Collagen Directly Stimulates Bladder Smooth Muscle Cell Growth In Vitro: Regulation by Extracellular Regulated Mitogen Activated Protein Kinase

Abstract

Bladders clinically subjected to excessive pressure or distention demonstrate an altered extracellular matrix (ECM) composition. We determined how an altered collagen substratum might affect bladder smooth muscle cell (bSMC) growth in vitro and probed the mechanism of this response. Primary culture rat bSMCs were seeded onto culture plates pre-coated with normal type I collagen (NC) or heat denatured type I collagen (DNC) under standard culture conditions. In separate experiments bSMCs from the 2 substrates were enzymatically released and changed to growth on normal collagen (NC-->NC or DNC-->NC) or denatured collagen (DNC-->DNC or NC-->DNC). At 24 hours proliferation was assessed by 3H-thymidine incorporation. Statistical significance in triplicate wells was determined by ANOVA. The proliferation of bSMCs on DNC was 5-fold greater than on NC (p <0.0001). Passage onto damaged collagen (DNC-->DNC) showed 2-fold further augmentation in proliferation (p <0.0001) but only a 50% decrease when NC was reintroduced (DNC-->NC) (p <0.001). Conversely replating on NC (NC-->NC) generated a 33% decrease in the already low proliferation rate (p <0.001) but 9-fold stimulation of proliferation when changed to damaged ECM (NC-->DNC) (p <0.0001). The mitogenic effect of damaged ECM on bSMC growth was abolished by specific inhibition of extracellular regulated kinase mitogen activated protein kinase signaling using PD98059. Damaged type I collagen (ECM) is mitogenic to bSMCs. The response is amplified by re-exposure to DNC. However, mitogenicity is only partially reversible by re-introducing NC. These results demonstrate striking bSMC responsiveness to ECM conformation. Signaling through the extracellular regulated kinase mitogen activated protein kinase pathway supports bSMC-ECM interaction. We speculate that remodeling the ECM in vivo may regulate bSMC growth.