泛素连接酶Fbw7负调控成年胰腺导管上皮细胞转分化为胰岛样细胞的研究

Adult pancreas has been highly differentiated, which the regenerative capability into islet cells is limited. Our preclinical observations showed that adult diabetic and non-diabetic pancreatic tissue had insulin-producing cells, suggesting the prospect of ductal cells differentiating into islet-like cells merit further study. This study intends to down-regulation of Fbw7 by siRNA on isolated adult mouse pancreatic ductal epithelial cells and to induce transdifferentiation in vitro, then identifying the morphology and function of transdifferentiated pancreatic islet-like cells. Functional islet cells are transplanted into type 1, type 2 diabetic mice, and then metabolic parameters, such as blood glucose and insulin, are evaluated. The same experimental steps are conducted in human pancreatic ductal epithelial cells (hTERT-HPNE) for transdifferentiation via Fbw7 silence. After identifying the morphological and physiological functional characteristics of differentiated islet-like cells, metabolic indexes in vivo are determined following islet transplantation. In addition, this project applies molecular biology techniques to confirm impact of Fbw7 regulatory factor on substrate protein levels during ubiquitination, as well as to investigate negative regulation of Fbw7 on expression of endocrine specific proteins/genes and critical regulator associated with Notch signaling pathway during cell differentiation. We aim to explore the physiological differentiation potential of adult ductal epithelial cells, and to provide a theoretical basis for transdifferentiation into islet-like cells, thereby offering new avenues for clinical islet transplantation.

成人胰腺已高度分化，其再生胰岛细胞的能力有限。我们的前期临床实践发现成年糖尿病及非糖尿病人胰腺导管具有胰岛素阳性表达的细胞，提示导管分化为胰岛样细胞的前景值得深入研究。本课题拟对分离纯化成年小鼠胰腺导管上皮细胞经siRNA下调Fbw7表达，采用诱导技术转分化为胰岛样细胞后进行形态学和功能鉴定。将功能性胰岛细胞移植入1型、2型糖尿病小鼠模型，进行术后血糖及胰岛素等代谢参数评价。同样的实验手段对人胰腺导管上皮细胞（hTERT-HPNE）进行Fbw7沉默后诱导转分化，鉴定分化胰岛样细胞的形态和生理功能特性，并进行胰岛移植后的体内代谢指标研究。本项目采用分子生物学技术，明确Fbw7调控因子对泛素化降解底物蛋白的影响，以及对特异性蛋白/基因和Notch通路关键调节因子表达的负调控作用。旨在探讨成年胰腺导管上皮细胞的分化潜能，为其转化为胰岛样细胞提供理论依据，进而为临床胰岛移植提供新的途径。

我们本课题在成功构建成年胰腺外分泌导管上皮细胞表型转分化为胰岛功能细胞及探讨其分子机制，为纠正糖尿病病情和自体胰岛移植提供新策略。同时在国家自然基金的资助下（No. 81560134），以第一作者及通讯作者完成发表SCI学术论文8篇（高被引论文1篇），专利1项。发表论文累积影响因子（IF）为29.594，他引次数为65，得到了国内同行的一致认可与研究成果传播。我们积极从事临床实践￫基础研究￫转化医学应用的多向研究体系，为其转化医学提供关键的理论依据。本团队成员通过本项目已经掌握了胰岛分离、细胞培养、免疫组化/荧光染色，荧光定量pcr检测，免疫印迹，流式细胞分选等实验技能，极大促进了本团队的科研技能。此外，我们积极对外需求合作伙伴。目前我们团队已经与香港城市大学Dr Ball Keng Po Lai博士以及日本九州大学Dr William Tse博士开展合作研究。我们已经共同发表了1篇高质量的SCI 学术论文。