Efficient production of bioactive insulin from human epidermal keratinocytes and tissue-engineered skin substitutes: implications for treatment of diabetes.
Efficient production of bioactive insulin from human epidermal keratinocytes and tissue-engineered skin substitutes: implications for treatment of diabetes.
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DOI：
10.1089/ten.2006.0210
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发表时间：
2007-08
影响因子：
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通讯作者：
P. Lei;A. Ogunade;K. Kirkwood;S. G. Laychock;S. Andreadis
中科院分区：
文献类型：
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作者：
P. Lei;A. Ogunade;K. Kirkwood;S. G. Laychock;S. Andreadis
尽管经过多年的研究，每日注射胰岛素仍然是糖尿病治疗的黄金标准。基因治疗可以通过赋予从异位部位分泌胰岛素的能力来提供替代策略。表皮是一种自我更新的组织，很容易接近，可以提供大量的自体细胞来产生分泌胰岛素的皮肤替代品。在这里，我们使用重组逆转录病毒来修饰人表皮角质形成细胞，其中编码人胰岛素原的基因在A-C和B-C连接处含有弗林蛋白酶识别序列。角质形成细胞能够加工胰岛素原并分泌促进葡萄糖摄取的活性胰岛素。原代表皮细胞比细胞系产生更高量的胰岛素，这表明胰岛素分泌可能取决于生产细胞的生理状态。修饰的细胞保持分层成在基底层和基底上层表达胰岛素的三维皮肤等同物的能力。弗林蛋白酶识别位点的修饰并没有改善胰岛素原的加工，但胰岛素原B链中的单个氨基酸取代分别使培养细胞和生物工程皮肤替代品的C肽分泌增强了10倍和28倍。这些结果表明，基因修饰的生物工程皮肤可能为糖尿病的治疗提供一种替代的胰岛素输送方式。
Despite many years of research, daily insulin injections remain the gold standard for diabetes treatment. Gene therapy may provide an alternative strategy by imparting the ability to secrete insulin from an ectopic site. The epidermis is a self-renewing tissue that is easily accessible and can provide large numbers of autologous cells to generate insulin-secreting skin substitutes. Here we used a recombinant retrovirus to modify human epidermal keratinocytes with a gene encoding for human proinsulin containing the furin recognition sequences at the A-C and B-C junctions. Keratinocytes were able to process proinsulin and secrete active insulin that promoted glucose uptake. Primary epidermal cells produced higher amounts of insulin than cell lines, suggesting that insulin secretion may depend on the physiological state of the producer cells. Modified cells maintained the ability to stratify into 3-dimensional skin equivalents that expressed insulin at the basal and suprabasal layers. Modifications at the furin recognition sites did not improve proinsulin processing, but a single amino acid substitution in the proinsulin B chain enhanced C-peptide secretion from cultured cells and bioengineered skin substitutes 10- and 28-fold, respectively. These results suggest that gene-modified bioengineered skin may provide an alternative means of insulin delivery for treatment of diabetes.