颗粒状骨髓间充质干细胞在关节镜热休克的有害影响下能得到更好的保护。
Pelleted Bone Marrow Derived Mesenchymal Stem Cells Are Better Protected from the Deleterious Effects of Arthroscopic Heat Shock.
作者信息
Kalamegam Gauthaman, Abbas Mohammed, Gari Mamdooh, Alsehli Haneen, Kadam Roaa, Alkaff Mohammed, Chaudhary Adeel, Al-Qahtani Mohammed, Abuzenadah Adel, Kafienah Wael, Mobasheri Ali
机构信息
Center of Excellence in Genomic Medicine Research, King Abdulaziz UniversityJeddah, Saudi Arabia; Sheik Salem Bin Mahfouz Scientific Chair for Treatment of Osteoarthritis by Stem Cells, King Abdulaziz UniversityJeddah, Saudi Arabia.
Sheik Salem Bin Mahfouz Scientific Chair for Treatment of Osteoarthritis by Stem Cells, King Abdulaziz UniversityJeddah, Saudi Arabia; Department of Orthopedic Surgery, Faculty of Medicine, King Abdulaziz University HospitalJeddah, Saudi Arabia.
出版信息
Front Physiol. 2016 May 24;7:180. doi: 10.3389/fphys.2016.00180. eCollection 2016.
INTRODUCTION
The impact of arthroscopic temperature on joint tissues is poorly understood and it is not known how mesenchymal stem cells (MSCs) respond to the effects of heat generated by the device during the process of arthroscopy assisted experimental cell-based therapy. In the present study, we isolated and phenotypically characterized human bone marrow mesenchymal stem cells (hBMMSCs) from osteoarthritis (OA) patients, and evaluated the effect of arthroscopic heat on cells in suspension and pellet cultures.
METHODS
Primary cultures of hBMMSCs were isolated from bone marrow aspirates of OA patients and cultured using DMEM supplemented with 10% FBS and characterized for their stemness. hBMMSCs (1 × 10(6) cells) cultured as single cell suspensions or cell pellets were exposed to an illuminated arthroscope for 10, 20, or 30 min. This was followed by analysis of cellular proliferation and heat shock related gene expression.
RESULTS
hBMMSCs were viable and exhibited population doubling, short spindle morphology, MSC related CD surface markers expression and tri-lineage differentiation into adipocytes, chondrocytes and osteoblasts. Chondrogenic and osteogenic differentiation increased collagen production and alkaline phosphatase activity. Exposure of hBMMSCs to an illuminated arthroscope for 10, 20, or 30 min for 72 h decreased metabolic activity of the cells in suspensions (63.27% at 30 min) and increased metabolic activity in cell pellets (62.86% at 10 min and 68.57% at 20 min). hBMMSCs exposed to 37, 45, and 55°C for 120 s demonstrated significant upregulation of BAX, P53, Cyclin A2, Cyclin E1, TNF-α, and HSP70 in cell suspensions compared to cell pellets.
CONCLUSIONS
hBMMSC cell pellets are better protected from temperature alterations compared to cell suspensions. Transplantation of hBMMSCs as pellets rather than as cell suspensions to the cartilage defect site would therefore support their viability and may aid enhanced cartilage regeneration.
引言
关节镜温度对关节组织的影响尚不清楚，并且在关节镜辅助的基于实验性细胞治疗过程中，间充质干细胞（MSCs）如何响应设备产生的热效应也不明确。在本研究中，我们从骨关节炎（OA）患者中分离出人类骨髓间充质干细胞（hBMMSCs）并对其进行表型特征分析，同时评估关节镜热对悬浮培养和贴壁培养细胞的影响。
方法
从OA患者的骨髓抽吸物中分离出hBMMSCs原代培养物，使用添加10%胎牛血清的DMEM进行培养，并对其干性进行特征分析。将培养为单细胞悬浮液或细胞团块的hBMMSCs（1×10⁶个细胞）暴露于带光源的关节镜下10、20或30分钟。随后分析细胞增殖和热休克相关基因表达。
结果
hBMMSCs具有活力，表现出群体倍增、短梭形形态、表达MSC相关的CD表面标志物，并能向脂肪细胞、软骨细胞和成骨细胞进行三系分化。软骨生成和成骨分化增加了胶原蛋白的产生和碱性磷酸酶活性。将hBMMSCs暴露于带光源的关节镜下10、20或30分钟，持续72小时，会降低悬浮培养细胞的代谢活性（30分钟时为63.27%），并增加贴壁培养细胞的代谢活性（10分钟时为62.86%，20分钟时为68.57%）。与贴壁培养细胞相比，暴露于37、45和55°C 120秒的hBMMSCs在悬浮培养细胞中BAX、P53、细胞周期蛋白A2、细胞周期蛋白E1、肿瘤坏死因子-α和热休克蛋白70显著上调。
结论
与细胞悬浮液相比，hBMMSC细胞团块对温度变化的保护更好。因此，将hBMMSCs作为团块而非细胞悬浮液移植到软骨缺损部位将有助于维持其活力，并可能有助于增强软骨再生。
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