Cultivation and identification of vascular smooth muscle cells from spiral modiolar artery.

Objective:To establish a method for primary cultures of vascular smooth muscle cells(VSMCs) from the spiral modiolar artery(SMA) by combining two common cultural methods in vitro.Methods:VSMCs were isolated from SMAs of guinea pigs.The arterial tissues were minced and enzymatically digested at 37℃ for 20 min using a trypsin solution(0.1%).After digestion,the fragments were explanted in a 35-mm culture dish.Contaminated fibroblasts were separated from VSMCs due to their different adhesion abilities.Results:The cells migrated from the explants within 7-10 d and grew to confluence in approximately3 weeks.Pure and viable VSMCs were obtained from the confluent third passage.The cells obtained based on this protocol were subsequently characterized by morphology,immunofluorescence and transmission electron microscopy analysis.The results demonstrated typical characteristics of VSMCs,including a "hill-and-valley" growth pattern and the expression of cell type-specific markers(α-smooth muscle actin and myosin) by morphological and immunofluorescence analysis,respectively.Conclusion:The multitude and purified cells obtained from this method could be a model to investigate the physiological mechanisms of VSMCs in the circulation disturbances of the inner ear.In addition,VSMCs were regarded to be an excellent model system for the evaluation of drugs in vitro.