骨髓间充质干细胞来源的膜微囊对谷氨酸损伤PC12细胞的保护作用  

Protective Effect of Bone Marrow Mesenchymal Stem Cell-derived Microvesicles on Glutamate Injured PC12 Cells

作　　者：林珊珊[1] 朱波[2] 郭子宽[3] 黄国志[1] 

机构地区：[1]南方医科大学珠江医院康复医学科,广东广州510282 [2]南方医科大学珠江医院骨科广东广州510282 [3]军事医学科学院放射与辐射医学研究所实验血液学研究室,北京100850

出　　处：《中国实验血液学杂志》2014年第4期1078-1083,共6页Journal of Experimental Hematology

摘　　要：本研究目的在于探讨骨髓间充质干细胞(BMMSC)来源的膜微囊(microvesicle,MV)对神经细胞的保护作用,以了解MSC对神经修复作用的机制。采用密度梯度离心联合贴壁法分离BMMSC,采用低温高速离心法提取BMMSC-MV。采用流式细胞术鉴定BMMSC,并在透射电子显微镜下观察MV形态。建立谷氨酸致PC12细胞损伤模型,实验分为对照组、谷氨酸损伤组以及BMMSC-MV共培养组。用MTT法检测MV对PC12细胞活性的影响;流式细胞术、Hoechst荧光标记法检测细胞凋亡率的变化。结果表明:分离提取的BMMSC高表达CD29和CD44,不表达或低表达CD31,CD34,CD45。在电子显微镜下MV呈典型的圆形囊泡状。与对照组相比,损伤组细胞活性显著下降,凋亡率增加;BMMSC-MV共培养显著提高了谷氨酸损伤的PC12细胞活性、降低了凋亡率。结论:BMMSC来源的MV对谷氨酸兴奋性损伤的细胞具有明显的保护作用,为MV应用于神经兴奋性损伤性疾病提供了初步的实验和理论依据。This study was aimed to investigate the protective effect of bone mesenchymal stem cell-derived microvesicles （BMMSC-MV） on glutamate injured PC12 cells so as to elucidate the mechanism of the neural damage repair. BMMSC were isolated and purified with density-gradient centrifugation method, BMMSC-MV were harvested from the supernatants of BMMSC by hypothermal ultracentrifugation method. The surface markers of BMMSC reacted against different antibodies were detected by flow cytometry. The morphology features of MV were observed under an electron microscope. Experiment was divided into three groups, one was a control group, and the other two were glutamate-injured group and co-culture group of BMMSC-MV and glutamate-damaged cells respectively. MTT test was used to evaluate the proliferative status of PC12 cells and the AnnexinV-FITC detecting kit and Hoechst33342 were used to detect the apoptosis of PC12 cells in different groups. The results showed that BMMSC isolated from rat bone marrow were highly positive for CD29, CD44 and negative for CD31, CD34 and CD45. The morphology of MV was round and the vesicles were homogenous in size. BMMSC-MV exhibited a protective effect on the excitotoxicity-injured PC12 cells, displaying increase of cell viability, decrease of Annexin-V/PI staining positive and nuclear condensed cells. It is concluded that BMMSC-MV can protect PC12 cells from glutamate-induced apoptosis, suggesting that BMMSC-MV may be a potential candidate for treatment of neurological diseases. This study provides the preliminary experimental and theoretical evidence for use of BMMSC-MV in treatment of neural excited damage.

关 键 词：骨髓间充质干细胞 膜微囊 PC12细胞 兴奋性损伤 

分 类 号：R456[医药卫生—治疗学]