Adaptation of Murine Inner Medullary Collecting Duct (IMCD3) Cell Cultures to Hypertonicity
Recently, we have adapted IMCD3 cell cultures to survive under increasing hypertonic conditions (i.e., 600 and 900 mOsmol/kg H2O). In adapted cells, ATPase activity is increased by one order of magnitude, while the expression of the ? and ? subunit is increased by a factor of 4 to 5 over controls (300 mOsmol/kg H2O). Corresponding increases in mRNAs were also detected. The ? subunit has been described as being uniquely expressed in some areas of the kidney, but never in cell cultures (even those derived from kidney tissues). However, the ? subunit was detected at the protein and mRNA levels in the adapted IMCD3 cells. In contrast to the ? and ? subunits, the levels of ? protein and mRNA expression continue to increase as a function of the media ion concentration. We have also demonstrated that signaling pathways that upregulate the ?, ?, and ? subunits are very different. Increasing concentrations of the PI3 kinase inhibitor, LY294002, resulted in a dose-dependent reduction in the expression of the ? subunit, with total abolition at 10 ?M. However, LY294002 had no significant effect on the expression of the ? subunit. Inhibition of the JNK2 (but not of the JNK1) pathways by dominant negative transfections abolished the upregulation of the ?, but not the a subunit. Failure to upregulate the expression of the ? subunit was associated with a marked decrease in cell viability upon stress.
signaling pathways cell adaptation ? subunit regulation