雪旺细胞对BMSCs来源内皮细胞分泌一氧化氮的作用研究

发布时间：2018-07-06 20:47

本文选题：骨组织工程 + BMSCs　；参考：《中国修复重建外科杂志》2014年08期

【摘要】：目的研究雪旺细胞(Schwann cells,SCs)对BMSCs来源内皮细胞分泌一氧化氮(nitric oxide,NO)功能的影响,为进一步探究组织工程骨成骨过程中神经因素对血管因素的作用奠定实验基础。方法取1日龄SD乳鼠坐骨神经及臂丛神经,体外分离获取SCs,采用S-100免疫组织化学染色鉴定;取2周龄SD大鼠胫骨及股骨骨髓,体外分离获取BMSCs,诱导分化成为内皮细胞,采用血管性血友病因子与CD31免疫荧光染色鉴定。采用Transwell共培养板对SCs与BMSCs来源内皮细胞进行非接触共培养作为实验组,单纯培养BMSCs来源内皮细胞作为对照组,分别于1、3、5、7 d检测培养液中NO浓度,1、3、7、10 d采用实时荧光定量PCR(real-time fl uorescence quantitative PCR,RT-qPCR)检测一氧化氮合酶2(nitric oxide synthetase 2,NOS2)、NOS3 mRNA表达。结果分离及诱导分化的细胞通过形态学及免疫组织化学和免疫荧光染色鉴定为SCs和内皮细胞。实验组及对照组内各时间点间释放NO浓度比较,差异均有统计学意义(P0.05);除3 d外,其余各时间点实验组释放NO浓度均显著高于对照组(P0.05)。RT-qPCR检测结果示,培养各时间点实验组NOS2 mRNA相对表达量均显著高于对照组(P0.05);两组内各时间点间比较差异均有统计学意义(P0.05)。除10 d外,其余各时间点实验组NOS3 mRNA相对表达量均显著高于对照组(P0.05);实验组内各时间点间NOS3 mRNA相对表达量比较差异均无统计学意义(F=6.673,P=0.062),对照组内各时间点间比较差异均有统计学意义(F=36.581,P=0.000)。结论 SCs可促进BMSCs来源内皮细胞释放NO,可能与SCs促进NOS活性有关。
[Abstract]:Objective to study the effect of Schwann cells (Schwann cells) on the secretion of nitric oxide (no) in endothelial cells derived from BMSCs. Methods the sciatic and brachial plexus nerves of 1-day-old SD rats were isolated and identified by S-100 immunohistochemical staining in vitro, and BMSCs were isolated from the tibia and femur of 2-week-old SD rats and differentiated into endothelial cells in vitro. Von Willebrand factor and CD31 immunofluorescence staining were used. Non-contact co-culture of endothelial cells derived from BMSCs and SCs was carried out on Transwell co-culture plate as experimental group, and BMSCs derived endothelial cells as control group. Nitric oxide synthase 2 (nitric oxide synthetase 2 no S2 mRNA expression in culture medium was detected by real time fluorescence quantitative polymerase chain reaction (real-time fl uorescence quantitative PCR) for 10 days. Results the cells isolated and differentiated were identified as SCs and endothelial cells by morphological, immunohistochemical and immunofluorescence staining. The concentration of no released from the experimental group and the control group was significantly higher than that of the control group (P0.05), except for 3 days, the concentration of no released in the experimental group was significantly higher than that in the control group (P0.05) .RT-qPCR results showed that the concentration of no in the experimental group was significantly higher than that in the control group (P0.05). The relative expression of NOS2 mRNA in the experimental group was significantly higher than that in the control group at each time point (P0.05), and the difference between the two groups at each time point was statistically significant (P0.05). Except for 10 days, At other time points, the relative expression of NOS3 mRNA in the experimental group was significantly higher than that in the control group (P0.05), but there was no significant difference in the relative expression of NOS3 mRNA between each time point in the experimental group (FF6.673P0. 062), and there was significant difference in the comparison between the control group and the control group at each time point (P0. 000). Conclusion SCs can promote the release of no from endothelial cells derived from BMSCs, which may be related to the promotion of NOS activity by SCs.
【作者单位】：郑州大学第一附属医院骨科;南方医科大学南方医院创伤骨科;中国医学科学院北京协和医学院肿瘤医院骨科;青岛大学医学院附属烟台毓璜顶医院手足骨科;第四军医大学西京医院全军骨科研究所;
【基金】：国家自然科学基金资助项目(31070866、81301570)~~
【分类号】：R329

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