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首页> 外文期刊>Journal of cellular biochemistry. >Non insulin producing cell line, MIA PaCa-2 is rendered insulin producing in vitro via mesenchymal epithelial transition
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Non insulin producing cell line, MIA PaCa-2 is rendered insulin producing in vitro via mesenchymal epithelial transition

机译：非胰岛素产生细胞系MIA PaCa-2通过间质上皮转化在体外产生胰岛素

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We used non-insulin producing pancreatic carcinoma cell line, MIA PaCa-2 and have modulated its culture conditions by using 1% matrigel as extracellular matrix, N2, B27 growth supplements and serum free conditions. Expression of markers was analyzed using qRT-PCR, immunofluorescence and in vitro functional assay for insulin and C-peptide release was assessed using insulin and C-peptide ELISA, respectively. The cells grown under this altered culture conditions have exhibited a transition in the morphology from mesenchymal to epithelial with extensive piling up of cells. A reduction in doubling time from 40 to 18 h, upregulation of beta islet specific markers like pancreatic duodenal homeobox-1 (Pdx-1), C-peptide, insulin, and disappearance of markers like vimentin were observed. On the functional level, the altered morphology bearing cells released high levels of insulin in response to 10 μM tolbutamide (an activator of insulin pathway) and reduced insulin secretion in response to 50 μM nifedipine (inhibitor of the pathway). On the contrary, the original cells (mesenchymal morphology) had failed to release any insulin in response to varying concentrations of glucose and also the activators and inhibitors of the insulin pathway. This investigation thus provides a basis for using this basic developmental biology phenomenon mesenchymal to epithelial transition as a strategy to generate a large number of functional islets from stem cells of mesenchymal origin.
机译：我们使用了非胰岛素产生的胰腺癌细胞系MIA PaCa-2，并通过使用1％基质胶作为细胞外基质，N2，B27生长补充剂和无血清条件来调节其培养条件。使用qRT-PCR分析标记物的表达，进行免疫荧光分析，并通过胰岛素和C肽ELISA分别评估胰岛素的体外功能测定和C肽释放。在这种改变的培养条件下生长的细胞已经表现出从间充质到上皮的形态转变，伴随着大量的细胞堆积。观察到倍增时间从40小时减少到18小时，观察到β胰岛特异性标志物（如胰十二指肠同源盒1（Pdx-1），C肽，胰岛素）的上调，以及诸如波形蛋白的标志物的消失。在功能水平上，改变形态的轴承细胞响应10μM甲苯磺丁酰胺（胰岛素途径的激活剂）释放高水平的胰岛素，响应50μM硝苯地平（该途径的抑制剂）而减少胰岛素分泌。相反，原始细胞（间质形态）未能响应于不同浓度的葡萄糖以及胰岛素途径的激活剂和抑制剂而释放任何胰岛素。因此，该研究提供了使用这种从骨髓间充质向上皮过渡的基本发育生物学现象作为从骨髓间充质来源的干细胞中产生大量功能性胰岛的策略的基础。

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