大鼠视神经损伤后大胶质细胞去分化及预变性腓总神经的诱导

DEDIFFERENTIATION OF MACROGLIAL CELLS AFTER OPTIC NERVE INJURY AND ITS INDUCTION WITH PREDEGENERATED COMMON PERONEAL NERVE IN RATS

目的研究视神经损伤后神经胶质细胞的去分化及其诱导。方法成年雄性SD大鼠,分为正常对照组、损伤组、移植组和压榨移植组,在视神经伤后3d、7d、14d和28d 4个不同时相点取视神经,用HE方法计数细胞数量,用免疫组织化学、免疫印迹、原位杂交组织化学结合计算机图像分析检测巢蛋白(Nestin)、胶质纤维酸性蛋白(GFAP)、髓鞘碱性蛋白(MBP)、神经丝(NF)、脑源性神经营养因子(BDNF)、Nogo-A及Nogo-A mRNA的表达,用免疫荧光双标组织化学法检测Nestin-GFAP和Nestin-MBP的共表达。结果损伤组细胞数量仅在伤后7d明显增多,Nestin、MBP、Nogo-A及其mRNA表达上调,GFAP、NF、BDNF下调,出现一些Nestin-GFAP双标细胞和少量Nestin-MBP双标细胞;与损伤组相比,移植组和压榨移植组一些时相点的细胞数量不同程度地增多,Nestin、GFAP、BDNF和NF表达上调,MBP、Nogo-A及其mRNA表达下调,Nestin-GFAP双标细胞有所增加,且28d组高于14d组。结论视神经损伤后,主要是星形胶质细胞发生去分化,大胶质细胞呈现一些不利于神经再生的基因表达变化;移植预变性周围神经能进一步诱导星形胶质细胞去分化,大胶质细胞的一些基因表达变化有利于神经再生。

Objective To investigate the dedifferentiation of neuroglial cells and its induction after optic nerve injury. Methods Adult male SD rats were randomly divided into 4 groups the normal control group, the injury group, the transplantation group and the microcrush and transplantation group. Optic nerves were harvested at days 3, 7, 14 and 28 after the operation. HE staining was used to count the number of neuroglial cells. Immunohistochemistry, Western blotting and in situ hybridization histochemistry were employed together with computerized image analysis to evaluate the expressions of Nestin, GFAP, MBP, NF, BDNF, Nogo-A and Nogo-A mRNA. Immunofluorescence double staining was used to detect the co-expression of Nestin and GFAP or Nestin and MBP. Results The number of cells only increased at day 7 after the nerve injury, the expressions of Nestin, MBP, Nogo-A and Nogo-A mRNA were upregulated, the expressions of GFAP, NF and BDNF were down-regulated, and some Nestin- GFAP positive cells and a few of Nestin-MBP positive cells were detected in the injury group. Compared with the injury group, the number of cells was increased sometime after the nerve injury; the expressions of Nestin, GFAP, BDNF and NF were upregulated, the expressions of MBP, Nogo-A and Nogo-A mRNA were down-regulated, and the number of Nestin-GFAP positive cells increased in the transplantation group and the microcrush and transplantation group. Conclusion After optic nerve injury, some astrocytes undergo dedifferentiation while the macroglial cells display a gene expression pattern that is unfavorable for nerve regeneration. Pre-degenerated peripheral nerves could enhance the dedifferentiation of astrocytes and induce the gene expression pattern of macroglial cells that is favorable for nerve regeneration.