改良组织块法培养SD大鼠阴茎海绵体平滑肌细胞

目的应用改良植块法体外培养大鼠阴茎海绵体平滑肌细胞。方法15只SD雄性大鼠,随机分成3组,每组5只,分别采用组织块法、酶消化法及改良组织块法分离大鼠阴茎海绵体平滑肌细胞,在含双抗及20%胎牛血清DMEM,37℃、5%CO2、95%空气的细胞培养箱静置中培养,相差显微镜观察细胞形态及扩增情况,用α-平滑肌肌动蛋白(a-SM-Actin)及结蛋白(desmin)免疫组织化学染色,鉴定细胞类型。结果α-平滑肌肌动蛋白在阴茎海绵体平滑肌细胞阳性率为96.3%,在成纤维细胞中阳性率为23.8%,结蛋白在阴茎海绵体平滑肌细胞阳性率为74.4%,在成纤维细胞中呈阴性。三组间结蛋白阳性细胞率具有显著差异,改良组织块法组结蛋白阳性细胞率高于组织块法和酶消化法组。结论结蛋白是鉴定海绵体平滑肌细胞的特异性指标,改良组织块法可获纯度更高、结构和功能良好的阴茎海绵体平滑肌细胞。

A modified tissue culture method for culturing corpus cavernosum smooth muscle cells from the penis of Sprague-Dawley rats

Objective To culture rat corpus cavernosum smooth muscle cells in vitro using a modified tissue culture method.Methods Fifteen male rats were randomized into 3 equal groups, namely enzyme digestion group, tissue culture group, and modified tissue culture group. The penis of the rats was separated carefully and cut into small pieces, and seeded onto culture flasks and cultured in complete medium consisting of DMEM containing 20% fetal calf serum at 37 ℃ in a humidified atmosphere with 5% carbon dioxide. The cells growth was observed under phase contrast microscope and the smooth muscle cell specific proteins α-SM-actin and desmin were identified immunohistochernically. Results The α-SM-actin-positive cell rate was 96.3% in rat corpus cavernosum smooth muscle and 23.8% in the fibroblasts, and the corpus cavernosum smooth muscle contained 74.4% desmin-positive cells while the fibroblasts showed no desmin positivity. Significant difference was found in the positive cell rate for desmin among the 3 groups, with the highest positive cell rate occurred in modified tissue culture group. Conclusion Desmin may serve as a marker for identifying corpus cavernosum smooth muscle cells. The modified tissue culture method can result in highly purified corpus cavernosum smooth muscle cells with intact structure and functions.