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目的 探讨从成年大白鼠坐骨神经中培养出纯的许旺细胞的方法。 方法 采用以下几种培养方法 :(1)常规方法进行大白鼠坐骨神经组织块培养 (A法 )。 (2 )应用常规的胰酶消化方法进行坐骨神经细胞培养 (B法 )。 (3 )是经过改良的细胞培养方法 (C法 )。实验组 :从坐骨神经束抽取单条神经纤维 ,剪碎后用高浓度的双酶 (0 5 %胰蛋白酶和 0 0 6%胶元蛋白酶 )经 80～ 90min的消化后进行细胞培养。对照组 :把抽取神经纤维后剩余的神经束膜和神经鞘膜 ,常规胰酶消化后进行细胞培养。用S10 0抗体和成纤维细胞抗体对培养物进行鉴定 ,并用MTT测定和H3 胸腺嘧啶核苷检测细胞的增殖能力。 结果 在A ,B法的培养物中 ,虽然有少量许旺细胞 ,但同时出现大量的成纤维细胞 ;用C法分离培养的细胞经鉴定证实全为许旺细胞 ,MTT和H3 测定证实生长良好 ,而对照组的培养物均全部为成纤维细胞。 结论 在C法中使用的抽取单条神经纤维和高浓度双酶长时间消化技术 ,能彻底清除成纤维细胞 ,从而保持许旺细胞的正常生长繁殖
Objective To explore a method of culturing pure Schwann cells from the sciatic nerve of adult rats. Methods The following methods of culture: (1) conventional method of rat sciatic nerve tissue block culture (A method). (2) Applying conventional trypsinization method to culture sciatic nerve cells (method B). (3) is a modified cell culture method (method C). Experimental group: A single nerve fiber was extracted from the sciatic nerve bundles. After shearing, the cells were digested with high concentration of double enzymes (0 5% trypsin and 0 0 6% collagenase) for 80-90 min. The control group: the nerve fibers and nerve sheaths after the remaining nerve fibers were removed and the cells were cultured by conventional trypsinization. Cultures were identified with SlO 0 antibody and fibroblast antibody and cell proliferation was assessed by MTT assay and H3 thymidine. Results A, B method of culture, although a small amount of Schwann cells, but at the same time a large number of fibroblasts; C method was isolated and cultured cells were identified as all Schwann cells, MTT and H3 assay confirmed good growth , While the control group cultures were all fibroblasts. Conclusion The single nerve fiber extracted in C method and high concentration of double enzyme long time digestion technology can completely eliminate fibroblasts so as to maintain the normal growth and reproduction of Schwann cells