SDF-1／CXCR4轴通过调控间充质干细胞定向分化修复缺氧缺血性脑损伤  

The SDF-1/CXCR4 axis repairs hypoxia-ischemia brain damage viaregulating oriented differentiation of mesenchymal stem cells

作　　者：余勤[1] 林洁[1] 刘丽珍[2] 王艳[3] 宣晓波[3] 王标[1] 单威[1] 周丽萍[1] 刘伟[1] 

机构地区：[1]浙江中医药大学,生物工程学院,浙江杭州310053 [2]浙江大学医学院附属第一医院骨髓移植中心,浙江杭州310003 [3]浙江中医药大学,第一临床医学院,浙江杭州310053

出　　处：《中国病理生理杂志》2012年第8期1424-1430,共7页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.30940028);浙江省科技计划项目(No.2010C33024);浙江省医药卫生科技计划(No.2010KYA147)

摘　　要：目的:探讨基质细胞衍生因子-1(stromal cell-derived factor-1,SDF-1)/CXC趋化因子受体4(CXC chemokine receptor 4,CXCR4)轴调控间充质干细胞定向分化、修复缺氧缺血性脑损伤的作用。方法:大鼠间充质干细胞(rat mesenchymal stem cells,rMSCs)经缺氧培养不同时点(0h、6h、12h、24h、48h、72h)或SDF-1α(10μg/L)孵育后,采用RT-PCR、Western blotting和流式细胞术检测其表面CXCR4表达的变化;建立大鼠缺氧缺血性脑损伤模型,运用RT-PCR和Western blotting检测造模后不同时点(1d、3d、5d、7d、14d、21d)大鼠脑部海马组织中SDF-1αmRNA转录和蛋白表达的改变情况;用AMD3100(CXCR4拮抗剂)拮抗rMSCs表面CXCR4后,免疫细胞化学和Western blotting检测rMSCs诱导向神经细胞分化中神经元特异性烯醇化酶(neuron-specific eno-lase,NSE)和胶质纤维酸性蛋白(glial fibrillary acidic protein,GFAP)等神经细胞特异性标志物的阳性率及表达变化情况。结果:低氧培养6 h及12 h的rMSCs CXCR4 mRNA及蛋白表达水平均较常氧培养组明显增加(P<0.01),10μg/L SDF-1α孵育后rMSCs的CXCR4表达水平明显增加(P<0.01);缺氧缺血性脑损伤模型的大鼠脑内SDF-1α蛋白表达显著增加(P<0.01);5 mg/L AMD3100处理后的rMSCs在向神经细胞诱导分化中NSE和GFAP蛋白的表达明显减少。结论:微小剂量的SDF-1α可诱导低氧培养的rMSCs表面CXCR4的表达,而在缺氧缺血性脑损伤模型的大鼠脑内SDF-1α表达增加,从而使SDF-1/CXCR4轴的生物学效应得以增强;CXCR4拮抗的rMSCs在分化中神经细胞特异性标志物NSE和GFAP的表达降低,表明SDF-1/CXCR4轴在rMSCs定向神经分化修复缺血缺氧脑损伤中具有重要的调控作用。To explore the effect of stromal cell - derived factor - 1 （ SDF - 1 ）/CXC chemokine receptor 4 （CXCR4）axis on the neural differentiation of rat mesenchymal stem cells （rMSCs） and recovery from hypoxia- ischemia brain damage （HIBD）. METHODS： The rMSCs were isolated from the rat bone marrow, and expanded in vitro. The mR- NA and protein levels of CXCR4 in rMSCs treated with SDF -let （10 μg/L） and hypoxia for0 h, 6 h, 12 h, 24 h, 48 h and 72 h were detected by RT - PCR, Western blotting and flow cytometry. The mRNA and protein levels of SDF - lot in the hippocampus of the rats with hypoxia - ischemia for 1 d, 3 d, 5 d, 7 d, 14 d and 21 d were also detected by the same methods. The protein levels of neuron- specific enolase （NSE） and glial fibrillary acidic protein （GFAP）, as well as thepositive rate of neural- induced rMSCs pretreated with AMD3100 （a CXCR4 antagonist） at dose of 5 mg/L were deter- mined by Western blotting and immunocytochemistry. RESULTS： Compared with the normal controls, both mRNA and protein levels of CXCR4 increased in rMSCs exposed to hypoxia for 6 h and 12 h, and the results were also confirmed by flow cytometry. As expected, the mRNA level of SDF - 1 ot in the hippocampus of HIBD rats was higher than that in normal control rats （P 〈 0.01 ）. Moreover, the mRNA expression of CXCR4 was extremely up- regulated in rMSCs treated with SDF - lot at concentration of 10 ~g/L, and the results were also confirmed by Western blotting and flow cytometry analysis （ P 〈 0.01 ）. The protein levels and positive cell numbers of NSE and GFAP were extremely decreased in rMSCs pretreated with AMD3100 at concentration of 5 mg/L. CONCLUSION： Compared with normal rats, SDF - lot level in the hippo- campus of the rats with hypoxia - ischemia is increased. Hypoxia and micro - dose of SDF - 1 ot induce the expression of CXCR4 in rMSCs, while CXCR4 antagonist reduces the neural differentiation of rMSCs, suggesting that SDF - 1/CXCR4 axis may be deeply involved in the n

关 键 词：间充质干细胞 细胞分化 基质细胞衍生因子-1Α CXC趋化因子受体4 

分 类 号：R741.05[医药卫生—神经病学与精神病学]