Characterization of the control of intracellular (Ca2+) and the contractile phenotype of cultured human detrusor smooth muscle cells.

摘要

PURPOSE: We measured the functional properties of cultured human detrusor myocytes with respect to their ability to regulate their intracellular [Ca2+] and generate force in collagen matrices. MATERIALS AND METHODS: Human detrusor biopsies were dissociated into single cells by collagenase treatment and used immediately or cultured in D-valine medium and subsequently used after culture trypsinization. Intracellular [Ca2+] was measured in Fura-2 loaded myocytes. Cell force development was measured by incorporating cells into a collagen gel and attaching it to an isometric strain gauge. RESULTS: Carbachol was equally effective in generating Ca transients in freshly isolated and cultured cells. Carbachol potency (pEC50) and the magnitude of Ca2+ transients were similar. Adenosine triphosphate potency was decreased in cultured cells and Ca2+ transients showed properties consistent with a purinoceptor shift from a purinergic subtype. Temporal restitution of Ca2+ transients was similar in the 2 groups, indicative of retained intracellular Ca2+ stores in cultured cells. Cultured cells (approximately 10(6)) embedded in collagen gel generated a force about 10 times greater than that generated by gel alone. The cell dependent force could be further increased by adding carbachol. CONCLUSIONS: Cultured cells retain the ability to generate agonist induced intracellular Ca2+ transients. There was no evidence that the cell culture altered the properties of muscarinic receptors, although purinoceptor mediated properties were altered. Restitution experiments indicated that functional intracellular Ca2+ stores were retained in cultured cells. Cultured cells also retained a contractile phenotype, especially in response to carbachol. The magnitude of force was attenuated, which may be a function of the biomechanical properties of the gel used to embed the cells.