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目的探讨人脐带间充质干细胞(hUCMSCs)对新生大鼠缺氧缺血性脑损伤(HIBD)的神经修复作用,为hUCMSCs治疗新生儿HIBD寻找实验依据。方法将7日龄SD大鼠制成HIBD动物模型,分为HIBD后24 h、72 h 2个时段干预组,每个时段选用3种不同的干预方法,分别为:静脉注入干细胞、腹腔注入干细胞、静脉注入干细胞+腹腔注入神经节苷脂,同时设立HIBD模型鉴定组和HIBD后24 h生理盐水对照组。将hUCMSCs进行Dil标记,分别通过颈静脉或腹腔将标记的hUCMSCs注入模型鼠体内。选用Morris水迷宫测试方法,了解干细胞干预后实验大鼠学习记忆力改善情况。采用免疫组化方法,了解干细胞在大鼠脑组织内的定值、分化。结果 HIBD后24 h、72 h静脉注入hUCMSCs组大鼠水迷宫测试逃避潜伏时间分别为(56.0±6.9)s、(76.4±6.7)s,较生理盐水组(98.8±9.0)明显缩短(P<0.05),且24 h组较72 h组逃避潜伏时间明显缩短(P<0.05);HIBD后24 h、72 h静脉注入hUCMSCs+腹腔注入神经节苷脂组大鼠逃避潜伏时间分别为(38.3±7.5)s、(50.0±7.2)s,较24 h、72 h单纯静脉注入hUCMSCs大鼠逃避潜伏时间(56.0±6.9)s和(76.4±6.7)s明显缩短(P<0.05)。进入大鼠体内的hUCMSCs能进入脑内并显示神经元前体细胞特性;hUCMSCs静脉移植组缺血区胶质纤维酸性蛋白阳性细胞(1276.7±551.8)个,明显低于对照组阳性细胞(2789.0±940.6)个(P<0.01)。结论静脉移植的hUCMSCs可以向脑内迁移并分化为神经元前体细胞,抑制神经胶质细胞增生,明显改善HIBD大鼠行为。大鼠HIBD制模后24 h输注hUCMSCs较72 h输注脑功能改善情况更好,移植hUCMSCs联合神经节苷脂对神经修复有促进作用。
Objective To investigate the effect of human umbilical cord mesenchymal stem cells (hUCMSCs) on the neurological repair of hypoxic-ischemic brain damage (HIBD) in neonatal rats and to find out the experimental basis for the treatment of neonatal HIBD by hUCMSCs. Methods The 7-day-old SD rats were made HIBD animal model and divided into two groups at 24 h and 72 h after HIBD. The intervention groups were divided into three groups according to different interventions: intravenous injection of stem cells and intraperitoneal injection of stem cells , Intravenous injection of stem cells + intraperitoneal injection of gangliosides, HIBD model established at the same time and HIBD 24 h saline control group. The hUCMSCs were labeled with Dil, and the labeled hUCMSCs were injected into the model mice through the jugular vein or abdominal cavity respectively. Morris water maze test method was used to understand the improvement of learning and memory of experimental rats after intervention of stem cells. Using immunohistochemical methods to understand the value of stem cells in rat brain tissue differentiation. Results The escape latency of rats in hUCMSCs injected with HUCMSCs 24 h and 72 h after HIBD were (56.0 ± 6.9) s and (76.4 ± 6.7) s, respectively, which were significantly shorter than that of the normal saline group (98.8 ± 9.0) (P < 0.05). The escape latency of 24 h and 72 h groups was significantly shorter than that of the 72 h group (P <0.05). The escape latency of hUCMSCs plus intraperitoneal injection of ganglioside at 24 h and 72 h after HIBD were (38.3 ± 7.5 ) s, (50.0 ± 7.2) s respectively. The escape latency (56.0 ± 6.9) s and (76.4 ± 6.7) s were significantly shorter than that of hUCMSCs injected intravenously at 24 h and 72 h (P <0.05). The hUCMSCs entering into the rat’s body could enter the brain and show the characteristics of neuronal precursor cells. The amount of glial fibrillary acidic protein positive cells (1276.7 ± 551.8) in hUCMSCs vein graft group was significantly lower than that in the control group (2789.0 ± 940.6) (P <0.01). Conclusion Intravenously transplanted hUCMSCs can migrate into the brain and differentiate into neuronal precursor cells, inhibit glial cell proliferation and significantly improve the behavior of HIBD rats. Transfected hUCMSCs at 24 h after model establishment in rats were better than brains injected at 72 h, and transplantation of hUCMSCs and gangliosides promoted neurorestoration.