Human Chronic Kidney Disease Research Articles

MO056: Alteration of Glycocalyx on Endothelium of Peritubular Capillaries in CKD

Abstract BACKGROUND AND AIMS The glycocalyx is a carbohydrate-rich gel-like mesh, which covers the luminal surface of cells, including the endothelium. The glycocalyx is involved in many regulatory functions of the endothelium, including vascular permeability. In chronic kidney disease (CKD), peritubular capillaries undergo anatomical, structural and functional alterations such as rarefaction, increased tortuosity and increased permeability. Here, we hypothesized that the glycocalyx of peritubular capillaries might be affected in CKD and investigated morphological and ultrastructural pathological alterations of the glycocalyx in different murine CKD models and human kidney tissue. METHOD We stained the glycocalyx of peritubular capillaries in kidney tissue of murine and human CKD specimen using different fluorescently labeled plant-derived lectins. Next, we established the Lanthanum Dysprosium Glycosamino Glycan adhesion (LaDy GAGa) staining technique to visualize the ultrastructure of the glycocalyx using transmission electron microscopy and to perform quantitative analyses. Finally, we analyzed the expression and regulation of glycocalyx components in primary murine endothelial cells. RESULTS Fluorescence stainings using different lectins with high affinity to components of the renal glycocalyx revealed a reduced binding to the endothelium in CKD. We found a similar change in a human kidney tissue. The LaDy GAGa staining technique visualized the ultrastructure of the glycocalyx and enabled quantitive analyses. We found a significant reduction of the glycocalyx thickness and density in two different models of renal fibrosis and CKD, i.e. unilateral ureteral obstruction and ischemia–reperfusion injury. Additionally, mRNA expression of proteins involved in glycocalyx biology, synthesis and turnover, i.e. syndecan 1 and glypican 1, which are main components of the glycocalyx, and exostosin 2, involved in the synthesis of the glycocalyx, were significantly upregulated in endothelial cells isolated from murine CKD models. CONCLUSION Visualization of glycocalyx using specific ultrastructural analyses allows qualitative and quantitative analyses, revealing significant pathological alterations in the glycocalyx of peritubular capillaries in CKD.

Apelinergic System: an in‐vitro Investigation of Inflammatory & Oxidative Stress in Chronic Kidney Disease

Over 37 million US adults have Chronic Kidney Disease (CKD). Of that population–35% are Black. With 4x higher predisposition to developing renal failure due to comorbidities (i.e. hypertension and diabetes), there is a dire need to address both preventative and therapeutic options in this population. Besides dialysis and transplant as reactionary treatments of CKD, vitamin E—an antioxidant—has been given to those in late stage CKD. Our study aims to address mechanisms behind CKD pathology. The 5/6 nephrectomy (5/6Nx) rat CKD model provides the opportunity to isolate the effects of impaired renal function without significant confounding comorbidities. A hallmark feature of both rat and human CKD is adverse vascular remodeling which is often preceded by endothelial cell (EC) dysfunction. With respect to EC dysfunction, inflammatory and oxidative imbalances are suspected to be involved. Our previous transcriptomic analysis of kidney tissues from the 5/6Nx rat identified a significant increase in apelin receptor (APLNR) expression compared to sham controls. In addition, the data revealed several pro‐inflammatory cytokines (IL‐6 and IL‐1β) preceding APLNR transcript increases. Treatment of Normal Rat Kidney epithelial (NRK) cells with IL‐6 caused a significant reduction in apelin (APLN) ligand, while IL‐1β augmented APLNR transcript levels. After further study of the 3 pathways, a common downstream target, protein kinase B (AKT), arose. Interestingly enough, AKT is responsible for various vascular cascades including interaction with well‐known reactive oxygen species (ROS) producer–NADPH Oxidase 4 (NOX4). Hydrogen Peroxide (H2O2), a ROS product of NOX4, coordinates vasoactivity through AKT signaling and has overlapping function with apelinergic (APJ) system in the vasculature. Due to the inflammatory and oxidative nature of CKD, we want to address the relationship between H2O2 and the APJ system. We hypothesize that pretreating NRK cells with APLN prior to administration of H2O2 will have 2 effects: oxidant attenuation and antioxidant augmentation. NRK cells were pretreated with 2.5ng/mL apelin or 30mg/mL vitamin E 48‐hours prior to addition of 20mM H2O2. At study termination, cells were collected/quantified using the Bradford protein assay. Protein expression was measured via Western blots against catalase, an antioxidant marker, and GPx, an oxidant protein. One‐Way ANOVA and Tukey Post‐Hoc test were used to establish statistical significance (p<0.05). Catalase: no significant expression changes after APLN or vitamin E treat (p = 0.10), but significant increases after H2O2 treat (p = 0.05) compared to control. GPx: significant increases in expression after APLN treat compared to [control (p=0.0002), vitamin E (p=0.04), and H2O2 + APLN (p=0.0001)] and vitamin E compared to H2O2 + Vitamin E treat (p=0.01). Data presented as mean ± SEM (n=3). These studies assessed functionality of the APJ system in an in‐vitro model of CKD. Both of the inflammatory cytokines are regulators of APJ system transcription, which is relevant for patients in a chronic inflammatory state. These studies also highlight the interaction between APJ system and oxidative stressors. Results here begin to lay the foundation for future studies involving the APJ system as a viable target for assessing CKD pathology.

Ovarian Hormone‐Dependent miR‐146b‐5p Regulation of Plasminogen Activator Inhibitor‐1 in Renal Epithelial Cells

Chronic kidney disease (CKD) progression differs in males and females and the influence of gonadal hormones on pathology are incompletely understood. We utilized the 5/6 nephrectomy (5/6Nx) rat model, which recapitulates hallmarks of human CKD, to study differences in regulation of CKD pathology between sexes. We previously found that genomic deletion of microRNA 146b‐5p (miR‐146b) results in a sex‐dependent phenotype in 5/6Nx rats. Male miR‐146b‐/‐ rats develop renal pathology similar to their WT counterparts. However, female miR‐146b‐/‐ rats experience markedly exacerbated renal pathology relative to WT, 5/6Nx, females as well as WT and miR‐146b‐/‐, male, 5/6Nx rats. These sex differences are abolished by ovary removal, suggesting that renoprotective effects of female gonadal hormones are miR‐146b dependent. Our objective is to identify a molecular mechanism behind these effects. PAI‐1 has previously been shown to be regulated by miR‐146b‐5p isomiR, miR‐146a‐5p, which shares a common seed sequence. PAI‐1 expression is also known to be ovarian hormone dependent, its renal expression is low in health, but elevated in many disease settings, where it may contribute to remodeling. We hypothesized that PAI‐1 expression in rat proximal tubular epithelial cells (NRK‐52E) will be dependent upon (1) the presence of miR‐146b, (2) ovarian hormones, and (3) transforming growth factor beta (TGFβ), which is elevated in our 5/6Nx rats and known to contribute to renal pathology. To aid in testing our hypothesis we used CRISPR‐Cas9 technology to generate miR‐146b knockout rat proximal tubule epithelial (NRK‐52E) cells. Results. PAI‐1 mRNA was highly elevated in untreated miR‐146b KO rat NRK‐52E cells relative to WT NRK‐52E cell controls (11.25 ± 5.3 vs. 1.00 ± 0.68, P<0.05). Conversely, WT NRK‐52E cells treated with pre‐miR‐146b had significantly reduced PAI‐1 mRNA expression (0.137 ± 0.14 vs. 1.00 ± 0.45, P<0.05). Treatment of WT NRK‐52E cells with β‐estradiol did not significantly alter PAI‐1 mRNA expression relative to untreated controls, whereas treatment with progesterone (P4) significantly increased it. Treatment of WT NRK‐52E cells with TGFβ resulted in a significant increase of PAI‐1 mRNA expression. Lastly, treatment of WT NRK‐52E cells with P4 and TGFβ combined resulted in a significant increase in PAI‐1 mRNA expression, and this expression was significantly reduced by transfection with pre‐miR‐146b. Collectively, these observations demonstrate that miR‐146b, P4, and TGFβ regulate PAI‐1 expression in NRK‐52E cells and thereby suggest PAI‐1 as a mechanistic candidate behind the sex‐specific effects of miR‐146b in CKD.

Effect of Lipocalin‐2 expression on mitochondrial function in renal tubular cells in chronic kidney disease

Kidney cells metabolic status requires high abundance of mitochondria. It has been reported in chronic kidney disease (CKD) a loss of mitochondrial mass and function leading to reactive oxygen species (ROS) production and subsequent tubular cell death. Hence, mitochondrial dysfunction participates to CKD onset. Nevertheless, the cellular mechanisms inducing mitochondrial dysfunction in kidney cells are poorly understood. We have recently shown that in CKD, Lipocalin‐2 (Lcn2) expression is strongly induced in kidney cells leading to ROS production and apoptosis. However, the mechanisms by which Lcn2 triggers these phenomena are unknown. Considering that mitochondria can induce oxidative stress and apoptosis, we hypothesized that Lcn2 participates to CKD onset via an effect on mitochondrial dependent mechanisms. Using renal tubular cell lines overexpressing (OE) or knock‐down (KD) for Lcn2, we aimed at studying the effect of Lcn2 protein expression on mitochondrial function. First, we demonstrated that Lcn2 regulates mitochondrial dynamics, mass and function. Indeed, we observed by immunofluorescence and electron microscopy that kidney cells expressing Lcn2 present smaller and rounded mitochondria while Lcn2 KD induces mitochondrial elongation. Moreover, Lcn2 expression increases mitochondrial fission marker abundance and decreases fusion marker expression. Mitotracker staining and mtDNA content measurement experiments demonstrated that Lcn2 expression decreased mitochondrial mass while Lcn2 KD increased it. Furthermore, we observed that Lcn2 expression decrease mitochondrial membrane potential and accordingly increased Lactate production compared to cell KD for Lcn2. Using iron chelator or Lcn2 mutant unable to transport iron, we demonstrated that Lcn2 expression induces mitochondrial dysfunction independently of its iron transport effect. In addition, we observed that extracellular Lcn2 did not rescue Lcn2 effects on mitochondria while intracellular Lcn2 did. Finally, using a subtotal nephrectomy mice model, that recapitulates different features of human CKD, we demonstrated that Lcn2 invalidation by knock‐out in mice decreased the mitochondrial fission and the subsequent mitochondrial fragmentation and mass loss in kidney tubular cells normally observed during CKD onset in wild‐type mice.Altogether, these results demonstrated that Lcn2 expression regulates mitochondrial dynamics, mass and function in kidney tubular cells. This suggests that Lcn2 driven mitochondrial dysfunction might by responsible at least in part of CKD onset.

The significance of NAD\u2009+\u2009metabolites and nicotinamide N-methyltransferase in chronic kidney disease

Dysregulation of nicotinamide adenine dinucleotide (NAD +) metabolism contributes to the initiation and progression of age-associated diseases, including chronic kidney disease (CKD). Nicotinamide N-methyltransferase (NNMT), a nicotinamide (NAM) metabolizing enzyme, regulates both NAD + and methionine metabolism. Although NNMT is expressed abundantly in the kidney, its role in CKD and renal fibrosis remains unclear. We generated NNMT-deficient mice and a unilateral ureter obstruction (UUO) model and conducted two clinical studies on human CKD to investigate the role of NNMT in CKD and fibrosis. In UUO, renal NNMT expression and the degraded metabolites of NAM increased, while NAD + and NAD + precursors decreased. NNMT deficiency ameliorated renal fibrosis; mechanistically, it (1) increased the DNA methylation of connective tissue growth factor (CTGF), and (2) improved renal inflammation by increasing renal NAD + and Sirt1 and decreasing NF-κB acetylation. In humans, along with CKD progression, a trend toward a decrease in serum NAD + precursors was observed, while the final NAD + metabolites were accumulated, and the level of eGFR was an independent variable for serum NAM. In addition, NNMT was highly expressed in fibrotic areas of human kidney tissues. In conclusion, increased renal NNMT expression induces NAD + and methionine metabolism perturbation and contributes to renal fibrosis.

In Vivo Assessment of Ferroptosis and Ferroptotic Stress in Mice.

Ferroptosis is iron-dependent, lipid peroxidation-driven, regulated cell death that is triggered when cellular glutathione peroxidase 4 (GPX4)-mediated cellular defense is insufficient to prevent pathologic accumulation of toxic lipid peroxides. Ferroptosis is implicated in various human pathologies, including neurodegeneration, chemotherapy-resistant cancers, ischemia-reperfusion injury, and acute and chronic kidney diseases. Despite the fact that the ferroptotic process has been rigorously interrogated in multiple preclinical models, the lack of specific and readily available biomarkers to detect ferroptosis in vivo in mouse models makes it challenging to delineate its contribution to key pathologic events in vivo. Critical steps to practically evaluate ferroptosis include, but are not limited to, detecting increased cell death and pathologic accumulation of toxic lipid peroxides and testing augmentation of observed pathologic events by genetic inhibition of the glutathione-GPX4 axis or mitigation of the pathologic process by ferroptosis inhibitors. Here, we describe methods to evaluate these key features of the ferroptotic process in mice in vivo. Specifically, we describe methods to detect toxic lipid peroxides (4-hydroxynonenal) and cell death (based on terminal deoxynucleotidyl transferase dUTP nick end labeling staining) as well as a protocol to pharmacologically inhibit ferroptotic stress using liproxstatin-1. These protocols provide tools for understanding the ferroptotic process in mouse genetic or disease models. © 2022 Wiley Periodicals LLC. Basic Protocol 1: How to use liproxstatin-1 Basic Protocol 2: How to evaluate ferroptosis in mouse kidneys.

The Signaling Pathway of TNF Receptors: Linking Animal Models of Renal Disease to Human CKD.

Chronic kidney disease (CKD) has been recognized as a global public health problem. Despite the current advances in medicine, CKD-associated morbidity and mortality remain unacceptably high. Several studies have highlighted the contribution of inflammation and inflammatory mediators to the development and/or progression of CKD, such as tumor necrosis factor (TNF)-related biomarkers. The inflammation pathway driven by TNF-α, through TNF receptors 1 (TNFR1) and 2 (TNFR2), involves important mediators in the pathogenesis of CKD. Circulating levels of TNFRs were associated with changes in other biomarkers of kidney function and injury, and were described as predictors of disease progression, cardiovascular morbidity, and mortality in several cohorts of patients. Experimental studies describe the possible downstream signaling pathways induced upon TNFR activation and the resulting biological responses. This review will focus on the available data on TNFR1 and TNFR2, and illustrates their contributions to the pathophysiology of kidney diseases, their cellular and molecular roles, as well as their potential as CKD biomarkers. The emerging evidence shows that TNF receptors could act as biomarkers of renal damage and as mediators of the disease. Furthermore, it has been suggested that these biomarkers could significantly improve the discrimination of clinical CKD prognostic models.

First genome-wide association study investigating blood pressure and renal traits in domestic cats

Hypertension (HTN) and chronic kidney disease (CKD) are common in ageing cats. In humans, blood pressure (BP) and renal function are complex heritable traits. We performed the first feline genome-wide association study (GWAS) of quantitative traits systolic BP and creatinine and binary outcomes HTN and CKD, testing 1022 domestic cats with a discovery, replication and meta-analysis design. No variants reached experimental significance level in the discovery stage for any phenotype. Follow up of the top 9 variants for creatinine and 5 for systolic BP, one SNP reached experimental-wide significance for association with creatinine in the combined meta-analysis (chrD1.10258177; P = 1.34 × 10–6). Exploratory genetic risk score (GRS) analyses were performed. Within the discovery sample, GRS of top SNPs from the BP and creatinine GWAS show strong association with HTN and CKD but did not validate in independent replication samples. A GRS including SNPs corresponding to human CKD genes was not significant in an independent subset of cats. Gene-set enrichment and pathway-based analysis (GSEA) was performed for both quantitative phenotypes, with 30 enriched pathways with creatinine. Our results support the utility of GWASs and GSEA for genetic discovery of complex traits in cats, with the caveat of our findings requiring validation.

Chronic kidney disease mediates cardiac dysfunction associated with increased resident cardiac macrophages

BackgroundThe leading cause of death in end-stage kidney disease is related to cardiovascular disease. Macrophages are known to be involved in both chronic kidney disease (CKD) and heart failure, however their role in the development of cardiorenal syndrome is less clear. We thus sought to investigate the role of macrophages in uremic cardiac disease.MethodsWe assessed cardiac response in two experimental models of CKD and tested macrophage and chemokine implication in monocytopenic CCR2−/− and anti-CXCL10 treated mice. We quantified CXCL10 in human CKD plasma and tested the response of human iPSC-derived cardiomyocytes and primary cardiac fibroblasts to serum from CKD donors.ResultsWe found that reduced kidney function resulted in the expansion of cardiac macrophages, in particular through local proliferation of resident populations. Influx of circulating monocytes contributed to this increase. We identified CXCL10 as a crucial factor for cardiac macrophage expansion in uremic disease. In humans, we found increased plasma CXCL10 concentrations in advanced CKD, and identified the production of CXCL10 in cardiomyocytes and cardiac fibroblasts.ConclusionsThis study provides new insight into the role of the innate immune system in uremic cardiomyopathy.

AIM2 inflammasome contributes to aldosterone-induced renal injury via endoplasmic reticulum stress.

Inflammatory response and renal fibrosis are the hallmarks of chronic kidney disease (CKD). However, the specific mechanism of aldosterone-induced renal injury in the progress of CKD requires elucidation. Emerging evidence has demonstrated that absent in melanoma 2 (AIM2)-mediated inflammasome activation and endoplasmic reticulum stress (ERS) play a pivotal role in the renal fibrosis. Here, we investigated whether overexpression or deficiency of AIM2 affects ERS and fibrosis in aldosterone-infused renal injury. Interestingly, we found that AIM2 was markedly expressed in the diseased proximal tubules from human and experimental CKD. Mechanically, overactivation of AIM2 aggravated aldosterone-induced ERS and fibrotic changes in vitro while knockdown of AIM2 blunted these effects in vivo and in vitro. By contrast, AIM2 deficiency ameliorated renal structure and function deterioration, decreased proteinuria levels and lowered systolic blood pressure in vivo; silencing of AIM2 blocked inflammasome-mediated signaling pathway, relieved ERS and fibrotic changes in vivo. Furthermore, mineralocorticoid receptor (MR) antagonist eplerenone and ERS inhibitor tauroursodeoxycholic acid (TUDCA) had nephroprotective effects on the basis of AIM2 overactivation in vitro, while they failed to produce a more remarkable renoprotective effect on the treatment of AIM2 silence in vitro. Notably, the combination of TUDCA with AIM2 knockdown significantly reduced proteinuria levels in vivo. Additionally, immunofluorescence assay identified that apoptosis-associated speck-like protein (ASC) recruitment and Gasdermin-D (GSDMD) cleavage respectively occurred in the glomeruli and tubules in vivo. These findings establish a crucial role for AIM2 inflammasome in aldosterone-induced renal injury, which may provide a novel therapeutic target for the pathogenesis of CKD.

Altered vitamin K biodistribution and metabolism in experimental and human chronic kidney disease

Chronic kidney disease (CKD) is accompanied with extensive cardiovascular calcification, in part correlating with functional vitamin K deficiency. Here, we sought to determine causes for vitamin K deficiency beyond reduced dietary intake. Initially, vitamin K uptake and distribution into circulating lipoproteins after a single administration of vitamin K1 plus K2 (menaquinone 4 and menaquinone 7, respectively) was determined in patients on dialysis therapy and healthy individuals. The patients incorporated very little menaquinone 7 but more menaquinone 4 into high density lipoprotein (HDL) and low-density lipoprotein particles than did healthy individuals. In contrast to healthy persons, HDL particles from the patients could not be spiked with menaquinone 7 invitro and HDL uptake was diminished in osteoblasts. A reduced carboxylation activity (low vitamin K activity) of uremic HDL particles spiked with menaquinone 7 vs. that of controls was confirmed in a bioassay using human primary vascular smooth muscle cells. Kidney menaquinone 4 tissue levels were reduced in 5/6-nephrectomized versus sham-operated C57BL/6 mice after four weeks of a vitamin K rich diet. From the analyzed enzymes involved in vitamin K metabolism, kidney HMG-CoA reductase protein was reduced in both rats and patients with CKD. In a trial on the efficacy and safety of atorvastatin in 1051 patients with type 2 diabetes receiving dialysis therapy, no pronounced vitamin K deficiency was noted. However, the highest levels of PIVKA-II (biomarker of subclinical vitamin K deficiency) were noted when a statin was combined with a proton pump inhibitor. Thus, profound disturbances in lipoprotein mediated vitamin K transport and metabolism in uremia suggest that menaquinone 7 supplementation to patients on dialysis therapy has reduced efficacy.

Tubule-derived INHBB promotes interstitial fibroblast activation and renal fibrosis.

Upstream stimuli for myofibroblast activation are of considerable interest for understanding the mechanisms underlying renal fibrosis. Activin B, a member of the TGF-β family, exists as a homodimer of inhibin subunit beta B (INHBB), but its role in renal fibrosis remains unknown. We found that INHBB expression was significantly increased in various renal fibrosis models and human chronic kidney disease specimens with renal fibrosis. Notably, the increase of INHBB occurred mainly in the tubular epithelial cells (TECs). In vivo, inhibiting INHBB blocked the activation of interstitial fibroblasts and ameliorated the renal fibrosis induced by unilateral ureteral obstruction or ischemia-reperfusion injury, while ectopic expression of INHBB in the TECs was able to activate interstitial fibroblasts and initiate interstitial fibrosis. In vitro, overexpression of INHBB in TECs led to the secretion of activin B, thereby promoting the proliferation and activation of interstitial fibroblasts through activin B/Smad signaling. Furthermore, inhibition of activin B/Smad signaling attenuated the fibrotic response caused by tubular INHBB. Mechanistically, the upregulation of INHBB depended on the transcription factor Sox9 in the injured TECs. Clinical analyses also identified a positive correlation between Sox9 and INHBB expression in human specimens, suggesting the Sox9/INHBB axis as a positive regulator of renal fibrosis. In conclusion, tubule-derived INHBB is implicated in the pathogenesis of renal fibrosis by activating the surrounding fibroblasts in a paracrine manner, thereby exhibiting as a potential therapeutic target. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

A key gene in Drosophila eye development provides protection for tubules and glomeruli upon injury

Dachshund homolog 1 (DACH1) is highly expressed in both the developing kidney, where it has a role in kidney development, and in the adult kidney, where we know very little about its function. Recently, 2 groups independently reported that DACH1 protects podocytes1 and tubules2 from experimental injury and provided promising evidence that their findings may be relevant to human chronic kidney disease.

Reduced urinary release of AQP1- and AQP2-bearing extracellular vesicles in patients with advanced chronic kidney disease.

Although several studies have shown that release of water channel proteins, aquaporin 1 (AQP1) and AQP2 in urinary extracellular vesicles (uEV‐AQP1 and ‐AQP2), were altered in experimental kidney injury models, their release in human chronic kidney disease (CKD) has been largely unexplored. The aim of the present study was to clarify whether the release of uEV‐AQP1 and ‐AQP2 is altered in patients with CKD. Urine samples were collected from 15 healthy volunteers (normal group) and 62 CKD patients who were categorized into six glomerular filtration rate (GFR) categories (G1, G2, G3a, G3b, G4, and G5) in between 2005 and 2016 at Miyazaki Prefectural Miyazaki Hospital, Japan. uEV‐proteins were evaluated by immunoblot analysis. The release of AQP1 and AQP2 were significantly decreased in patients with both CKD G4 and G5, in comparison with the normal group. The area under the receiver operating characteristic (ROC) curve (AUC) values for AQP1 and AQP2 in patients with CKD G4 and G5 were 0.926 and 0.881, respectively. On the other hand, the AUC values in patients with CKD G1‐G3 were 0.512 for AQP1 and 0.680 for AQP2. Multiple logistic regression analysis showed that AQP1 and AQP2 in combination were useful for detecting CKD G4 and G5, with a higher AUC value of 0.945. These results suggest that the release of uEV‐AQP1 and ‐AQP2 was decreased in patients with CKD G4 and G5, and these proteins might be helpful to detect advanced CKD.

Association Between PAHs and Chronic Kidney Disease in the US Adult Population

BACKGROUND AND AIM: Polycyclic aromatic hydrocarbons (PAHs) are a group of chemicals produced from the incomplete combustion of carbon-containing products. Exposure to PAHs is linked to respiratory disease, heart disease, liver disease, and cancer. Chronic kidney disease (CKD) is irreversible, progressive, and linked with higher cardiovascular risk. Very few studies have investigated the association between exposure to PAHs and CKD in humans. The rationale of this study was to assess seven types of urinary PAHs (1-hydroxynaphthalene, 2-hydroxynaphthalene, 3-hydroxyfluorene, 2-hydroxyfluorene, 1-hydroxyphenanthrene, 1-hydroxypyrene, and 2 & 3-hydroxyphenanthrene) concentrations and the correlation with CKD in adult US population. METHODS: A cross-sectional analyses using the 2015-2016 National Health and Nutrition Examination Survey (NHANES) dataset was conducted. A specialized complex survey design analysis package was used in analyzing the NHANES data. Chronic kidney disease was defined as estimated glomerular filtration rate (eGFR) 60 ml/min/1.73m2 or albumin to creatinine ratio (ACR) 30 mg/gm. Prior to analysis, the urinary PAH concentrations were normalized by the urinary creatinine concentration to account for dilution effects. Multivariate Logistic Regression Analysis of Complex Survey models were used to study the correlation between urinary PAH concentrations (seven types) and kidney disease associated with abnormal eGFR or ACR. Models were adjusted for demographic and lifestyle factors. RESULTS:The total study population analyzed included 4,117 adults aged 20 years and older, of whom 49.6% were males and 50.4% females. The analysis showed urinary 2-Hydroxynaphthalene [Odds Ratio (OR): 1.60, 95% Confidence Interval (CI): 1.41, 2.24] was significantly associated with increased odds of chronic kidney diseases. The other six types of urinary PAHs were not associated with CKD. CONCLUSIONS:The study concluded that urinary 2-Hydroxynaphthalene form of PAHs is significantly associated with CKD. KEYWORDS: polycyclic aromatic hydrocarbons, chronic kidney disease, urinary 2-Hydroxynaphthalene, NHANES, environmental pollution

A generic crystallopathic model for chronic kidney disease progression.

Chronic kidney disease (CKD) has reached epidemic proportions globally. The natural course of chronic kidney disease is almost uniformly progressive, albeit at different rates in different individuals. The downhill course appears to pervade kidney diseases of all etiologies and seems to spiral down a self-perpetuating vortex, even if the original insult is ameliorated or controlled. In this issue of the JCI, Shiizaki, Tsubouchi, and colleagues proposed a model of renal tubule luminal calcium phosphate crystallopathy that accounts for renal function demise. Calcium phosphate crystals attached to TLR4 and underwent endocytosis at the brush border, triggering inflammation and fibrosis. This mechanism might operate in different kinds of kidney disease, with a theoretical phosphate concentration threshold in the proximal tubular lumen, beyond which is triggered undesirable downstream effects that eventuate in loss of renal function. If this model parallels human CKD, clinicians may focus efforts on determining phosphate exposure in the proximal tubular lumen.

Reduced Expression of Metallothionein-I/II in Renal Proximal Tubules Is Associated with Advanced Chronic Kidney Disease.

Chronic kidney disease (CKD) is a commonly occurring complex renal syndrome that causes overall mortality in many diseases. The clinical manifestations of CKD include renal tubulointerstitial fibrosis and loss of renal function. Metallothionein-I/II (MT-I/II) is potentially expressed in the liver and kidney, and possesses antioxidant and metal detoxification properties. However, whether MT-I/II expression is associated with the prognosis of nephropathy remains unknown. In this study, we investigated the MT-I/II level in human CKD, using immunohistochemistry. MT-I/II is located on the proximal tubules and is notably reduced in patients with CKD. MT-I/II expression was significantly correlated with the functional and histological grades of CKD. In an aristolochic acid (AAI)-induced nephropathy mouse model, MT-I/II was abundantly increased after AAI injection for 7 days, but decreased subsequently compared to that induced in the acute phase when injected with AAI for 28 days. Furthermore, we found that ammonium pyrrolidinedithiocarbamate (PDTC) restored AAI-induced MT-I/II reduction in HK2 cells. The injection of PDTC ameliorated AAI-induced renal tubulointerstitial fibrosis and reduced the concentrations of blood urea nitrogen and creatinine in mouse sera. Taken together, our results indicate that MT-I/II reduction is associated with advanced CKD, and the retention of renal MT-I/II is a potential therapeutic strategy for CKD.

Long-term B cell depletion associates with regeneration of kidney function.

BackgroundChronic kidney disease (CKD) is a common condition that increases mortality and the risk of cardiovascular and other morbidities regardless of underlying renal condition. Chronic inflammation promotes renal fibrosis. Recently, renal B cell infiltrates were described in chronic kidney disease of various etiologies beyond autoimmunity.MethodsWe here investigated B cells and indicators of tertiary lymphoid structure formation in human renal biopsies. Renal function was studied during long‐term B cell depletion in human patients with membranous nephropathy and with CKD of unknown origin.ResultsCytokine profiles of tertiary lymphoid structure formation were detected in human renal interstitium in a range of kidney diseases. Complex B cell structures consistent with tertiary lymphoid organ formation were evident in human membranous nephropathy. Here, B cell density did not significantly associate with proteinuria severity, but with worse excretory renal function. Proteinuria responses mostly occurred within the first 6 months of B cell depletion. In contrast, recovery of excretory kidney function was observed only after 18 months of continuous therapy, consistent with a structural process. Renal tertiary lymphatic structures were also detected in the absence of autoimmune kidney disease. To start to address whether B cell depletion may affect CKD in a broader population, we assessed kidney function in neurologic patients with CKD of unknown origin. In this cohort, eGFR significantly increased within 24 months of B cell depletion.ConclusionLong‐term B cell depletion associated with significant improvement of excretory kidney function in human CKD. Kinetics and mechanisms of renal B cell aggregation should be investigated further to stratify the impact of B cells and their aggregates as therapeutic targets.

Hepatic cysteine sulphinic acid decarboxylase depletion and defective taurine metabolism in a rat partial nephrectomy model of chronic kidney disease

BackgroundTaurine depletion occurs in patients with end-stage chronic kidney disease (CKD). In contrast, in the absence of CKD, plasma taurine is reported to increase following dietary L-glutamine supplementation. This study tested the hypothesis that taurine biosynthesis decreases in a rat CKD model, but is rectified by L-glutamine supplementation.MethodsCKD was induced by partial nephrectomy in male Sprague-Dawley rats, followed 2 weeks later by 2 weeks of 12% w/w L-glutamine supplemented diet (designated NxT) or control diet (NxC). Sham-operated control rats (S) received control diet.ResultsTaurine concentration in plasma, liver and skeletal muscle was not depleted, but steady-state urinary taurine excretion (a measure of whole-body taurine biosynthesis) was strongly suppressed (28.3 ± 8.7 in NxC rats versus 78.5 ± 7.6 μmol/24 h in S, P < 0.05), accompanied by reduced taurine clearance (NxC 0.14 ± 0.05 versus 0.70 ± 0.11 ml/min/Kg body weight in S, P < 0.05). Hepatic expression of mRNAs encoding key enzymes of taurine biosynthesis (cysteine sulphinic acid decarboxylase (CSAD) and cysteine dioxygenase (CDO)) showed no statistically significant response to CKD (mean relative expression of CSAD and CDO in NxC versus S was 0.91 ± 0.18 and 0.87 ± 0.14 respectively). Expression of CDO protein was also unaffected. However, CSAD protein decreased strongly in NxC livers (45.0 ± 16.8% of that in S livers, P < 0.005). L-glutamine supplementation failed to rectify taurine biosynthesis or CSAD protein expression, but worsened CKD (proteinuria in NxT 12.5 ± 1.2 versus 6.7 ± 1.5 mg/24 h in NxC, P < 0.05).ConclusionIn CKD, hepatic CSAD is depleted and taurine biosynthesis impaired. This is important in view of taurine’s reported protective effect against cardio-vascular disease - the leading cause of death in human CKD.

Pro-cachectic factors link experimental and human chronic kidney disease to skeletal muscle wasting programs.

Skeletal muscle wasting is commonly associated with chronic kidney disease (CKD), resulting in increased morbidity and mortality. However, the link between kidney and muscle function remains poorly understood. Here, we took a complementary interorgan approach to investigate skeletal muscle wasting in CKD. We identified increased production and elevated blood levels of soluble pro-cachectic factors, including activin A, directly linking experimental and human CKD to skeletal muscle wasting programs. Single-cell sequencing data identified the expression of activin A in specific kidney cell populations of fibroblasts and cells of the juxtaglomerular apparatus. We propose that persistent and increased kidney production of pro-cachectic factors, combined with a lack of kidney clearance, facilitates a vicious kidney/muscle signaling cycle, leading to exacerbated blood accumulation and, thereby, skeletal muscle wasting. Systemic pharmacological blockade of activin A using soluble activin receptor type IIB ligand trap as well as muscle-specific adeno-associated virus-mediated downregulation of its receptor ACVR2A/B prevented muscle wasting in different mouse models of experimental CKD, suggesting that activin A is a key factor in CKD-induced cachexia. In summary, we uncovered a crosstalk between kidney and muscle and propose modulation of activin signaling as a potential therapeutic strategy for skeletal muscle wasting in CKD.