去细胞异种神经移植后T细胞亚群及细胞内细胞因子的实验研究

Study of effects on T lymphocyte subsets and intracellular cytokines after transferring of chemical acellular xenograft nerve

目的为化学去细胞异种周围神经移植的临床应用提供进一步的免疫学实验依据。方法 128只BALB/C小鼠随机分为假手术组、自体神经移植组、新鲜同种异体神经移植组和化学去细胞异种神经移植组,每组32只。对各实验组分别进行相应的手术。分别在术后3、7、14、28d将各组8只小鼠脾淋巴细胞进行分离,特异性荧光标记的单克隆抗体作用后,经流式细胞仪检测T淋巴细胞亚群及细胞内细胞因子的水平及其变化趋势。结果在各时间点中,化学去细胞异种神经移植组CD3+、CD4+、CD8+、CD25+阳性细胞率以及IL-2、IFN-γ、TNF-α阳性细胞率与假手术组、自体神经移植组相比无显著性差异。而新鲜异体神经移植组与其他3组比较有显著性差异。结论化学去细胞处理的异种神经的免疫原性等于或接近于自体神经,明显低于新鲜同种异体神经。

Objective To observe the immune mechanism of rejection of chemical extracted acellular xenograft nerve for future clinical application. Methods 128 BALB/C mice were randomly divided into negative contrast group（NC, 32 mice）,fresh autograft group （ AG,32 mice）, fresh allogeneic nerve group （FN,32 mice）and chemical extracted acellular xenogeneic nerve group （ CEXN,32 mice）. Then we put various kinds of nerve grafts into the thigh muscle of BALB/C mice in each group. By the time of 3、7、14、28 days after operation, the mice（8 in each group ） were killed, and the spleens were taken out. Spleens were grinded into paste, erythrocyte and other cells were burst by distilled water, and T lymphocytes were collected up. Then monoclonal antibodies（CD3、CD4、CD8、CD25、IL-2、IFN-γ、TNF-α） were added into the dilution. FACS was used to determine the positive rates of cells combined with above monoclonal antibodies. Results There were no statistically significant differences between the CEXN group, the NC group and the AG group. But above parameters was significantly higher in the FN group than those of the other three groups at the corresponding time point. Conclusion There is no evident immune rejections of chemical extracted acellular xenograft nerve when compared with the fresh autograft nerve.