Chandravanshi Bhawna, Bhonde Ramesh
School of Regenerative Medicine, Manipal University, GKVK Post, Bellary Road, Allalasandra, Yelahanka, 560065 Bangalore, India.
School of Regenerative Medicine, Manipal University, GKVK Post, Bellary Road, Allalasandra, Yelahanka, 560065 Bangalore, India.
Eur J Pharmacol. 2016 Nov 15;791:424-432. doi: 10.1016/j.ejphar.2016.09.012. Epub 2016 Sep 8.
Transplantation of pancreatic islets is the most reliable treatment for Type 1 diabetes. However cell death mediated by hypoxia is considered as one of the main difficulties hindering success in islet transplantation. The aim of our experiment was to investigate the role of small molecules in survival of Islet like cell aggregates (ICAs) engineered from umbilical cord matrix under oxygen deprived condition (<5% O). ICAs were analyzed for cell death via fluoroscein diacetate/propidium iodide (FDA/PI) staining, estimation of Caspase 3 and free radical release in presence and absence of small molecules. The samples were also analyzed for the presence of hypoxia inducible factor 1α (HIF1α) at both transcriptional and translational level. The addition of small molecules showed profound defensive effect on ICAs under hypoxic environment as evidenced by their viability and insulin secretion compared to untreated ICAs. The combinations of Eicosapentaenoic acid (EPA), Docosahexaenoic acid(DHA) and metformin and EPA, DHAandγ amino butyric acid (GABA) acted as anti-apoptotic agents for human ICAs when exposed to 1% O for 48h. The combinations of the small molecules reduced the total reactive oxygen species and malonaldehyde (MDA) levels and enhanced the production of glutathione peroxidise (GPx) enzyme under hypoxic conditions. Finally the increase in HIF1α at both protein and gene level confirmed the defensive effect of the additives in hypoxia. These results suggest that the combination of small molecules maintained the viability and functionality of the ICAs in hypoxia by up-regulating HIF1α expression and down regulating the Caspase 3 activity.
胰岛移植是1型糖尿病最可靠的治疗方法。然而，缺氧介导的细胞死亡被认为是阻碍胰岛移植成功的主要困难之一。我们实验的目的是研究小分子在缺氧条件（<5% O₂）下由脐带基质工程化构建的胰岛样细胞聚集体（ICA）存活中的作用。通过荧光素二乙酸酯/碘化丙啶（FDA/PI）染色、在有无小分子存在的情况下对Caspase 3和自由基释放的评估来分析ICA的细胞死亡情况。还在转录和翻译水平上分析了样本中缺氧诱导因子1α（HIF1α）的存在情况。与未处理的ICA相比，小分子的添加对缺氧环境下的ICA显示出显著的保护作用，这通过它们的活力和胰岛素分泌得到证明。当暴露于1% O₂ 48小时时，二十碳五烯酸（EPA）、二十二碳六烯酸（DHA）与二甲双胍的组合以及EPA、DHA与γ-氨基丁酸（GABA）的组合对人ICA起到抗凋亡剂的作用。在缺氧条件下，小分子组合降低了总活性氧和丙二醛（MDA）水平，并增强了谷胱甘肽过氧化物酶（GPx）的产生。最后，蛋白质和基因水平上HIF1α的增加证实了添加剂在缺氧中的保护作用。这些结果表明，小分子组合通过上调HIF1α表达和下调Caspase 3活性，在缺氧条件下维持了ICA的活力和功能。
