Pyruvate-induced long-term maintenance of glutathione S-transferase in rathepatocyte cultures

The addition of pyruvate to the culture medium has been reported to improve the maintenance of P450-dependent enzyme expression in primary rat hepatoc yte cultures. In this study, the effects of 30mM pyruvate on cell morpholog y, albumin secretion and glutathione S-transferase (GST) expression were in vestigated as a function of the time in culture. The effect of triiodothyro nine (T3) exposure on GST expression was also measured in pyruvate-treated cultures. Transmission electron microscopy showed that untreated hepatocyte s deteriorated after culture for 7 days, whereas the morphology of the pyru vate-treated cells was similar to that observed in intact liver tissue. The albumin secretion rate was significantly higher in rat hepatocytes exposed to pyruvate than in control cells. In the presence of pyruvate, mu and alp ha class GST activities were well maintained, whereas GST pi activity was i ncreased over the entire culture period. HPLC analysis revealed that the co mplement of GST subunits present in hepatocytes is altered during culture w ith pyruvate: mu class proteins remained relatively constant, whereas a dec rease in the alpha class content was accompanied by a strong increase in GS T subunit P1 (GSTP1). The induction of GSTP1 was confirmed at the mRNA leve l. In control cultures, pi class GST activity was increased, but total, mu, and alpha class GST ctivities continuously declined as a function of cultu re time and became undetectable beyond 7 days in culture. At the protein an d mRNA levels, a much smaller increase in GSTP1 was observed than in the py ruvate cultures. When the pyruvate-treated cell cultures were exposed to T3 , an inhibitory effect on GST activities and proteins was found. These resu lts indicate that this simple culture model could be useful for studying th e expression and regulation of GST.