NDGA对移植入损伤脊髓后的化学去细胞肌肉支架中的羊膜上皮细胞存活的影响

Effect of NDGA on survival of amniotic epithelial cells in chemical acellular muscle scaffold after transplanted into injured spinal cord

目的:观察NDGA对植入大鼠脊髓损伤处的化学去细胞肌肉支架中的羊膜上皮细胞(AECs)存活的影响,为NDGA在脊髓损伤中的进一步应用提供实验依据。方法:制备化学去细胞肌肉支架并联合Dil标记的6只孕鼠AECs用于大鼠脊髓半横断损伤。将72只模型鼠随机分成生理盐水组(n=18)、20mg·kg-1 NDGA组(n=18)、30mg·kg-1 NDGA组(n=18)和40mg·kg-1 NDGA组(n=18),各组大鼠分别于术后1周每天注射生理盐水和20、30、40mg·kg-1 NDGA,并分别于术后1、2和4周取材,荧光显微镜下观察并比较各组大鼠AECs存活的数量;采用免疫组织化学方法观察各组大鼠损伤脊髓新生血管阳性面积百分比和NG2阳性内源性神经干细胞的数量。结果:各组大鼠支架中AECs的数量随着时间的延长均不断减少,移植后1、2和4周各时间点,30mg·kg-1 NDGA组和40mg·kg-1 NDGA组大鼠支架中AECs的存活数量明显高于生理盐水组和20mg·kg-1 NDGA组(P<0.05);移植1周后,30mg·kg-1 NDGA组和40mg·kg-1 NDGA组大鼠新生血管阳性面积百分比明显低于生理盐水组和20mg·kg-1 NDGA组(P<0.05);移植1周后,各组大鼠间NG2阳性细胞数量差异无统计学意义(P>0.05)。结论:NDGA能够明显增加大鼠脊髓损伤后的化学去细胞肌肉支架中移植AECs的存活数量,对损伤脊髓新生血管具有抑制作用,而对于内源性神经干细胞无影响。

Objective To observe the effect of NDGA on the survival of amniotic epithelial cells （AECs） in the chemical aeellular muscle scaffold after transplanted into injured spinal cord, and to provide experimental basis for the application of DNGA in spinal cord injury. Methods Chemical acellular muscle scaffold combined with Dil labeled AECs of 6 pregnant rats were used for spinal cord hemisection injury model. 72 adult male rats were randomly divided into saline group （n= 18）, 20 mg · kg 1NDGA group （n= 18）, 30 mg · kg-1NDGA group （n=18）, and 40 mg · kg 1NDGA group （n=18）. The rats in saline group were given intraperitoneal injection of saline. The rats in each NDGA groups were given intraperitoneal injection of NDGA by the concentration of 20, 30, and 40 mg · kg-1 , respectively. And 6 rats in each group were separately sacrificed on the 1st, 2nd, and 4th week after operation. The number of the survival AECs waas observed under fluorescence microscope; the percentage of positive blood vessel area and the number of NG2 positive cells of the rats in various groups were observed by immunohistochemical method. Results The number of AECs of the rats in various groups was decreased with the prolongation of time. 1, 2, and 4 weeks after transplantation, the number of AECs in 30 mg · kg-1NDGA group and 40 mg · kg-1NDGA group was significantly higher than those in saline group and 20 mg · kg-1NDGA group （P〈0.05）. 1 week after transplantation, the percentages of positive blood vessel area of the rats in 30 mg · kg-1 NDGA group and 40 mg · kg-1 NDGA group were significantly lower than those in saline group and 20 mg · kg NDGA group （P〈0.05）. 1 week after transplantation, the number of the NG2 positive cells of the rats had no significant difference between various groups （P〈0.05）. Conclusion NDGA is able to significantly extend the survival of the transplanted AECs in the chemical acellular muscle scaffold after spinal cord injury. NDGA can inhibit the new blood vessels of the injured spinal cord, and dose not affect the endogenous neural stem cells.