Podocyte-targeted therapies - progress and future directions.

Podocytes are the key target cells for injury across the spectrum of primary and secondary proteinuric kidney disorders, which account for up to 90% of cases of kidney failure worldwide. Seminal experimental and clinical studies have established a causative link between podocyte depletion and the magnitude of proteinuria in progressive glomerular disease. However, no substantial advances have been made in glomerular disease therapies, and the standard of care for podocytopathies relies on repurposed immunosuppressive drugs. The past two decades have seen a remarkable expansion in understanding of the mechanistic basis of podocyte injury, with prospects increasing for precision-based treatment approaches. Dozens of disease-causing genes with roles in the pathogenesis of clinical podocytopathies have been identified, as well as a number of putative glomerular permeability factors. These achievements, together with the identification of novel targets of podocyte injury, the development of potential approaches to harness the endogenous podocyte regenerative potential of progenitor cell populations, ongoing clinical trials of podocyte-specific pharmacological agents and the development of podocyte-directed drug delivery systems, contribute to an optimistic outlook for the future of glomerular disease therapy.

Rationale and Design of a Phase 2, Double-blind, Placebo-Controlled, Randomized Trial Evaluating AMP Kinase-Activation by Metformin in Focal Segmental Glomerulosclerosis.

Introduction: Focal segmental glomerulosclerosis (FSGS), the most common primary glomerular disease leading to end-stage kidney disease (ESKD), is characterized by podocyte injury and depletion, whereas minimal change disease (MCD) has better outcomes despite podocyte injury. Identifying mechanisms capable of preventing podocytopenia during injury could transform FSGS to an "MCD-like" state. Preclinical data have reported conversion of an MCD-like injury to one with podocytopenia and FSGS by inhibition of AMP-kinase (AMPK) in podocytes. Conversely, in FSGS, AMPK-activation using metformin (MF) mitigated podocytopenia and azotemia. Observational studies also support beneficial effects of MF on proteinuria and chronic kidney disease (CKD) outcomes in diabetes. A randomized controlled trial (RCT) to test MF in podocyte injury with FSGS has not yet been conducted. Methods: We report the rationale and design of phase 2, double-blind, placebo-controlled RCT evaluating the efficacy and safety of MF as adjunctive therapy in FSGS. By randomizing 30 patients with biopsy-confirmed FSGS to MF or placebo (along with standard immunosuppression), we will study mechanistic biomarkers that correlate with podocyte injury or depletion and evaluate outcomes after 6 months. We specifically integrate novel urine, blood, and tissue markers as surrogates for FSGS progression along with unbiased profiling strategies. Results and Conclusion: Our phase 2 trial will provide insight into the potential efficacy and safety of MF as adjunctive therapy in FSGS-a crucial step to developing a larger phase 3 study. The mechanistic assays here will guide the design of other FSGS trials and contribute to understanding AMPK activation as a potential therapeutic target in FSGS. By repurposing an inexpensive agent, our results will have implications for FSGS treatment in resource-poor settings.

Podocyte-derived soluble RARRES1 drives kidney disease progression through direct podocyte and proximal tubular injury.

Retinoic acid receptor responder protein-1 (RARRES1) is a podocyte-enriched transmembrane protein whose increased expression correlates with human glomerular disease progression. RARRES1 promotes podocytopenia and glomerulosclerosis via p53-mediated podocyte apoptosis. Importantly, the cytopathic actions of RARRES1 are entirely dependent on its proteolytic cleavage into a soluble protein (sRARRES1) and subsequent podocyte uptake by endocytosis, as a cleavage mutant RARRES1 exerted no effects in vitro or in vivo. As RARRES1 expression is upregulated in human glomerular diseases, here we investigated the functional consequence of podocyte-specific overexpression of RARRES1 in mice in the experimental focal segmental glomerulosclerosis and diabetic kidney disease. We also examined the effects of long-term RARRES1 overexpression on slowly developing aging-induced kidney injury. As anticipated, the induction of podocyte overexpression of RARRES1 (Pod-RARRES1WT) significantly worsened glomerular injuries and worsened kidney function in all three models, while overexpression of RARRES1 cleavage mutant (Pod-RARRES1MT) did not. Remarkably, direct uptake of sRARRES1 was also seen in proximal tubules of injured Pod-RARRES1WT mice and associated with exacerbated tubular injuries, vacuolation, and lipid accumulation. Single cell RNA sequence analysis of mouse kidneys demonstrated RARRES1 led to a marked deregulation of lipid metabolism in proximal tubule subsets. We further identified matrix metalloproteinase 23 (MMP23) as a highly podocyte-specific metalloproteinase and responsible for RARRES1 cleavage in disease settings, as adeno-associated virus 9-mediated knockdown of MMP23 abrogated sRARRES1 uptake in tubular cells in vivo. Thus, our study delineates a previously unrecognized mechanism by which a podocyte-derived protein directly facilitates podocyte and tubular injury in glomerular diseases and suggests that podocyte-specific functions of RARRES1 and MMP23 may be targeted to ameliorate glomerular disease progression in vivo.

Integrated multiomics implicates dysregulation of ECM and cell adhesion pathways as drivers of severe COVID-associated kidney injury.

COVID-19 has been a significant public health concern for the last four years; however, little is known about the mechanisms that lead to severe COVID-associated kidney injury. In this multicenter study, we combined quantitative deep urinary proteomics and machine learning to predict severe acute outcomes in hospitalized COVID-19 patients. Using a 10-fold cross-validated random forest algorithm, we identified a set of urinary proteins that demonstrated predictive power for both discovery and validation set with 87% and 79% accuracy, respectively. These predictive urinary biomarkers were recapitulated in non-COVID acute kidney injury revealing overlapping injury mechanisms. We further combined orthogonal multiomics datasets to understand the mechanisms that drive severe COVID-associated kidney injury. Functional overlap and network analysis of urinary proteomics, plasma proteomics and urine sediment single-cell RNA sequencing showed that extracellular matrix and autophagy-associated pathways were uniquely impacted in severe COVID-19. Differentially abundant proteins associated with these pathways exhibited high expression in cells in the juxtamedullary nephron, endothelial cells, and podocytes, indicating that these kidney cell types could be potential targets. Further, single-cell transcriptomic analysis of kidney organoids infected with SARS-CoV-2 revealed dysregulation of extracellular matrix organization in multiple nephron segments, recapitulating the clinically observed fibrotic response across multiomics datasets. Ligand-receptor interaction analysis of the podocyte and tubule organoid clusters showed significant reduction and loss of interaction between integrins and basement membrane receptors in the infected kidney organoids. Collectively, these data suggest that extracellular matrix degradation and adhesion-associated mechanisms could be a main driver of COVID-associated kidney injury and severe outcomes.

HIPK2 C-terminal domain inhibits NF-κB signaling and renal inflammation in kidney injury.

HIPK2 is a multifunctional kinase that acts as a key pathogenic mediator of chronic kidney disease and fibrosis. It acts as a central effector of multiple signaling pathways implicated in kidney injury, such as TGF-ß/Smad3-mediated extracellular matrix accumulation, NF-κB-mediated inflammation, and p53-mediated apoptosis. Thus, a better understanding of the specific HIPK2 regions necessary for distinct downstream pathway activation is critical for optimal drug development for CKD. Our study now shows that caspase-6-mediated removal of the C-terminal region of HIPK2 (HIPK2-CT) lead to hyperactive p65 NF-κB transcriptional response in kidney cells. In contrast, the expression of cleaved HIPK2-CT fragment could restrain the NF-κB transcriptional activity by cytoplasmic sequestration of p65 and the attenuation of IκBα degradation. Therefore, we examined whether HIPK2-CT expression can be exploited to restrain renal inflammation in vivo. The induction of HIPK2-CT overexpression in kidney tubular cells attenuated p65 nuclear translocation, expression of inflammatory cytokines, and macrophage infiltration in the kidneys of mice with unilateral ureteral obstruction and LPS-induced acute kidney injury. Collectively, our findings indicate that the HIPK2-CT is involved in the regulation of nuclear NF-κB transcriptional activity and that HIPK2-CT or its analogs could be further exploited as potential antiinflammatory agents to treat kidney disease.

Edge roughness quantifies impact of physician variation on training and performance of deep learning auto-segmentation models for the esophagus.

Manual segmentation of tumors and organs-at-risk (OAR) in 3D imaging for radiation-therapy planning is time-consuming and subject to variation between different observers. Artificial intelligence (AI) can assist with segmentation, but challenges exist in ensuring high-quality segmentation, especially for small, variable structures, such as the esophagus. We investigated the effect of variation in segmentation quality and style of physicians for training deep-learning models for esophagus segmentation and proposed a new metric, edge roughness, for evaluating/quantifying slice-to-slice inconsistency. This study includes a real-world cohort of 394 patients who each received radiation therapy (mainly for lung cancer). Segmentation of the esophagus was performed by 8 physicians as part of routine clinical care. We evaluated manual segmentation by comparing the length and edge roughness of segmentations among physicians to analyze inconsistencies. We trained eight multiple- and individual-physician segmentation models in total, based on U-Net architectures and residual backbones. We used the volumetric Dice coefficient to measure the performance for each model. We proposed a metric, edge roughness, to quantify the shift of segmentation among adjacent slices by calculating the curvature of edges of the 2D sagittal- and coronal-view projections. The auto-segmentation model trained on multiple physicians (MD1-7) achieved the highest mean Dice of 73.7 ± 14.8%. The individual-physician model (MD7) with the highest edge roughness (mean ± SD: 0.106 ± 0.016) demonstrated significantly lower volumetric Dice for test cases compared with other individual models (MD7: 58.5 ± 15.8%, MD6: 67.1 ± 16.8%, p < 0.001). A multiple-physician model trained after removing the MD7 data resulted in fewer outliers (e.g., Dice ≤ 40%: 4 cases for MD1-6, 7 cases for MD1-7, Ntotal = 394). While we initially detected this pattern in a single clinician, we validated the edge roughness metric across the entire dataset. The model trained with the lowest-quantile edge roughness (MDER-Q1, Ntrain = 62) achieved significantly higher Dice (Ntest = 270) than the model trained with the highest-quantile ones (MDER-Q4, Ntrain = 62) (MDER-Q1: 67.8 ± 14.8%, MDER-Q4: 62.8 ± 15.7%, p < 0.001). This study demonstrates that there is significant variation in style and quality in manual segmentations in clinical care, and that training AI auto-segmentation algorithms from real-world, clinical datasets may result in unexpectedly under-performing algorithms with the inclusion of outliers. Importantly, this study provides a novel evaluation metric, edge roughness, to quantify physician variation in segmentation which will allow developers to filter clinical training data to optimize model performance.

Additive renal protective effects between arctigenin and puerarin in diabetic kidney disease.

Recent studies have shown that the combined use of renin angiotensin system inhibitor, SGLT2 inhibitors and/or mineralocorticoid receptor antagonist provides additional renal protection for patients with diabetic kidney disease (DKD). Similarly, in traditional Chinese medicine, the synergistic application of multiple herbs often brings more significant therapeutic effects. However, the synergistic or additive mechanisms of traditional Chinese medicine in combination therapy are not fully understood. In our previous studies, we show that arctigenin (ATG), a major component of Fructus Arctii, attenuates proteinuria and renal injury in diabetic mice by activating PP2A, and puerarin (a class of known isoflavones) can also reduce proteinuria and renal injury in diabetic mice via activation of Sirt1. Here, we further explored the potential additive renal protection of these two compounds in diabetic mice. Research has found that ATG and puerarin have a synergistic effect in reducing albuminuria in db/db mice. Mechanistically, we found that ATG reduced NF-κB p65 phosphorylation likely through activation of PP2A while puerarin reduced p65 acetylation via Sirt1 activation. Therefore, ATG and puerarin have additive inhibitory effects on the NF-κB activation, which is a key inflammatory pathway in DKD. RNA-sequencing analysis revealed distinct pathways activated by ATG and puerarin in the diabetic kidney, which may provide an additional mechanism for their additive effects in DKD. Our study suggests that ATG and puerarin could be a new combination therapy for DKD and reveals its underlined mechanisms.

Chronic kidney disease in a murine model of non-alcoholic steatohepatitis (NASH).

Clinical studies suggest that non-alcoholic steatohepatitis (NASH) is an independent risk factor for chronic kidney disease (CKD), but causality and mechanisms linking these two major diseases are lacking. To assess whether NASH can induce CKD, we have characterized kidney function, histological features, transcriptomic and lipidomic profiles in a well-validated murine NASH model. Mice with NASH progressively developed significant podocyte foot process effacement, proteinuria, glomerulosclerosis, tubular epithelial cell injury, lipid accumulation, and interstitial fibrosis. The progression of kidney fibrosis paralleled the severity of the histologic NASH-activity score. Significantly, we confirmed the causal link between NASH and CKD by orthotopic liver transplantation, which attenuated proteinuria, kidney dysfunction, and fibrosis compared with control sham operated mice. Transcriptomic analysis of mouse kidney cortices revealed differentially expressed genes that were highly enriched in mitochondrial dysfunction, lipid metabolic process, and insulin signaling pathways in NASH-induced CKD. Lipidomic analysis of kidney cortices further revealed that phospholipids and sphingolipids were the most significantly changed lipid species. Notably, we found similar kidney histological changes in human NASH and CKD. Thus, our results confirm a causative role of NASH in the development of CKD, reveal potential pathophysiologic mechanisms of NASH-induced kidney injury, and established a valuable model to study the pathogenesis of NASH-associated CKD. This is an important feature of fatty liver disease that has been largely overlooked but has clinical and prognostic importance.

Peripheral Transcriptomics in Acute and Long-Term Kidney Dysfunction in SARS-CoV2 Infection.

Background: Acute kidney injury (AKI) is common in hospitalized patients with SARS-CoV2 infection despite vaccination and leads to long-term kidney dysfunction. However, peripheral blood molecular signatures in AKI from COVID-19 and their association with long-term kidney dysfunction are yet unexplored. Methods: In patients hospitalized with SARS-CoV2, we performed bulk RNA sequencing using peripheral blood mononuclear cells(PBMCs). We applied linear models accounting for technical and biological variability on RNA-Seq data accounting for false discovery rate (FDR) and compared functional enrichment and pathway results to a historical sepsis-AKI cohort. Finally, we evaluated the association of these signatures with long-term trends in kidney function. Results: Of 283 patients, 106 had AKI. After adjustment for sex, age, mechanical ventilation, and chronic kidney disease (CKD), we identified 2635 significant differential gene expressions at FDR<0.05. Top canonical pathways were EIF2 signaling, oxidative phosphorylation, mTOR signaling, and Th17 signaling, indicating mitochondrial dysfunction and endoplasmic reticulum (ER) stress. Comparison with sepsis associated AKI showed considerable overlap of key pathways (48.14%). Using follow-up estimated glomerular filtration rate (eGFR) measurements from 115 patients, we identified 164/2635 (6.2%) of the significantly differentiated genes associated with overall decrease in long-term kidney function. The strongest associations were 'autophagy', 'renal impairment via fibrosis', and 'cardiac structure and function'. Conclusions: We show that AKI in SARS-CoV2 is a multifactorial process with mitochondrial dysfunction driven by ER stress whereas long-term kidney function decline is associated with cardiac structure and function and immune dysregulation. Functional overlap with sepsis-AKI also highlights common signatures, indicating generalizability in therapeutic approaches. SIGNIFICANCE STATEMENT: Peripheral transcriptomic findings in acute and long-term kidney dysfunction after hospitalization for SARS-CoV2 infection are unclear. We evaluated peripheral blood molecular signatures in AKI from COVID-19 (COVID-AKI) and their association with long-term kidney dysfunction using the largest hospitalized cohort with transcriptomic data. Analysis of 283 hospitalized patients of whom 37% had AKI, highlighted the contribution of mitochondrial dysfunction driven by endoplasmic reticulum stress in the acute stages. Subsequently, long-term kidney function decline exhibits significant associations with markers of cardiac structure and function and immune mediated dysregulation. There were similar biomolecular signatures in other inflammatory states, such as sepsis. This enhances the potential for repurposing and generalizability in therapeutic approaches.

Viral Glomerulopathy.

Background: The association between viral infections and glomerular diseases, commonly known as "viral glomerulopathies," has been described in various clinical scenarios for decades. Despite advancements in diagnostic tools, it remains challenging to establish a causative link fully. Summary: Data from mouse models have substantiated clinical observations and implicate direct viral infection in the pathogenesis of viral glomerulopathy, particularly in human immunodeficiency virus-associated nephropathy. In addition to the traditional concept of direct viral effects on kidneys, other factors such as APOL1 risk alleles can further modify the clinical outcomes or presentations of different viral glomerulopathies. Newly developed antiviral drugs are now applicable to a wider range of patients with lower kidney function and fewer side effects. Key Message: Efforts focusing on vaccines and antiviral treatments have significantly reduced the incidence of viral glomerulopathies. However, the most recent pandemic caused by severe acute respiratory syndrome coronavirus 2 infection complicated by COVID-associated nephropathy illustrates our susceptibility to novel viruses. Ongoing research is pivotal to deciphering the mechanisms behind viral glomerulopathies and discovering therapeutics in a collaborative approach.

Deep learning on electronic medical records identifies distinct subphenotypes of diabetic kidney disease driven by genetic variations in the Rho pathway.

Kidney disease affects 50% of all diabetic patients; however, prediction of disease progression has been challenging due to inherent disease heterogeneity. We use deep learning to identify novel genetic signatures prognostically associated with outcomes. Using autoencoders and unsupervised clustering of electronic health record data on 1,372 diabetic kidney disease patients, we establish two clusters with differential prevalence of end-stage kidney disease. Exome-wide associations identify a novel variant in ARHGEF18, a Rho guanine exchange factor specifically expressed in glomeruli. Overexpression of ARHGEF18 in human podocytes leads to impairments in focal adhesion architecture, cytoskeletal dynamics, cellular motility, and RhoA/Rac1 activation. Mutant GEF18 is resistant to ubiquitin mediated degradation leading to pathologically increased protein levels. Our findings uncover the first known disease-causing genetic variant that affects protein stability of a cytoskeletal regulator through impaired degradation, a potentially novel class of expression quantitative trait loci that can be therapeutically targeted.

Multiscale genetic architecture of donor-recipient differences reveals intronic LIMS1 mismatches associated with kidney transplant survival.

Donor-recipient (D-R) mismatches outside of human leukocyte antigens (HLAs) contribute to kidney allograft loss, but the mechanisms remain unclear, specifically for intronic mismatches. We quantified non-HLA mismatches at variant-, gene-, and genome-wide scales from single nucleotide polymorphism (SNP) data of D-Rs from 2 well-phenotyped transplant cohorts: Genomics of Chronic Allograft Rejection (GoCAR; n = 385) and Clinical Trials in Organ Transplantation-01/17 (CTOT-01/17; n = 146). Unbiased gene-level screening in GoCAR uncovered the LIMS1 locus as the top-ranked gene where D-R mismatches associated with death-censored graft loss (DCGL). A previously unreported, intronic, LIMS1 haplotype of 30 SNPs independently associated with DCGL in both cohorts. Haplotype mismatches showed a dosage effect, and minor-allele introduction to major-allele-carrying recipients showed greater hazard of DCGL. The LIMS1 haplotype and the previously reported LIMS1 SNP rs893403 are expression quantitative trait loci (eQTL) in immune cells for GCC2 (not LIMS1), which encodes a protein involved in mannose-6-phosphase receptor (M6PR) recycling. Peripheral blood and T cell transcriptome analyses associated the GCC2 gene and LIMS1 SNPs with the TGF-ß1/SMAD pathway, suggesting a regulatory effect. In vitro GCC2 modulation impacted M6PR-dependent regulation of active TGF-ß1 and downstream signaling in T cells. Together, our data link LIMS1 locus D-R mismatches to DCGL via GCC2 eQTLs that modulate TGF-ß1-dependent effects on T cells.

GPR56 Promotes Diabetic Kidney Disease Through eNOS Regulation in Glomerular Endothelial Cells.

Although glomerular endothelial dysfunction is well recognized as contributing to the pathogenesis of diabetic kidney disease (DKD), the molecular pathways contributing to DKD pathogenesis in glomerular endothelial cells (GECs) are only partially understood. To uncover pathways that are differentially regulated in early DKD that may contribute to disease pathogenesis, we recently conducted a transcriptomic analysis of isolated GECs from diabetic NOS3-null mice. The analysis identified several potential mediators of early DKD pathogenesis, one of which encoded an adhesion G protein-coupled receptor-56 (GPR56), also known as ADGRG1. Enhanced glomerular expression of GPR56 was observed in human diabetic kidneys, which was negatively associated with kidney function. Using cultured mouse GECs, we observed that GPR56 expression was induced with exposure to advanced glycation end products, as well as in high-glucose conditions, and its overexpression resulted in decreased phosphorylation and expression of endothelial nitric oxide synthase (eNOS). This effect on eNOS by GPR56 was mediated by coupling of Gα12/13-RhoA pathway activation and Gαi-mediated cAMP/PKA pathway inhibition. The loss of GPR56 in mice led to a significant reduction in diabetes-induced albuminuria and glomerular injury, which was associated with reduced oxidative stress and restoration of eNOS expression in GECs. These findings suggest that GPR56 promotes DKD progression mediated, in part, through enhancing glomerular endothelial injury and dysfunction.

Relationship between Macrophages and Tissue Microenvironments in Diabetic Kidneys.

Diabetic nephropathy (DN) is the leading cause of end-stage kidney disease. Increasing evidence has suggested that inflammation is a key microenvironment involved in the development and progression of DN. Studies have confirmed that macrophage accumulation is closely related to the progression to human DN. Macrophage phenotype is highly regulated by the surrounding microenvironment in the diabetic kidneys. M1 and M2 macrophages represent distinct and sometimes coexisting functional phenotypes of the same population, with their roles implicated in pathological changes, such as in inflammation and fibrosis associated with the stage of DN. Recent findings from single-cell RNA sequencing of macrophages in DN further confirmed the heterogeneity and plasticity of the macrophages. In addition, intrinsic renal cells interact with macrophages directly or through changes in the tissue microenvironment. Macrophage depletion, modification of its polarization, and autophagy could be potential new therapies for DN.

Comparative evaluation of glomerular morphometric techniques reveals differential technical artifacts between focal segmental glomerulosclerosis and normal glomeruli.

Morphometric estimates of mean or individual glomerular volume (MGV, IGV) have biological implications, over and above qualitative histologic data. However, morphometry is time-consuming and requires expertise limiting its utility in clinical cases. We evaluated MGV and IGV using plastic- and paraffin-embedded tissue from 10 control and 10 focal segmental glomerulosclerosis (FSGS) mice (aging and 5/6th nephrectomy models) using the gold standard Cavalieri (Cav) method versus the 2-profile and Weibel-Gomez (WG) methods and a novel 3-profile method. We compared accuracy, bias and precision, and quantified results obtained when sampling differing numbers of glomeruli. In both FSGS and controls, we identified an acceptable precision for MGV of 10-glomerular sampling versus 20-glomerular sampling using the Cav method, while 5-glomerular sampling was less precise. In plastic tissue, 2- or 3-profile MGVs showed greater concordance with MGV when using Cav, versus MGV with WG. IGV comparisons using the same glomeruli reported a consistent underestimation bias with both 2- or 3-profile methods versus the Cav method. FSGS glomeruli showed wider variations in bias estimation than controls. Our 3-profile method offered incremental benefit to the 2-profile method in both IGV and MGV estimation (improved correlation coefficient, Lin's concordance and reduced bias). In our control animals, we quantified a shrinkage artifact of 52% from tissue processed for paraffin-embedded versus plastic-embedded tissue. FSGS glomeruli showed overall reduced shrinkage albeit with variable artifact signifying periglomerular/glomerular fibrosis. A novel 3-profile method offers slightly improved concordance with reduced bias versus 2-profile. Our findings have implications for future studies using glomerular morphometry.

Basin record of a Miocene lithosphere drip beneath the Colorado Plateau.

The sinking of gravitationally unstable lithosphere beneath high-elevation plateaus is proposed to be a key driver of their uplift. Numerical geodynamic models predict that lithosphere removal can lead to transient, dynamic topographic changes that could be preserved in the surface record, particularly in sedimentary deposits of lakes or playas that are subsequently inverted. However, few such examples have been documented. Here we show that the Miocene Bidahochi Basin, which was partially and intermittently filled by the Hopi Paleolake, preserves a record of the quasi-elliptical surface response to a viscous drip of lithosphere >100 km beneath the Colorado Plateau. New detrital zircon U-Pb, Lu-Hf, and trace-element data reveal systematic isotopic, geochemical, temperature and fO2 transitions in magmatism proximal to the basin. Integration of geophysical, geochemical, and geological evidence supports a spatially and temporally varying record of subsidence and uplift that is consistent with models of progressive dripping beneath plateaus with thick lithosphere. We demonstrate that dynamic topography at the scale of individual lithosphere drips can be recognized on the Colorado Plateau, despite the strength of its lithosphere.

HCK induces macrophage activation to promote renal inflammation and fibrosis via suppression of autophagy.

Renal inflammation and fibrosis are the common pathways leading to progressive chronic kidney disease (CKD). We previously identified hematopoietic cell kinase (HCK) as upregulated in human chronic allograft injury promoting kidney fibrosis; however, the cellular source and molecular mechanisms are unclear. Here, using immunostaining and single cell sequencing data, we show that HCK expression is highly enriched in pro-inflammatory macrophages in diseased kidneys. HCK-knockout (KO) or HCK-inhibitor decreases macrophage M1-like pro-inflammatory polarization, proliferation, and migration in RAW264.7 cells and bone marrow-derived macrophages (BMDM). We identify an interaction between HCK and ATG2A and CBL, two autophagy-related proteins, inhibiting autophagy flux in macrophages. In vivo, both global or myeloid cell specific HCK-KO attenuates renal inflammation and fibrosis with reduces macrophage numbers, pro-inflammatory polarization and migration into unilateral ureteral obstruction (UUO) kidneys and unilateral ischemia reperfusion injury (IRI) models. Finally, we developed a selective boron containing HCK inhibitor which can reduce macrophage pro-inflammatory activity, proliferation, and migration in vitro, and attenuate kidney fibrosis in the UUO mice. The current study elucidates mechanisms downstream of HCK regulating macrophage activation and polarization via autophagy in CKD and identifies that selective HCK inhibitors could be potentially developed as a new therapy for renal fibrosis.