The importance of the interaction between hepatocyte and hepatic stellate cells in fibrogenesis induced by fatty accumulation.

Non-alcoholic fatty liver disease is characterized by an initial accumulation of triglycerides that can progress to non-alcoholic steatohepatitis, which can ultimately evolve to cirrhosis and hepatocellular carcinoma. Hepatic stellate cells play a key role in liver fibrogenesis by an increased activation and an altered profile of genes involved in the turnover of extracellular matrix components. To reproduce in-vitro the functional cell connections observed in vivo it is essential to consider cell-to-cell proximity and interaction. The aim of this study was to determine the response to free fatty acids in a simultaneous co-culture model of hepatocytes and hepatic stellate cells. Simultaneous co-culture model and monoculture of each cell type (control) were exposed to FFA for 24 up to 144 h. Quantification of steatosis; stellate cell activation; assessment of fibrogenic response; expression and activity of metalloproteinases as well as collagen biosynthesis were evaluated. Free fatty acids induced comparable steatosis in simultaneous co-culture and monoculture. However, the activation of the stellate cells assessed by alpha-smooth muscle actin expression is greater when cells were in close contact. Furthermore, a time-dependent increment of tissue inhibitor metalloproteinase-2 protein was observed, which was inversely correlated with protein expression and activity of matrix-metalloproteinases, suggesting enhanced collagen biosynthesis. This behavior was absent in cell monoculture. These data indicate that cell-to-cell proximity between hepatocytes and stellate cells is necessary for the initiation of the fibrotic process.