Chandravanshi Bhawna, Bhonde Ramesh Ramchandra
School of Regenerative Medicine, Manipal University, Bangalore, Karnataka, India.
J Cell Biochem. 2017 Sep;118(9):2672-2683. doi: 10.1002/jcb.25885. Epub 2017 May 15.
In the present study we focused on the improvisation of islet survival in hypoxia.The Islet like cell aggregates (ICAs) derived from wharton's jelly mesenchymal stem cells (WJ MSC) were cultured with and without WJ MSC for 48 h in hypoxia and normoxia and tested for their direct trophic effect on β cell survival. The WJ MSCs themselves secreted insulin upon glucose challenge and expressed the pancreatic markers at both transcription and translational level (C-peptide, Insulin, Glucagon, and Glut 2). Direct contact of MSCs with ICAs facilitated highest viability under hypoxia as evidenced by fluorescein diacetate/propidium iodide and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The cytokine analysis of the co-cultured ICAs revealed amplification of anti-inflammatory cytokine like TGFβ and TNFα accompanied by depletion of pro-inflammatory cytokines. The increment in VEGF and PDGFa was also seen showing their ability to vascularize upon transplantation. This was further accompanied by reduction in total reactive oxygen species, nitric oxide, and super oxide ions and down regulation of Caspase3, Caspase8, p53, and up regulation of Bcl2 confirming prevention of apoptosis in ICAs. The western blot analysis confirmed the cytoprotective effect of WJ MSC on ICAs as they enhanced the anti-apoptotic marker BCL2 and reduced the expression of apoptotic markers, Annexin 5 and Caspase 3. There was a significant reduction in the expression of p38 protein in the presence of MSCs making the ICAs responsive to glucose. Taken together our data demonstrate for the first time that the WJ MSC expressed pancreatic markers and their supplementation protected engineered islets against hypoxia and oxidative stress. J. Cell. Biochem. 118: 2672-2683, 2017. © 2017 Wiley Periodicals, Inc.
在本研究中，我们着重于提高胰岛在缺氧环境下的存活率。将源自沃顿胶间充质干细胞（WJ MSC）的胰岛样细胞聚集体（ICA）在缺氧和常氧条件下分别与WJ MSC共同培养或单独培养48小时，并检测其对β细胞存活的直接营养作用。WJ MSC自身在葡萄糖刺激下分泌胰岛素，并在转录和翻译水平均表达胰腺标志物（C肽、胰岛素、胰高血糖素和葡萄糖转运蛋白2）。通过荧光素二乙酸酯/碘化丙啶和3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐（MTT）检测可知，在缺氧条件下，MSC与ICA的直接接触促进了最高的存活率。对共培养的ICA进行细胞因子分析发现，抗炎细胞因子如转化生长因子β和肿瘤坏死因子α增加，同时促炎细胞因子减少。还观察到血管内皮生长因子和血小板衍生生长因子A增加，表明它们在移植后具有血管生成能力。这进一步伴随着总活性氧、一氧化氮和超氧离子的减少，以及半胱天冬酶3、半胱天冬酶8、p53的下调和Bcl2的上调，证实了ICA中细胞凋亡的预防。蛋白质印迹分析证实了WJ MSC对ICA的细胞保护作用，因为它们增强了抗凋亡标志物BCL2的表达，并降低了凋亡标志物膜联蛋白5和半胱天冬酶3的表达。在MSC存在的情况下，p38蛋白的表达显著降低，使ICA对葡萄糖有反应。综上所述，我们的数据首次证明WJ MSC表达胰腺标志物，并且它们的补充可保护工程化胰岛免受缺氧和氧化应激的影响。《细胞生物化学杂志》2017年第118卷：2672 - 2683页。© 2017威利期刊公司。
