Establishment of Primary Cell Culture from Stria Vascularis Explants: Morphological and Functional Characterization

Abstract

Klm H-N, Chang MS, Chung MH, Park K. Establishment of primary cell culture from stria vascularis explants. Morphological and functional characterization. Acta Otolaryngol (Stockh) 1996; 116: 805-811.

To provide the prerequisite for long-term study of the inner ear related to structural and functional integrity, tissue of stria vascularis with spiral ligament was isolated from Wistar rat cochleas and cultured using the explant-culture technique. the following culture media were used: EMEM with Hepes buffer, hydrocortisone (400 ng/ml), transferrin (5 μg/ml), triiodothyronine (10⁻⁹ M), cholera toxin (10⁻¹⁰ M), insulin (5 μg/ml), and epidermal growth factor (10 ng/ml). to characterize the cells growing out from the explant, immunofluorescence with cytokeratin (cytokeratin 18) and ultrastrucural examination with SEM and TEM were performed. the marginal cell function was investigated by expression of Na⁺, K⁺-ATPase antisera against β2 subunit of rat Na ⁺, K⁺-ATPase and P-NPPase. We were able to maintain the cultured cells for 3 weeks or more. Monolayered marginal cells were observed beyond 14 days in vitro and the expression of cytokeratin 18 was especially enhanced. the cultured marginal cells were almost identical to in vivo cells both as regards ultrastructural features and Na⁺, K⁺-ATPase activity. the present results suggest that the primary explant culture technique is a reliable in vitro model of strial marginal cells. However, establishment of the' cell line is needed for long-term study.

Key Words:

• strial marginal cells
• explant culture technique
• cytokeratin 18. ullrastructures Na⁺K⁺-ATPase