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背景:通过组织工程方法构建神经桥接体修复神经损伤,需要大量纯化体外培养的许旺细胞。目的:对比观察预损伤法和改良传代法获取许旺细胞的纯度与质量。方法:①预损伤法:预损伤SD乳鼠坐骨神经,3d后取出坐骨神经,分离神经外膜,用胰酶、胶原酶消化,差速贴壁除去成纤维细胞,接种培养。②改良传代法:直接获取SD乳鼠坐骨神经,分离神经外膜,运用双酶消化法结合单酶消化法进行许旺细胞原代培养,5～7d后采用单酶快速消化离心法行传代培养,同时纯化许旺细胞。结果与结论:预损伤法和改良传代法体外培养的许旺细胞纯度均达95%以上,两种方法获得的许旺细胞纯度差异无显著性意义(P>0.05)。两种方法获取的许旺细胞形态正常,数量及纯度高,增殖旺盛,说明预损伤法和改良传代法都是体外获取高质量与高纯度许旺细胞的理想方法。
BACKGROUND: To construct nerve bridge by tissue engineering method to repair nerve injury requires extensive purification of Schwann cells cultured in vitro. OBJECTIVE: To compare the purity and quality of Schwann cells with pre-injury method and improved passaging method. Methods: ① Pre-injury method: The sciatic nerve of SD neonatal rats were pre-injured. After 3 days, the sciatic nerve was taken out and the epineurium was removed. The cells were digested with trypsin and collagenase and fibroblasts were removed by differential adherence. ② improved passaging method: obtained directly from the sciatic nerve of SD neonatal rats, the separation of the epineurium, the use of double enzyme digestion method combined with single enzyme digestion of Schwann cells primary culture, 5-7 days after the rapid digestion by single-enzyme centrifugation line subculture, Simultaneously purified Schwann cells. RESULTS AND CONCLUSION: The purity of Schwann cells cultured by pre-injury method and modified passage method were more than 95%. The purity of Schwann cells obtained by two methods had no significant difference (P> 0.05). The Schwann cells obtained by the two methods showed normal morphology, high number and purity, and strong proliferation, indicating that the pre-injury method and the improved passaging method are ideal methods for obtaining high-quality and high-purity Schwann cells in vitro.