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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。
OBJECTIVE: To culture rat corpus cavernosum smooth muscle cells in vitro by modified explant method. Methods Fifteen male Sprague-Dawley rats were randomly divided into 3 groups with 5 rats in each group. The smooth muscle cells of rat corpus cavernosum were isolated by tissue block method, enzymatic digestion method and modified tissue block method respectively. Serum DMEM, 37 ℃, 5% CO2, 95% air cell incubator were cultured in stationary phase contrast microscope observation of cell morphology and amplification, with α-smooth muscle actin (a-SM- Actin) and desmin desmin) immunohistochemical staining to identify cell types. Results The positive rate of α-smooth muscle actin in corpus cavernosum smooth muscle cells was 96.3%, that in fibroblasts was 23.8%, that of desmin was 74.4% in corpus cavernosum smooth muscle cells, and in fibroblasts negative. There was a significant difference between the three groups in the number of positive protein in the desmin. The rate of positive cells in the modified tissue block was higher than that in the tissue block and enzyme digestion. Conclusion The expression of desmin is a specific index for the identification of spontaneous smooth muscle cells. The improved tissue block method can obtain the smooth muscle cells of the corpus cavernosum with higher purity, structure and function.