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目的研究骨髓间充质干细胞(MSCs)植入缺氧缺血性脑损伤(HIBD)新生大鼠脑后,植入MSCs在脑组织中的迁移、神经细胞抗原分化率的变化,探讨丹参联合MSCs移植治疗新生儿HIE的可行性。方法将36只7日龄新生SD大鼠随机分为正常对照组(8只)、HIE组(8只)、MSCs移植组(10只)、MSCs移植+丹参组(10只)。在MSCs移植后18d取脑组织后做石蜡切片,免疫组织化学法检测并计数进入脑组织的MSCs,观察植入细胞在脑组织中的分布、迁移;免疫荧光双标记法检测植入细胞神经元巢蛋白(Nestin)、神经元特异性烯醇化酶(NSE)、胶质纤维酸性蛋白(GFAP)的表达,观察其向神经细胞的分化,并进行组间比较。结果植入MSCs主要分布于HIBD组左侧大脑皮质区,MSCs移植+丹参组标本中MSCs更多向右侧大脑半球迁移,且分布范围更广,与MSCs移植组比较,不同脑组织层面、两侧大脑半球MSCs计数均有统计学差异(Pa<0.05);免疫荧光双标记法观察MSCs主要在左侧大脑皮质、海马等部位分化为神经细胞,Nestin、NSE、GFAP均可表达。MSCs移植+丹参组植入MSCs的Nestin、NSE分化率更高,且有统计学意义;MSCs移植组和MSCs移植+丹参组GFAP分化率比较差异无统计学意义。结论 MSCs植入HIBD新生大鼠脑组织后能存活,并在脑组织中移行,植入MSCs主要在HIBD新生鼠左侧脑皮质、海马等部位分化为神经细胞,表达Nestin、NSE及GFAP,加用丹参后可促进植入MSCs表达Nestin及NSE,对GFAP表达无影响。
Objective To investigate the migration of bone marrow mesenchymal stem cells (MSCs) implanted into the brain of neonatal rats with hypoxic-ischemic brain damage (HIBD) and the changes of differentiation rate of neural cells in MSCs. Feasibility of transplantation for neonatal HIE. Methods Thirty - six newborn SD rats of 7 days old were randomly divided into normal control group (n = 8), HIE group (n = 8), MSCs transplantation group (n = 10) and MSCs transplantation + Danshen group (n = 10). The MSCs were harvested at 18 days after transplantation and paraffin sections were obtained. MSCs were harvested and counted in the brain tissue by immunohistochemistry to observe the distribution and migration of the implanted cells in the brain. The immunofluorescence double labeling method was used to detect the number of implanted neurons Nestin, NSE and GFAP expression were observed. Differentiation into neurons was observed and compared between groups. Results The MSCs were mainly distributed in the left cortex of HIBD group. MSCs transplantation + Danshen group migrated more to the right hemisphere and distributed more widely. Compared with MSCs transplantation group, The counts of MSCs in the lateral hemispheres were significantly different (Pa <0.05). The immunofluorescence double labeling method was used to observe the differentiation of MSCs into neurons in the left cerebral cortex and hippocampus. Nestin, NSE and GFAP were all expressed. The MSCs transplantation + Salvia miltiorrhiza implantation of MSCs Nestin, NSE higher differentiation rate, and statistically significant; MSCs transplantation group and MSCs transplantation + Salvia miltiorrhiza group GFAP rate was no significant difference. Conclusion The MSCs can survive in the brain tissue of HIBD neonatal rats and migrate in the brain tissue. MSCs mainly differentiate into neurons in the left cerebral cortex and hippocampus of HIBD neonates, which express Nestin, NSE and GFAP Salvia miltiorrhiza could promote the expression of Nestin and NSE in MSCs, but had no effect on the expression of GFAP.