Acute kidney injury increases morbidity and mortality and previous studies have shown that remote ischemic preconditioning (RIPC) reduces the risk of acute kidney injury after cardiac surgery. RIPC increases urinary HMGB1 (high mobility group box protein-1) levels in patients which correlates with kidney protection. Here, we show that RIPC reduces renal ischemia-reperfusion injury and improves kidney function in mice. Mechanistically, RIPC increases HMGB1 levels in the plasma and urine and HMGB1 binds to Toll-like receptor 4 (TLR4) on renal tubular epithelial cells, inducing transcriptomic modulation of renal tubular epithelial cells and providing renal protection, whereas TLR4 activation on non-renal cells was shown to contribute to renal injury. This protection is mediated by activation of induction of AMPK⍺ and NF-kB, which induces the upregulation of Sema5b that triggers a transient, protective G1 cell-cycle arrest. In cardiac surgery patients at high risk for postoperative acute kidney injury, increased HMGB1 and Sema5b levels after RIPC were associated with renal protection after surgery. The results may help to develop future clinical treatment options for acute kidney injury.
Jan Rossaint, Melanie Meersch, Katharina Thomas, Sina Mersmann, Martin Lehmann, Jennifer Skupski, Tobias Tekath, Peter Rosenberger, John A. Kellum, Hermann Pavenstädt, Alexander Zarbock
Although macrophages are undoubtedly attractive therapeutic targets for acute kidney injury (AKI) because of their critical roles in renal inflammation and repair, the underlying mechanisms of macrophage phenotype switching and efferocytosis in the regulation of inflammatory responses during AKI are still largely unclear. The present study was to elucidate the role of JAML (junctional adhesion molecule-like protein) in the pathogenesis of AKI. We found that JAML was significantly up-regulated in the kidney from two different murine AKI models including renal ischemia/reperfusion injury (IRI) and cisplatin-induced AKI. By generation of bone marrow chimeric mice, macrophage-specific and tubular-specific Jaml conditional knockout mice, we demonstrated JAML promotes AKI mainly via a macrophage-dependent mechanism and found that JAML-mediated macrophage phenotype polarization and efferocytosis is one of critical signal transduction pathways linking inflammatory responses to AKI. Mechanistically, the effects of JAML on the regulation of macrophages was at least in part, associated with a Mincle-dependent mechanism. Collectively, our studies for the first time explore new biological functions of JAML in macrophages and conclude that JAML is an important mediator and biomarker of AKI. Pharmacologic targeting of JAML mediated signaling pathways at multiple levels may provide a novel therapeutic strategy for patients with AKI.
Wei Huang, Bi-Ou Wang, Yunfeng Hou, Yi Fu, Sijia Cui, Jinghan Zhu, Xinyu Zhan, Rongkun Li, Wei Tang, Jichao Wu, Ziying Wang, Mei Wang, Xiaojie Wang, Yan Zhang, Min Liu, Yusheng Xie, Yu Sun, Fan Yi
Nephrolithiasis is a common and recurrent disease affecting 9% of the US population. Hyperoxaluria is major risk factor for calcium oxalate kidney stones, which constitutes two-thirds of all kidney stones. Gastrointestinal epithelia play an important role in oxalate handling due to the presence of SLC26A family anion exchangers that facilitate oxalate transport. SLC26A3 (originally named DRA, down-regulated in adenoma) is an anion exchanger of chloride, bicarbonate and oxalate thought to facilitate intestinal oxalate absorption, as evidenced by ~70% reduced urine oxalate excretion in knock-out mice. We previously identified, by high-throughput screening and medicinal chemistry, a small molecule SLC26A3 inhibitor (DRAinh-A270) that selectively inhibited SLC26A3-mediated chloride/bicarbonate exchange (IC50 ~ 35 nM), and, as found here, oxalate/chloride exchange (IC50 ~ 60 nM). In colonic closed loops in mice, luminal DRAinh-A270 inhibited oxalate absorption by 70%. In a model of acute hyperoxaluria produced by oral sodium oxalate loading, DRAinh-A270 largely prevented the ~2.5-fold increase in urine oxalate/creatinine ratio. In a mouse model of oxalate nephropathy produced by a high-oxalate low-calcium diet, vehicle-treated mice by day 14 developed marked hyperoxaluria with elevated serum creatinine, renal calcium oxalate crystal deposition and renal injury, which were largely prevented by DRAinh-A270 (10 mg/kg twice daily). DRAinh-A270 administered over 7 days to healthy mice did not show significant toxicity as assessed by CBC, serum chemistries and tissue histology. Our findings support a major role of SLC26A3 in intestinal oxalate absorption and suggest the therapeutic utility of SLC26A3 inhibition for treatment of hyperoxaluria and prevention of calcium oxalate nephrolithiasis.
Onur Cil, Qi Tifany Chu, Sujin Lee, Peter M. Haggie, Alan S. Verkman
COVID-19 infection causes collapse of glomerular capillaries and loss of podocytes, terminating in a severe kidney disease called COVID-19 associated nephropathy (COVAN). The underlying mechanism of COVAN is unknown. We hypothesized that cytokines induced by COVID-19 trigger expression of pathogenic APOL1 via JAK-STAT signaling, resulting in podocyte loss and COVAN phenotype. Here, based on nine biopsy-proven COVAN cases, we demonstrated for the first time that APOL1 protein is abundantly expressed in podocytes and glomerular endothelial cells (GECs) of COVAN kidneys but not in controls. Moreover, a majority (77.8%) of COVAN patients carried two APOL1 risk alleles. We showed that recombinant cytokines induced by SARS-CoV-2 act synergistically to drive APOL1 expression through the JAK-STAT pathway in primary human podocytes, GECs, and kidney micro-organoids derived from a carrier of two APOL1 risk alleles but was blocked by JAK1/2-inhibitor, baricitinib. We demonstrated for the first time that cytokine-induced JAK-STAT-APOL1 signaling reduced the viability of kidney organoid podocytes but was rescued by baricitinib. Together, our results support the conclusion that COVID-19-induced cytokines are sufficient to drive COVAN-associated podocytopathy via JAK-STAT-APOL1 signaling and that JAK-inhibitor could block this pathogenic process. These findings suggest that JAK-inhibitors may have therapeutic benefits for managing cytokine-induced APOL1-mediated podocytopathy.
Sarah E. Nystrom, Guojie Li, Somenath Datta, Karen Soldano, Daniel Silas, Astrid Weins, Gentzon Hall, David B. Thomas, Opeyemi A. Olabisi
Inherited kidney diseases are the fifth most common cause of the end-stage renal disease. Mitochondrial dysfunction plays a vital role in the progression of inherited kidney diseases, while mt-tRNA variants and their pathogenic contributions to the kidney disease remain largely unclear. In this study, we identified the pathogenic mitochondrial tRNAPhe 616T>C mutation in three families; and documented that m.616T>C showed a high pathogenic threshold, with both heteroplasmy and homoplasmy leading to isolated chronic kidney disease and hyperuricemia without hematuria, proteinuria, or renal cyst formation. Moreover, one proband with homoplamic m.616T>C presented with end-stage renal disease as a child. No symptoms of nervous system evolvement were observed in these families. Lymphoblast cells bearing m. 616T>C exhibited swollen mitochondria, underwent active mitophagy and showed respiratory deficiency, leading to reduced mitochondrial ATP production, diminished membrane potential, and overproduction of mitochondrial reactive oxygen species. Pathogenic m. 616T>C abolished a highly conserved base-pair (A31-U39) in the anticodon stem-loop which altered the structure of mt-tRNAPhe, as confirmed by a decreased melting temperature and slower electrophoretic mobility of the mutant tRNA. Furthermore, the unstable structure of mt-tRNAPhe contributed to a shortage of steady-state mt-tRNAPhe and enhanced aminoacylation efficiency, which resulted in impaired mitochondrial RNA translation and a significant decrease in mt-DNA encoded polypeptides. Collectively, these findings provide new insights into the pathogenesis underlying inherited kidney disease caused by mitochondrial variants.
Chengxian Xu, Lingxiao Tong, Jia Rao, Qing Ye, Yuxia Chen, Yingying Zhang, Jie Xu, Xiaoting Mao, Feilong Meng, Huijun Shen, Zhihong Lu, Xiaohui Cang, Haidong Fu, Shugang Wang, Weiyue Gu, En Yin Lai, Minxin Guan, Pingping Jiang, Jianhua Mao
UMOD is a major risk gene for monogenic and complex forms of kidney disease. The encoded kidney-specific protein uromodulin is highly abundant in urine and related to chronic kidney disease, hypertension, and pathogen defense. To gain insights into potential systemic roles, we performed genome-wide screens of circulating uromodulin using complementary antibody-based (N=13,985) and aptamer-based (N=18,070) assays. We detected 3 and 10 distinct significant (p<5e-8) loci, respectively. Integration of antibody-based results at the UMOD locus with functional genomics data (RNA-seq, ATAC-seq, Hi-C) of primary human kidney tissue highlights an upstream variant with differential accessibility and transcription in uromodulin-synthesizing kidney cells as underlying the observed cis effect. Shared association patterns with complex traits, including chronic kidney disease and blood pressure, place the PRKAG2 locus in the same pathway as UMOD. Experimental validation of the third antibody-based locus, B4GALNT2, shows that the p.Cys466Arg variant of the encoded N-acetylgalactosaminyltransferase has a loss-of-function effect leading to higher serum uromodulin levels. Aptamer-based results point to enzymes writing glycan marks present on uromodulin and to their receptors in the circulation, suggesting that this assay permits investigating uromodulin’s complex glycosylation rather than its quantitative levels. Overall, our study provides new insights into circulating uromodulin and its emerging functions.
Yong Li, Yurong Cheng, Francesco Consolato, Guglielmo Schiano, Michael R. Chong, Maik Pietzner, Ngoc Quynh H. Nguyen, Nora Scherer, Mary L. Biggs, Marcus E. Kleber, Stefan Haug, Burulça Göçmen, Marie Pigeyre, Peggy Sekula, Inga Steinbrenner, Pascal Schlosser, Christina B. Joseph, Jennifer A. Brody, Morgan E. Grams, Caroline Hayward, Ulla T. Schultheiss, Bernhard K. Krämer, Florian Kronenberg, Annette Peters, Jochen Seissler, Dominik Steubl, Cornelia Then, Matthias Wuttke, Winfried März, Kai-Uwe Eckardt, Christian Gieger, Eric Boerwinkle, Bruce M. Psaty, Josef Coresh, Peter J. Oefner, Guillaume Pare, Claudia Langenberg, Jürgen E. Scherberich, Bing Yu, Shreeram Akilesh, Olivier Devuyst, Luca Rampoldi, Anna Köttgen
Atrial natriuretic peptide (ANP), encoded by Nppa, is a vasodilatory hormone that promotes salt excretion. Genome-wide association studies identified Nppa as a causative factor of blood pressure development, and in humans, ANP levels were suggested as an indicator of salt sensitivity. This study aimed to provide insights into the effects of ANP on cardiorenal function in salt-sensitive hypertension. To address this question, hypertension was induced in SSNPPA-/- (knockout of Nppa in the Dahl Salt-Sensitive (SS) rat background) or SSWT (wild type Dahl SS) rats by a high salt diet challenge (HS, 4% NaCl for 21 days). Chronic infusion of ANP in SSWT rats attenuated the increase in blood pressure and cardiorenal damage. Overall, SSNPPA-/- strain demonstrated higher blood pressure and intensified cardiac fibrosis (with no changes in ejection fraction) compared to SSWT rats. Furthermore, SSNPPA-/- rats exhibited kidney hypertrophy and higher glomerular injury scores, reduced diuresis, and lower sodium and chloride excretion than SSWT when fed a HS diet. Additionally, the activity of epithelial Na+ channel (ENaC) was found to be increased in the collecting ducts of the SSNPPA-/- rats. Taken together, these data show promise for the therapeutic benefits of ANP and ANP-increasing drugs for treating salt-sensitive hypertension.
Daria V. Ilatovskaya, Vladislav Levchenko, Kristen Winsor, Gregory R. Blass, Denisha R. Spires, Elizaveta Sarsenova, Iuliia Polina, Adrian Zietara, Mark Paterson, Alison J. Kriegel, Alexander Staruschenko
In this investigation, a potentially novel signaling pathway in gentamicin-induced acute kidney injury—worsened by overexpression of proximal tubular enzyme, myo-inositol oxygenase (MIOX)—was elucidated. WT, MIOX-transgenic (MIOX-Tg), and MIOX-KO mice were used. Gentamicin was administered to induce tubular injury. MIOX-Tg mice had severe tubular lesions associated with increased serum creatinine and proteinuria. Lesions were relatively mild, with no rise in serum creatinine and no albuminuria in MIOX-KO mice. Transfection of HK-2 cells with MIOX-pcDNA led to increased gentamicin-induced reactive oxygen species (ROS). Marked increase of ROS-mediated lipid hydroperoxidation was noted in MIOX-Tg mice, as assessed by 4-HNE staining. This was associated with increased expression of arachidonate 12-lipoxygenase (ALOX-12) and generation of 12-hydroxyeicosatetraenoic acid (12-HETE). In addition, notable monocyte/macrophage influx, upregulation of NF-κB and inflammatory cytokines, and apoptosis was observed in MIOX-Tg mice. Treatment of cells with ALOX-12 siRNA abolished gentamicin-mediated induction of cytokines and 12-HETE generation. HETE-12 treatment promoted this effect, along with upregulation of various signaling kinases and activation of GPCR31. Similarly, treatment of cells or mice with the ALOX-12 inhibitor ML355 attenuated inflammatory response, kinase signaling cascade, and albuminuria. Collectively, these studies highlight a potentially novel mechanism (i.e., the ROS/ALOX-12/12-HETE/GPR31 signaling axis) relevant to gentamicin-induced nephrotoxicity modulated by MIOX.
Isha Sharma, Yingjun Liao, Xiaoping Zheng, Yashpal S. Kanwar
The importance of healthy mitochondrial function is implicated in the prevention of chronic/diabetic kidney diseases (CKD/DKD). Sex differences also play an important role in DKD. Our previous studies revealed that mitochondrial substrate overload (modeled by homozygous deletion of carnitine acetyl-transferase – CrAT) in proximal tubules causes renal injury. Here we demonstrate the importance of intact mitochondrial substrate efflux by titrating the amount of overload through the generation of a heterozygous CrAT knockout model (“PT-CrATHET” mouse). Intriguingly, these animals developed renal injury similarly to their homozygous counterparts. Mitochondria were structurally and functionally impaired in both sexes. Transcriptomic analyses, however, revealed striking sex differences. Male mice shut down fatty acid oxidation and several other metabolism-related pathways. Females had a significantly weaker transcriptional response in metabolism but activation of inflammatory pathways was prominent. Proximal tubular cells from PT-CrATHET mice of both sexes exhibited a shift towards a more glycolytic phenotype, but females were still able to oxidize fatty acid-based substrates. Our results demonstrate that maintaining mitochondrial substrate metabolism balance is crucial to satisfy proximal tubular energy demand. Our findings have potentially broad implications as both the glycolytic shift and the sexual dimorphisms discovered herein offer new modalities for future interventions for treating kidney disease.
Allison McCrimmon, Kerin M. Cahill, Claudia Kruger, Margaret E. Mangelli, Emily Bouffard, Timothy Dobroski, Kelly N. Michanczyk, Susan J. Burke, Robert C. Noland, Daria V. Ilatovskaya, Krisztian Stadler
Kidney fibrosis is the final common pathway of progressive kidney diseases, the underlying mechanisms of which is not fully understood. The purpose of the current study is to investigate a role of Piezo1, a mechanosensitive nonselective cation channel, in kidney fibrosis. In human fibrotic kidneys, Piezo1 protein expression was markedly upregulated. The abundance of Piezo1 protein in kidneys of mice with UUO or with folic-acid treatment was significantly increased. Inhibition of Piezo1 with GsMTx4 markedly ameliorated UUO or folic acid-induced kidney fibrosis. Mechanical stretch, compression or stiffness induced Piezo1 activation and pro-fibrotic responses in human HK2 cells and primary cultured mouse proximal tubular cells (mPTCs), which were greatly prevented by inhibition or silence of Piezo1. TGFβ-1 induced increased Piezo1 expression and pro-fibrotic phenotypic alterations in HK2 cells and mPTCs, which was again markedly prevented by inhibition of Piezo1. Activation of Piezo1 by Yoda1, a Piezo1 agonist, caused calcium influx and profibrotic responses in HK2 cells and induced calpain2 activation, followed by talin1 cleavage and upregulation of integrinβ1. Also, Yoda1 promoted the link between ECM and integrinβ1. In conclusion, Piezo1 is involved in the progression of kidney fibrosis and pro-fibrotic alterations in renal proximal tubular cells, likely through activating calcium-calpain2-integrinβ1 pathway.
Xiaoduo Zhao, Yonglun Kong, Baien Liang, Jinhai Xu, Yu Lin, Nan Zhou, Jing Li, Bin Jiang, Jianding Cheng, Chunling Li, Weidong Wang
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