Kazeminia Sara, Kaur Barnit, Shivam Kumar, Zhu Xiang-Yang, Tang Hui, Jordan Kyra L, Bajpai Shivam, Xue Ailing, Chade Alejandro R, Irazabal Maria V, Lerman Lilach O, Eirin Alfonso
Division of Nephrology and Hypertension, Department of Medicine, Mayo Clinic, Rochester, Minnesota, USA.
Department of Medical Pharmacology and Physiology, University of Missouri-Columbia, Columbia, Missouri, USA.
Am J Nephrol. 2025 Apr 10:1-17. doi: 10.1159/000545795.
CD24+/CD133+ scattered tubular-like cells (STCs) are surviving renal cells that acquire progenitor-like characteristics to repair other damaged kidney cells. Renal artery stenosis (RAS) impairs the reparative capacity of STCs, but the underlying mechanisms remain unknown. STCs contain abundant endoplasmic reticulum (ER), but its capacity to fold proteins could become saturated (ER stress), leading to STC dysfunction. We hypothesized that RAS alters the expression of genes implicated in ER stress in swine STCs.
STCs were harvested from pig kidneys after 10 weeks of RAS or sham (n = 6 each) and expression of ER stress genes was assessed using mRNA-seq (n = 3 each). To elucidate mechanisms regulating ER stress genes in RAS-STCs, integrated mRNA-seq/microRNA (miRNA)-seq and transcription factor (TF) prediction analysis were performed. STC ER stress was assessed in vitro using Western blotting, serial block-face electron microscopy, and mass spectrometry. The involvement of ER stress in regulating the STC-protective effects was also assessed in vitro by their capacity to improve viability of injured human tubular epithelial cells.
RAS pigs developed significant renal dysfunction. mRNA-seq identified 25 ER stress genes upregulated and 30 downregulated in RAS-STCs versus normal-STCs. miRNAs were found to target over a third of all differentially expressed ER stress genes, and almost half of genes encoding for the top 50 TFs involved in regulation of ER stress genes were dysregulated in RAS-STCs. RAS-STCs exhibited higher ER stress compared to normal-STCs, reflected in significant ER dilation and formation of ER-mitochondria contacts and increased levels of ER stress-related amino acids. Importantly, ER stress inhibition improved the reparative capacity of RAS-STCs in vitro.
Renal ischemia alters expression of ER stress-related genes in swine STCs, likely through post-transcriptional- and TF-regulatory mechanisms, which induces ER stress and impairs their reparative potency. These alterations may limit the potential of STCs to repair damaged kidneys in subjects with RAS.
CD24+/CD133+散在管状样细胞（STCs）是存活的肾细胞，具有祖细胞样特征以修复其他受损的肾细胞。肾动脉狭窄（RAS）会损害STCs的修复能力，但其潜在机制尚不清楚。STCs含有丰富的内质网（ER），但其蛋白质折叠能力可能会饱和（内质网应激），导致STC功能障碍。我们假设RAS会改变猪STCs中与内质网应激相关的基因表达。
在RAS或假手术10周后从猪肾中收集STCs（每组n = 6），并使用mRNA测序评估内质网应激基因的表达（每组n = 3）。为了阐明RAS-STCs中调节内质网应激基因的机制，进行了整合的mRNA测序/微小RNA（miRNA）测序和转录因子（TF）预测分析。使用蛋白质印迹、连续块面电子显微镜和质谱在体外评估STC内质网应激。还通过它们改善受损人肾小管上皮细胞活力的能力在体外评估内质网应激在调节STC保护作用中的参与情况。
RAS猪出现明显的肾功能障碍。mRNA测序确定，与正常STCs相比，RAS-STCs中有25个内质网应激基因上调，30个下调。发现miRNA靶向所有差异表达的内质网应激基因的三分之一以上，并且在RAS-STCs中，参与内质网应激基因调节的前50个TF中，几乎一半的编码基因失调。与正常STCs相比，RAS-STCs表现出更高的内质网应激，这反映在内质网明显扩张、内质网与线粒体接触的形成以及内质网应激相关氨基酸水平的增加。重要的是，内质网应激抑制改善了RAS-STCs在体外的修复能力。
肾缺血可能通过转录后和TF调节机制改变猪STCs中内质网应激相关基因的表达，从而诱导内质网应激并损害其修复能力。这些改变可能会限制STCs在RAS患者中修复受损肾脏的潜力。
