Establishing primary cultures of vascular smooth muscle cells from the spiral modiolar artery.

Objective: This article is reporting a method for establishment primary cultures of vascular smooth muscle cells (VSMCs) from the spiral modiolar artery (SMA).

Methods: VSMCs were isolated from guinea pig SMAs. Arterial tissues were cut and enzymatically digested at 37 °C for 20 min using a 0.1% trypsin solution. After digestion, tissue fragments were explanted in a 35-mm culture dish. Contaminated fibroblasts were separated from VSMCs because of their different adhesion abilities. The cells migrated from the explants within 7-10 days and grew to confluence in approximately 4 weeks.

Results: We obtained pure and viable VSMCs from the confluent third passage. The morphological and immunofluorescence analyses demonstrated a "hill-and-valley" growth pattern that is characteristics of VSMCs, and the expression of cell type-specific markers (α-smooth muscle actin and myosin), respectively. The change of intracellular calcium concentration induced by angiotensin II and CaCl(2) showed that the VSMCs had good cell viability.

Conclusion: We obtained purified VSMCs using this method. All cell cultures expressed smooth muscle markers (α-SM actin, and myosin) and were negative for vWF. This article provides a simple method to obtain VSMCs for in vitro studies of physiology and pathophysiology in the circulation disturbances of the inner ear. In addition, VSMCs are regarded to be an excellent model to evaluate drugs in vitro.