A multi-trait GWAS identifies novel genes influencing albuminuria.

BACKGROUND: Albuminuria is common and associated with increased risks of end-stage kidney disease and cardiovascular diseases, yet its underlying mechanism remains obscure. Previous genome-wide association studies (GWAS) for albuminuria did not consider gene pleiotropy and primarily focused on European ancestry populations. This study adopted a multi-trait analysis of GWAS (MTAG) approach to jointly analyze two vital kidney traits, estimated glomerular filtration rate (eGFR) and urine albumin-to-creatinine ratio (UACR) to identify and prioritize the genes associated with UACR. METHODS: Data from the Taiwan Biobank from 2012 to 2023 were analyzed. GWAS of UACR and eGFR were performed separately and the summary statistics from these GWAS were jointly analyzed using MTAG. The polygenic risk scores (PRS) of UACR were constructed for validation. The UACR-associated loci were further fine-mapped and prioritized based on their deleteriousness, eQTL associations, and relatedness to Mendelian kidney diseases. RESULTS: MTAG analysis of the UACR revealed 15 genetic loci, including 12 novel loci. The PRS for UACR was significantly associated with urinary albumin level (P < 0.001) and microalbuminuria (P = 0.001 ∼ 0.045). A list of priority genes was generated. Twelve genes with high priority included the albumin endocytic receptor gene LRP2 and ciliary genes  IFT172. CONCLUSIONS: The findings of this multi-trait GWAS suggest that primary cilia play a role in sensing mechanical stimuli, leading to albumin endocytosis. The priority list of genes warrants further translational investigation to reduce albuminuria.

Update of pericytes function and their roles in kidney diseases.

Studies have highlighted the significant involvement of kidney pericytes in renal fibrosis. Kidney pericytes, classified as interstitial mesenchymal cells, are extensively branched, collagen-producing cells that closely interact with endothelial cells. This article aims to provide an overview of the recent advancements in understanding the physiological functions of pericytes and their roles in kidney diseases. In a healthy kidney, pericytes have essential physiological function in angiogenesis, erythropoietin (EPO) production, and the regulation of renal blood flow. Nevertheless, pericyte-myofibroblast transition has been identified as the primary cause of disease progression in acute kidney injury (AKI)-to-chronic kidney disease (CKD) continuum. Our recent research has demonstrated that hypoxia-inducible factor-2α (HIF-2α) regulates erythropoietin production in pericytes. However, this production is repressed by EPO gene hypermethylation and HIF-2α downregulation which were induced by transforming growth factor-ß1-activated DNA methyltransferase and activin receptor-like kinase-5 signaling pathway during renal fibrosis, respectively. Additionally, AKI induces epigenetic modifications in pericytes, rendering them more prone to extracellular matrix production, cell migration and proliferation, thereby contributing to subsequent capillary rarefaction and renal fibrosis. Further investigation into the specific functions and roles of different subpopulations of pericytes may contribute for the development of targeted therapies aimed at attenuating kidney disease and mitigating their adverse effects.

Humeral and cellular immune responses to SARS-CoV-2 vaccination in patients on peritoneal dialysis.

BACKGROUND: Patients with chronic kidney disease are at high risk for coronavirus disease 2019. Little is known about immune response to severe acute respiratory syndrome coronavirus 2 vaccination in patients on peritoneal dialysis (PD). METHOD: We prospectively enrolled 306 PD patients receiving two doses of vaccines (ChAdOx1-S: 283, mRNA-1273: 23) from July 2021 at a medical center. Humeral and cellular immune responses were assessed by anti-spike IgG concentration and blood T cell interferon-γ production 30 days after vaccination. Antibody ≥0.8 U/mL and interferon-γ ≥ 100 mIU/mL were defined as positive. Antibody was also measured in 604 non-dialysis volunteers (ChAdOx1-S: 244, mRNA-1273: 360) for comparison. RESULT: PD patients had less adverse events after vaccinations than volunteers. After the first dose of vaccine, the median antibody concentrations were 8.5 U/mL and 50.4 U/mL in ChAdOx1-S group and mRNA-1273 group of PD patients, and 66.6 U/mL and 195.3 U/mL in ChAdOx1-S group and mRNA-1273 group of volunteers, respectively. And after the second dose of vaccine, the median antibody concentrations were 344.8 U/mL and 9941.0 U/mL in ChAdOx1-S group and mRNA-1273 group of PD patients, and 620.3 U/mL and 3845.0 U/mL in ChAdOx1-S group and mRNA-1273 group of volunteers, respectively. The median IFN-γ concentration was 182.8 mIU/mL in ChAdOx1-S group, which was substantially lower than the median concentration 476.8 mIU/mL in mRNA-1273 group of PD patients. CONCLUSION: Both vaccines were safe and resulted in comparable antibody seroconversion in PD patients when compared with volunteers. However, mRNA-1273 vaccine induced significantly higher antibody and T cell response than ChAdOx1-S in PD patients. Booster doses are recommended for PD patients after two doses of ChAdOx1-S vaccination.

Targeting ER protein TXNDC5 in hepatic stellate cell mitigates liver fibrosis by repressing non-canonical TGFß signalling.

BACKGROUND AND OBJECTIVES: Liver fibrosis (LF) occurs following chronic liver injuries. Currently, there is no effective therapy for LF. Recently, we identified thioredoxin domain containing 5 (TXNDC5), an ER protein disulfide isomerase (PDI), as a critical mediator of cardiac and lung fibrosis. We aimed to determine if TXNDC5 also contributes to LF and its potential as a therapeutic target for LF. DESIGN: Histological and transcriptome analyses on human cirrhotic livers were performed. Col1a1-GFPTg , Alb-Cre;Rosa26-tdTomato and Tie2-Cre/ERT2;Rosa26-tdTomato mice were used to determine the cell type(s) where TXNDC5 was induced following liver injury. In vitro investigations were conducted in human hepatic stellate cells (HSCs). Col1a2-Cre/ERT2;Txndc5fl/fl (Txndc5cKO ) and Alb-Cre;Txndc5fl/fl (Txndc5Hep-cKO ) mice were generated to delete TXNDC5 in HSCs and hepatocytes, respectively. Carbon tetrachloride treatment and bile duct ligation surgery were employed to induce liver injury/fibrosis in mice. The extent of LF was quantified using histological, imaging and biochemical analyses. RESULTS: TXNDC5 was upregulated markedly in human and mouse fibrotic livers, particularly in activated HSC at the fibrotic foci. TXNDC5 was induced by transforming growth factor ß1 (TGFß1) in HSCs and it was both required and sufficient for the activation, proliferation, survival and extracellular matrix production of HSC. Mechanistically, TGFß1 induces TXNDC5 expression through increased ER stress and ATF6-mediated transcriptional regulation. In addition, TXNDC5 promotes LF by redox-dependent JNK and signal transducer and activator of transcription 3 activation in HSCs through its PDI activity, activating HSCs and making them resistant to apoptosis. HSC-specific deletion of Txndc5 reverted established LF in mice. CONCLUSIONS: ER protein TXNDC5 promotes LF through redox-dependent HSC activation, proliferation and excessive extracellular matrix production. Targeting TXNDC5, therefore, could be a potential novel therapeutic strategy to ameliorate LF.

Angiopoietin-2 inhibition attenuates kidney fibrosis by hindering chemokine C-C motif ligand 2 expression and apoptosis of endothelial cells.

Plasma levels of angiopoietin-2 are increased in patients with chronic kidney disease (CKD). Moreover, mouse models of progressive kidney disease also demonstrate increased angiopoietin-2 in both plasmas and kidneys. The role of dysregulated angiopoietins in the progression of kidney disease has not been thoroughly investigated. Here, we found in a cohort of 319 patients with CKD that plasma angiopoietin-2 and angiopoietin-2/angiopoietin-1 ratios were positively associated with the development of kidney failure. In mice with progressive kidney disease induced by either ureteral obstruction or ischemia-reperfusion injury, overexpression of human angiopoietin-1 in the kidney tubules not only reduced macrophage infiltration in the initial stage post-injury but also attenuated endothelial cell apoptosis, microvascular rarefaction, and fibrosis in the advanced disease stage. Notably, angiopoietin-1 attenuated chemokine C-C motif ligand 2 (CCL2) expression in the endothelial cells of the fibrosing kidneys, and these protective effects led to attenuation of functional impairment. Mechanistically, angiopoietin-1 reduced CCL2-activated macrophage migration and protected endothelial cells against cell apoptosis induced by angiopoietin-2 and Wnt ligands. Based on this, we applied L1-10, an angiopoietin-2 inhibitor, to the mouse models of progressive kidney disease and found inhibitory effects on macrophage infiltration, microvascular rarefaction, and fibrosis. Thus, we defined the detrimental impact of increased angiopoietin-2 on kidney survival of patients with CKD which appears highlighted by angiopoietin-2 induced endothelial CCL2-activated macrophage infiltration and endothelial cell apoptosis in their kidneys undergoing fibrosis.

Kidney pericyte hypoxia-inducible factor regulates erythropoiesis but not kidney fibrosis.

Prolyl hydroxylase domain enzyme (PHD) inhibitors are effective in the treatment of chronic kidney disease (CKD)-associated anemia by stabilizing hypoxia inducible factor (HIF), thereby increasing erythropoietin and consequently erythropoiesis. However, concern for CKD progression needs to be addressed in clinical trials. Although pre-clinical studies showed an anti-inflammatory effect in kidney disease models, the effect of PHD inhibitors on kidney fibrosis was inconsistent probably because the effects of HIF are cell type and context dependent. The major kidney erythropoietin-producing cells are pericytes that produce erythropoietin through HIF-2α-dependent gene transcription. The concern for the impact of HIF in pericytes on kidney fibrosis arises from the fact that pericytes are the major precursor cells of myofibroblasts in CKD. Since cells expressing Gli1 fulfill the morphologic and anatomic criteria for pericytes, we induced Gli1+ cell-specific HIF stabilization or knockout to study the impact of HIF in pericytes on kidney pathology of mice with or without fibrotic injury induced by unilateral ureteral obstruction. Compared with the littermate controls, mice with pericyte-specific HIF stabilization due to von Hippel-Lindau protein or PHD2 knockout showed increased serum erythropoietin and polycythemia rather than a discernible difference in kidney fibrosis. Compared with Gli1+ pericytes sorted from littermate controls, Gli1+ pericytes sorted from PHD2 knockout mice showed increased erythropoietin gene expression rather than discernible changes in Col1a1 or Acta2 expression. Furthermore, pericyte-specific knockout of HIF-1α or HIF-2α did not affect kidney fibrosis. Thus, our study supports the absence of negative effects of PHD inhibitors on kidney fibrosis of mice despite HIF stabilization in pericytes.

Transforming growth factor-ß1 decreases erythropoietin production through repressing hypoxia-inducible factor 2α in erythropoietin-producing cells.

BACKGROUND: Renal erythropoietin (EPO)-producing (REP) cells produce EPO through hypoxia-inducible factor (HIF) 2α-activated gene transcription. Insufficient EPO production leads to anemia in patients with chronic kidney disease. Although recombinant EPO is effective to improve anemia, no reliable REP cell lines limit further progress of research and development of novel treatment. METHODS: We screened Epo mRNA expression in mouse fibroblast cell lines. Small interfering RNA specific for HIF1α or HIF2α was transfected to study Epo expression in C3H10T1/2 cells. The effect of transforming growth factor-ß1 (TGF-ß1) on HIF-EPO axis was studied in C3H10T1/2 cells and mice. RESULTS: Similar to mouse REP pericytes, C3H10T1/2 cells differentiated to α-smooth muscle actin+ myofibroblasts after exposure to TGF-ß1. Specific HIF knockdown demonstrated the role of HIF2α in hypoxia-induced Epo expression. Sustained TGF-ß1 exposure increased neither DNA methyltransferase nor methylation of Epas1 and Epo genes. However, TGF-ß1 repressed HIF2α-encoding Epas1 promptly through activating activin receptor-like kinase-5 (ALK5), thereby decreasing Epo induction by hypoxia and prolyl hydroxylase domain inhibitor roxadustat. In mice with pro-fibrotic injury induced by ureteral obstruction, upregulation of Tgfb1 was accompanied with downregulation of Epas1 and Epo in injured kidneys and myofibroblasts, which were reversed by ALK5 inhibitor SB431542. CONCLUSION: C3H10T1/2 cells possessed the property of HIF2α-dependent Epo expression in REP pericytes. TGF-ß1 induced not only the transition to myofibroblasts but also a repressive effect on Epas1-Epo axis in C3H10T1/2 cells. The repressive effect of TGF-ß1 on Epas1-Epo axis was confirmed in REP pericytes in vivo. Inhibition of TGF-ß1-ALK5 signaling might provide a novel treatment to rescue EPO expression in REP pericytes of injured kidney.

Inflammatory macrophages switch to CCL17-expressing phenotype and promote peritoneal fibrosis.

Peritoneal fibrosis remains a problem in kidney failure patients treated with peritoneal dialysis. Severe peritoneal fibrosis with encapsulation or encapsulating peritoneal sclerosis is devastating and life-threatening. Although submesothelial fibroblasts as the major precursor of scar-producing myofibroblasts in animal models and M2 macrophage (Mϕ)-derived chemokines in peritoneal effluents of patients before diagnosis of encapsulating peritoneal sclerosis have been identified, attenuation of peritoneal fibrosis is an unmet medical need partly because the mechanism for cross talk between Mϕs and fibroblasts remains unclear. We use a sodium hypochlorite-induced mouse model akin to clinical encapsulated peritoneal sclerosis to study how the peritoneal Mϕs activate fibroblasts and fibrosis. Sodium hypochlorite induces the disappearance of CD11bhigh F4/80high resident Mϕs but accumulation of CD11bint F4/80int inflammatory Mϕs (InfMϕs) through recruiting blood monocytes and activating local cell proliferation. InfMϕs switch to express chemokine (C-C motif) ligand 17 (CCL17), CCL22, and arginase-1 from day 2 after hypochlorite injury. More than 75% of InfMϕs undergo genetic recombination by Csf1r-driven Cre recombinase, providing the possibility to reduce myofibroblasts and fibrosis by diphtheria toxin-induced Mϕ ablation from day 2 after injury. Furthermore, administration of antibody against CCL17 can reduce Mϕs, myofibroblasts, fibrosis, and improve peritoneal function after injury. Mechanistically, CCL17 stimulates migration and collagen production of submesothelial fibroblasts in culture. By breeding mice that are induced to express red fluorescent protein in Mϕs and green fluorescence protein (GFP) in Col1a1-expressing cells, we confirmed that Mϕs do not produce collagen in peritoneum before and after injury. However, small numbers of fibrocytes are found in fibrotic peritoneum of chimeric mice with bone marrow from Col1a1-GFP reporter mice, but they do not contribute to myofibroblasts. These data demonstrate that InfMϕs switch to pro-fibrotic phenotype and activate peritoneal fibroblasts through CCL17 after injury. CCL17 blockade in patients with peritoneal fibrosis may provide a novel therapy. © 2019 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Endoplasmic Reticulum Protein TXNDC5 Augments Myocardial Fibrosis by Facilitating Extracellular Matrix Protein Folding and Redox-Sensitive Cardiac Fibroblast Activation.

RATIONALE: Cardiac fibrosis plays a critical role in the pathogenesis of heart failure. Excessive accumulation of extracellular matrix (ECM) resulting from cardiac fibrosis impairs cardiac contractile function and increases arrhythmogenicity. Current treatment options for cardiac fibrosis, however, are limited, and there is a clear need to identify novel mediators of cardiac fibrosis to facilitate the development of better therapeutics. Exploiting coexpression gene network analysis on RNA sequencing data from failing human heart, we identified TXNDC5 (thioredoxin domain containing 5), a cardiac fibroblast (CF)-enriched endoplasmic reticulum protein, as a potential novel mediator of cardiac fibrosis, and we completed experiments to test this hypothesis directly. OBJECTIVE: The objective of this study was to determine the functional role of TXNDC5 in the pathogenesis of cardiac fibrosis. METHODS AND RESULTS: RNA sequencing and Western blot analyses revealed that TXNDC5 mRNA and protein were highly upregulated in failing human left ventricles and in hypertrophied/failing mouse left ventricle. In addition, cardiac TXNDC5 mRNA expression levels were positively correlated with those of transcripts encoding transforming growth factor ß1 and ECM proteins in vivo. TXNDC5 mRNA and protein were increased in human CF (hCF) under transforming growth factor ß1 stimulation in vitro. Knockdown of TXNDC5 attenuated transforming growth factor ß1-induced hCF activation and ECM protein upregulation independent of SMAD3 (SMAD family member 3), whereas increasing expression of TXNDC5 triggered hCF activation and proliferation and increased ECM protein production. Further experiments showed that TXNDC5, a protein disulfide isomerase, facilitated ECM protein folding and that depletion of TXNDC5 led to ECM protein misfolding and degradation in CF. In addition, TXNDC5 promotes hCF activation and proliferation by enhancing c-Jun N-terminal kinase activity via increased reactive oxygen species, derived from NAD(P)H oxidase 4. Transforming growth factor ß1-induced TXNDC5 upregulation in hCF was dependent on endoplasmic reticulum stress and activating transcription factor 6-mediated transcriptional control. Targeted disruption of Txndc5 in mice (Txndc5-/-) revealed protective effects against isoproterenol-induced cardiac hypertrophy, reduced fibrosis (by ≈70%), and markedly improved left ventricle function; post-isoproterenol left ventricular ejection fraction was 59.1±1.5 versus 40.1±2.5 (P<0.001) in Txndc5-/- versus wild-type mice, respectively. CONCLUSIONS: The endoplasmic reticulum protein TXNDC5 promotes cardiac fibrosis by facilitating ECM protein folding and CF activation via redox-sensitive c-Jun N-terminal kinase signaling. Loss of TXNDC5 protects against ß agonist-induced cardiac fibrosis and contractile dysfunction. Targeting TXNDC5, therefore, could be a powerful new therapeutic approach to mitigate excessive cardiac fibrosis, thereby improving cardiac function and outcomes in patients with heart failure.

Rejuvenation: Turning back the clock of aging kidney.

Aging is inevitable in life. It is defined as impaired adaptive capacity to environmental or internal stresses with growing rates of disease and death. Aging is also an important risk factor for various kidney diseases such as acute kidney injury and chronic kidney disease. Patients older than 65 years have nearly 28% risk of failing recovery of kidney function when suffering from acute kidney injury. It is reported that more than a third of population aged 65 years and older have chronic kidney disease in Taiwan, and the occurrence of multiple age-related disorders is predicted to increase in parallel. Renal aging is a complex, multifactorial process characterized by many anatomical and functional changes. Several factors are involved in renal aging, such as loss of telomeres, cell cycle arrest, chronic inflammation, activation of renin-angiotensin system, decreased klotho expression, and development of tertiary lymphoid tissues. These changes can also be observed in many other different types of renal injury. Recent studies suggested that young blood may rejuvenate aged organs, including the kidneys. In order to develop new therapeutic strategies for renal aging, the mechanisms underlying renal aging and by which young blood can halt or reverse aging process warrants further study.

Angiopoietin 1 influences ischemic reperfusion renal injury via modulating endothelium survival and regeneration.

BACKGROUND: Damage to the endothelium due to ischemia reperfusion injury (IRI) leads to a disruption of the microvasculature, which could be influenced by angiopoietin 1 via its effects on endothelium. We investigated the physiological and therapeutic roles of angiopoietin 1 in renal IRI using angiopoietin 1 knockout and over-expression mice. METHODS: Renal IRI was induced by clamping the right renal artery seven days after left uninephrectomy for 25 min followed by reperfusion. A whole body angiopoietin 1 knockout was achieved by induction with tamoxifen. The renal tubule over-expression of angiopoietin 1 was induced by doxycycline. RESULTS: In the normal mice, the renal expression of angiopoietin 1 increased 7 days to 14 days after IRI. The angiopoietin 1 knockout caused a delay in the recovery of renal function, less tubular regeneration and more residual tubular necrosis. The endothelial density was lower and the VE-cadherin protein loss was greater in the knockout mice. The over-expression of angiopoietin 1 attenuated the tubular necrosis and renal function impairment 1 and 3 days after IRI. The loss of the endothelium was ameliorated in the over-expression mice. This protective effect was associated with the up-regulation of the gene expression of epidermal growth factor, hepatocyte growth factor, and insulin like growth factor-1 and less tubular apoptosis. The over-expression of angiopoietin 1 stimulated tumor necrosis factor-α, C-C chemokine receptor type 2 and CX3C chemokine receptor 1 inflammatory gene expression, but did not influence macrophage infiltration. CONCLUSIONS: Altogether, the augmentation and downregulation of angiopoietin 1 attenuated renal damage and impaired renal recovery, respectively, by influencing the survival/regeneration of the endothelium. The manipulation of angiopoietin 1 represents a novel therapeutic approach for the treatment of ischemic kidney injury.

Heart rate variability as a predictor of rapid renal function deterioration in chronic kidney disease patients.

AIM: Autonomic dysfunction contributes to cardiovascular morbidity/mortality and can be evaluated with heart rate variability (HRV). This study is to evaluate the prognostic significance of HRV on renal function in non-dialysis chronic kidney disease (CKD) patients. METHODS: We enrolled 326 non-dialysis CKD patients in this prospective observational study. The median follow-up period was 2.02 years. Five-minutes of electrocardiography recordings obtained at enrolment were reprocessed to assess HRV. Five frequency-domain measures and one time-domain measures were obtained. Rapid CKD progression was defined as annual estimated glomerular filtration rate (eGFR) loss over 30% per year or eGFR decline rate over 3 mL/min per 1.73 m2 per year. The prevalence of abnormal HRV, associated factors of HRV and impact of HRV on the risk of CKD progression were analyzed. RESULTS: The abnormality of HRV increased along with the severity of CKD. In patients with stage 5 CKD, the proportion of abnormal ln(low frequency power) (LF), ln(high frequency power) (HF), lnLF/HF were 69.5, 52.8 and 50%, respectively. Associated factors of HRV included advanced CKD, diabetes mellitus, serum albumin, severe proteinuria, Beck Anxiety Inventory score, erythropoietin use, renin-angiotensin system inhibitors and heart failure. Multivariate logistic regression model analysis revealed lower lnLF/HF, hypertension and severe proteinuria were the risk factors of rapid CKD progression. CONCLUSION: The prevalence of autonomic dysfunction measured by HRV among each stage CKD patients is different. Most patients in advanced CKD stage have reduced values of HRV parameters. The estimation of lnLF/HF also provided prognostic information on CKD progression in addition to classical risk factors.

Emergency department utilization and resuscitation rate among patients receiving maintenance hemodialysis.

BACKGROUND: End-stage renal disease (ESRD) is a growing global health concern with increased disease burden and high medical costs. Utilization of the emergency department (ED) among dialyzed patients and the associated risk factors remain unknown. METHODS: Participants of this study, selected from the National Health Insurance Database in Taiwan, were aged 19-90 years and received maintenance hemodialysis from January 1, 2010, to December 31, 2010. A control group consisting of individuals who did not receive dialysis, selected from the same data source, were matched for age, sex, and the Charlson Comorbidity Index (CCI). Subgroup analysis with hemodialysis frequency was also performed. ED utilization among enrolled individuals was assessed in 2012. Generalized estimating equations with multiple variable adjustments were used to identify risk factors associated with resuscitation during ED visits. RESULTS: One group of 2985 individuals who received maintenance hemodialysis, and another group of 2985 patients that did not receive hemodialysis, between January 1, 2010, and December 31, 2010, were included in this study. There were 4822 ED visits in the hemodialysis group, and 1755 ED visits in the non-dialysis group between January 1, 2012, and December 31, 2012. Analysis of multivariable generalized estimating equations identified the risk associated with resuscitation during ED visits to be greater in individuals who were receiving maintenance hemodialysis, aged older than 55 years, hospitalized in the past year, and assigned first and second degree of triage. CONCLUSION: Patients receiving maintenance hemodialysis had higher ED utilization and a significantly higher risk of resuscitation during ED visits than those without hemodialysis.

Erythropoietin modulates macrophages but not post-ischemic acute kidney injury in mice.

BACKGROUND/PURPOSE: Substantial progress was made in acute kidney injury (AKI) over the past 10 years, but no therapeutic interventions have been shown to prevent AKI or accelerate functional recovery after injury. A large number of preclinical studies supports the use of recombinant human erythropoietin (rHuEPO) to prevent AKI, but the clinical trial data are inconclusive. To address concerns about preclinical study design and reporting in AKI, we here presented our rigorous experiments on the use of rHuEPO in a mouse model simulating the most common post-ischemic AKI in patients. METHODS: Use of saline vehicle or rHuEPO (100 or 1000 U/KgBW) in mice subjected to AKI induced by ischemia-reperfusion injury of left kidney 2 weeks after right nephrectomy (NX + IRI). RESULTS: NX + IRI resulted in a reproducible AKI model. Use of rHuEPO as a pretreatment or posttreatment did not affect AKI severity, functional recovery, and mouse survival regardless of gender, injury severity, or doses of rHuEPO. Administering rHuEPO with 1000 U/KgBW did increase hematocrit and modulate AKI kidney macrophages by Nos2 downregulation and Ccl17 upregulation. Active expression of erythropoietin receptor (EPOR) was not identified in renal cells by lineage tracing study, whereas expression of colony-stimulating factor 2 receptor ß (CSF2Rß) was identified in kidney macrophages and upregulated after AKI. Both EPOR and CSF2Rß were identified in cultured bone marrow derived macrophages, possibly mediated the robust inhibition of cytokine-induced phenotype switching by rHuEPO. CONCLUSION: Use of rHuEPO can modulate macrophage function but not the post-ischemic AKI severity, functional recovery and survival in mice.

Role of renin-angiotensin system in acute kidney injury-chronic kidney disease transition.

Acute kidney injury (AKI) can increase the risk of developing incident chronic kidney disease (CKD). The severity, frequency and duration of AKI are crucial predictors of poor renal outcome. A repair process after AKI can be adaptive and kidney recovers completely after a mild injury. However, severe injury will lead to a maladaptive repair, which frequently progresses to nephron loss, vascular rarefaction, chronic inflammation and fibrosis. Although different mechanisms underlying AKI-CKD transition have been extensively discussed, no definite intervention has been proved effective to block or to retard the transition until recently. In CKD, renin-angiotensin system (RAS) inhibitor has been proved effective to slow down disease progression. Furthermore, RAS needs to be highlighted again in AKI-CKD transition because recent animal studies have shown the activation of intra-renal RAS after AKI, and RAS blockade can reduce the ensuing CKD and mortality. In patients with the complete renal recovery after AKI, administration of RAS inhibitor is associated with reduced risk of subsequent CKD as well. In this article, we will demonstrate the role of RAS in AKI-CKD transition comprehensively. We will then emphasize the promising effect of RAS inhibitor on CKD prevention in patients recovering from AKI based on evidence from the bench to clinical research. All of these discussions will contribute to the establishment of reliable monitoring and therapeutic strategies for patients with functional recovery from AKI who can be most easily ignored.

Physiology and pathophysiology of renal erythropoietin-producing cells.

Anemia is a common complication and contributes to increased morbidity and mortality in chronic kidney disease (CKD) patients. Whereas there has been a significant improvement of understanding the underlying mechanism of erythropoiesis, the treatment of renal anemia is still restricted to erythropoietin (EPO)-stimulating agents. The purpose of this article is to review the physiology of erythropoiesis, functional role of EPO and underlying molecular and cellular basis that regulate EPO production. Regulation of EPO production is at mRNA level. When anemia or hypoxia occurs, transcriptional factor, hypoxia-inducible factor (HIF), binds to EPO 5' hypoxic response element and EPO gene transcription increases. The renal EPO is mainly produced by pericytes. In CKD, pericytes transdifferentiate to myofibroblasts, and subsequently the ability of EPO production decreases, leading to renal anemia. Recent experimental and clinical studies show the promising efficacy of prolyl hydroxylase inhibitors in renal anemia through increasing EPO production by stabilizing HIF. Recent advances on epigenetics create a new field to study EPO gene expression at chromatin level. We will discuss the role of demethylating agent on restoring EPO expression, providing a novel approach to the treatment of renal anemia.

Therapeutic efficacy of pentoxifylline on proteinuria and renal progression: an update.

Blood pressure control with renin-angiotensin system (RAS) blockade has remained the gold standard for treating patients with proteinuric chronic kidney disease (CKD) up to date. Nevertheless, RAS blockade slows but does not halt the progression of kidney disease, thus highlighting the need to search for additional therapeutic approaches. The nonselective phosphodiesterase (PDE) inhibitor pentoxifylline (PTX) is an old drug that exhibits prominent anti-inflammatory, anti-proliferative and anti-fibrotic activities both in vitro and in vivo. Studies in human subjects have shown that PTX monotherapy decreases urinary protein excretion, and add-on therapy of PTX to background RAS blockade additively reduces proteinuria in patients with CKD of various etiology. More recent studies find that PTX combined with RAS blockade delays the decline of glomerular filtration rate in diabetic patients with mild to moderate CKD, and reduces the risk of end-stage renal disease in diabetic and non-diabetic patients in late stage of CKD with high proteinuria levels. In this review, we update the clinical trial results of PTX as monotherapy, or in conjunction or in comparison with RAS blockade on patients with proteinuria and CKD, and propose a mechanistic scheme explaining the renoprotective activities of this drug.

The C-terminal disulfide bonds of Helicobacter pylori GroES are critical for IL-8 secretion via the TLR4-dependent pathway in gastric epithelial cells.

Helicobacter pylori GroES (HpGroES), a potent immunogen, is a secreted virulence factor that stimulates production of proinflammatory cytokines and may contribute to gastric carcinogenesis. HpGroES is larger than other bacterial orthologs because of an additional C-terminal region, known as domain B. We found that the HpGroES-induced IL-8 release by human gastric epithelial cells was dependent on activation of the MAPK and NF-κB pathways. HpGroES lacking domain B was unable to induce IL-8 release. Additionally, a TLR4 inhibitor significantly inhibited IL-8 secretion and reduced HpGroES-induced activation of MAPKs. Furthermore, HpGroES-induced IL-8 release by primary gastric epithelial cells from TLR4(-/-) mice was significantly lower than from wild-type mice. We also found that HpGroES bound to TLR4 in cell lysates and colocalized with TLR4 on the cell membrane only when domain B was present. We then constructed two deletion mutants lacking C-terminal regions and mutants with point mutations of two of the four cysteine residues, C111 and C112, in domain B and found that the deletion mutants and a double mutant lacking the C94-C111 and C95-C112 disulfide bonds were unable to interact with TLR4 or induce IL-8 release. We conclude that HpGroES, in which a unique conformational structure, domain B, is generated by these two disulfide bonds, induces IL-8 secretion via a TLR4-dependent mechanism.

Circadian CLOCK Mediates Activation of Transforming Growth Factor-ß Signaling and Renal Fibrosis through Cyclooxygenase 2.

The circadian rhythm regulates blood pressure and maintains fluid and electrolyte homeostasis with central and peripheral clock. However, the role of circadian rhythm in the pathogenesis of tubulointerstitial fibrosis remains unclear. Here, we found that the amplitudes of circadian rhythm oscillation in kidneys significantly increased after unilateral ureteral obstruction. In mice that are deficient in the circadian gene Clock, renal fibrosis and renal parenchymal damage were significantly worse after ureteral obstruction. CLOCK-deficient mice showed increased synthesis of collagen, increased oxidative stress, and greater transforming growth factor-ß (TGF-ß) expression. TGF-ß mRNA expression oscillated with the circadian rhythms under the control of CLOCK-BMAL1 heterodimers. The expression of cyclooxygenase 2 was significantly higher in kidneys from CLOCK-deficient mice with ureteral obstruction. Treatment with a cyclooxygenase 2 inhibitor celecoxib significantly improved renal fibrosis in CLOCK-deficient mice. Taken together, these data establish the importance of the circadian rhythm in tubulointerstitial fibrosis and suggest CLOCK/TGF-ß signaling as a novel therapeutic target of cyclooxygenase inhibition.