Abstract

Non-alcoholic fatty liver, as the initial link in the development of non-alcoholic steatohepatitis, its mechanism of occurrence and development is of great significance for the treatment of clinical non-alcoholic fatty liver disease. The aim of this study was to establish a stable and reliable co-culture model of primary mature adipocytes and hepatocytes in non-alcoholic fatty liver model rats for in-depth study of adipocytes-hepatocyte interaction, revealing the pathophysiological mechanism of adipocyte effects on hepatocyte steatosis lays a pharmacological research foundation. 8 w old male Sprague-Dawley rats were randomly divided into control group and non-alcoholic fatty liver group. Control group was given normal diet and non-alcoholic fatty liver group was given high fat diet. After 6 w of continuous feeding, hematoxylin and eosin, oil red O staining and non-alcoholic fatty liver disease activity score were performed on liver tissue sections. Serum triglyceride, total cholesterol, alanine aminotransferase and aspartate aminotransferase contents were measured by kits to verify non-alcoholic fatty liver rat model which was successfully established. Rat primary mature adipocytes and hepatocytes were extracted and differentiated by collagenase in situ perfusion and collagenase digestion, and the co-culture system was constructed by transwell chambers. Oil red O staining and triglyceride kit were used to detect lipid accumulation in primary hepatocytes before and after co-culture; quantitative real-time polymerase chain reaction and Western blot were used to detect the expression levels of p38 mitogen-activated protein kinase, phosphorylated p38 mitogen activated protein kinases and aquaporin 9 in primary hepatocytes before and after co-culture for 48 h. The co-culture model of primary mature adipocytes and hepatocytes in non-alcoholic fatty liver model rats was successfully established and the co-culture of mature adipocytes in non-alcoholic fatty liver pathological state may be involved in hepatocyte steatosis by up-regulating the expression of aquaporin 9 through activating p38 mitogen activated protein kinase pathway.