Bioactive Polymers of Effective Islet Delivery System

本提案是针对R 01 DK 56884的竞争性续期申请，支持4年。生物活性聚合物已被设计和测试用于通过与聚（乙二醇）（Hb-C）交联的血红蛋白改善氧转运到包封的胰岛，以及通过磺酰脲（SU）和胰高血糖素样肽-1（GLP-1）的水溶性聚合物缀合物刺激胰岛。实验结果表明，聚合物是有效的促进功能和延长封装胰岛的寿命。在该提案中，将追求源自Hb和GLP-1的生物活性聚合物的功能性的稳定和扩展，旨在用于更长的持续时间。四聚体Hb易于氧化（metHb产生）、解离和变性。GLP-1也会发生去稳定化， 在生理环境中变性。这方面的中心研究主题将放在基于Hb和GLP-1的生物活性聚合物的稳定化上。对于Hb-C，通过使用分子内交联和/或吡哆醛化的Hb以及将抗氧化酶引入Hb-C，可以提高Hb-C的稳定性。此外，通过直接测量氧分压和Hb-C在胰岛囊中的扩散率，将优化氧结合特性以改善胰岛的氧供应，并详细研究促进氧转运的机制。对于GLP-1结合物，将采用锌诱导结晶或沉淀方法。然后使沉淀物与生物活性物质平衡。 可溶性GLP-1缀合物，并将成为胰岛胶囊中GLP-1缀合物的连续供应源。 本研究的一个新课题是构建一种有效的多聚体胰岛基因载体，用于抗凋亡基因Bcl-2的转移。当通过Bcl-2表达移植时，保护包囊化的胰岛免受早期细胞因子攻击是决定胰岛存活的另外的关键因素。这一挑战依赖于我们的观察结果，即SU-缀合物被主动内化到表达SU受体的细胞（胰岛和胰岛素瘤细胞系）中。由于所设计的生物活性聚合物是基于不同的机制，用于改善胰岛功能和延长寿命的相同目标，因此将评估生物活性聚合物的组合效应，以验证是否存在协同效应、累加效应或单组分主导效应。

This proposal is for competing renewal application of R01 DK56884 supported for 4 years. Bioactive polymers have been designed and tested for improved oxygen transport to the encapsulated islets by hemoglobin crosslinked with poly(ethylene glycol) (Hb-C) and for islet stimulation by water-soluble polymer conjugates of sulfonylurea (SU) and glucagon-like peptide-1 (GLP-1). The experimental results demonstrated that the polymers are effective in promoting the functioning and prolonging the life-span of encapsulated islets. In this proposal, the stabilization and expansion of the functionality of the bioactive polymers derived from Hb and GLP-1 intended for a longer duration will be pursued. Tetrameric Hb is prone to oxidation (metHb production), dissociation and denaturation. GLP-1 is also subject to destabilization and denaturation in physiological environment. The central research theme in this aspect will be placed on the stabilization of Hb and GLP-1 based bioactive polymers. For Hb-C, by using intramolecularly crosslinked and/or pyridoxalated Hb and introducing antioxidant enzymes to Hb-C, the stability of Hb-C could be promoted. In addition, the oxygen binding property will be optimized for improved oxygen supply to islets with a detailed mechanistic study of facilitated oxygen transport by directly measuring the profile of oxygen partial _ressure and Hb-C diffusivity in islet capsules. For GLP-1 conjugate, zinc-induced crystallization or precipitation approach will be employed. The precipitate will then be allowed to equilibrate with bioactive soluble GLP-1 conjugates and will be the source of continuous supply of GLP-1 conjugate in islet capsules. A new subject in this research is centered on the construction of effective polymeric gene carriers to pancreatic islets for the transfer of anti-apoptotic Bcl-2 gene. The protection of encapsulated islets from early cytokine assault when transplanted by Bcl-2 expression is an additional critical factor for determining islet survival. This challenge relies on our observation that SU-conjugate was actively internalized into the cells (islets and insulinoma cell line) that express SU receptors. Because the bioactive polymers designed were based on different mechanisms for the same goals of improved islet functionality and prolonged life-span, the combined effects of the bioactive polymers will be evaluated to verify if there is synergistic, summing up or one-component dominant effect.