The thiazide-sensitive NaCl cotransporter (NCC) is important for renal salt handling and blood-pressure homeostasis. The canonical NCC-activating pathway consists of With-No-Lysine (WNK) kinases and their downstream effector kinases SPAK and OSR1, which phosphorylate NCC directly. The upstream mechanisms that connect physiological stimuli to this system remain obscure. Here, we have shown that aldosterone activates SPAK/OSR1 via WNK1. We identified 2 alternatively spliced exons embedded within a proline-rich region of WNK1 that contain PY motifs, which bind the E3 ubiquitin ligase NEDD4-2. PY motif–containing WNK1 isoforms were expressed in human kidney, and these isoforms were efficiently degraded by the ubiquitin proteasome system, an effect reversed by the aldosterone-induced kinase SGK1. In gene-edited cells, WNK1 deficiency negated regulatory effects of NEDD4-2 and SGK1 on NCC, suggesting that WNK1 mediates aldosterone-dependent activity of the WNK/SPAK/OSR1 pathway. Aldosterone infusion increased proline-rich WNK1 isoform abundance in WT mice but did not alter WNK1 abundance in hypertensive
Ankita Roy, Lama Al-Qusairi, Bridget F. Donnelly, Caroline Ronzaud, Allison L. Marciszyn, Fan Gong, Y.P. Christy Chang, Michael B. Butterworth, Núria M. Pastor-Soler, Kenneth R. Hallows, Olivier Staub, Arohan R. Subramanya
Chronic kidney disease is characterized by interstitial fibrosis and proliferation of scar-secreting myofibroblasts, ultimately leading to end-stage renal disease. The hedgehog (Hh) pathway transcriptional effectors GLI1 and GLI2 are expressed in myofibroblast progenitors; however, the role of these effectors during fibrogenesis is poorly understood. Here, we demonstrated that GLI2, but not GLI1, drives myofibroblast cell-cycle progression in cultured mesenchymal stem cell–like progenitors. In animals exposed to unilateral ureteral obstruction, Hh pathway suppression by expression of the GLI3 repressor in GLI1+ myofibroblast progenitors limited kidney fibrosis. Myofibroblast-specific deletion of
Rafael Kramann, Susanne V. Fleig, Rebekka K. Schneider, Steven L. Fabian, Derek P. DiRocco, Omar Maarouf, Janewit Wongboonsin, Yoichiro Ikeda, Dirk Heckl, Steven L. Chang, Helmut G. Rennke, Sushrut S. Waikar, Benjamin D. Humphreys
Macrophages (Mø) are integral in ischemia/reperfusion injury–incited (I/R-incited) acute kidney injury (AKI) that leads to fibrosis and chronic kidney disease (CKD). IL-34 and CSF-1 share a receptor (c-FMS), and both cytokines mediate Mø survival and proliferation but also have distinct features. CSF-1 is central to kidney repair and destruction. We tested the hypothesis that IL-34–dependent, Mø-mediated mechanisms promote persistent ischemia-incited AKI that worsens subsequent CKD. In renal I/R, the time-related magnitude of Mø-mediated AKI and subsequent CKD were markedly reduced in IL-34–deficient mice compared with controls. IL-34, c-FMS, and a second IL-34 receptor, protein-tyrosine phosphatase ζ (PTP-ζ) were upregulated in the kidney after I/R. IL-34 was generated by tubular epithelial cells (TECs) and promoted Mø-mediated TEC destruction during AKI that worsened subsequent CKD via 2 distinct mechanisms: enhanced intrarenal Mø proliferation and elevated BM myeloid cell proliferation, which increases circulating monocytes that are drawn into the kidney by chemokines. CSF-1 expression in TECs did not compensate for IL-34 deficiency. In patients, kidney transplants subject to I/R expressed IL-34, c-FMS, and PTP−ζ in TECs during AKI that increased with advancing injury. Moreover, IL-34 expression increased, along with more enduring ischemia in donor kidneys. In conclusion, IL-34-dependent, Mø-mediated, CSF-1 nonredundant mechanisms promote persistent ischemia-incited AKI that worsens subsequent CKD.
Jea-Hyun Baek, Rui Zeng, Julia Weinmann-Menke, M. Todd Valerius, Yukihiro Wada, Amrendra K. Ajay, Marco Colonna, Vicki R. Kelley
Kidney size adaptively increases as mammals grow and in response to the loss of 1 kidney. It is not clear how kidneys size themselves or if the processes that adapt kidney mass to lean body mass also mediate renal hypertrophy following unilateral nephrectomy (UNX). Here, we demonstrated that mice harboring a proximal tubule–specific deletion of
Jian-Kang Chen, Kojiro Nagai, Jianchun Chen, David Plieth, Masayo Hino, Jinxian Xu, Feng Sha, T. Alp Ikizler, C. Chad Quarles, David W. Threadgill, Eric G. Neilson, Raymond C. Harris
Steroid-resistant nephrotic syndrome (SRNS) is a frequent cause of progressive renal function decline and affects millions of people. In a recent study, 30% of SRNS cases evaluated were the result of monogenic mutations in 1 of 27 different genes. Here, using homozygosity mapping and whole-exome sequencing, we identified recessive mutations in
Heon Yung Gee, Fujian Zhang, Shazia Ashraf, Stefan Kohl, Carolin E. Sadowski, Virginia Vega-Warner, Weibin Zhou, Svjetlana Lovric, Humphrey Fang, Margaret Nettleton, Jun-yi Zhu, Julia Hoefele, Lutz T. Weber, Ludmila Podracka, Andrej Boor, Henry Fehrenbach, Jeffrey W. Innis, Joseph Washburn, Shawn Levy, Richard P. Lifton, Edgar A. Otto, Zhe Han, Friedhelm Hildebrandt
Autosomal dominant polycystic kidney disease (ADPKD) is characterized by renal cyst formation, inflammation, and fibrosis. Macrophages infiltrate cystic kidneys, but the role of these and other inflammatory factors in disease progression are poorly understood. Here, we identified macrophage migration inhibitory factor (MIF) as an important regulator of cyst growth in ADPKD. MIF was upregulated in cyst-lining epithelial cells in polycysitn-1–deficient murine kidneys and accumulated in cyst fluid of human ADPKD kidneys. MIF promoted cystic epithelial cell proliferation by activating ERK, mTOR, and Rb/E2F pathways and by increasing glucose uptake and ATP production, which inhibited AMP-activated protein kinase signaling. MIF also regulated cystic renal epithelial cell apoptosis through p53-dependent signaling. In polycystin-1–deficient mice, MIF was required for recruitment and retention of renal macrophages, which promoted cyst expansion, and
Li Chen, Xia Zhou, Lucy X. Fan, Ying Yao, Katherine I. Swenson-Fields, Mihaela Gadjeva, Darren P. Wallace, Dorien J.M. Peters, Alan Yu, Jared J. Grantham, Xiaogang Li
Podocytes are specialized epithelial cells in the kidney glomerulus that play important structural and functional roles in maintaining the filtration barrier. Nephrotic syndrome results from a breakdown of the kidney filtration barrier and is associated with proteinuria, hyperlipidemia, and edema. Additionally, podocytes undergo changes in morphology and internalize plasma proteins in response to this disorder. Here, we used fluid-phase tracers in murine models and determined that podocytes actively internalize fluid from the plasma and that the rate of internalization is increased when the filtration barrier is disrupted. In cultured podocytes, the presence of free fatty acids (FFAs) associated with serum albumin stimulated macropinocytosis through a pathway that involves FFA receptors, the Gβ/Gγ complex, and RAC1. Moreover, mice with elevated levels of plasma FFAs as the result of a high-fat diet were more susceptible to Adriamycin-induced proteinuria than were animals on standard chow. Together, these results support a model in which podocytes sense the disruption of the filtration barrier via FFAs bound to albumin and respond by enhancing fluid-phase uptake. The response to FFAs may function in the development of nephrotic syndrome by amplifying the effects of proteinuria.
Jun-Jae Chung, Tobias B. Huber, Markus Gödel, George Jarad, Björn Hartleben, Christopher Kwoh, Alexander Keil, Aleksey Karpitskiy, Jiancheng Hu, Christine J. Huh, Marina Cella, Richard W. Gross, Jeffrey H. Miner, Andrey S. Shaw
Thiazide diuretics are used to treat hypertension; however, compensatory processes in the kidney can limit antihypertensive responses to this class of drugs. Here, we evaluated compensatory pathways in SPAK kinase–deficient mice, which are unable to activate the thiazide-sensitive sodium chloride cotransporter NCC (encoded by
P. Richard Grimm, Yoskaly Lazo-Fernandez, Eric Delpire, Susan M. Wall, Susan G. Dorsey, Edward J. Weinman, Richard Coleman, James B. Wade, Paul A. Welling
Miguel Verbitsky, Simone Sanna-Cherchi, David A. Fasel, Brynn Levy, Krzysztof Kiryluk, Matthias Wuttke, Alison G. Abraham, Frederick Kaskel, Anna Köttgen, Bradley A. Warady, Susan L. Furth, Craig S. Wong, Ali G. Gharavi
Familial forms of focal segmental glomerulosclerosis (FSGS) have been linked to gain-of-function mutations in the gene encoding the transient receptor potential channel C6 (TRPC6). GPCRs coupled to Gq signaling activate TRPC6, suggesting that Gq-dependent TRPC6 activation underlies glomerular diseases. Here, we developed a murine model in which a constitutively active Gq α subunit (GqQ209L, referred to herein as GqQ>L) is specifically expressed in podocytes and examined the effects of this mutation in response to puromycin aminonucleoside (PAN) nephrosis. We found that compared with control animals, animals expressing GqQ>L exhibited robust albuminuria, structural features of FSGS, and reduced numbers of glomerular podocytes. Gq activation stimulated calcineurin (CN) activity, resulting in CN-dependent upregulation of TRPC6 in murine kidneys. Deletion of TRPC6 in GqQ>L-expressing mice prevented FSGS development and inhibited both tubular damage and podocyte loss induced by PAN nephrosis. Similarly, administration of the CN inhibitor FK506 reduced proteinuria and tubular injury but had more modest effects on glomerular pathology and podocyte numbers in animals with constitutive Gq activation. Moreover, these Gq-dependent effects on podocyte injury were generalizable to diabetic kidney disease, as expression of GqQ>L promoted albuminuria, mesangial expansion, and increased glomerular basement membrane width in diabetic mice. Together, these results suggest that targeting Gq/TRPC6 signaling may have therapeutic benefits for the treatment of glomerular diseases.
Liming Wang, Grant Jirka, Paul B. Rosenberg, Anne F. Buckley, Jose A. Gomez, Timothy A. Fields, Michelle P. Winn, Robert F. Spurney
The HSP40 cochaperone SEC63 is associated with the SEC61 translocon complex in the ER. Mutations in the gene encoding SEC63 cause polycystic liver disease in humans; however, it is not clear how altered SEC63 influences disease manifestations. In mice, loss of SEC63 induces cyst formation both in liver and kidney as the result of reduced polycystin-1 (PC1). Here we report that inactivation of SEC63 induces an unfolded protein response (UPR) pathway that is protective against cyst formation. Specifically, using murine genetic models, we determined that SEC63 deficiency selectively activates the IRE1α-XBP1 branch of UPR and that SEC63 exists in a complex with PC1. Concomitant inactivation of both SEC63 and XBP1 exacerbated the polycystic kidney phenotype in mice by markedly suppressing cleavage at the G protein–coupled receptor proteolysis site (GPS) in PC1. Enforced expression of spliced XBP1 (XBP1s) enhanced GPS cleavage of PC1 in SEC63-deficient cells, and XBP1 overexpression in vivo ameliorated cystic disease in a murine model with reduced PC1 function that is unrelated to SEC63 inactivation. Collectively, the findings show that SEC63 function regulates IRE1α/XBP1 activation, SEC63 and XBP1 are required for GPS cleavage and maturation of PC1, and activation of XBP1 can protect against polycystic disease in the setting of impaired biogenesis of PC1.
Sorin V. Fedeles, Jae-Seon So, Amol Shrikhande, Seung Hun Lee, Anna-Rachel Gallagher, Christina E. Barkauskas, Stefan Somlo, Ann-Hwee Lee
Kidney injury molecule 1 (KIM-1, also known as TIM-1) is markedly upregulated in the proximal tubule after injury and is maladaptive when chronically expressed. Here, we determined that early in the injury process, however, KIM-1 expression is antiinflammatory due to its mediation of phagocytic processes in tubule cells. Using various models of acute kidney injury (AKI) and mice expressing mutant forms of KIM-1, we demonstrated a mucin domain–dependent protective effect of epithelial KIM-1 expression that involves downregulation of innate immunity. Deletion of the mucin domain markedly impaired KIM-1–mediated phagocytic function, resulting in increased proinflammatory cytokine production, decreased antiinflammatory growth factor secretion by proximal epithelial cells, and a subsequent increase in tissue macrophages. Mice expressing KIM-1Δmucin had greater functional impairment, inflammatory responses, and mortality in response to ischemia- and cisplatin-induced AKI. Compared with primary renal proximal tubule cells isolated from KIM-1Δmucin mice, those from WT mice had reduced proinflammatory cytokine secretion and impaired macrophage activation. The antiinflammatory effect of KIM-1 expression was due to the interaction of KIM-1 with p85 and subsequent PI3K-dependent downmodulation of NF-κB. Hence, KIM-1–mediated epithelial cell phagocytosis of apoptotic cells protects the kidney after acute injury by downregulating innate immunity and inflammation.
Li Yang, Craig R. Brooks, Sheng Xiao, Venkata Sabbisetti, Melissa Y. Yeung, Li-Li Hsiao, Takaharu Ichimura, Vijay Kuchroo, Joseph V. Bonventre
Maintenance of mitochondrial structure and function is critical for preventing podocyte apoptosis and eventual glomerulosclerosis in the kidney; however, the transcription factors that regulate mitochondrial function in podocyte injury remain to be identified. Here, we identified Krüppel-like factor 6 (KLF6), a zinc finger domain transcription factor, as an essential regulator of mitochondrial function in podocyte apoptosis. We observed that podocyte-specific deletion of
Sandeep K. Mallipattu, Sylvia J. Horne, Vivette D’Agati, Goutham Narla, Ruijie Liu, Michael A. Frohman, Kathleen Dickman, Edward Y. Chen, Avi Ma’ayan, Agnieszka B. Bialkowska, Amr M. Ghaleb, Mandayam O. Nandan, Mukesh K. Jain, Ilse Daehn, Peter Y. Chuang, Vincent W. Yang, John C. He
The lymphocyte adaptor protein LNK (also known as SH2B3) is primarily expressed in hematopoietic and endothelial cells, where it functions as a negative regulator of cytokine signaling and cell proliferation. Single-nucleotide polymorphisms in the gene encoding LNK are associated with autoimmune and cardiovascular disorders; however, it is not known how LNK contributes to hypertension. Here, we determined that loss of LNK exacerbates angiotensin II–induced (Ang II–induced) hypertension and the associated renal and vascular dysfunction. At baseline, kidneys from
Mohamed A. Saleh, William G. McMaster, Jing Wu, Allison E. Norlander, Samuel A. Funt, Salim R. Thabet, Annet Kirabo, Liang Xiao, Wei Chen, Hana A. Itani, Danielle Michell, Tianxiao Huan, Yahua Zhang, Satoshi Takaki, Jens Titze, Daniel Levy, David G. Harrison, Meena S. Madhur
Acute kidney injury (AKI) is a public health concern with an annual mortality rate that exceeds those of breast and prostate cancer, heart failure, and diabetes combined. Oxidative stress and mitochondrial damage are drivers of AKI-associated pathology; however, the pathways that mediate these events are poorly defined. Here, using a murine cisplatin-induced AKI model, we determined that both oxidative stress and mitochondrial damage are associated with reduced levels of renal sirtuin 3 (SIRT3). Treatment with the AMPK agonist AICAR or the antioxidant agent acetyl-
Marina Morigi, Luca Perico, Cinzia Rota, Lorena Longaretti, Sara Conti, Daniela Rottoli, Rubina Novelli, Giuseppe Remuzzi, Ariela Benigni
Fibrosis underlies the loss of renal function in patients with chronic kidney disease (CKD) and in kidney transplant recipients with chronic allograft nephropathy (CAN). Here, we studied the effect of an intronic SNP in
Madhav C. Menon, Peter Y. Chuang, Zhengzhe Li, Chengguo Wei, Weijia Zhang, Yi Luan, Zhengzi Yi, Huabao Xiong, Christopher Woytovich, Ilana Greene, Jessica Overbey, Ivy Rosales, Emilia Bagiella, Rong Chen, Meng Ma, Li Li, Wei Ding, Arjang Djamali, Millagros Saminego, Philip J. O’Connell, Lorenzo Gallon, Robert Colvin, Bernd Schroppel, John Cijiang He, Barbara Murphy
The epithelial Na+ channel (ENaC) is essential for Na+ homeostasis, and dysregulation of this channel underlies many forms of hypertension. Recent studies suggest that mTOR regulates phosphorylation and activation of serum/glucocorticoid regulated kinase 1 (SGK1), which is known to inhibit ENaC internalization and degradation; however, it is not clear whether mTOR contributes to the regulation of renal tubule ion transport. Here, we evaluated the effect of selective mTOR inhibitors on kidney tubule Na+ and K+ transport in WT and
Catherine E. Gleason, Gustavo Frindt, Chih-Jen Cheng, Michael Ng, Atif Kidwai, Florian Lang, Michel Baum, Lawrence G. Palmer, David Pearce
MicroRNA-21 (miR-21) contributes to the pathogenesis of fibrogenic diseases in multiple organs, including the kidneys, potentially by silencing metabolic pathways that are critical for cellular ATP generation, ROS production, and inflammatory signaling. Here, we developed highly specific oligonucleotides that distribute to the kidney and inhibit miR-21 function when administered subcutaneously and evaluated the therapeutic potential of these anti–miR-21 oligonucleotides in chronic kidney disease. In a murine model of Alport nephropathy, miR-21 silencing did not produce any adverse effects and resulted in substantially milder kidney disease, with minimal albuminuria and dysfunction, compared with vehicle-treated mice. miR-21 silencing dramatically improved survival of Alport mice and reduced histological end points, including glomerulosclerosis, interstitial fibrosis, tubular injury, and inflammation. Anti–miR-21 enhanced PPARα/retinoid X receptor (PPARα/RXR) activity and downstream signaling pathways in glomerular, tubular, and interstitial cells. Moreover, miR-21 silencing enhanced mitochondrial function, which reduced mitochondrial ROS production and thus preserved tubular functions. Inhibition of miR-21 was protective against TGF-β–induced fibrogenesis and inflammation in glomerular and interstitial cells, likely as the result of enhanced PPARα/RXR activity and improved mitochondrial function. Together, these results demonstrate that inhibition of miR-21 represents a potential therapeutic strategy for chronic kidney diseases including Alport nephropathy.
Ivan G. Gomez, Deidre A. MacKenna, Bryce G. Johnson, Vivek Kaimal, Allie M. Roach, Shuyu Ren, Naoki Nakagawa, Cuiyan Xin, Rick Newitt, Shweta Pandya, Tai-He Xia, Xueqing Liu, Dorin-Bogdan Borza, Monica Grafals, Stuart J. Shankland, Jonathan Himmelfarb, Didier Portilla, Shiguang Liu, B. Nelson Chau, Jeremy S. Duffield
The most severe form of autosomal dominant polycystic kidney disease occurs in patients with mutations in the gene (
Yiqiang Cai, Sorin V. Fedeles, Ke Dong, Georgia Anyatonwu, Tamehito Onoe, Michihiro Mitobe, Jian-Dong Gao, Dayne Okuhara, Xin Tian, Anna-Rachel Gallagher, Zhangui Tang, Xiaoli Xie, Maria D. Lalioti, Ann-Hwee Lee, Barbara E. Ehrlich, Stefan Somlo
Ischemia is a leading cause of acute kidney injury. Kidney ischemia is associated with loss of cellular ion homeostasis; however, the pathways that underlie ion homeostasis dysfunction are poorly understood. Here, we evaluated the nonselective cation channel transient receptor potential melastatin 2 (TRPM2) in a murine model of kidney ischemia/reperfusion (I/R) injury. TRPM2-deficient mice were resistant to ischemic injury, as reflected by improved kidney function, reduced histologic damage, suppression of proapoptotic pathways, and reduced inflammation. Moreover, pharmacologic TRPM2 inhibition was also protective against I/R injury. TRPM2 was localized mainly in kidney proximal tubule epithelial cells, and studies in chimeric mice indicated that the effects of TRPM2 are due to expression in parenchymal cells rather than hematopoietic cells. TRPM2-deficient mice had less oxidative stress and lower levels of NADPH oxidase activity after ischemia. While RAC1 is a component of the NADPH oxidase complex, its relation to TRPM2 and kidney ischemic injury is unknown. Following kidney ischemia, TRPM2 promoted RAC1 activation, with active RAC1 physically interacting with TRPM2 and increasing TRPM2 expression at the cell membrane. Finally, inhibition of RAC1 reduced oxidant stress and ischemic injury in vivo. These results demonstrate that TRPM2-dependent RAC1 activation increases oxidant stress and suggest that therapeutic approaches targeting TRPM2 and/or RAC1 may be effective in reducing ischemic kidney injury.
Guofeng Gao, Weiwei Wang, Raghu K. Tadagavadi, Nicole E. Briley, Michael I. Love, Barbara A. Miller, W. Brian Reeves