下尿路梗阻与膀胱癌复发相关性的生物力学研究

Bladder cancer is a highly recurrent malignant tumor of the urinary system. The recurrence rate of bladder cancer is significantly higher in the case of lower urinary tract obstruction. Lower urinary tract obstruction can lead to changes of macro structure and mechanical characterization of bladder tissue, which will cause the changes of biomechanical parameters of bladder and can be closely related to the recurrence of cancer. Based on the highly recurrence rate of bladder cancer with lower urinary tract obstruction, use biomechanical numerical simulation model of the anatomical structure and function of the lower urinary tract system and molecular biology methods (1) establish the urinary system biomechanical numerical model simulation of Chinese people and Wistar rat (2) with the help of microfluidic chip and animal model, to test and verify that the changes of biomechanical parameters in bladder was correlated with the malignant behavior of cancer cells. And then, analyse the results (3) calculate the key mechanical parameters and mechanical threshold, screening the key factors of signal transduction pathway, and then, explore the molecular mechanism that the change of biomechanics factors can influence the biological behavior of malignant cells. In this study, the relationship between lower urinary tract obstruction and recurrence of bladder cancer was analyzed by computational biomechanics. The obiective is to provide new evidence for finding the opportunity to release obstruction and reduce the recurrence rate of bladder cancer.

膀胱癌是泌尿系统极易复发的恶性肿瘤，而合并下尿路梗阻时膀胱癌复发率明显升高。下尿路梗阻能导致膀胱宏观结构及生物力学表征发生变化，继发力学响应异变，且认为该因素与膀胱癌的复发密切相关。项目以下尿路梗阻导致膀胱癌复发率增高为研究背景，通过下尿路系统解剖结构及功能的生物力学数值模型与仿真、分子生物学等学科交叉手段（1）采集合并下尿路梗阻复发性膀胱癌患者及大鼠的力学表征基线资料，建立国人和大鼠泌尿系统生物力学仿真数值模型（2）采用微流控芯片实验平台体外细胞三维培养及动物实验，验证膀胱内力学响应异变能够对膀胱癌细胞的恶性生物学行为产生影响，并进行定量分析（3）提取关键力学参数及阈值，筛选力学信号传导通路关键因子，探索生物力学因素影响膀胱癌细胞恶性生物学行为的相关分子机制。本研究通过生物力学研究手段分析下尿路梗阻与膀胱癌复发相关性，旨在为临床及时解除梗阻进而降低膀胱癌复发率的手术时机选择提供新的依据。

生物力学因素对膀胱癌恶性行为具有重要的作用，合并下尿路梗阻时膀胱癌复发率明显升高，且下尿路梗阻能导致膀胱宏观结构及力学表征发生变化，致使膀胱壁明显增厚，肿瘤基底宽，浸润深度增加，呈扁平样浸润型生长，形态上反映了肿瘤侵袭能力增强。通过组织弹性超声检查肿瘤弹性模量，发现合并下尿路梗阻患者膀胱肿瘤刚度大，肿瘤间质含量明显增加。我们根据HE病理切片上膀胱肿瘤与间质的比例与形态将患者进一步分组，结合患者临床资料发现间质高且间质与肿瘤相混合的膀胱肿瘤易出现转移复发，结合基因组测序数据我们发现这组患者高表达免疫炎症、表皮间充质转变相关基因。因此，下尿路梗阻有可能通过影响膀胱肿瘤间质的含量及混合状态从而影响膀胱癌复发。我们进一步比较各组的转录组数据，发现ZBTB7A可能与膀胱癌间质改变及恶性生物学行为相关。.通过免疫组化我们发现合并下尿路梗阻患者肿瘤细胞中ZBTB7A表达明显增加，我们通过构建稳转细胞系、平板克隆、Transwell、裸鼠成瘤实验，发现敲减ZBTB7A能抑制膀胱癌细胞的生长和迁移，并能降低裸鼠成瘤的大小和重量。通过基因测序、双荧光素酶、染色质免疫共沉淀等实验，我们发现ZBTB7A能特异性的与HIC1的启动子结合，从而抑制HIC1的转录，在膀胱癌中起到促癌作用。miR-144-3p能通过靶向结合ZBTB7A 3’UTR，抑制ZBTB7A的翻译，miR144-3p的靶向基因ZBTB7A通过下调HIC1的表达促进膀胱肿瘤的生长迁移。根据研究结果我们推测解除下尿路梗阻能减少膀胱癌的间质含量，从而降低肿瘤刚度，延长膀胱肿瘤复发时间，而miR-144-3p/ZBTB7A/HIC1信号轴在膀胱癌进展中发挥重要作用，为膀胱癌的治疗提供新的思路。