Cellular and Molecular Therapeutics Laboratory, Centre for Biomaterials Cellular and Molecular Theranostics (CBCMT), Vellore Institute of Technology (VIT), Vellore, TN, 632014, India.
Stem Cell Rev Rep. 2018 Aug;14(4):558-573. doi: 10.1007/s12015-018-9810-4.
Hypoxia-reperfusion (H/R) emblems a plethora of pathological conditions which is potent in contributing to the adversities encountered by human mesenchymal stem cells (hMSCs) in post-transplant microenvironment, resulting in transplant failure. D-Alanine 2, Leucine 5 Enkephaline (DADLE)-mediated delta opioid receptor (DOR) activation is well-known for its recuperative properties in different cell types like neuronal and cardiomyocytes. In the current study its effectiveness in assuaging hMSC mortality under H/R-like insult has been delineated. The CoCl mimicked H/R conditions in vitro was investigated upon DOR activation, mediated via DADLE. hMSCs loss of viability, reactive oxygen species (ROS) production, inflammatory responses and disconcerted unfolded protein response (UPR) were assessed using AnnexinV/PI flow cytometry, fluorescence imaging, mitochondrial complex 1 assay, quantitative PCR, immunoblot analysis and ELISA. H/R like stress induced apoptosis of hMSCs was significantly mitigated by DADLE via modulation of the apoptotic regulators (Bcl-2/Bax) along with significant curtailment of ROS and mitochondrial complex 1 activity. DADLE concomitantly repressed the misfolded protein aggregation, alongside the major UPR sensors: PERK/BiP/IRE-1α /ATF-6, evoked due to the H/R mimicked endoplasmic reticulum stress. Undermined phosphorylation of the Akt signalling pathway was observed, which concerted its effect onto regulating both the pro and anti-inflammatory cytokines, actuated as a response to the H/R-like insult. The effects of DADLE were subdued by naltrindole (specific DOR antagonist) reaffirming the involvement of DOR in the process. Taken together these results promulgate the role of DADLE-induced DOR activation on improved hMSC survival, which signifies the plausible implications of DOR-activation in cell-transplantation therapies and tissue engineering aspect.
缺氧再灌注（H/R）标志着多种病理状况，这些状况在人类间充质干细胞（hMSC）移植后微环境中遇到的逆境中起着重要作用，导致移植失败。D-丙氨酸 2，亮氨酸 5 脑啡肽（DADLE）介导的δ阿片受体（DOR）激活在神经元和心肌细胞等不同细胞类型中具有恢复特性。在本研究中，研究了其在减轻 hMSC 死亡率方面的有效性H/R 样损伤。通过 DADLE 介导的 CoCl 模拟体外 H/R 条件。使用 AnnexinV/PI 流式细胞术、荧光成像、线粒体复合物 1 测定、定量 PCR、免疫印迹分析和 ELISA 评估 hMSC 活力丧失、活性氧（ROS）产生、炎症反应和失调的未折叠蛋白反应（UPR）。DADLE 通过调节凋亡调节剂（Bcl-2/Bax）显著减轻 H/R 样应激诱导的 hMSC 凋亡，同时显著减少 ROS 和线粒体复合物 1 活性。DADLE 同时抑制错误折叠蛋白聚集，以及主要的 UPR 传感器：PERK/BiP/IRE-1α/ATF-6，这是由于模拟内质网应激的 H/R 引起的。观察到 Akt 信号通路的磷酸化减弱，这协调了其调节促炎和抗炎细胞因子的作用，作为对 H/R 样损伤的反应。纳曲吲哚（特异性 DOR 拮抗剂）削弱了 DADLE 的作用，这证实了 DOR 在该过程中的参与。总之，这些结果表明 DADLE 诱导的 DOR 激活对改善 hMSC 存活的作用，这表明 DOR 激活在细胞移植治疗和组织工程方面具有潜在的意义。
