Regulated insulin secretion from tissue engineered skin*

描述（由申请人提供）：1型糖尿病是一种以胰腺细胞自身免疫破坏导致胰岛素缺乏为特征的疾病。虽然胰岛素注射提供了生存所需的激素，但它们往往不能提供严格的血糖水平调节，从而导致神经病变和心血管疾病等长期并发症。在这里，我们建议使用基因治疗作为一种替代治疗方式，以提供恒定的胰岛素基线水平，并根据需求调节胰岛素分泌。具体来说，我们建议开发由转基因角质形成细胞制成的组织工程皮肤，作为体内分泌胰岛素治疗糖尿病的输送装置。我们将构建逆转录病毒载体，通过外源性小有机分子调控胰岛素分泌。使用最佳的基因转移方案，我们将转导表皮角质形成细胞，并从培养细胞和分层表皮组织中检测活性胰岛素的分泌。将转基因皮肤移植到胸腺糖尿病小鼠身上，将用于检测角化细胞胰岛素分泌对血糖水平的影响，并确定胰岛素调节对外源性药物的影响。组织工程皮肤将经过基因改造，以提供餐间胰岛素的基线量和餐前胰岛素的较高量，以模拟β细胞对营养刺激的胰岛素分泌反应。我们的最终目标是创造一种基于细胞的设备，以更好地控制血糖水平，克服糖尿病的破坏性长期影响。我们的团队由一个组织工程师（STA），一个分子生物学家（KLK）和一个糖尿病专家（SGL）组成。这种专业知识的结合对于这个涉及分子生物学、组织工程、组织移植和控制药物输送的多方面项目的成功至关重要，该项目旨在设计一种基于细胞的设备，用于调节重组胰岛素的输送。

DESCRIPTION (provided by applicant): Type 1 diabetes is a disease characterized by lack of insulin due to autoimmune destruction of pancreatic beta-cells. Although insulin injections supply the amount of hormone necessary for survival, they often fail to provide tight regulation of blood glucose levels resulting in long-term complications such as neuropathy and cardiovascular disease. Here we propose to use gene therapy as an alternative therapeutic modality to supply constant baseline levels of insulin and regulate insulin secretion according to demand. Specifically, we propose to develop tissue engineered skin made of genetically modified keratinocytes as an in vivo delivery device to secrete insulin for the treatment of diabetes. We will construct retroviral vectors that regulate insulin secretion through exogenous administration of small organic molecules. Using optimal gene transfer protocols, we will transduce epidermal keratinocytes and examine secretion of active insulin from cells in culture and from stratified epidermal tissues. Transplantation of genetically modified skin equivalents onto athymic diabetic mice will be used to examine the effects of keratinocyte insulin secretion on blood glucose levels and determine the efficacy of insulin regulation in response to exogenous agents. Tissue engineered skin will be genetically modified to provide baseline amounts of insulin between meals and higher amounts before meals to mimic the insulin secretory response of beta-cells to nutrient stimulation. Our ultimate goal is to create a cell-based device to provide better control of blood glucose levels and overcome the devastating long-term effects of diabetes. Our team is composed of a tissue engineer (STA), a molecular biologist (KLK) and a diabetes expert (SGL). This combination of expertise is essential for the success of this multifaceted project, which involves molecular biology, tissue engineering, tissue transplantation and controlled drug delivery to engineer a cell-based device for regulated delivery of recombinant insulin.