OBJECTIVETo explore whether the function of the CL-1 hepatocytes, co-cultured with human hepatic stellate cells (HSC) on microcarriers was better than that cultured without HSC.

METHODSCL-1 hepatocytes were divided into 2 groups. The co-culture group was cultured with HSC in DMEM culture medium containing 10% fetal bovine serum, and HSC were not added in single-culture group. The cytomorphology was observed by inverted microscope and scanning electron microscopy. The effects of different culture method on the proliferation in vitro were analyzed by MTT assay. The function of hepatocytes was evaluated through measuring the concentration of ALT and albumin.

RESULTSThe inverted microscope and the MTT staining results showed that the quantity and viability of the human hepatocyte (C3A) in bidirectional bioreactor group were much better than the RCMW group. The growth curve results showed that the density of the human hepatocyte (C3A) was firstly increased and then decreased in both groups, and the peak of the curve appeared in day 5. The density of human liver cells in the second generation of bi-directional rotation and perfusion microgravity bioreactor group was significantly higher than the RCMW group from day1 to day 7 (P<0.05). The functional results showed that the albumin and urea concentration, which reached the peak on day 5, also gone up firstly and then gradually gone down in both teams. And the albumin and urea concentration in bidirectional bioreactor group was significantly higher than RCMW group from day 1 to day7 (P<0.01). Besides, the concentration of ALT and AST in bidirectional bioreactor group was significantly lower than RCMW group from day 1 to day 7 (P<0.05).

CONCLUSIONThis study shows that this new culture method is advantageous in enhancing cell viability and function. It indicates that co-culturing hepatocytes with HSC has a good application prospect in the development of artificial liver technology.