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摘要：
目的 探讨骨髓间充质干细胞（mesenchymal stem cell, MSC）鞘内移植能否抑制缺血再灌注损伤脊髓受体相互作用蛋白激酶1（receptor interacting protein kinase 1, RIPK1）表达促进神经功能修复。 方法 用SD大鼠制作脊髓缺血再灌注损伤（spine cord ischemia reperfusion injury, SCII）模型。50只SD大鼠按随机数字表法分为5组（每组10只）：正常组（Sham组），大鼠开腹游离腹主动脉后立即关腹，不做腹主动脉阻断；对照组（Con组），造成SCII后鞘内不做任何处理；PBS组，造成SCII后鞘内注射等容量PBS液；坏死性凋亡抑制剂（necrostatin‑1, Nec‑1）组，造成SCII后静脉注射Nec‑1；MSC组，造成SCII后鞘内注射MSC。采用流式细胞仪对MSC表面细胞标志CD29、CD90、CD34、CD45进行鉴定；SCII后第2天，采用比索比蒂布雷斯纳汉下肢运动功能评分法和下肢体感诱发电位（sensory evoked potentials, SSEP）监测大鼠脊髓神经功能，采用免疫组织化学法观察大鼠脊髓RIPK1阳性神经元数目，采用Western blot法检测大鼠脊髓RIPK1蛋白水平。 结果 CD29表达率为100%，CD90表达率为99.6%，CD34表达率为0.7%，CD45表达率为1.0%。与Sham组比较，Con组、PBS组、Nec‑1组和MSC组下肢运动功能评分下降，脊髓RIPK1阳性神经元数目增多，脊髓RIPK1蛋白水平升高，SSEP的P1波和N1波潜伏期延长，波幅降低（P<0.05）。与Con组比较，Nec‑1组和MSC组下肢运动功能评分增加，脊髓RIPK1阳性神经元数目减少，脊髓RIPK1蛋白水平降低，SSEP的P1波和N1波潜伏期缩短，波幅升高（P<0.05）。与Nec‑1组比较，MSC组下肢运动功能评分增加，脊髓RIPK1阳性神经元数目增多，脊髓RIPK1蛋白水平升高，SSEP的P1波和N1波潜伏期缩短，波幅升高（P<0.05）。 结论 鞘内移植MSC可能通过抑制缺血再灌注损伤脊髓RIPK1表达促进神经功能修复。
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Abstract: Objective To explore whether intrathecal transplantation of mesenchymal stem cell (MSC) inhibits the expression of receptor interacting protein kinase 1 (RIPK1) in the spinal cord of rats with ischemia reperfusion injury and promotes neural repair. Methods A spinal cord ischemia reperfusion injury (SCII) model of SD rats was established. According to the random number table method, 50 rats were divided into five groups (n=10): a sham‑operated (Sham) group, a control (Con) group, a PBS group, a necrostatin‑1 (Nec‑1) group and a MSC group. In the Sham group, the abdominal aorta of rats was exposed before immediate closure of the abdomen without abdominal aorta occlusion. In the Con group, no intrathecal treatment was performed after SCII. For the PBS group, an equal‑volume of PBS was intrathecally injected after SCII. Rats in the Nec‑1 group were intravenously injected Nec‑1 after SCII. For the MSC group, MSC were intrathecally injected after SCII. Then, the cell markers CD29, CD90, CD34 and CD45 on the surface of MSC were identified by flow cytometry. The second day after SCII, Bisobiti Bresnahan lower extremity motor function score and lower extremity sensory evoked potential were used to monitor the spinal cord nerve function. The number of RIPK1 neurons in the spinal cord was observed by immunohistochemistry and the level of RIPK1 protein in the spinal cord was detected by Western blot. Results The expression rate was 100% for CD29, 99.6% for CD90, 0.7% for CD34 and 1.0% for CD45. Compared with the Sham group, the Con, PBS, Nec‑1 and MSC groups showed decreases in the lower limb motor function score, increases in the number of RIPK1 positive neurons in the spinal cord and the content of spinal cord RIPK1 protein, prolonged P1 and N1 latency of SSEP, and decreased wave amplitude (P<0.05). Compared with the Con group, the Nec‑1 and MSC groups presented increases in the lower limb motor function score, decreases in the number of RIPK1 positive neurons in the spinal cord and the content of spinal cord RIPK1 protein, shortened P1 and N1 latency of SSEP and increased wave amplitude (P<0.05). Compared with the Nec‑1 group, the MSC group showed increases in the lower limb motor function score, the number of RIPK1 positive neurons in the spinal cord and the content of RIPK1 protein in the spinal cord, shortened P1 and N1 latency of SSEP, and increased wave amplitude (P<0.05). Conclusion Intrathecal transplantation of MSC may promote the repair of nerve function by inhibiting the expression of RIPK1 in the injured spinal cord after ischemia reperfusion.
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