腺相关病毒对糖尿病膀胱基因治疗的应用及机制研究

Diabetic cystopathy (DCP) is the most common complication of diabetes in the urinary system, the late manifestations of DCP is that the detrusor muscle fascicle arranged in disorder and detrusor myocyte appear a apoptosis-like changes. Studies have confirmed that SERCA2a is a class of ATP-dependent calcium channel protein, and its down expression were closely related to detrusor contraction disfunction and detrusor cell apoptosis, but the function of SERCA2a in Diabetic cystopathy is not clear.Our previous study found that with the progression of diabetes, miR-25 expression in detrusor cells gradually increased, SERCA2a protein expression decreased, and detrusor cells injury worsened. Therefore, we hypothesize that the miR-25 / SERCA2a disorder affects detrusor contractility and promotes detrusor cells apoptosis by regulating intracellular calcium concentration. This project will be combined with in vitro and in vivo studies to reveal the upstream and downstream signal that how miR-25 / SERCA2a regulates detrusor contractility and inhibits detrusor myocytes apoptosis. Adeno-associated virus was also used as a vector to induce the expression of miR-25 / SERCA2a in bladder detrusor cells .The dynamic changes of urodynamics and detrusor contraction in the animal model were observed to provide new theoretical basis and experimental basis for the individualized treatment of diabetic cystopathy patients.

糖尿病膀胱是糖尿病在泌尿系统最主要的并发症，其晚期主要表现为膀胱逼尿肌肌束排列紊乱，逼尿肌细胞出现凋亡样改变。有研究证实SERCA2a是一类ATP依赖的钙离子通道蛋白，其表达失衡与肌肉收缩功能受损及肌细胞凋亡密切相关, 但其在糖尿病膀胱的作用尚不明确。本课题组前期研究发现，随着糖尿病病程进展，逼尿肌细胞中的miR-25表达量逐步升高，SERCA2a蛋白表达量降低，逼尿肌损伤加重。因此我们推测miR-25/SERCA2a紊乱通过调控细胞内钙离子浓度，影响逼尿肌收缩功能并促进肌细胞凋亡。本项目将结合体内外研究，揭示miR-25/SERCA2a调控逼尿肌收缩功能并抑制逼尿肌细胞凋亡的上下游机制，并使用腺相关病毒作为载体，以膀胱灌注的方式干预逼尿肌细胞中miR-25/SERCA2a的表达，观察动物模型尿动力学变化、膀胱逼尿肌收缩变化情况，以期为糖尿病膀胱患者的个体化治疗提供新的理论依据和实验支持。

糖尿病膀胱是糖尿病在泌尿系统最主要的并发症，其晚期主要表现为膀胱逼尿肌肌束排列紊乱，逼尿肌细胞出现凋亡样改变。本研究发现在大鼠糖尿病模型中，随病程进展，其膀胱逼尿肌细胞中的miR-25表达量逐步升高，SERCA2a蛋白表达量逐步降低，且其逼尿肌细胞内钙离子浓度负荷增加，细胞明显凋亡。分离糖尿病大鼠的膀胱逼尿肌并向其中转染SERCA2a蛋白后，可见细胞内钙离子浓度负荷降低，细胞凋亡减少。本研究同时使用生物信息学分析及荧光素酶报告基因实验证明，miR-25可以负向调控SERCA2a蛋白表达，从而抑制逼尿肌细胞生长，增强其细胞内钙离子负荷，诱导细胞凋亡，促进糖尿病膀胱的形成。该研究为糖尿病膀胱患者的个体化治疗提供了一个非常有价值的思路及实验依据。