Does the mTOR system regulate the renin progenitor cell niche of the juxtaglomerular apparatus under physiological conditions and after mesangial cell injury?

通过建立特异性转基因小鼠模型，我们可以在先前的出版物中证实，肾小球旁细胞龛募集肾素阳性祖细胞对于系膜细胞损伤后的系膜细胞再生和愈合是重要的。祖细胞在进入肾小球内区域时失去肾素阳性，并开始表达全新的典型系膜标记蛋白。因此，我们完全不知道肾素细胞中的哪些信号对肾小球内募集和迁移是重要的。在我们的再生模型之前和过程中，通过转录组分析对肾素谱系细胞进行了无偏见的表征，特别是确定了mTORC1激活蛋白Rheb是肾素细胞在损伤后开始迁移到肾小球的可能触发因素。这一发现也得到了许多额外的试点实验的支持。根据我们的各种试点数据，在本拨款申请中，主要假设将被测试mTORC1激活是肾小球旁肾素系细胞的一个中心细胞信号，这对于这些细胞募集和迁移到肾小球以安排系膜细胞损伤后的成功修复是必要的。同时，我们还将研究mTOR系统在调节/维持肾小球祖细胞生态位中的局部作用，不仅在系膜溶解后再生期间，而且在生理条件下，因为已知mTOR也参与其他前体细胞生态位的细胞死亡/存活。因此，我们探讨了肾小球旁器官肾素阳性前体细胞中特异性raptor （mTORC1途径）或rictor （mTORC2途径）缺乏或两者缺位的影响，以及遗传诱导的（TSC-1缺位）mTORC1的过度激活，可能导致肾小球旁细胞壁龛的不受控制的增加/修饰，甚至不受控制的向肾小球内部位募集。此外，通过肾脏切片共聚焦显微镜和标记肾素谱系细胞迁移过程的日常活体显微镜，比较了局部肾脏mTOR调节与依维莫司系统性mTORC1阻断的影响。这些问题的答案应该在未来以一种更有区别的方式影响重要的mTOR系统，就像持续的系统性mTOR阻断一样。肾素前体细胞中受mTOR调节影响的通路可以通过转录组和蛋白质组分析（结果依赖）进一步分化，为未来的研究开发更具体的靶点。使用肾素特异性细胞系，mTOR调节的表型变化/影响将被表征。此外，mTOR在肾小球旁生态位和肾小球内区域的激活将在不同的人类肾脏疾病中进行研究，以探索其与人类疾病的潜在相关性。

By creation of specific transgenic mouse models, we could verify in previous publications that recruitment of renin-positive progenitor cells from a juxtaglomerular cell niche is important for mesangial cell repopulation and healing after mesangial cell injury. The progenitor cells lose their renin positivity when entering the intraglomerular area and start to express de novo typical mesangial marker proteins. Hereby, it is completely unknown which signals in the renin cells are important for intraglomerular recruitment and immigration.An unbiased approach to characterize the renin lineage cells via transcriptome analysis before and during our regeneration model in particular identified the mTORC1 activator protein Rheb as the possible trigger for the renin cells to start immigration into the glomerulus after injury. This finding is also supported by many additional pilot experiments for this grant application.According to our various pilot data, in this grant application the main hypothesis will be tested that mTORC1 activation is one central cellular signal in the juxtaglomerular renin-lineage cells which is necessary for recruitment and immigration of these cells into the glomerulus to arrange a successful repair after mesangial cell injury. Concurrently, we will also examine the local role of the mTOR system in regulating/maintaining the glomerular progenitor cell niche not only during regeneration after mesangiolysis but also under physiological conditions, since mTOR is known to be involved in cell death/survival of other precursor cell niches as well. Hereby, we explore the effects of specific raptor (mTORC1 pathway) or rictor (mTORC2 pathway) deficiency or both in renin-positive precursor cells of the juxtaglomerular apparatus, as well as a genetically induced (TSC-1 deficiency) hyperactivation of mTORC1, possibly leading to an uncontrolled increase/modification of the juxtaglomerular cell niche or even an uncontrolled recruitment towards intraglomerular sites. In addition, the effects of local renal mTOR modulations are compared with a systemic mTORC1 blockade via everolimus by confocal microscopy of kidney sections and longitudinally via daily intravital microscopy of the immigration process of the labelled renin lineage cells. The answers to these questions are supposed to influence the important mTOR system in a more differentiated way in the future as just continuing systemic mTOR blockade. The pathways influenced by mTOR modulation in the renin precursor cells can be further differentiated by transcriptome and proteome analysis (outcome-dependent) to develop more specific targets for future studies. Using renin specific cell lines the phenotypic changes/effects of mTOR modulation will be characterized. Furthermore, mTOR activation in the juxtaglomerular niche and intraglomerular area will be investigated in different human renal diseases to explore potential relevance for human disease.