusionIn the case of cholestasis,conjugated bile acids can activate the NF-,kB signaling pathway,directly down-regulate the expression of AQP10 in the liver,and reduce the water transported into the liver cells via the AQP10 protein,allowing bile acids to accumulate in the liver cells.To sum up,we found that when bile stasis occurs,bile acid can activate the NF-κB signaling pathway and down-regulate the expression of AQP10 in the liver.After the down-regulation of AQP10 expression,the amount of water that can enter the liver cells through AQP10 protein decreases,aggravating the deposition of bile acid in the liver cells and increasing the liver damage caused by bile stasis.