Regulation of extracellular matrix RNA levels in cultured smooth muscle cells: Relationship to cellular quiescence

Abstract

We have examined how the alpha 2(I), alpha 1(III), alpha 1(IV), and alpha 2(V) collagen genes as well as the thrombospondin and fibronectin genes are regulated by conditions that modulate the proliferation of smooth muscle cells in vitro. We find that smooth muscle cells (SMC) arrested by low serum containing media continue to express similar or slightly lower levels of mRNA for all the genes studied except type III collagen which exhibited a 5-fold increase. By contrast, when SMC are density-arrested, a coordinate increase in mRNA levels for all the extracellular matrix genes studied is observed. The increases range from approximately 1.5-fold for fibronectin to 5-fold for type III collagen. The mRNA levels for beta-tubulin and vimentin, however, are decreased in these density-arrested cells. Interestingly, the time course of increase for type IV collagen and fibronectin can be distinguished from the other extracellular matrix genes and best correlates with an increase in cell-cell contact. The increase in steady-state RNA levels for alpha 1(IV) collagen is not shared by fibroblasts similarly treated and may provide a useful marker for SMC in culture. Heparin, a potent inhibitor of SMC growth, increased the level of type III collagen mRNA about 2.5-fold and also increased the level of alpha 2(V) collagen and fibronectin. Transforming growth factor-beta is found to be a cell density-dependent modulator of SMC growth. Surprisingly, transforming growth factor-beta had little effect on most of the extracellular matrix genes studied. However, transforming growth factor-beta treatment of SMC increased thrombospondin mRNA levels about 4-fold, whereas type IV collagen mRNA exhibited a 2-fold increase. We conclude that type III collagen mRNA levels are positively and tightly coupled to SMC quiescence. The effect of cellular quiescence on other extracellular matrix mRNAs is complex but, in general, positive. This is particularly evident in SMC arrested by high cell density.