Effect of Trichostatin a on the expression of connexins in organotypical hepatocyte cultures.

Loss of intercellular communication is a major cause of dedifferentiation in cultured he-patocytes. Direct intercellular communication is mediated by gap junctions, which are composed of two connexin hexameric structures. Expression of connexins occurs in a cell-specific manner, and is shown to be cell cycle related, as a reduction of Cx32, the major liver connexin, is observed during the G1/S transition. Eukaryotic gene expression is mainly controlled by transcription factor binding. Recently, structural modifications, such as reversible histone acetylation, have been shown to play a role in gene regulation as well. Histone hyperacetylation is related with transcriptional activation. This process depends on a equilibrum between histone acetyltranferase (HAT) and histone deacetylase (HDAC). Inhibition of the latter is correlated with cell growth arrest and induction of dif-ferentiation and/or apoptosis. Trichostatin A (TSA) is a reversible and potent HDAC in-hibitor, which acts by inducing p53-mediated p21 gene expression. Since p21 is involved in controlling the G1/S cell cycle transition, TSA is thought to increase Cx32 expression. The in vitro effect of TSA on gap junctional intercellular communication in organotypical hepatocyte cultures (monolayer-, collagengel- en co-culture) is examined at the activity level (scrape-loading/dye transfer assay), at the translational level (Western blotting, im-munohistochemistry, HPLC) and at the transcriptional level (Northern blotting, PCR). Upregulation of intercellular communication provides a promising strategy to reduce de-differentiation of cultured hepatocytes. This contributes to further optimization of liver-based in vitro systems for pharmaco-toxicological research.