Nephrology
Abstract
We recently showed that cell surface translocation of the endoplasmic reticulum–resident protein GRP78, when bound by activated α 2-macroglobulin (α2M*), induces pro-fibrotic responses in glomerular mesangial cells in response to high glucose and regulates activation of the pro-fibrotic cytokine transforming growth factor-β1 (TGF-β1), implicating a pathogenic role in glomerulosclerosis. Interstitial fibrosis, largely mediated by proximal tubular epithelial cells (PTEC) and renal fibroblasts, develops later in kidney disease and correlates with functional decline. Here we investigated whether interstitial fibrosis was mediated by cell surface GRP78 (csGRP78)/α2M*. High glucose and TGF-β1 increased csGRP78 and α2M* in PTEC and renal fibroblasts, and their inhibition prevented fibrotic protein production. Interestingly, for TGF-β1, this depended on inhibition of noncanonical signaling through YAP/TAZ, with Smad3 activation unaffected. In vivo, type 1 diabetic Akita mice overexpressing TGF-β1 were treated with either a neutralizing antibody for csGRP78 (C38) or α2M* (Fα2M) or an inhibitory peptide blocking csGRP78/α2M* interaction, and mice with unilateral ureteral obstruction were treated with Fα2M or inhibitory peptide. Consistently, inhibition by antibody or peptide attenuated fibrosis and pro-fibrotic signaling. These findings show an important role for csGRP78/α2M* in mediating tubulointerstitial fibrosis in both diabetic and nondiabetic kidney disease and support their inhibition as a potential antifibrotic therapeutic intervention.
Authors
Jackie Trink, Ifeanyi Kennedy Nmecha, Katrine Pilely, Renzhong Li, Zi Yang, Sydney Kwiecien, Melissa MacDonald, Bo Gao, Mariam A. Mamai, Chao Lu, Urooj F. Bajwa, Nikhil Uppal, James C. Fredenburgh, Masao Kakoki, Salvatore V. Pizzo, Anthony F. Rullo, Matthew B. Lanktree, Jeffrey I. Weitz, Yaseelan Palarasah, Joan C. Krepinsky
Abstract
Kidney organoids are an emerging tool for disease modeling, especially genetic diseases. Among these diseases, X-linked Alport syndrome (XLAS) is a hematuric nephropathy affecting the glomerular basement membrane (GBM) secondary to pathogenic variations in the COL4A5 gene encoding the α5 subunit of type IV collagen [α5(IV)]. In patients carrying pathogenic variations affecting splicing, the use of antisense oligonucleotides (ASOs) offers immense therapeutic hope. In this study, we develop a framework combining the use of patient-derived cells and kidney organoids to provide evidence of the therapeutic efficacy of ASOs in XLAS patients. Using multiomics analysis, we describe the development of GBM in wild-type and mutated human kidney organoids. We show that GBM maturation is a dynamic process, which requires long organoid culture. Then, using semi-automated quantification of α5(IV) at basement membranes in organoids carrying the splicing variants identified in patients, we demonstrate the efficacy of ASO treatment for α5(IV) restoration. These data contribute to our understanding of the development of GBM in kidney organoids and pave the way for a therapeutic screening platform for patients.
Authors
Hassan Saei, Bruno Estebe, Nicolas Goudin, Mahsa Esmailpour, Julie Haure, Olivier Gribouval, Christelle Arrondel, Vincent Moriniere, Pinyuan Tian, Rachel Lennon, Corinne Antignac, Geraldine Mollet, Guillaume Dorval
Abstract
Vaso-occlusive episodes (VOEs) in the setting of hyperhemolysis can rapidly evolve into multiorgan failure in sickle cell disease (SCD). Although the mechanisms for rapid progression to multiorgan failure are unclear, a systemic vasculopathy with thrombotic microangiopathy-type features has been described. Reduced thrombomodulin (TM) function is implicated in some thrombotic microangiopathy syndromes. We observed a greater decline in platelet count and hemoglobin concentration and increase in vascular injury biomarkers within 24-hours of admission for a VOE in 12 SCD patients with versus 12 without multiorgan failure. We observed decreased TM expression on the lung and kidney vasculature of three additional SCD patients with multiorgan failure and an autopsy performed compared to a non-SCD control. Transgenic SCD mice challenged with cell-free hemoglobin had reduced TM function, increased vascular injury biomarkers, and reduced renal cortical blood flow. Infusion of recombinant TM 2- or 24-hours after the challenge restored cortical blood flow, mitigated increases in vascular injury, complement activation, and tubular injury biomarkers, and protected against acute kidney and lung injury. We demonstrated that impaired TM function may be involved in the systemic vasculopathy of SCD-related multiorgan failure and infusion of recombinant TM may restore vascular function and protect against acute organ damage.
Authors
Guohui Ren, Dustin R. Fraidenburg, Suman Setty, Jiwang Chen, Janae Gonzales, Maria Armila Ruiz, Zalaya Ivy, Najmeh Eskandari, Richard D. Minshall, James P. Lash, Victor R. Gordeuk, Santosh L. Saraf
Abstract
Urinary tract infections (UTIs) are the most common severe bacterial infections in young children, often associated with vesicoureteral reflux (VUR). To explore host genetic-microbiota interactions and their clinical implications, we analyzed the urinary microbiota (urobiota) and conducted genome-wide association studies (GWAS) for bacterial abundance traits in pediatric UTI and VUR patients from the RIVUR and CUTIE cohorts. We identified four urobiota community types based on relative abundance, characterized by the genera Enterococcus, Prevotella, Pseudomonas, and Escherichia/Shigella, and their associations with VUR, age, and toilet training. Children with VUR exhibited decreased microbial diversity and increased abundance of genera that included opportunistic pathogens, suggesting a disrupted urobiota. We detected genome-wide significant genetic associations with urinary bacterial relative abundances, in or near candidate genes including CXCL12, ABCC1, and ROBO1, which are implicated in urinary tract development and response to infection. We showed that Cxcl12 is induced 12 hours after uropathogenic bacterial infection in mouse bladder. The association with CXCL12 suggests a genetic link between UTI, VUR and cardiovascular phenotypes later in life. These findings provide the first characterization of host genetic influences on the pediatric urobiota in UTI and VUR, offering insights into the interplay between disease, host genetics and the urobiota composition.
Authors
Miguel Verbitsky, Pavan Khosla, Daniel Bivona, Atlas Khan, Yask Gupta, Heekuk Park, Tian H. Shen, Aryan Ghotra, Katherine Xu, Iman A. Ghavami, Priya Krithivasan, Jeremiah Martino, Tanya Sezin, Tze Y. Lim, Victoria Kolupaeva, Nita A. Limdi, Yuan Luo, Hakon Hakonarson, Simone Sanna-Cherchi, Krzysztof Kiryluk, Cathy L. Mendelsohn, Anne-Catrin Uhlemann, Jonathan Barasch, Ali G. Gharavi
Abstract
Mitochondrial dysfunction is a major mechanism of acute kidney injury (AKI), and increased circulating interleukin 6 (IL-6) is associated with systemic inflammation and death due to sepsis. We tested whether kidney mitochondrial DNA (mtDNA) contributes to IL-6 release in sepsis-associated AKI via Toll-like receptor 9 (TLR9). In a murine model of sepsis via cecal ligation and puncture (CLP), we used next-generation sequencing of plasma mtDNA to inform the design of optimal target sequences for quantification by droplet digital PCR, and to identify single-nucleotide polymorphisms (SNPs) to infer tissue origin. We found significantly higher concentrations of plasma mtDNA after CLP versus shams and that plasma mtDNA SNPs matched kidney SNPs more than other organs. Kidney mtDNA contributed directly to IL-6 and mtDNA release from dendritic cells in vitro and kidney mitochondria solution led to higher IL-6 concentrations in vivo. IL-6 release was mitigated by a TLR9 inhibitor. Finally, plasma mtDNA was significantly higher in septic patients with AKI compared with those without AKI and correlated significantly with plasma IL-6. We conclude that AKI contributes to increased circulating IL-6 in sepsis via mtDNA release. Targeting kidney mitochondria and mtDNA release are potential translational avenues to decrease mortality from sepsis-associated AKI.
Authors
Avnee J. Kumar, Katharine Epler, Jing Wang, Alice Shen, Negin Samandari, Mark L. Rolfsen, Laura A. Barnes, Gerald S. Shadel, Alexandra G. Moyzis, Alva G. Sainz, Karlen Ulubabyan, Kefeng Li, Kristen Jepsen, Xinrui Li, Mark M. Fuster, Roger G. Spragg, Roman Sasik, Volker Vallon, Helen Goodluck, Joachim H. Ix, Prabhleen Singh, Mark L. Hepokoski
Abstract
Renal polycystins (PKD1, PKD2) are ion channel–forming subunits that traffic to principal cell primary cilia. Variants in these proteins cause approximately 95% of autosomal dominant polycystic kidney disease (ADPKD), a common, lethal genetic disorder that lacks effective drug treatments. We assessed the mechanistic impact and pathogenic propensity of 2 disease-associated PKD2 truncating variants, R803X and R654X. Worldwide, hundreds of individuals with ADPKD harbor these germline mutations, including the R803X founder variant first identified within the patient population of Taiwan. Our biochemical, electrophysiological, and super-resolution imaging analyses demonstrated that the pore-truncating R654X variant abolished channel assembly and ciliary trafficking, whereas the R803X variant retained partial cilia trafficking and channel function. To assess disease impact, we generated transgenic mice with analogous truncation mutations. Homozygous mutants were embryonic lethal, whereas heterozygous mice expressing both variant and conditional Pkd2 repression alleles developed pronounced renal cysts. Cyst progression was slower in mice carrying the equivalent Taiwan mutation, reflecting the milder clinical course observed in patients. These findings revealed that the degree of impaired PKD2 channel trafficking to primary cilia correlated with cystic disease severity, providing insight into variant-specific ADPKD pathogenesis and newly developed animal models expressing clinically relevant variants for therapeutic testing.
Authors
Louise F. Kimura, Orhi Esarte Palomero, Megan Larmore, Paul G. DeCaen, Thuy N. Vien
Abstract
Prolonged and dysregulated neutrophilic inflammation causes tissue damage in chronic inflammatory diseases, including antibody-mediated glomerulonephritis (AGN). An increase in glycolysis, supported by enhanced glucose uptake, is a hallmark of hyperneutrophilic inflammation. Neutrophils upregulate glucose transporter 1–mediated (Glut1-mediated) glucose incorporation for renal antimicrobial activities. However, little is known about the role of neutrophil-specific Glut1 function in the pathogenesis of AGN. Using a well-vetted mouse model of AGN, we show that neutrophils upregulate Glut1 expression and function in the nephritic kidney. We demonstrate that Glut1 function in the hematopoietic cells during the early stage of the disease is necessary for kidney pathology. Most importantly, neutrophil-intrinsic Glut1 function is critical for AGN. While neutrophil-specific Glut1 ablation diminished the expression of tissue-damaging effector molecules in both the early and late stages, renal cytokines’ and chemokines’ production were compromised only in the late stage of the disease. Consequently, Glut1 inhibitor treatment ameliorated renal pathology in AGN mice. These data identify a Glut1-driven inflammatory circuit in neutrophils, which is amenable to therapeutic targeting in AGN.
Authors
Hossein Rahimi, Wonseok Choi, Doureradjou Peroumal, Shuxia Wang, Partha S. Biswas
Abstract
The mechanisms underlying cyst growth and progression in Autosomal Dominant Polycystic Kidney Disease (ADPKD) remain unresolved. Since cyst expansion requires epithelial salt and water secretion likely involving basolateral membrane K+ recycling, we investigated the role of KCNN4-encoded K+ channel KCa3.1, inhibited by the potent, pharmacospecific, well-tolerated antagonist, senicapoc. We hypothesized that genetic and/or pharmacological inactivation of KCNN4/KCa3.1 would slow PKD progression. KCNN4 was upregulated in kidneys of patients with ADPKD and of mechanistically distinct PKD mouse models. Cyst expansion in Pkd1–/– murine metanephroi was stimulated by KCa3.1 agonist and was prevented/reversed by senicapoc. In rapidly and/or slowly progressive mouse Pkd1 models, Kcnn4 inactivation slowed renal cyst growth; attenuated PKD-stimulated cAMP and ERK/Myc signaling pathways; reduced PKD-associated ciliary elongation, cell proliferation, and fibrosis; improved renal function; and prolonged survival. Importantly, senicapoc treatment of Pkd1 mouse models phenocopied most effects of Kcnn4 inactivation. This first study on the efficacy of KCa3.1 inhibition in PKD progression recommends senicapoc as a clinical trial candidate for ADPKD.
Authors
Guanhan Yao, Almira Kurbegovic, Camila Parrot, William Foley, William Roman, Seth L. Alper, Marie Trudel
Abstract
Glomerular inflammation and podocyte loss are the hallmarks of chronic kidney disease (CKD) progression. Understanding how podocytes and their microenvironment regulate inflammation is critical for developing effective therapies. In this study, we identified C-C chemokine ligand 5 (CCL5) as an inflammatory mediator elevated in injured podocytes, based on analyses of both human kidney biopsies and mouse models of CKD. We discovered that CCL5 exerts paradoxical effects in nephropathy: while it protects podocytes in vitro, it exacerbates glomerular injury in vivo. Recombinant CCL5 and podocyte-specific CCL5 overexpression promoted cell survival and reduced apoptosis in cultured podocytes. However, in Adriamycin-induced nephropathy, CCL5 worsened glomerular injury, increasing proteinuria, glomerulosclerosis, and podocyte loss. Bone marrow (BM) transplantation experiments revealed that CCL5 in BM-derived cells—not kidney-resident cells—drove disease progression. CCL5 deficiency in BM-derived cells conferred protection by increasing reparative M2 macrophages, whereas endogenous CCL5 promoted M1 polarization, inhibited M2 differentiation, and triggered M2-to-M1 transition. These findings demonstrate that while CCL5 supports podocyte survival, its expression in BM-derived cells promotes inflammatory macrophage phenotypes and glomerular injury. The harmful immune effects of CCL5 in BM-derived cells outweigh its podocyte-protective role, highlighting the importance of cell-targeted strategies to mitigate kidney damage.
Authors
Ika N. Kadariswantiningsih, Issei Okunaga, Kaho Yamasaki, Maulana A. Empitu, Hiroyuki Yamada, Shin-ichi Makino, Akitsu Hotta, Hideo Yagita, Masashi Aizawa, Ryo Koyama-Nasu, Motoko Y. Kimura, Narihito Tatsumoto, Katsuhiko Asanuma
Abstract
Normothermic machine perfusion (NMP) has become a valuable tool to expand the pool of transplantable organs. However, the application of NMP to kidneys presents substantial challenges, mostly due to high variability in the composition of currently used perfusion solutions. Here, we provide a multimodal cross-species cellular atlas of kidney injury associated with NMP using a literature-based consensus buffer. This resource provided a systematic framework that was used to develop a metabolite-enhanced perfusion solution, which protected renal proximal tubular cells, improving cellular viability and transplantation outcomes across species, including human kidneys.
Authors
Jan Czogalla, Fabian Hausmann, Simon Lagies, Sydney E. Gies, Sabrina Christiansen, Nico Kaiser, Fabian Haas, Yusuke Okabayashi, Dominik Kylies, Smilla Hofmann, Rossana Franzin, Niklas Sabra, Sarah Bouari, Yitian Fang, Gisela Ambagtsheer, Ilka Edenhofer, Silvia Chilla, Anne K. Mühlig, Marina Zimmermann, Milagros N. Wong, Takashi Yokoo, Oliver Kretz, Maja Lindenmeyer, Florian Grahammer, Martin J. Hoogduijn, Ron de Bruin, Malte Kuehl, Sonja Hänzelmann, Bernd Kammerer, Loreto Gesualdo, Stefan Bonn, Robert C. Minnee, Tobias B. Huber, Victor G. Puelles
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