Morphology and function of primarily cultured rat hepatocytes

AIM: To observe the process of the alteration of cell morphology and function in primarily cultured rat hepatocytes and to investigate their optimal functional condition for hepatocyte transplantation and bioartificial liver support system. METHODS: Rat hepatocytes were isolated by the modified two step method described by Seglen. The yield and viability were assessed by trypan blue exclusion. Hepatocytes were inoculated in the culture medium consisting of Williams’E supplemented with insulin,glycogen, transferrin,dexamethasone,nicotinamide and 10 g·L -1 fetal bovine serum. The morphologic changes of cultured hepatocytes were observed; the ultrastructural study was conducted and the concentrations of albumin, urea and LDH (lactate dehydrogenase) in the supernatant in different cultural period were examined. RESULTS: The average yield of rat hepatocytes was 1.21×10 8 cells per rat with an average viability of 93.6%. Hepatocytes were observed under light microscope, polygon hepatocytes,some of which were binuclear, were aggregated and arranged in groups or cords. Under electron microscope, abundance of organelles and particular ultrastructure of hepatocytes were found. Our data showed that LDH leakage, albumin synthesis and urea level had fluctuating changes in one week,but the third day found less LDH leakage and higher albumin synthesis and urea level. CONCLUSION: Primarily cultured rat hepatocytes function well on the third day, which may be the optimal time for transplantation and bioartificial liver support system.