Detection of aging mechanisms of nephron degeneration using nanoscale pathology

总结 随着发达国家老年人口的迅速增长，以及 随着移植需求的增加，迫切需要了解衰老相关的生物学， 器官退化在整个生命过程中器官功能的丧失包括实质的进行性丧失， 纤维化结缔组织的积聚。在衰老的肾脏中，肾单位萎缩导致肾组织中的 组织质量和整体过滤能力下降。在整个过程中，近端小管退化 伴随着过量的细胞外基质沉积在基底膜周围， 小管随着年龄的增长导致肾小管间质变性的机制在很大程度上仍然未知，这是 由于缺乏对上皮细胞衰老机制的了解， 由于近曲小管上皮细胞的老化和小管周围纤维化的进展， 是密切相关的事件，在这里，我们建议研究上皮细胞衰老和影响， 衰老相关的分泌表型的管周软骨。首先，我们建议研究衰老- 在来自年轻、成熟、中年和老年小鼠的PTEC中检测相关的核DNA事件，以鉴定 启动和发展DNA损伤成为异染色质沉默复合物，称为衰老， 相关异染色质灶（SAHF）。其次，我们将研究肾小管周膜的变化， 与上皮细胞采用促纤维化衰老相关分泌表型有关。到 为此，我们将研究肾小管周围成纤维细胞的行为和细胞外基质成分的变化， 衰老上皮细胞中含有结缔组织生长因子的分泌囊泡的形成 细胞为了获得对结构染色质变化以及细胞外基质的新见解， 改变，我们将采用一种新的显微镜技术，称为扩展显微镜（ExM），这使得 物理放大复杂的亚细胞结构，允许纳米级分辨率成像， 大分子复合物这项试点研究所产生的信息将大大有助于我们 了解衰老相关的肾单位衰老的机制。

Summary With a rapidly growing elderly population in developed societies, and an organ shortage crisis caused by increasing transplantation demands, there is a pressing need to understand the biology of aging-associated organ deterioration. Loss of organ function throughout life encompasses progressive loss of parenchyma and accumulation of fibrosing connective tissue. In the aging kidney, nephron atrophy results in significant loss of tissue mass and overall decline in filtration capacity. Throughout the process, proximal tubule deterioration occurs concomitantly with excess extracellular matrix deposition in the basement membrane surrounding the tubules. The mechanisms that lead to tubulointerstitial degeneration with age remain largely unknown, this is due to the lack of understanding of the epithelial cell aging mechanisms and the molecular and structural changes associated with it. Because proximal tubule epithelial cell aging and progression of peritubular fibrosis are intimately related events, here we propose to study epithelial cell senescence and the impact of a senescence-associated secretory phenotype on the peritubular interstitium. Firstly, we propose to study aging- associated nuclear DNA events in PTECs from young, mature, middle aged and old mice to identify the initiation and progression of DNA damage into heterochromatin silencing complexes known as senescence- associated heterochromatin foci (SAHFs). Secondly, we will investigate changes in the peritubular interstitium in relation to the adoption of a profibrotic senescence-associated secretory phenotype by epithelial cells. To this end, we will study the behavior of peritubular fibroblasts and changes in extracellular matrix composition in relation to the formation of secretory vesicles containing connective tissue growth factor in senescing epithelial cells. To gain a novel insight into the structural chromatin changes as well as the extracellular matrix alterations, we will employ a novel microscopy technique termed Expansion Microscopy (ExM), which enables physical magnification of complex subcellular structures, allowing nanoscale resolution imaging of macromolecular complexes. The information generated in this pilot study will greatly contribute to our understanding of the mechanisms of aging-associated nephron senescence.