STROMAL/EPITHELIAL INTERACTIONS IN THE BLADDER

该提案的主要目标是研究生长因子的作用 作为膀胱发育过程中基质-上皮相互作用的介质 功能障碍。 我们之前的工作表明膀胱平滑肌（SM） 是由膀胱上皮从未分化的膀胱间充质诱导而来。 该提议基于以下假设：膀胱之间的相互作用 上皮和间质细胞调节和维持SM细胞 发育中和功能障碍的膀胱的分化和生长。 这些 细胞与细胞之间的相互作用是由局部产生和作用介导的 生长因子和其他旁分泌作用介质。 这个假设将 使用尿道阻塞实验模型进行测试，其中 膀胱 SM 通过肥大做出反应。 我们还创建了一个 膀胱增大的实验模型，其中移植的无细胞 膀胱组织基质作为天然组织向内生长的支架 膀胱平滑肌和尿路上皮细胞。 在正常膀胱发育期间和两个实验系统中，细胞 调节正常和异常生长的信号机制 膀胱 SM 的分化将通过追求 遵循特定目标。 （具体目标“1）生长因子的作用 正常膀胱发育中细胞与细胞相互作用的介质 在膀胱圆形修复期间。 生长因子（TGF-α、EGF、 KGF、TGFbeta1、beta2 和 beta3) 和生长因子受体将 在正常膀胱发育和膀胱伤口修复过程中定义 RNase 保护和通过原位杂交定位 免疫组织化学。 （具体目标#2）生长因子和细胞的作用 膀胱肥大发病机制中的细胞信号传导 膀胱部分梗阻。 这一具体目标将由 定义生长因子和生长因子受体表达如下 大鼠部分膀胱梗阻。 这一特定目标的假设 SM 细胞是物理信号的直接目标 纤维肌肥大和弥散性生长因子。 （转化生长因子α， EGF、KGF、TGFbeta1、beta2 和 beta3) 都参与其中。 为了实现这一点 特定目标的嵌合膀胱将通过手术移植整个膀胱来制造 胚胎膀胱或新生儿或成人膀胱条（上皮 完整或移除）进入成人膀胱的逼尿肌下空间。 这个 独特的实验设计将允许研究 SM 的发展和 转基因小鼠膀胱肥大，其中生长因子基因 已被删除或受损（TGFα 敲除、EGF 受体敲除、 TGFbeta1 敲除和 FGFR2 显性失活（角蛋白 14 启动子）小鼠）。 （具体目标#3）使用无细胞膀胱基质贴片来研究正常情况 以及膀胱发育异常。 这一具体目标的策略 是为了表征逼尿肌新生和纤维肌肥大 无细胞膀胱基质移植后。 额外的研究将 评估生长因子表达的区域和时间差异 移植有无细胞膀胱基质的膀胱的膀胱再生。 最后，转基因小鼠膀胱移植的表现 基质驱动的膀胱再生将阐明生长的作用 膀胱再生的因素。 本研究的长期目标 是开发治疗膀胱的有效治疗策略 基于基质-上皮相互作用的功能障碍。

The main goal of this proposal is to investigate the role of growth factors as mediators of stromal-epithelial interactions in bladder development and dysfunction. Our previous work has shown that bladder smooth muscle (SM) is induced from undifferentiated bladder mesenchyme by bladder epithelium. This proposal is based on the hypothesis that interactions between bladder epithelial and mesenchymal cells regulate and maintain SM cell differentiation and growth in developing and dysfunctional bladders. These cell-cell interactions are mediated by the local production and action of growth factors an other paracrine acting mediators. This hypothesis will be tested using an experimental model of urethral obstruction in which bladder SM responds by undergoing hypertrophy. We have also created an experimental model of bladder augmentation in which a grafted acellular bladder tissue matrix serves as a scaffold for the ingrowth of native bladder smooth muscle and urothelial cells. During normal bladder development and in two experimental systems, cellular signaling mechanisms regulating both normal and abnormal growth and differentiation of bladder SM will be examined through pursuit of the following specific aims. (Specific aim "1) Role of growth factors as mediators of cell-cell interactions in normal bladder development and during bladder round repair. Expression of growth factors (TGF-alpha, EGF, KGF, and TGFbeta1, beta2 and beta3) and growth factor receptors will be defined in normal bladder development and during bladder wound repair by RNase protection and localized by in-situ hybridization and immunohistochemistry. (Specific aim #2) Role of growth factors and cell- cell signaling in pathogenesis of bladder hypertrophy resulting from partial obstruction of the bladder. This specific aim will be pursued by defining growth factor and growth factor receptor expression following partial bladder obstruction in rats. The hypothesis of this specific aim is that SM cells are direct targets of the physical signal eliciting fibromuscular hypertrophy and that diffusible growth factors. (TGFalpha, EGF, KGF, TGFbeta1, beta2 and beta3) are involved. To accomplish this specific aim chimeric bladders will be surgically created by grafting whole embryonic bladders or strips of neonatal or adult bladders (epithelium intact or removed) into the sub-detrusor space of adult bladders. This unique experimental design will allow study of SM development and hypertrophy in bladders from transgenic mice in which growth factor genes have been deleted or impaired (TGFalpha knockout, EGF receptor knockout, TGFbeta1 knockout and FGFR2 dominant negative (Keratin 14 promoter) mice). (Specific aim #3) Use of an acellular bladder matrix patch to study normal and abnormal development of the bladder. The strategy of this specific aim is to characterize detrusor neo-formation and fibromuscular hypertrophy following grafting of acellular bladder matrix. Additional studies will assess regional and temporal differences in growth factor expression during bladder regeneration in bladders grafted with acellular bladder matrix. Finally, the performance of transgenic mouse bladder transplants during matrix-driven bladder regeneration will elucidate the role of growth factors in bladder regeneration. The long-term objective of this research is to develop effective therapeutic strategies for treating bladder dysfunction based upon stromal-epithelial interactions.