title: Isolation and culture of endothelial cells, pericytes and perivascular resident macrophage-like melanocytes from the young mouse ear.
abstract: This protocol describes a growth medium-based approach for obtaining cochlear endothelial cells (ECs), pericytes (PCs) and perivascular resident macrophage-like melanocytes (PVM/Ms) from the stria vascularis of mice aged between P10 and P15 (P, postnatal day). The procedure does not involve mechanical or enzymatic digestion of the sample tissue. Explants of stria vascularis, 'mini-chips', are selectively cultured in growth medium, and primary cell lines are obtained in 7-10 d. The method is simple and reliable, and it provides high-quality ECs, PVM/Ms and PCs with a purity >90% after two passages. This protocol is suitable for producing primary culture cells from organs and tissues of small volume and high anatomical complexity, such as the inner ear capillaries. The highly purified primary cell lines enable cell culture-based in vitro modeling of cell-cell interactions, barrier control function and drug action.
authors.affiliation: Oregon Hearing Research Center, Department of Otolaryngology/Head and Neck Surgery, Oregon Health & Science University, Portland, Oregon, USA.
authors.forename: Lingling
authors.initials: L
authors.lastname: Neng
authors.forename: Wenjing
authors.initials: W
authors.lastname: Zhang
authors.forename: Ahmed
authors.initials: A
authors.lastname: Hassan
authors.forename: Marcin
authors.initials: M
authors.lastname: Zemla
authors.forename: Allan
authors.initials: A
authors.lastname: Kachelmeier
authors.forename: Anders
authors.initials: A
authors.lastname: Fridberger
authors.forename: Manfred
authors.initials: M
authors.lastname: Auer
authors.forename: Xiaorui
authors.initials: X
authors.lastname: Shi
mesh_terms: D000818:Animals; D018929:Cell Culture Techniques; D002460:Cell Line; D002469:Cell Separation / Q000379:methods; D015925:Cryopreservation / Q000379:methods; D003470:Culture Media; D004423:Ear; D042783:Endothelial Cells / Q000166:cytology*; D005260:Female; D008264:Macrophages / Q000166:cytology*; D008297:Male; D008544:Melanocytes / Q000166:cytology*; D051379:Mice; D008810:Mice, Inbred C57BL; D020286:Pericytes / Q000166:cytology*; D013316:Stria Vascularis / Q000166:cytology*
chemical_list: D003470:Culture Media