Bioactive Polymers of Effective Islet Delivery System

本建议书用于R01 DK56884的竞争性续展申请，支持期限为4年。已经设计和测试了生物活性聚合物，以改善由聚乙二醇(HB-C)交联的血红蛋白向被包裹的胰岛的氧气转运，以及通过磺酰尿素(SU)和胰升糖素样肽-1(GLP-1)的水溶性聚合物偶联物刺激胰岛。实验结果表明，聚合物能有效地促进包膜胰岛的功能，延长胰岛的寿命。在这项建议中，将寻求从HB和GLP-1衍生的生物活性聚合物的功能的稳定和扩展，旨在更长的持续时间。四聚体Hb容易氧化(metHb产生)、解离和变性。GLP-1也容易失稳和 生理环境中的变性。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性上。对于Hb-C，采用分子内交联和/或吡哆醇基化的Hb，并在Hb-C中引入抗氧化酶，可以提高Hb-C的稳定性。此外，通过直接测量胰岛胶囊中氧分压和HB-C扩散系数的分布，对促进氧转运的机理进行了详细的研究，从而优化了氧结合性能，以改善胰岛的氧气供应。对于GLP-1偶联物，将采用锌诱导结晶或沉淀法。然后沉淀物将被允许与生物活性平衡 可溶性GLP-1可作为胰岛胶囊中GLP-1结合物的持续供应来源。 本研究的一个新课题是构建有效的胰岛多聚体基因载体，将抗细胞凋亡基因转移到胰岛。被包裹的胰岛在移植时不受早期细胞因子攻击的保护是决定胰岛存活的另一个关键因素。这一挑战依赖于我们的观察，即SU结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。由于设计的生物活性聚合物基于不同的机制，以实现改善胰岛功能和延长寿命的相同目标，因此将对生物活性聚合物的综合效应进行评估，以验证是否存在协同效应、总和效应或单组分显性效应。

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.