Proliferative glomerulonephritis is a severe condition often leading to kidney failure. There is a significant lack of effective treatment for these disorders. Here, following the identification of a somatic PIK3CA gain-of-function mutation in podocytes of a patient, we demonstrate using multiple genetically engineered mouse models, single-cell RNA sequencing and spatial transcriptomics the crucial role played by this pathway for proliferative glomerulonephritis development by promoting podocyte proliferation, dedifferentiation and inflammation. Additionally, we show that alpelisib, a PI3Kα inhibitor, improves glomerular lesions and kidney function in different mouse models of proliferative glomerulonephritis and lupus nephritis by targeting podocytes. Surprisingly, we determined that pharmacological inhibition of PI3Kα affects B and T lymphocyte population in lupus nephritis mouse models with decrease in the production of proinflammatory cytokines, autoantibodies and glomerular complement deposition, which are all characteristic features of PI3K delta (PI3Kδ) inhibition, the primary PI3K isoform expressed in lymphocytes. Importantly, PI3Kα inhibition does not impact lymphocyte function under normal conditions. These findings were then confirmed in human lymphocytes isolated from patients with active lupus nephritis. In conclusion, we demonstrate the major role played by PI3Kα in proliferative glomerulonephritis and show that in this condition, alpelisib acts on both podocytes and the immune system.
Junna Yamaguchi, Pierre Isnard, Noémie Robil, Pierre de la Grange, Clément Hoguin, Alain Schmitt, Aurélie Hummel, Jérôme Mégret, Nicolas Goudin, Marine Luka, Mickaël M. Ménager, Cécile Masson, Mohammed Zarhrate, Christine Bôle-Feysot, Michalina Janiszewska, Kornelia Polyak, Julien Dairou, Sara Baldassari, Stéphanie Baulac, Christine Broissand, Christophe Legendre, Fabiola Terzi, Guillaume Canaud
The smoothened (Smo) receptor facilitates hedgehog signaling between kidney fibroblasts and tubules during acute kidney injury (AKI). Tubule-derived hedgehog is protective in AKI, but the role of fibroblast-selective Smo is unclear. Here, we report that Smo-specific ablation in fibroblasts reduced tubular cell apoptosis and inflammation, enhanced perivascular mesenchymal cells activities, and preserved kidney function after AKI. Global proteomics of these kidneys identified extracellular matrix proteins, and nidogen-1 glycoprotein in particular, as key response markers to AKI. Intriguingly, Smo was bound to nidogen-1 in cells, suggesting that loss of Smo could impact nidogen-1 accessibility. Phosphoproteomics revealed that the ‘AKI protector’ Wnt signaling pathway was activated in these kidneys. Mechanistically, nidogen-1 interacted with integrin β1 to induce Wnts in tubules to mitigate AKI. Altogether, our results support that fibroblast-selective Smo dictates AKI fate through cell-matrix interactions, including nidogen-1, and offers a robust resource and path to further dissect AKI pathogenesis.
Yuan Gui, Haiyan Fu, Zachary Palanza, Jianling Tao, Yi-Han Lin, Wenjian Min, Yi Qiao, Christopher Bonin, Geneva Hargis, Yuanyuan Wang, Peng Yang, Donald L. Kreutzer, Yanlin Wang, Yansheng Liu, Yanbao Yu, Youhua Liu, Dong Zhou
Renal interstitial fibrosis is an important mechanism in the progression of chronic kidney disease (CKD) to end-stage kidney disease. However, we lack specific treatments to slow or halt renal fibrosis. Ribosome profiling identified upregulation of a secreted micropeptide, C4orf48 (Cf48), in mouse diabetic nephropathy. Cf48 RNA and protein levels were upregulated in tubular epithelial cells in human and experimental CKD. Serum Cf48 levels were increased in human CKD and correlated with loss of kidney function, increasing CKD stage, and the degree of active interstitial fibrosis. Cf48 overexpression in mice accelerated renal fibrosis, while Cf48 gene deletion or knockdown by antisense oligonucleotides significantly reduced renal fibrosis in CKD models. In vitro, recombinant Cf48 (rCf48) enhanced TGF-β1-induced fibrotic responses in renal fibroblasts and epithelial cells independent of Smad3 phosphorylation. Cellular uptake of Cf48 and its pro-fibrotic response in fibroblasts operated via the transferrin receptor. RNA immunoprecipitation-sequencing identified Cf48 binding to mRNA of genes involved in the fibrotic response, including Serpine1, Acta2, Ccn2, and Col4a1. rCf48 binds to the 3′-untranslated region of Serpine1 and increases mRNA half-life. We identify the secreted Cf48 micropeptide as a potential enhancer of renal fibrosis which operates as an RNA-binding peptide to promote the production of extracellular matrix.
Jiayi Yang, Hongjie Zhuang, Jinhua Li, Ana B. Nunez-Nescolarde, Ning Luo, Huiting Chen, Andy Li, Xinli Qu, Qing Wang, Jinjin Fan, Xiaoyan Bai, Zhiming Ye, Bing Gu, Yue Meng, Xingyuan Zhang, Di Wu, Youyang Sia, Xiaoyun Jiang, Wei Chen, Alexander N. Combes, David J. Nikolic-Paterson, Xueqing Yu
Tissue regeneration is limited in several organs including the kidney, contributing to the high prevalence of kidney disease globally. However, evolutionary and physiological adaptive responses and the presence of renal progenitor cells suggest existing remodeling capacity. This study uncovered endogenous tissue remodeling mechanisms in the kidney that were activated by the loss of body fluid and salt and regulated by a unique niche of a minority renal cell type called the macula densa (MD). Here we identified neuronal differentiation features of MD cells that sense the local and systemic environment, secrete angiogenic, growth and extracellular matrix remodeling factors, cytokines and chemokines, and control resident progenitor cells. Serial intravital imaging, MD nerve growth factor receptor and Wnt mouse models and transcriptome analysis revealed cellular and molecular mechanisms of these MD functions. Human and therapeutic translation studies illustrated the clinical potential of MD factors including CCN1 as a urinary biomarker and therapeutic target in chronic kidney disease. The concept that a neuronally differentiated key sensory and regulatory cell type responding to organ-specific physiological inputs controls local progenitors to remodel or repair tissues may be applicable to other organs and diverse tissue regenerative therapeutic strategies.
Georgina Gyarmati, Urvi Nikhil Shroff, Anne Riquier-Brison, Dorinne Desposito, Wenjun Ju, Sean D. Stocker, Audrey Izuhara, Sachin Deepak, Alejandra Becerra Calderon, James L. Burford, Hiroyuki Kadoya, Ju-Young Moon, Yibu Chen, Markus M. Rinschen, Nariman Ahmadi, Lester Lau, Daniel Biemesderfer, Aaron W. James, Liliana Minichiello, Berislav Zlokovic, Inderbir S. Gill, Matthias Kretzler, János Peti-Peterdi
Uncontrolled accumulation of extracellular matrix leads to tissue fibrosis and loss of organ function. We previously demonstrated in vitro that the DNA/RNA-binding protein fused in sarcoma (FUS) promotes fibrotic responses by translocating to the nucleus, where it initiates collagen gene transcription. However, it is still not known whether FUS is profibrotic in vivo and whether preventing its nuclear translocation might inhibit development of fibrosis following injury. We now demonstrate that levels of nuclear FUS are significantly increased in mouse models of kidney and liver fibrosis. To evaluate the direct role of FUS nuclear translocation in fibrosis, we used mice that carry a mutation in the FUS nuclear localization sequence (FUSR521G) and the cell-penetrating peptide CP-FUS-NLS that we previously showed inhibits FUS nuclear translocation in vitro. We provide evidence that FUSR521G mice or CP-FUS-NLS–treated mice showed reduced nuclear FUS and fibrosis following injury. Finally, differential gene expression analysis and immunohistochemistry of tissues from individuals with focal segmental glomerulosclerosis or nonalcoholic steatohepatitis revealed significant upregulation of FUS and/or collagen genes and FUS protein nuclear localization in diseased organs. These results demonstrate that injury-induced nuclear translocation of FUS contributes to fibrosis and highlight CP-FUS-NLS as a promising therapeutic option for organ fibrosis.
Manuel Chiusa, Youngmin A. Lee, Ming-Zhi Zhang, Raymond C. Harris, Taylor Sherrill, Volkhard Lindner, Craig R. Brooks, Gang Yu, Agnes B. Fogo, Charles R. Flynn, Jozef Zienkiewicz, Jacek Hawiger, Roy Zent, Ambra Pozzi
Ca2+-activated BK channels in renal intercalated cells (ICs) mediate luminal flow–induced K+ secretion (FIKS), but how ICs sense increased flow remains uncertain. We examined whether PIEZO1, a mechanosensitive Ca2+-permeable channel expressed in the basolateral membranes of ICs, is required for FIKS. In isolated cortical collecting ducts (CCDs), the mechanosensitive cation-selective channel inhibitor GsMTx4 dampened flow-induced increases in intracellular Ca2+ concentration ([Ca2+]i), whereas the PIEZO1 activator Yoda1 increased [Ca2+]i and BK channel activity. CCDs from mice fed a high-K+ (HK) diet exhibited a greater Yoda1-dependent increase in [Ca2+]i than CCDs from mice fed a control K+ diet. ICs in CCDs isolated from mice with a targeted gene deletion of Piezo1 in ICs (IC-Piezo1-KO) exhibited a blunted [Ca2+]i response to Yoda1 or increased flow, with an associated loss of FIKS in CCDs. Male IC-Piezo1-KO mice selectively exhibited an increased blood [K+] in response to an oral K+ bolus and blunted urinary K+ excretion following a volume challenge. Whole-cell expression of BKα subunit was reduced in ICs of IC-Piezo1-KO mice fed an HK diet. We conclude that PIEZO1 mediates flow-induced basolateral Ca2+ entry into ICs, is upregulated in the CCD in response to an HK diet, and is necessary for FIKS.
Rolando Carrisoza-Gaytan, Stephanie M. Mutchler, Francisco Carattino, Joanne Soong, Marianela G. Dalghi, Peng Wu, WenHui Wang, Gerard Apodaca, Lisa M. Satlin, Thomas R. Kleyman
Maggie E. Jones-Carr, Huma Fatima, Vineeta Kumar, Douglas J. Anderson, Julie Houp, Jackson C. Perry, Gavin A. Baker, Leigh McManus, Andrew J. Shunk, Paige M. Porrett, Jayme E. Locke
Two coding variants of apolipoprotein L1 (APOL1) called G1 and G2 explain much of the excess risk of kidney disease in African Americans. While various cytotoxic phenotypes have been reported in experimental models, the proximal mechanism by which G1 and G2 cause kidney disease is poorly understood. Here, we leveraged three experimental models and a recently reported small molecule blocker of APOL1 protein, VX-147, to identify the upstream mechanism of G1-induced cytotoxicity. In HEK293 cells, we demonstrated that G1-mediated Na+ import/K+ efflux triggered activation of G protein-coupled receptor (GPCR)-IP3-mediated calcium release from the endoplasmic reticulum (ER), impaired mitochondrial ATP production, and impaired translation, which were all reversed by VX-147. In human podocyte-like epithelial cells (HUPEC), we demonstrated that G1 caused cytotoxicity that was again reversible by VX-147. Finally, in podocytes isolated from APOL1 G1 transgenic mice, we showed that Interferon gamma (IFNγ)-mediated induction of G1 caused K+ efflux, activation of GPCR-IP3 signaling, and inhibition of translation, podocyte injury, and proteinuria, all reversed by VX-147. Together, these results establish APOL1-mediated Na+/K+ transport as the proximal driver of APOL1-mediated kidney disease.
Somenath Datta, Brett M. Antonio, Nathan H. Zahler, Jonathan W. Theile, Doug Krafte, Hengtao Zhang, Paul B. Rosenberg, Alec B. Chaves, Deborah M. Muoio, Guofang Zhang, Daniel Silas, Guojie Li, Karen Soldano, Sarah Nystrom, Davis Ferreira, Sara E. Miller, James R. Bain, Michael J. Muehlbauer, Olga Ilkayeva, Thomas C. Becker, Hans-Ewald Hohmeier, Christopher B. Newgard, Opeyemi A. Olabisi
Worldwide, over 800 million people are affected by kidney disease, yet its pathogenesis remains elusive, hindering the development of novel therapeutics. In this study, we employed kidney-specific expression of quantitative traits and single-nuclear open chromatin analysis to show that genetic variants linked to kidney dysfunction on chromosome 20 target the acyl-CoA synthetase short-chain family 2 (ACSS2). By generating ACSS2 knock-out mice, we demonstrated their protection from kidney fibrosis in multiple disease models. Our analysis of primary tubular cells revealed that ACSS2 regulates de novo lipogenesis (DNL), causing NADPH depletion and increasing ROS levels, ultimately leading to NLRP3-dependent pyroptosis. Additionally, we discovered that pharmacological inhibition or genetic ablation of fatty acid synthase safeguarded kidney cells against profibrotic gene expression and prevented kidney disease in mice. Lipid accumulation and the expression of genes related to DNL were elevated in the kidneys of patients with fibrosis. Our findings pinpoint ACSS2 as a critical kidney disease gene and reveal the role of DNL in kidney disease.
Dhanunjay Mukhi, Lingzhi Li, Hongbo Liu, Tomohito Doke, Lakshmi P. Kolligundla, Eunji Ha, Konstantin A. Klötzer, Amin Abedini, Sarmistha Mukherjee, Junnan Wu, Poonam Dhillon, Hailong Hu, Dongyin Guan, Katsuhiko Funai, Kahealani Uehara, Paul M. Titchenell, Joseph A. Baur, Kathryn E. Wellen, Katalin Susztak
Mineralocorticoid excess commonly leads to hypertension and kidney disease. In our study, we employed single-cell expression and chromatin accessibility tools to characterize the mineralocorticoid target genes and cell types. We demonstrated that mineralocorticoid effects are established through open chromatin and target gene expression, primarily in principal and connecting tubule cells, and to a lesser extent, in segments of the distal convoluted tubule cells. We examined the kidney-protective effects of steroidal and non-steroidal mineralocorticoid antagonists (MRAs), as well as amiloride, an epithelial sodium channel inhibitor, in a rat model of deoxycorticosterone acetate, unilateral nephrectomy, and high salt consumption-induced hypertension and cardiorenal damage. All antihypertensive therapies protected from cardiorenal damage. However, finerenone was particularly effective in reducing albuminuria and improving gene expression changes in podocytes and proximal tubule cells, even with equivalent blood pressure reduction. There was a strong correlation between the accumulation of injured/profibrotic tubule cells expressing Spp1, Il34, and Pdgfb and the degree of fibrosis in rat kidneys. This gene signature also showed potential for classifying human kidney samples. Our multi-omics approach provides fresh insights into the possible mechanisms underlying hypertension associated kidney disease, the target cell types, and the protective effects of steroidal, non-steroidal MRAs, and amiloride.
Amin Abedini, Andrea Sanchez-Navarro, Junnan Wu, Konstantin A. Klötzer, Ziyuan Ma, Bibek Poudel, Tomohito Doke, Michael S. Balzer, Julia Frederick, Hana Cernecka, Hongbo Liu, Xiujie Liang, Steven Vitale, Peter Kolkhof, Katalin Susztak