Nephrologies
Abstract
Despite recent progress in the identification of mediators of podocyte injury, mechanisms underlying podocyte loss remain poorly understood, and cell-specific therapy is lacking. We previously reported that KIBRA, KIdney and BRAin expressed protein, encoded by WWC1, promotes podocyte injury in vitro through activation of the Hippo signaling pathway. KIBRA expression is increased in the glomeruli of patients with focal segmental glomerulosclerosis (FSGS), and KIBRA depletion in vivo is protective against acute podocyte injury. Here, we tested the consequences of transgenic podocyte-specific WWC1 expression in immortalized human podocytes and in mice, and we explored the association between glomerular WWC1 expression and glomerular disease progression. We found that KIBRA overexpression in immortalized human podocytes promoted cytoplasmic localization of YAP (Yes-associated protein), induced actin cytoskeletal reorganization, and altered focal adhesion expression and morphology. Transgenic WWC1 (KIBRA OE) mice were more susceptible to acute and chronic glomerular injury, with evidence of YAP inhibition in vivo. Of clinical relevance, glomerular WWC1 expression negatively correlated with renal survival among patients with primary glomerular diseases. These findings highlight the importance of KIBRA-YAP signaling to the regulation of podocyte structural integrity and identify KIBRA-mediated injury as a potential target for podocyte-specific therapy in glomerular disease.
Authors
Kristin Meliambro, Yanfeng Yang, Marina de Cos, Estefania Rodriguez-Ballestas, Caroline Malkin, Jonathan C. Haydak, John R. Lee, Fadi Salem, Laura H. Mariani, Ronald E. Gordon, John M. Basgen, Huei Hsun Wen, Jia Fu, Evren U. Azeloglu, John Cijiang He, Jenny S. Wong, Kirk N. Campbell
Abstract
We examine whether calcineurin or protein-phosphatase-2B (PP2B) regulates the basolateral Kir4.1/Kir5.1 in distal-convoluted-tubule (DCT). Application of tacrolimus (FK506) or cyclosporine-A (CsA) increased whole-cell-Kir4.1/Kir5.1-mediated-K+-currents and hyperpolarized DCT-membrane. Moreover, FK506-induced-stimulation of Kir4.1/Kir5.1 was absent in kidney-tubule-specific 12-kDa-FK506-binding-protein knockout-mice (Ks-FKBP-12-KO). In contrast, CsA still stimulated Kir4.1/Kir5.1 of the DCT in Ks-FKBP-12-KO mice, suggesting that FK506-induced stimulation of Kir4.1/Kir5.1 was due to inhibiting PP2B. Single-channel-patch-clamp experiments demonstrated that FK506 or CsA stimulated the basolateral Kir4.1/Kir5.1-activity of DCT, defined by NPo (a product of channel-Number- and- Open-Probability). However, this effect was absent in the DCT treated with Src-family-protein-tyrosine-kinase (SFK) inhibitor or hydroxyl-peroxide which stimulates SFK. Fluorescence-image demonstrated that CsA-treatment increased membrane-staining-intensity of Kir4.1 in the DCT of Kcnj10flox/flox mice. Moreover, CsA-treatment has no obvious effect on pNCC expression in Ks-Kir4.1-KO mice. Immunoblotting showed that acute FK506 treatment increased pNCC-expression in Kcnj10flox/flox mice, but this effect was attenuated in Ks-Kir4.1-KO mice. In vivo measurement of thiazide-induced-renal-Na+ excretion demonstrated that FK506 enhanced thiazide-induced natriuresis. This effect was absent in Ks-FKBP-12-KO mice and blunted in Ks-Kir4.1-KO mice. We conclude that inhibition of PP2B stimulates Kir4.1/Kir5.1 of DCT and NCC, and that PP2B-inhibition-induced stimulation of NCC is partially achieved by stimulation of the basolateral Kir4.1/Kir5.1.
Authors
Dan-Dan Zhang, Xin-Peng Duan, Kerim Mutig, Franziska Rausch, Yu Xiao, Jun-Ya Zheng, Dao-Hong Lin, Wen-Hui Wang
Abstract
The transcription factor c-Maf has been widely studied and has been reported to play a critical role in embryonic kidney development; however, the postnatal functions of c-Maf in adult kidneys remain unknown as c-Maf null C57BL/6J mice exhibit embryonic lethality. In this study, we investigated the role of c-Maf in adult mouse kidneys by comparing the phenotypes of tamoxifen (TAM)-inducible c-Maf knockout mice (c-Maf flox/flox; CAG-Cre-ERTM mice named “c-Maf ΔTAM”) with c-Maf flox/flox control mice10 days after TAM injection (TAM(10d)). In addition, we examined the effects of c-Maf deletion on diabetic conditions by injecting the mice with streptozotocin (STZ) four weeks before TAM injection. c-Maf ΔTAM mice displayed primary glycosuria caused by Sglt2 and Glut2 downregulation in the kidneys without diabetes, as well as morphological changes and life-threatening injuries in the kidneys on TAM(10d). Under diabetic conditions, c-Maf deletion promoted recovery from hyperglycemia and suppressed albuminuria and diabetic nephropathy by causing similar effects to Sglt2 knockout and SGLT2 inhibitors. In addition to demonstrating the unique gene regulation of c-Maf, these findings highlight the renoprotective effects of c-Maf deficiency under diabetic conditions and suggest that c-Maf could be a novel therapeutic target gene for treating diabetic nephropathy.
Authors
Mitsunori Fujino, Naoki Morito, Takuto Hayashi, Masami Ojima, Shun Ishibashi, Akihiro Kuno, Seizo Koshiba, Kunihiro Yamagata, Satoru Takahashi
Abstract
Acute kidney failure and chronic kidney disease are global health issues steadily rising in incidence and prevalence. Animal models on a single genetic background have so far failed to recapitulate the clinical presentation of human nephropathies. Here, we used a simple model of folic acid–induced kidney injury in 7 highly diverse mouse strains. We measured plasma and urine parameters, as well as renal histopathology and mRNA expression data, at 1, 2, and 6 weeks after injury, covering the early recovery and long-term remission. We observed an extensive strain-specific response ranging from complete resistance of the CAST/EiJ to high sensitivity of the C57BL/6J, DBA/2J, and PWK/PhJ strains. In susceptible strains, the severe early kidney injury was accompanied by the induction of mitochondrial stress response (MSR) genes and the attenuation of NAD+ synthesis pathways. This is associated with delayed healing and a prolonged inflammatory and adaptive immune response 6 weeks after insult, heralding a transition to chronic kidney disease. Through a thorough comparison of the transcriptomic response in mouse and human disease, we show that critical metabolic gene alterations were shared across species, and we highlight the PWK/PhJ strain as an emergent model of transition from acute kidney injury to chronic disease.
Authors
Jean-David Morel, Maroun Bou Sleiman, Terytty Yang Li, Giacomo von Alvensleben, Alexis M. Bachmann, Dina Hofer, Ellen Broeckx, Jing Ying Ma, Vinicius Carreira, Tao Chen, Nabil Azhar, Romer A. Gonzalez-Villalobos, Matthew Breyer, Dermot Reilly, Shannon Mullican, Johan Auwerx
Abstract
The glomerular endothelial glycocalyx (GEnGlx) forms the first part of the glomerular filtration barrier. Previously we showed that mineralocorticoid receptor (MR) activation caused GEnGlx damage and albuminuria. Here we investigated whether MR antagonism could limit albuminuria in diabetes and studied the site of action. Streptozotocin-induced diabetic Wistar rats developed albuminuria, increased glomerular albumin permeability (Ps’alb) and increased glomerular matrix metalloproteinase (MMP) activity with corresponding GEnGlx loss. MR antagonism prevented albuminuria progression, restored Ps’alb, preserved GEnGlx and reduced MMP activity. Enzymatic degradation of the GEnGlx negated the benefits of MR antagonism, confirming their dependence on GEnGlx integrity. Exposing human glomerular endothelial cells (GEnC) to diabetic conditions in vitro increased MMPs and caused glycocalyx damage. Amelioration of these effects confirmed a direct effect of MR antagonism on GEnC. To confirm relevance to human disease, we used a novel confocal imaging method to show loss of GEnGlx in renal biopsy specimens from patients with diabetic nephropathy (DN). In addition, DN patients randomised to receive an MR antagonist had reduced urinary MMP2 activity and albuminuria compared with placebo and baseline levels. Taken together our work suggests MR antagonists reduce MMP activity and thereby preserve GEnGlx resulting in reduced glomerular permeability and albuminuria in diabetes.
Authors
Michael Crompton, Joanne K. Ferguson, RainaD. Ramnath, Karen L. Onions, Anna S. Ogier, Monica Gamez, Colin J. Down, Laura J. Skinner, Kitty H.F. Wong, Lauren Kari Dixon, Judit Sutak, Steven J. Harper, Paola Pontrelli, Loreto Gesualdo, Hiddo L. Heerspink, Robert D. Toto, Gavin I. Welsh, Rebecca R. Foster, Simon C. Satchell, Matthew J. Butler
Abstract
Acute kidney injury (AKI) is one of the most important complications in COVID-19 patients and is considered a negative prognostic factor with respect to patient survival. The occurrence of direct infection of the kidney by SARS-CoV-2, and its contribution to the renal deterioration process, remains a controversial issue. By studying 32 renal biopsies from COVID-19 patients we confirmed that the major pathological feature of COVID-19 is acute tubular injury (ATI). Using smFISH, we showed that the SARS-CoV-2 infects living renal cells and that infection, which parallels renal ACE2 expression levels, is associated to increase death. Mechanistically, a transcriptomic analysis uncovered specific molecular signatures in SARS-CoV-2 infected kidneys as compared to healthy kidneys and non-COVID-19 ATI kidneys. On the other hand, we demonstrated that SARS-CoV-2 and Hantavirus, two RNA viruses, activated different genetic networks despite they triggered the same pathological lesions. Finally, we identified XAF1 as a critical target of SARS-CoV-2 infection. In conclusion, this study demonstrates that SARS-CoV2 can directly infect living renal cells and identified specific druggable molecular targets that can potentially aid in the design of novel therapeutic strategies to preserve renal function in severely affected COVID-19 patients.
Authors
Pierre Isnard, Paul Vergnaud, Serge Garbay, Matthieu Jamme, Maeva Eloudzeri, Alexandre Karras, Dany Anglicheau, Valerie Galantine, Arwa Jalal Eddine, Clément Gosset, Franck Pourcine, Mohammed Zarhrate, Jean-Baptiste Gibier, Elena Rensen, Stefano Pietropaoli, Giovanna Barba-Spaeth, Jean-Paul Duong-Van-Huyen, Thierry J. Molina, Florian Mueller, Christophe Zimmer, Marco Pontoglio, Fabiola Terzi, Marion Rabant
Abstract
Dietary potassium (K+) supplementation is associated with a blood pressure (BP) lowering effect, but not all studies agree. Here we examined the effects of short and long-term K+ supplementation on BP in mice, whether differences depend on the accompanying anion or the sodium (Na+) intake and molecular alterations in the kidney that may underlie BP changes. Relative to the control diet, BP was higher in mice fed a high NaCl (1.57% Na+) for 7 weeks or 2 weeks with a K+-free diet. BP was highest on a K+-free/high NaCl diet. Commensurate with increased abundance and phosphorylation of the thiazide sensitive sodium-chloride-cotransporter (NCC) on the K+-free/high NaCl diet, BP returned to normal with thiazides. Three weeks of a high K+ diet (5% K+) increased BP (predominantly during night-time) independently of dietary Na+ or anion intake. Conversely, 4 days of KCl feeding reduced BP. Both feeding periods resulted in lower NCC levels, but increased levels of cleaved (active) α and γ subunits of the epithelial Na+ channel ENaC. The elevated BP after chronic K+ feeding was reduced by amiloride but not thiazide. Our results suggest that dietary K+ has an optimal threshold where it may be most effective for cardiovascular health.
Authors
Robert Little, Sathish K. Murali, Søren B. Poulsen, Paul R. Grimm, Adrienne Assmus, Lei Cheng, Jessica R. Ivy, Ewout J. Hoorn, Vladimir V. Matchkov, Paul A. Welling, Robert A. Fenton
Abstract
Organic anion transporter 1 (OAT1/SLC22A6, NKT) is a multispecific drug transporter in the kidney with numerous substrates, including pharmaceuticals, endogenous metabolites, natural products, and uremic toxins. Here, we show that OAT1 regulates levels of gut microbiome–derived metabolites. We depleted the gut microbiome of Oat1-KO and WT mice and performed metabolomics to analyze the effects of genotype (KO versus WT) and microbiome depletion. OAT1 is an in vivo intermediary between the host and the microbes, with 40 of the 162 metabolites dependent on the gut microbiome also impacted by loss of Oat1. Chemoinformatic analysis revealed that the altered metabolites (e.g., indoxyl sulfate, p-cresol sulfate, deoxycholate) had more ring structures and sulfate groups. This indicates a pathway from gut microbes to liver phase II metabolism, to renal OAT1–mediated transport. The idea that multiple gut-derived metabolites directly interact with OAT1 was confirmed by in vitro transport and magnetic bead binding assays. We show that gut microbiome–derived metabolites dependent on OAT1 are impacted in a chronic kidney disease (CKD) model and human drug-metabolite interactions. Consistent with the Remote Sensing and Signaling Theory, our results support the view that drug transporters (e.g., OAT1, OAT3, OATP1B1, OATP1B3, MRP2, MRP4, ABCG2) play a central role in regulating gut microbe–dependent metabolism, as well as interorganismal communication between the host and microbiome.
Authors
Jeffry C. Granados, Vladimir Ermakov, Koustav Maity, David R. Vera, Geoffrey Chang, Sanjay K. Nigam
Abstract
Obesity is a major risk factor for end-stage kidney disease. We previously found that lysosomal dysfunction and impaired autophagic flux contributed to lipotoxicity in obesity-related kidney disease, both in humans and experimental animal models. However, the regulatory factors involved in countering renal lipotoxicity are largely unknown. Here we found that palmitic acid (PA) strongly promoted dephosphorylation and nuclear translocation of transcription factor EB (TFEB) by inhibiting the mechanistic target of rapamycin kinase complex 1 (MTORC1) pathway in a Rag GTPase–dependent manner, although these effects gradually diminished after extended treatment. We then investigated the role of TFEB in the pathogenesis of obesity-related kidney disease. Proximal tubular epithelial cell (PTEC)-specific Tfeb-deficient mice fed a high-fat diet (HFD) exhibited greater phospholipid accumulation in enlarged lysosomes, which manifested as multilamellar bodies (MLBs). Activated TFEB mediated lysosomal exocytosis of phospholipids, which help reduce MLB accumulation in PTECs. Furthermore, HFD-fed PTEC-specific Tfeb-deficient mice showed autophagic stagnation and exacerbated injury upon renal ischemia–reperfusion. Finally, higher body mass index was associated with increased vacuolation and decreased nuclear TFEB in the proximal tubules of chronic kidney disease patients. These results indicate a critical role of TFEB-mediated lysosomal exocytosis in counteracting renal lipotoxicity.
Authors
Jun Nakamura, Takeshi Yamamoto, Yoshitsugu Takabatake, Tomoko Namba-Hamano, Satoshi Minami, Atsushi Takahashi, Jun Matsuda, Shinsuke Sakai, Hiroaki Yonishi, Shihomi Maeda, Sho Matsui, Isao Matsui, Takayuki Hamano, Masatomo Takahashi, Maiko Goto, Yoshihiro Izumi, Takeshi Bamba, Miwa Sasai, Masahiro Yamamoto, Taiji Matsusaka, Fumio Niimura, Motoko Yanagita, Shuhei Nakamura, Tamotsu Yoshimori, Andrea Ballabio, Yoshitaka Isaka
Abstract
Preterm birth results in low nephron endowment and increased risk of acute kidney injury (AKI) and chronic kidney disease (CKD). To understand the pathogenesis of AKI and CKD in preterm humans, we generated novel mouse models with a 30-70% reduction in nephron number by inhibiting or deleting Ret tyrosine kinase in the developing ureteric bud. These mice developed glomerular and tubular hypertrophy followed by the transition to CKD, recapitulating the renal pathological changes seen in humans born preterm. We injected neonatal mice with gentamicin, a ubiquitous nephrotoxic exposure in preterm infants, and detected more severe proximal tubular injury in mice with low nephron number compared to controls with normal nephron number. Mice with low nephron number have reduced proliferative repair with more rapid development of CKD. Furthermore, mice had more profound inflammation with highly elevated levels of MCP-1 and CXCL10, produced in part by damaged proximal tubules. Our study directly links low nephron endowment with postnatal renal hypertrophy, which in this model is maladaptive and results in CKD. Underdeveloped kidneys are more susceptible to gentamicin-induced AKI, suggesting that AKI in the setting of low nephron number is more severe and further increases the risk of CKD in this vulnerable population.
Authors
Pamela I. Good, Ling Li, Holly A. Hurst, Ileana M. Serrano-Herrera, Katherine Xu, Meenakshi Rao, David A. Bateman, Qais Al-Awqati, Vivette D. D'Agati, Franklin Costantini, Fangming Lin
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