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[1]肖权,徐长明,王永超,等.间充质干细胞源性外泌体调控NF-κB信号通路抑制髓核细胞凋亡[J].医学研究与战创伤救治(原医学研究生学报),2020,22(5):449-455.[doi:10.3969/j.issn.1672-271X.2020.05.001]
　XIAO Quan,XU Chang-ming,WANG Yong-chao,et al.Mesenchymal stem cell-derived exosomes inhibit nucleus pulposus cell apoptosis by regulating NF-κB signaling pathway[J].JOURNAL OF MEDICALRESEARCH —COMBAT TRAUMA CARE,2020,22(5):449-455.[doi:10.3969/j.issn.1672-271X.2020.05.001]
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间充质干细胞源性外泌体调控NF-κB信号通路抑制髓核细胞凋亡()
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《医学研究与战创伤救治》（原医学研究生学报）[ISSN:1672-271X/CN:32-1713/R]

卷:
第22卷
期数:
2020年5期
页码:
449-455
栏目:
基础研究
出版日期:
2020-09-09

文章信息/Info

Title:
Mesenchymal stem cell-derived exosomes inhibit nucleus pulposus cell apoptosis by regulating NF-κB signaling pathway
作者:
肖权; 徐长明; 王永超; 周全
作者单位：223002 淮安，徐州医科大学附属淮安医院骨科(肖权、周全)；223400涟水，南京医科大学康达学院附属涟水县人民医院骨科（徐长明、王永超）
Author(s):
XIAO Quan1;  XU Chang-ming2;  WANG Yong-chao2;  ZHOU Quan1
(1.Department of Orthopaedics,the Affiliated Huai’an Hospital of Xuzhou Medical Universty, Huai’an 223002, Jiangsu, China;2.Department of Orthopaedics, the Affiliated Lianshui County People’s Hospital of Kangda College of Nanjing Medical Universty, Lianshui 223400, Jiangsu, China)
关键词:
间充质干细胞; 髓核细胞; 凋亡; 外泌体; 核因子-κB
Keywords:
mesenchymal stem cells;  nucleus pulposus cell;  apoptosis;  exosomes;  NF-κB
分类号:
R681.53；R318.04；R496
DOI:
10.3969/j.issn.1672-271X.2020.05.001
文献标志码:
A
摘要:
目的探究骨髓间充质干细胞源性外泌体（BMSC-Exo）对髓核细胞凋亡的影响及其机制。方法分离培养大鼠骨髓间充质干细胞（BMSCs），提取BMSC-Exo。分离培养髓核细胞，先取部分细胞分为内毒素组、内毒素+二硫氨基甲酸肽吡咯烷（PDTC）组，分别加入内毒素500 μg/mL、内毒素500 μg/mL+PDTC 5 μmol/L孵育24 h。流式细胞仪检测细胞凋亡率。另取细胞分为对照组、退变组、外泌体组、佛波醇酯组，分别采用基础培养基，内毒素500 μg/mL刺激，内毒素和外泌体20 μg刺激，内毒素、外泌体和佛波醇酯50 μg/mL刺激培养24 h。流式细胞仪检测细胞凋亡率；Western blot检测Cleaved Caspase-3、磷酸化NF-κB抑制蛋白（p-IκBα）表达情况。结果成功分离培养BMSCs并提取BMSC-Exo。内毒素+PDTC组细胞凋亡率[（11.55±0.86）%]较内毒素组[（28.63±2.86）%]降低（t=9.909，P=0.001）。对照组凋亡率[（9.05±1.61）%]较退变组[（29.48±2.83）%]显著升高（t=-10.874，P=0.000）；与退变组相比，外泌体组下降至（15.08±1.65）%，差异有统计学意义（t=7.620，P=0.002）；与外泌体组相比，佛波醇酯组升高至（25.41±1.38）%，差异有统计学意义（t=-8.323，P=0.001）。与对照组比较，退变组p-IκBα蛋白表达升高（t=-10.575，P=0.000），Cleaved Caspase-3蛋白表达升高（t=-8.506，P=0.001）；与退变组比较，外泌体组p-IκBα蛋白表达降低（t=10.052，P=0.001），Cleaved Caspase-3蛋白表达降低（t=7.419，P=0.002）；与外泌体组比较，佛波醇酯组p-IκBα蛋白表达升高（t=-12.125，P=0.000），Cleaved Caspase-3蛋白表达升高（t=-8.421，P=0.001）。结论BMSC-Exo可抑制内毒素诱导的髓核细胞凋亡，其作用可能是通过抑制NF-κB信号通路实现的。
Abstract:
ObjectiveTo investigate the effect of bone marrow mesenchymal stem cell-derived exosomes (BMSC-Exo) on the apoptosis of nucleus pulposus cells and its mechanism.MethodsBone mesenchymal stem cells (BMSCs) were isolated and cultured, and BMSC-Exo were extracted. Nucleus pulposus cells were isolated, cultured, and divided into two groups: endotoxin group and endotoxin + pyrrolidine dithiocarbamate (PDTC) group. Endotoxin 500 μg/mL, endotoxin 500 μg/mL + PDTC 5 μmol/L wereadded to incubate for 24 hours, respectively. The apoptosis rates were detected by flow cytometry. Then, cells were divided into control group, degenerative group, exosome group and phorbol ester group. Cells were stimulated by basic medium, endotoxin 500 μg/mL, endotoxin and exosomes 20 μg, endotoxin, exosomes and phorbol ester 50 μg/mL for 24 hours, respectively. The apoptosis rates were detected by flow cytometry, and the expressions of cleaved Caspase-3 and phosphorylated NF-κBα inhibitor protein (p-IκBα) were detected by Western blotting.ResultsBMSCs were successfully isolated and cultured and BMSC-Exo were extracted. The apoptosis rate of endotoxin + PDTC group [(11.55±0.86)%] was lower than that of endotoxin group [(28.63±2.86)%] (t=9.909, P=0.001). The apoptotic rate in control group (9.05±1.61)% was increased than that in in degenerative group (29.48±2.83)% (t=-10.874, P=0.000). Compared with degenerative group, the apoptotic rate in exosome group was decreased (15.08±1.65)% (t=7.620, P=0.002). Compared with exosome group, the apoptotic rate in phorbol ester group was increased (25.41%±1.38%) (t=-8.323, P=0.001). Compared with control group, the expression of p-IκBα protein was increased (t=-10.575, P=0.000) and cleaved Caspase-3 protein was increased (t=-8.506, P=0.001) in degenerative group. Compared with degenerative group, the expression of p-IκBα protein was decreased (t=10.052, P=0.001) and cleaved Caspase-3 protein was decreased (t=7.419, P=0.002) in exosome group. Compared with exosome group, the expression of p-IκBα protein was increased in phorbol ester group (t=-12.125, P=0.000), and Cleaved Caspase-3 protein expression was increased (t=-8.421, P=0.001).ConclusionBMSC-Exo can inhibit LPS induced apoptosis of nucleus pulposus cells via inhibiting NF-κB signaling pathway.

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备注/Memo

备注/Memo:
基金项目：江苏省自然科学基金（BK20171265）
更新日期/Last Update: 2020-09-09