Co-culture of Hepatocytes,Kuffer Cell of SD Rat in Vitro / 昆明医科大学学报

ABSTRACT

Objective Through co-culture Kupffer cells with hepatocyte in vitro,to understand the influence of Kupffer cells on the growth、morphology and metabolism of hepatocyte.Methods In-situ IV collagenase two-step perfusion method to digest SD rat liver,apply low speed centrifugalization to isolate parenchymal hepatic cells and densitygradient centrifugation by percoll fluid to isolate Kupffer cells respectively.Inoculate hepatocyte cells into 6 holes culture plate to culture alone and/or co-culture with Kupffer cells in the proportion six to one,the growth,morphous of hepatocyte are observed under the light microscope and the level of albumin and glucose in the culture supernatant are detected by automatic biochemistry meter,and compare synthesis function and metabolism of hepatocytes culture alone and coculture with Kupffer cells.Results The hepatocytes of culture alone their growth and developed to normal hepatocytes morphous quickly,2 weeks later the cells death occurs,after 21 days the hepatocytes culture alone are totaly death,when hepatocytes co-culture with kupffer cells that proliferate slower than that culture alone.The hepatocites beginning to death 48 hours later, and totally death after 10 days co-culture.The culture supernatant was collected and tested the level of albumin and glucose at 24 hours intervals.In culture alone group,the albumin level is significant higher than in co-culture group at 48h、96h、120h、144h、168h(P