米诺环素对脊髓损伤大鼠凋亡机制探索和神经红蛋白表达的影响

目的 探讨米诺环素对脊髓横断损伤大鼠凋亡蛋白 Bcl-2、Bax和 Caspase-3的表达及神经红蛋白 Ngb表 达的影响。方法 36只健康成年雄性 Wistar大鼠随机分为假手术组、脊髓损伤组、米诺环素干预组，每组 12只。假 手术组只暴露脊髓，脊髓损伤组和米诺环素干预组制备胸3~4脊髓节段全横断损伤模型。米诺环素干预组于术前腹 腔注射米诺环素（90 mg/kg），1次/d，连续5 d，脊髓损伤组和假手术组腹腔注射等体积生理盐水。术后22 d处死所有 大鼠，采用免疫组化染色和 Western blot法检测脊髓组织 Bcl-2、Bax和 Caspase-3蛋白的表达；Hoechst 染色检测凋亡 细胞；免疫荧光染色检测神经红蛋白（Ngb）的表达。结果 与假手术组相比，脊髓损伤组脊髓组织中 Bcl-2表达增 多，Bax和 Caspase-3蛋白的表达增多，凋亡细胞数明显增多，Ngb荧光强度明显增强（均P＜0.05）。与脊髓损伤组相 比，米诺环素干预组Bcl-2表达进一步增高（P＜0.05），Bax和Caspase-3表达显著降低（P＜0.05），而凋亡细胞数有所 减少，Ngb的平均荧光强度较其明显增强。结论 米诺环素干预可通过降低 Bax和 Caspase-3的表达，升高 Bcl-2的 表达来抑制细胞凋亡，提高神经红蛋白表达。

Effects of minocycline pretreatment on apoptotic mechanism and expression of Ngb protein in rats with spinal cord injury

Objective To investigate the effect of minocycline (MINO) on expressions of apoptotic proteins Bcl-2, Bax, Caspase-3 and expression of Ngb in rats with spinal cord transection injury. Methods A total of 36 healthy adult male Wistar rats were randomly divided into Sham group, spinal cord injury (SCI) group and MINO group, with 12 rats in each group. Rats in Sham group were only exposed spinal cord without transecting. SCI group and MINO group were established the T3-4 spinal cord segment transection injury model. MINO group was given intraperitoneal injection of minocycline (90 mg/kg) once a day for 5 consecutive days before operation. While SCI group and Sham group were given equal volume of normal saline once a day for 5 consecutive days. After 22 days of operation, all the rats were killed. Immunohistochemical stainning and Western blot assay were used to determine the expressions of Bcl-2, Bax and Caspase-3 proteins. Hoechst staining was used to analyze the positive cells of apoptosis. Ngb protein expression was detected by immunofluorescence method. Results Compared to Sham group, the expressions of Bcl-2, Bax, Caspase-3 and apoptotic cells were increased in SCI group, and the fluorescence intensity of Ngb increased significantly (P＜0.05). Compared to SCI group, the expression of Bcl-2 was obvious increased in MINO group (P＜0.05) while the expressions of Bax and Caspase-3 were decreased and the number of apoptotic cells decreased significantly (P＜0.05). The mean fluorescence intensity of Ngb was significantly higher in MINO group than that of SCI group. Conclusion Minocycline can inhibit apoptosis and increase neuroglobin expression by downregulating the expressions of Bax and Caspase-3, upregulating the expression of Bcl-2 in SCI rats.