Nephrology
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
Abstract
BACKGROUND. Active vitamin D metabolites, including 25-hydroxyvitamin D (25D) and 1,25-dihydroxyvitamin D (1,25D), have potent immunomodulatory effects that attenuate acute kidney injury (AKI) in animal models. METHODS. We conducted a phase 2, randomized, double-blind, multiple-dose, 3-arm clinical trial comparing oral calcifediol (25D), calcitriol (1,25D), and placebo among 150 critically ill adult patients at high-risk of moderate-to-severe AKI. The primary endpoint was a hierarchical composite of death, kidney replacement therapy (KRT), and kidney injury (baseline-adjusted mean change in serum creatinine), each assessed within 7 days following enrollment using a rank-based procedure. Secondary endpoints included new or progressive AKI and a composite of KRT or death. Hypercalcemia was the key safety endpoint. We also performed RNA sequencing on circulating CD14+ monocytes collected immediately prior to randomization and two days later. RESULTS. The global rank score for the primary endpoint was similar among calcifediol (n = 51) vs. placebo (n = 49) treated patients (P = 0.85) and for calcitriol (n = 50) versus placebo-treated patients (P = 0.58). Secondary endpoints also occurred at similar rates across groups. Hypercalcemia occurred in one patient in the calcifediol group (1.7%), one patient in the calcitriol group (2.0%), and none of the patients in the placebo group. Compared to placebo, calcitriol upregulated more individual genes and pathways in circulating monocytes than did calcifediol, including pathways involving interferon (IFN)-α, IFN-γ, oxidative phosphorylation, DNA repair, and heme metabolism. CONCLUSION. Treatment with calcifediol or calcitriol in critically ill adults upregulated multiple genes and pathways involving immunomodulation, DNA repair, and heme metabolism, but did not attenuate AKI. TRIAL REGISTRATION. ClinicalTrials.gov (NCT02962102). FUNDING. NIH/NIDDK grant K23DK106448 (Leaf) and NIH/NHLBI grant R01HL16687 (Kim)
Authors
David E. Leaf, Tushar Shenoy, Kevin Zinchuk, Shruti Gupta, Julie-Alexia Dias, Daniel Sanchez-Almanzar, Adit A. Ginde, Humra Athar, Changde Cheng, Tomoyoshi Tamura, Edy Y. Kim, Sushrut S. Waikar
Abstract
Mutations on genes encoding polycystin-1 (PC1) and -2 (PC2) cause autosomal-dominant polycystic kidney disease. How these two proteins work together to exert anti-cystogenesis remains elusive. PC1 resembles adhesion G-protein coupled receptors and undergoes autocleavage in the extracellular N-terminus to expose a hidden “stalk” region, which is hypothesized to act as a “tethered agonist”. Here, we showed that wildtype PC1 and PC2 formed functional heteromeric channel complexes in Xenopus oocytes with different biophysical properties from PC2 homomeric channels. Deletion of PC1 N-terminus, which exposed the stalk, increased calcium permeability in PC1/PC2 heteromers that required the presence of stalk. Extracellular application of synthetic stalk peptide increased calcium permeation in stalkless PC1/PC2. Application of Wnt9B protein increased calcium permeability in PC1/PC2, but not in heteromers containing cleavage-resistant mutant PC1. Wnt9B interacted with N-terminal leucine-rich repeat (LRR) of PC1. Pretreatment with LRR blunted the increase in calcium permeability by Wnt9B. Thus, PC1 and PC2 form receptor-channel complexes that is activated by exposure of the stalk region following ligand binding to the PC1 N-terminus. The stalk peptide acts as a tethered agonist to activate PC1/PC2 by impacting ion selectivity of the complexes.
Authors
Runping Wang, Danish Idrees, Mohammad Amir, Biswajit Padhy, Jian Xie, Chou-Long Huang
Abstract
Mutations in the transcription factor TFAP2A are linked to congenital anomalies of the kidney and urinary tract in humans. While Tfap2a knockout (KO) in mouse collecting ducts leads to tubular epithelial abnormalities, its precise molecular functions in kidney tubules remain unclear. To investigate Tfap2a-dependent gene regulatory networks in the mouse kidney collecting ducts, we employed conditional knockout (Hoxb7-Cre; Tfap2aflox/flox) models combined with transcriptomics. Histomorphological and physiological assessments of Tfap2a knockout mice revealed progressive postnatal dilation of the outer medullary collecting ducts. Integrating bulk and single-nucleus RNA sequencing with in silico motif mapping in ATAC-seq datasets demonstrated that Tfap2a is highly expressed and active in normal collecting duct principal cells. Comparative transcriptomics between 3-month-old Tfap2a KO and control mice identified dysregulated genes associated with cell adhesion and WNT signaling, including Alcam and Wnt9b. These alterations were confirmed by in situ hybridization. Our findings reveal that Tfap2a regulates medullary collecting duct diameter by orchestrating a transcriptional network involving Wnt9b and Alcam, providing new insights into its role in kidney structural integrity.
Authors
Janna Leiz, Karen I. López-Cayuqueo, Shuang Cao, Louisa M. S. Gerhardt, Christian Hinze, Kai M. Schmidt-Ott
Abstract
BK virus nephropathy is a severe, graft-threatening complication of kidney transplantation that requires an effective T cell response. It typically emerges in the kidney medulla. Elevated osmolyte concentrations that dynamically respond to loop diuretic therapy characterize this environment. BK-viremia development in kidney graft recipients negatively correlated with loop diuretic therapy. The association remained significant in multivariable and propensity score matched analyses. Kidney function was better preserved and CD8+ T cell abundance higher in loop diuretic-exposed allografts. CD8+ T cell densities in healthy human and murine kidney medulla were lower than in cortex and increased upon loop diuretic therapy in mice. As a potential underlying mechanism, kidney medullary NaCl and urea concentrations decreased primary human CD8+ T cell numbers in vitro by induction of cell death and limitation of proliferation, respectively. Both osmolytes downregulated interferon-related gene expression. NaCl induced p53-dependent apoptosis and upregulated Na+-transporter SLC38A2, which promoted caspase 3 activation. Both decreased T cell response and cytokine secretion in response to viral peptide and allogenic tubular epithelial cell killing, components of anti-BKV response in the kidney allograft. Our results propose osmolyte-mediated mitigation of CD8+ T cell function as a what we believe to be novel mechanism that impairs immune response to BK virus, therapeutic potential of which is testable.
Authors
Peyman Falahat, Adrian Goldspink, Lucia Oehler, Jessica Schmitz, Julia Miranda, Islem Gammoudi, Jan Hinrich Bräsen, Niklas Klümper, Olena Babyak, Christian Kurts, Herrmann Haller, Marieta Toma, Sibylle von Vietinghoff
Abstract
Nonresolving inflammation and maladaptive renal repair contribute to the pathogenesis of acute kidney injury (AKI) transition to chronic kidney disease (CKD). Few therapies have been identified that can modulate these injurious pathways following AKI. Spleen tyrosine kinase (SYK) is an immune regulator expressed in the kidney and a potential therapeutic target for AKI. The effect of the selective SYK inhibitor entospletinib was studied in AKI-to-CKD transition. Entospletinib was administered to mice undergoing unilateral renal ischemia-reperfusion injury (IRI), with kidneys analyzed over 14 days. Single-cell RNA sequencing, digital spatial profiling, intravital microscopy, and flow cytometry were employed to study renal phenotypes. Entospletinib administered before and after IRI protected ischemic kidneys and significantly attenuated the transition to CKD. Entospletinib targeted leukocyte-expressed SYK and prevented neutrophil/monocyte recruitment to the kidney. Entospletinib reduced nonresolving tubulointerstitial inflammation after AKI by blocking activation of mannose receptor-1– and C-type lectin domain family 7 member A–expressing proinflammatory macrophages. The resolution of renal inflammation mediated by entospletinib was associated with a reciprocal increase in resident macrophages, reparative gene expression, preserved tubular integrity, and reduced renal fibrosis. The SYK inhibitor entospletinib resolves renal inflammation and promotes repair following AKI.
Authors
Esteban E. Elias, Arthur Lau, Sisay Getie Belay, Afshin Derakhshani, Graciela Andonegui, Craig N. Jenne, Antoine Dufour, Nathan A. Bracey, Justin Chun, Daniel A. Muruve
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