Efficacy and safety of Yiqi Peiyuan granules for improving the short-term prognosis of patients with acute kidney injury: A multicenter, double-blind, placebo-controlled, randomized trial.

BACKGROUND: Yiqi Peiyuan (YQPY) prescription, a composite prescription of traditional Chinese medicine, has been used to prevent or delay the continued deterioration of renal function after acute kidney injury (AKI) in some institutions and has shown considerable efficacy. OBJECTIVE: This is the first randomized controlled trial to assess efficacy and safety of YQPY for improving short-term prognosis in adult patients with AKI. DESIGN, SETTING, PARTICIPANTS AND INTERVENTIONS: This is a prospective, double-blind, multicenter, randomized, and placebo-controlled clinical trial. A total of 144 enrolled participants were randomly allocated to two groups according to a randomization schedule. Participants, caregivers and investigators assessing the outcomes were blinded to group assignment. Patients in the YQPY group received 36 g YQPY granules twice a day for 28 days. Patients in the placebo group received a placebo in the same dose as the YQPY granules. MAIN OUTCOME MEASURES: The primary outcome was the change in the estimated glomerular filtration rate (eGFR) between baseline and after 4 and 24 weeks of treatment. The secondary outcomes were the change of serum creatinine (Scr) level between baseline and after treatment, and the incidence of endpoint events, defined as eGFR increasing by more than 25% above baseline, eGFR >75 mL/min per 1.73 m2 or the composite endpoint, which was defined as the sum of patients meeting either of the above criteria. RESULTS: Data from a total of 114 patients (59 in the YQPY group and 55 in the control group) were analyzed. The mean changes in eGFR and Scr in weeks 4 and 24 had no difference between the two groups. In further subgroup analysis (22 in the YQPY group and 31 in the control group), the mean change in eGFR after treatment for 4 weeks was 27.39 mL/min per 1.73 m2 in the YQPY group and 5.78 mL/min per 1.73 m2 in the placebo group, and the mean difference between groups was 21.61 mL/min per 1.73 m2 (P < 0.001). Thirteen (59.1%) patients in the YQPY group and 5 (16.1%) in the placebo group reached the composite endpoints (P = 0.002). During the intervention, 2 and 4 severe adverse events were reported in the YQPY and placebo groups, respectively. CONCLUSION: The YQPY granules can effectively improve the renal function of patients 4 weeks after the onset of AKI, indicating that it has good efficacy for improving short-term renal outcomes in patients with AKI. The YQPY granules may be a promising therapy for adults with AKI. TRIAL REGISTRATION: Chinese Clinical Trial Registry ChiCTR2100051723. Please cite this article as: Wu JJ, Zhang TY, Qi YH, Zhu MY, Fang Y, Qi CJ, Cao LO, Lu JF, Lu BH, Tang LM, Shen JX, Mou S. Efficacy and safety of Yiqi Peiyuan granules for improving the short-term prognosis of patients with acute kidney injury: a multicenter, double-blind, placebo-controlled, randomized trial. J Integr Med. 2024; 22(3): 279-285.

Activation of lipophagy is required for RAB7 to regulate ferroptosis in sepsis-induced acute kidney injury.

Sepsis-induced acute kidney injury (S-AKI) is the most common type of acute kidney injury (AKI), accompanied by elevated morbidity and mortality rates. This study investigated the mechanism by which lipid droplets (LDs) degraded via autophagy (lipophagy)required for RAB7 regulated ferroptosis in the pathogenesis of S-AKI. Here, we constructed the S-AKI model in vitro and in vivo to elucidate the potential relationship of lipophagy and ferroptosis, and we first confirmed that the activation of lipophagy promoted renal tubular epithelial cell ferroptosis and renal damage in S-AKI. The results showed that lipopolysaccharide (LPS) induced a marked increase in lipid peroxidation and ferroptosis, which were rescued by ferrstain-1 (Fer-1), an inhibitor of ferroptosis. In addition, LPS induced the remarkable activation of RAB7-mediated lipophagy. Importantly, silencing RAB7 alleviated LPS-induced lipid peroxidation and ferroptosis. Thus, the present study demonstrated the potential significant role of ferroptosis and lipophagy in sepsis-induced AKI, and contributed to better understanding of the pathogenesis and treatment targets of AKI.

Integrative ATAC-seq and RNA-seq analysis associated with diabetic nephropathy and identification of novel targets for treatment by dapagliflozin.

Dapagliflozin (DAPA) are clinically effective in improving diabetic nephropathy (DN). However, whether and how chromatin accessibility changed by DN responds to DAPA treatment is unclear. Therefore, we performed ATAC-seq, RNA-seq, and weighted gene correlation network analysis to identify the chromatin accessibility, the messenger RNA (mRNA) expression, and the correlation between clinical phenotypes and mRNA expression using kidney from three mouse groups: db/m mice (Controls), db/db mice (case group), and those treated with DAPA (treatment group). RNA-Seq and ATAC-seq conjoint analysis revealed many overlapping pathways and networks suggesting that the transcriptional changes of DN and DAPA intervention largely occured dependently on chromatin remodeling. Specifically, the results showed that some key signal transduction pathways, such as immune dysfunction, glucolipid metabolism, oxidative stress and xenobiotic and endobiotic metabolism, were repeatedly enriched in the analysis of the RNA-seq data alone, as well as combined analysis with ATAC-seq data. Furthermore, we identified some candidate genes (UDP glucuronosyltransferase 1 family, Dock2, Tbc1d10c, etc.) and transcriptional regulators (KLF6 and GFI1) that might be associated with DN and DAPA restoration. These reversed genes and regulators confirmed that pathways related to immune response and metabolism pathways were critically involved in DN progression.

The presence of exudative thickening of Bowman's capsule predict poor prognosis in diabetic kidney disease.

BACKGROUND: The relationship between Bowman's capsule thickening and progression of diabetic kidney disease (DKD) remains uncertain. METHODS: Renal biopsy specimens from 145 DKD patients and 20 control subjects were evaluated for Bowman's capsule thickness. Immunohistochemical staining assessed col4α2, laminin ß1, and albumin expression. In a discovery cohort of 111 DKD patients with eGFR ≥ 30 ml/min/1.73 m2, thickening was classified as fibrotic or exudative. The composite endpoint included CKD stage 5, dialysis initiation, and renal disease-related death. Prognosis was analyzed using Kaplan-Meier and Cox regression analyses. Two validation cohorts were included. RESULTS: Three types of thickening were observed: fibrotic, exudative, and periglomerular fibrosis. Parietal epithelial cell matrix protein accumulation contributed to fibrotic thickening, while albumin was present in exudative thickening. Bowman's capsule was significantly thicker in DKD patients (5.74 ± 2.09 µm) compared to controls (3.38 ± 0.43 µm, P < 0.01). In discovery cohort, the group of exudative thickning had a poorer prognosis(median time 20 months vs 57 months, P = 0.000). Cox multivariate analysis revealed that exudative thickening of Bowman's capsule were associated with a poor prognosis. The validation cohorts confirmed the result. CONCLUSIONS: Various mechanisms contribute to Bowman's capsule thickening in DKD. The proportion of exudative thickening may serve as a valuable prognostic indicator for DKD patients.

DDRGK1-mediated ER-phagy attenuates acute kidney injury through ER-stress and apoptosis.

Acute kidney injury (AKI) constitutes a prevalent clinical syndrome characterized by elevated morbidity and mortality rates, emerging as a significant public health issue. This study investigates the interplay between endoplasmic reticulum (ER) stress, unfolded protein response (UPR), and ER-associated degradation (ER-phagy) in the pathogenesis of AKI. We employed four distinct murine models of AKI-induced by contrast media, ischemia-reperfusion injury, cisplatin, and folic acid-to elucidate the relationship between ER-phagy, ER stress, and apoptosis. Our findings reveal a marked decrease in ER-phagy coinciding with an accumulation of damaged ER, elevated ER stress, and increased apoptosis across all AKI models. Importantly, overexpression of DDRGK1 in HK-2 cells enhanced ER-phagy levels, ameliorating contrast-induced ER stress and apoptosis. These findings unveil a novel protective mechanism in AKI, wherein DDRGK1-UFL1-mediated ER-phagy mitigates ER stress and apoptosis in renal tubular epithelial cells. Our results thereby contribute to understanding the molecular underpinnings of AKI and offer potential therapeutic targets for its treatment.

Tubular basement membrane deposits after allogeneic hematopoietic stem cell transplantation.

BACKGROUND: Extraglomerular immune complex deposition is rare and only a few membranous nephropathy cases with tubular basement membrane deposits have been reported following allogeneic hematopoietic stem cell transplantation. CASE PRESENTATION: We reported a 56-year-old man with increased serum creatinine after allogeneic hematopoietic stem cell transplantation who underwent a renal biopsy. Tubular interstitial nephritis was identified on light microscope. The unique histologic features were diffuse tubular basement membrane immune complex deposition detected by both immunofluorescence and electron microscopy, while the glomerular involvement was inconspicuous. The differential diagnosis from other forms of tubular basement membrane deposition is discussed. CONCLUSION: Diffuse granular tubular basement membrane immune complex deposition with minimal glomerular involvement is also a manifestation of renal complication in hematopoietic stem cell transplantation recipient. However, the exact mechanism and target antigen remains unknown.

Mitophagy alleviates cisplatin-induced renal tubular epithelial cell ferroptosis through ROS/HO-1/GPX4 axis.

Cisplatin is widely recommended in combination for the treatment of tumors, thus inevitably increasing the incidence of cisplatin-induced acute kidney injury. Mitophagy is a type of mitochondrial quality control mechanism that degrades damaged mitochondria and maintains cellular homeostasis. Ferroptosis, a new modality of programmed cell death, is characterized by iron-dependent phospholipid peroxidation and oxidative membrane damage. However, the role of mitophagy in ferroptosis in kidney disease is unclear. Here, we investigated the mechanism underlying both BNIP3-mediated and PINK1-PARK2-mediated mitophagy-induced attenuation of ferroptosis in cisplatin-induced acute kidney injury. The results showed that cisplatin induced mitochondrial injury, ROS release, intracellular iron accumulation, lipid peroxidation and ferroptosis in the kidney, which were aggravated in Bnip3 knockout, Pink1 knockout or Park2 knockout cisplatin-treated mice. Ferrstatin-1, a synthetic antioxidative ferroptosis inhibitor, rescued iron accumulation, lipid peroxidation and ferroptosis caused by inhibition of mitophagy. Thus, the present study elucidated a novel mechanism by which both BNIP3-mediated and PINK1-PARK2-mediated mitophagy protects against cisplatin-induced renal tubular epithelial cell ferroptosis through the ROS/HO1/GPX4 axis.

Renal histopathological lesions after liver transplantation: What can we find besides calcineurin inhibitor-induced nephrotoxicity?

BACKGROUND: Chronic kidney disease (CKD) is a common complication after liver transplantation and is traditionally considered to be secondary to calcineurin inhibitors (CNIs). However, several studies have reported that the etiology of CKD after liver transplantation is broad and may only be assessed accurately by renal biopsy. The current study aimed to explore the usefulness of renal biopsies in managing CKD after liver transplantation in daily clinical practice. METHOD: This retrospective analysis enrolled all post-liver transplantation patients who had a renal biopsy in a single center from July 2018 to February 2021. RESULTS: Fourteen renal biopsies were retrieved for review from 14 patients at a median of 35.7 (minimum-maximum: 2.80-134.73) months following liver transplantation. The male-to-female ratio was 13:1 (age range, 31-75 years). The histomorphological alterations were varied. The predominant glomerular histomorphological changes included focal segmental glomerular sclerosis (FSGS) (n = 4), diabetic glomerulopathy (n = 4), and membranoproliferative glomerulonephritis (n = 4). Thirteen (92.9%) patients had renal arteriolar sclerosis. Immune complex nephritis was present in six patients, of whom only two had abnormal serum immunological indicators. Despite interstitial fibrosis and tubular atrophy being present in all the patients, only six (42.9%) presented with severe interstitial injury. No major renal biopsy-related complications occurred. After a mean follow-up of 11.8 months (range: 1.2-29.8), three patients progressed to end-stage renal disease (ESRD). CONCLUSION: The etiology of CKD after liver transplantation might be more complex than originally thought and should not be diagnosed simply as calcineurin inhibitors(CNI)-related nephropathy. Renal biopsy plays a potentially important role in the diagnosis and treatment of CKD after liver transplantation and might not be fully substituted by urine or blood tests. It may help avoid unnecessary changes to the immunosuppressants and inadequate treatment of primary diseases.

Downregulation of PPARα mediates FABP1 expression, contributing to IgA nephropathy by stimulating ferroptosis in human mesangial cells.

Immunoglobulin A nephropathy (IgAN) is the commonest primary glomerulonephritis, and a major cause of end-stage renal disease; however, its pathogenesis requires elucidation. Here, a hub gene, FABP1, and signaling pathway, PPARα, were selected as key in IgAN pathogenesis by combined weighted gene correlation network analysis of clinical traits and identification of differentially expressed genes from three datasets. FABP1 and PPARα levels were lower in IgAN than control kidney, and linearly positively correlated with one another, while FABP1 levels were negatively correlated with urinary albumin-to-creatinine ratio, and GPX4 levels were significantly decreased in IgAN. In human mesangial cells (HMCs), PPARα and FABP1 levels were significantly decreased after Gd-IgA1 stimulation and mitochondria appeared structurally damaged, while reactive oxygen species (ROS) and malondialdehyde (MDA) were significantly increased, and glutathione and GPX4 decreased, relative to controls. GPX4 levels were decreased, and those of ACSL4 increased on siPPARα and siFABP1 siRNA treatment. In PPARα lentivirus-transfected HMCs stimulated by Gd-IgA1, ROS, MDA, and ACSL4 were decreased; glutathione and GPX4, and immunofluorescence colocalization of PPARα and GPX4, increased; and damaged mitochondria reduced. Hence, PPARα pathway downregulation can reduce FABP1 expression, affecting GPX4 and ACSL4 levels, causing HMC ferroptosis, and contributing to IgAN pathogenesis.

C4d as a Screening Tool and an Independent Predictor of Clinical Outcomes in Lupus Nephritis and IgA Nephropathy.

BACKGROUND: As an indispensable marker of complement cascades activation, C4d was confirmed of its crucial role in the pathogenesis of both lupus nephritis (LN) and IgA nephropathy (IgAN). While the studies directly comparing the diagnostic value, and outcomes predicting function of C4d between LN and IgAN are still absent. METHODS: A cohort of 120 LN patients, 120 IgAN patients who were diagnosed by renal biopsy between January 2015 and December 2017 and 24 healthy age matched controls were prospectively analyzed. The patients were followed till December 2020. The outcomes were adverse disease treatment response (disease relapse) and kidney disease progression event (decline of estimated glomerular filtration rate by more than 20% or end-stage kidney disease). The renal C4d deposition proportion and pattern were compared between IgAN and LN patients. In addition, the relationship between renal C4d deposition and disease subtypes, disease relapse as well as disease progression for LN and IgAN patients were also analyzed. RESULTS: The LN, IgAN patients and healthy controls were well matched in ages. The follow-up period was 38.5 (30.3-60.8) months for LN patients and 45.0 (30.5-57.0) months for IgAN patients. 78 patients (65.0%) with LN had renal C4d deposition, compared with only 39 IgAN patients (32.5%) with C4d deposition in renal tissues (P < 0.001). The LN patients shared different renal C4d distribution patterns with IgAN patients. Compared with IgAN patients, the C4d deposition in LN patients was significantly more in renal glomerulus (P < 0.001) and less in renal tubules (P = 0.003). For disease subtypes, renal C4d deposition was especially strong in class V membranous LN and IgAN with tubulointerstitial fibrosis (T1/T2) lesions. Renal C4d deposition was independently correlated with the disease relapse of LN patients (HR = 1.007, P = 0.040), and acted as an independent predictor of disease progression during the follow-up period for IgAN patients (HR = 1.821, P = 0.040). CONCLUSIONS: Renal C4d distribution proportion and pattern differed between LN and IgAN patients. The presence of C4d in renal tissue acted as an independent predictor of relapse for LN patients and disease progression for IgAN patients.

Downregulation of Renal Hsa-miR-127-3p Contributes to the Overactivation of Type I Interferon Signaling Pathway in the Kidney of Lupus Nephritis.

MicroRNAs are involved in the pathogenesis of systemic lupus erythematosus (SLE) and dysregulated in the kidneys of lupus nephritis (LN) patients, but their pathogenic roles in LN are largely unknown. Janus Kinase 1 (JAK1) mediates the activation of the downstream signaling pathways of many inflammatory cytokines, including type I interferon (IFN-I) signaling pathway which is critical to the development of SLE and LN. Thus, JAK1 is a potent therapeutic target for these autoimmune diseases. However, there are few studies demonstrating the dysregulation of JAK1 in autoimmune diseases and the molecular mechanism behind it. In this concise report, we show an inhibitory effect of hsa-miR-127-3p, a microRNA that is downregulated in the renal tissues of LN patients, on IFN-I signaling. We found the overexpression of hsa-miR-127-3p could inhibit the induction of ISRE and GAS mediated gene expression, the phosphorylation of STAT1 and STAT2, and the upregulation of IFN stimulated genes (ISGs) stimulated by IFN-I. While, hsa-miR-127-3p antagonist enhanced the activation of IFN-I signaling in primary renal mesangial cells. Subsequently, we identified JAK1 as a bona fide target gene of hsa-miR-127-3p and showed hsa-miR-127-3p targeted JAK1 through binding to its 3'UTR and coding region. Consistently, we found the downregulation of hsa-miR-127-3p was associated with the upregulation of JAK1 and ISGs in kidney tissues of LN patients. Our data indicate renal downregulation of hsa-miR-127-3p contributes to the overactivation of IFN-I signaling pathway in the kidneys of LN patients through promoting the expression of JAK1, suggesting hsa-miR-127-3p mimics may be used to inhibit JAK1 and IFN-I signaling pathway in LN.

Inhibition of microRNA-130b Alleviated Podocyte Injury Induced by Puromycin Aminonucleoside in vitro.

OBJECTIVE: To investigate the effect of microRNA-130b (miR-130b) on podocyte injury induced by puromycin aminonucleoside (PAN) and its possible mechanisms. METHODS: The immortalized podocytes (HPC) were treated by 25, 50, or 100 µg/mL PAN, then real-time polymerase chain reaction (PCR) was used to detect the expression of miR-130b. The HPC were transfected with miR-130b inhibitor or normal control (NC) inhibitor, and then the cells were stimulated with 100 µg/mL PAN for 24h. Western blot was used to detect the protein expression of synaptopodin and nephrin. Phalloidin dying was used to detect the changes in the cytoskeleton. Flow cytometry was used to measure podocyte apoptosis. Luciferase reporter gene assays were performed to explore the interaction between miR-130b and PGC1α. RESULTS: PAN significantly upregulated the expression of miR-130b. The western blot showed that inhibition of miR-130b increased the protein expression of synaptopodin and nephrin compared to the negative control inhibitor group. The phalloidin dying showed that inhibition of miR-130b alleviated cytoskeletal remodeling of podocytes induced by PAN. Flow-cytometric analysis showed that apoptosis was decreased after miR-130b silencing. The miR-130b mimic could significantly down-regulate the protein expression of PGC1α, and the dual luciferase reporter assay showed that miR-130b induced a decrease in PGC1α 3'-UTR luciferase activity compared to the control mimic group, but there was no significant difference between the control mimic group and the mut·PGC1α 3'-UTR group. CONCLUSION: miR-130b ameliorates podocyte injury induced by PAN through inhibiting the expression of PGC1α.

EZH2 Inhibition Interferes With the Activation of Type I Interferon Signaling Pathway and Ameliorates Lupus Nephritis in NZB/NZW F1 Mice.

Enhancer of zeste homolog 2 (EZH2) is a histone-lysine N-methyltransferase mediating trimethylation of H3K27, which represses gene expression and is critical to immune regulation. Inhibition of EZH2 is proved to have the potential of treating many diseases. However, whether inhibition of EZH2 affects type I interferon (IFN-I) signaling pathway, the abnormality of which is an important pathogenic mechanism for SLE, is still elusive. Here, we report, unexpectedly, a positive regulatory function of EZH2 in IFN-I signaling pathway, which contributes to the overactivation of IFN-I signaling pathway in SLE. We show that the expression of EZH2 was upregulated and positively correlated with the overexpression of interferon stimulated genes (ISGs) in both peripheral blood mononuclear cells and renal tissues of SLE patients. In vitro inhibition of EZH2 by either siRNAs or chemical inhibitors reduced the phosphorylation of STAT1 and the induction of ISGs stimulated by IFN-I. Additionally, inhibition of EZH2 interfered with the in vivo and ex vivo activation of IFN-I signaling pathway elicited by intravenous injection of adenovirus vector expressing mouse IFN-α5 and exogeneous stimulation with IFN-α, respectively. We evaluated the therapeutic effects of EZH2 inhibitor in NZB/NZW F1 mice which depend on IFN-I signaling pathway for the lupus-like disease development. Administration of EZH2 inhibitor prolonged the survival, reduced the levels of anti-dsDNA autoantibodies, and improved lupus nephritis of the mice. What's more, EZH2 inhibitor attenuated the expression of ISGs in the kidneys of these mice. In summary, we show that excessive EZH2 contributes to the overactivation of IFN-I signaling pathway in SLE. EZH2 inhibitor has the potential to inhibit IFN-I signaling pathway and alleviate lupus nephritis. Additionally, diverse disease driving pathways exist among systemic lupus erythematosus (SLE) patient, and even in the same patients. Common regulators of different pathogenic pathways can be multivalent therapeutic targets. Together with previous studies showing EZH2 is involved in T-cell and B-cell mediated immune responses, EZH2 could be a potent multivalent therapeutic target for SLE.

The key candidate genes in tubulointerstitial injury of chronic kidney diseases patients as determined by bioinformatic analysis.

Pathologic changes such as renal tubular atrophy and interstitial fibrosis are common in chronic kidney disease (CKD), which in turn, leads to loss of renal function. The aims of present study were to screen critical genes with tubulointerstitial lesion in CKD by weighted gene correlation network analysis (WGCNA). Gene expression data including 169 tubulointerstitial samples of CKD and 21 controls downloaded from Gene Expression Omnibus (GEO) database. Totally 294 differentially expressed genes (DEGs) were screened, including 180 upregulated and 114 downregulated genes. Meanwhile, 90 expression data of tubulointerstitial samples combined with clinic information were applied to explore the potential mechanisms of tubulointerstitial lesion. As a consequence, the blue, brown and yellow modules which included the most DEGs compared to the other modules and exhibited strongly association with eGFR, were significantly enriched in several signalling pathways that have been reported involved in pathogenesis of CKD. Furthermore, it was found that the four genes (PLG, ITGB2, CTSS and CCL5) was one of the DEGs which also be identified as hub genes according to Kwithin. Finally, the Nephroseq online tool showed that the tubulointerstitial expression levels of PLG significantly positively correlated with the estimated glomerular filtration rate (eGFR), while ITGB2, CTSS and CCL5 connected negatively to the eGFR of CKD patients. Taken together, WGCNA is an efficient approach to system biology. By this procedure, the present study enhanced the understanding of the transcriptome status of CKD and might shed a light on the further investigation on the mechanisms of renal tubulointerstitial injury in CKD. SIGNIFICANCE OF STUDY: Traditional molecular biology can only explain the local part of the biological system, and difficult to make comprehensive exploration of the whole biological system in the chronic kidney disease (CKD). In this study, we gave an explicit elucidation of dysregulated protein coding genes by the analysis of microarray datasets in GEO database. We have presented a novel approach using weighted gene correlation network analysis (WGCNA) to explore the DEGs which implicated in CKD process. In this study, we conducted WGCNA to explore the potential mechanisms of renal tubular damage, and provided novel biomarkers associated with the molecular mechanisms underlying renal tubulointerstitial injury in CKD.

Drp1-regulated PARK2-dependent mitophagy protects against renal fibrosis in unilateral ureteral obstruction.

Mitophagy is a principle mechanism to degrade damaged mitochondria through PARK2-dependent or PARK2-independent pathway. Mitophagy has been identified to play an important role in acute kidney disease, whereas its role in renal fibrosis remains ill-defined. We sought to investigate the involvement and regulation of mitophagy in renal tubular epithelial cell(RTEC) injury and renal fibrosis after unilateral ureteral obstruction(UUO). Mitochondrial damageand mitochondrial reactive oxygen species (ROS) production was increased in kidney after obstruction of the left ureter. Mitophagy was increased in kidneys following UUO and HK-2 cells under hypoxia exposure, assessed by electron microscopy of mitophagosome, colocalization of MitotrackerRed-stained mitochondria and LC3 staining. The upregulation of PINK1, PARK2, and LC3 II in mitochondrial fraction was observed in the obstructed kidney and hypoxia-exposed HK-2 cells. Pink1 or Park2 gene deletion markedly increased mtROS production, mitochondrial damage, TGFß1 expression in RTEC, and renal fibrosis in UUO. Mitochondrial recruitment of Drp1 was also induced after UUO. The Drp1 inhibitor, Mdivi-1, decreased mitochondrial PINK1, PARK2 and LC3II level, increased mtROS production both in vivo and in vitro, activated TGFß1-Smad2/3 signaling in HK-2 cells under hypoxia and worsened renal fibrosis following UUO. The upregulation of TGFß1 signaling in hypoxia-treated HK-2 cells due to PINK1 or PARK2 silencing, or worsened renal fibrosis after UUO due to Pink1-or Park2-KO mice was rescued by mitoTEMPO, a mitochondria-targeted antioxidant. The findings of this study suggest that Drp1-regulated PARK2-dependent mitophagy plays a critical role in hypoxia-induced renal tubular epithelial cell injury and renal fibrosis in UUO.

Investigation of the Mechanism Underlying Calcium Dobesilate-Mediated Improvement of Endothelial Dysfunction and Inflammation Caused by High Glucose.

BACKGROUND/AIMS: Diabetic kidney disease (DKD) is a leading cause of end-stage renal disease. Calcium dobesilate (CaD) is widely used to treat diabetic retinopathy. Recent studies have demonstrated that CaD exerts protective effects against diabetic nephropathy. The aim of this study was to elucidate the molecular and cellular mechanisms underlying the protective effects of CaD. METHODS: Human umbilical vein endothelial cells (HUVECs) were cultured with different D-glucose concentrations to determine the effects of high glucose on HUVEC gene expression. HUVECs were also incubated with CaD (25 µM, 50 µM, and 100 µM) for 3 days to determine the effects of CaD on HUVEC viability. db/db mice were treated with CaD. 2-[(Aminocarbonyl)amino]-5-(4-fluorophenyl)-3-thiophenecarboxamide (TPCA-1) blocked the nuclear factor-κB (NF-κB) pathway in HUVECs. A pentraxin 3 (PTX3) small interfering RNA (siRNA) intervention experiment was performed in the cells. An adenovirus-encapsulated PTX3 siRNA intervention experiment was performed in db/db mice. Western blot and real-time PCR analyses were used to detect PTX3, p-IKBa/IKBa (I-kappa-B-alpha), and p-eNOS/eNOS (endothelial nitric oxide synthase) expression in mice and HUVECs. Hematoxylin-eosin (HE) staining and periodic acid-Schiff (PAS) staining were used to observe renal tissue damage in mice. PTX3 expression was observed by immunohistochemical staining. RESULTS: CaD downregulated the expression of PTX3 and p-IKBa/IKBa and upregulated the expression of p-eNOS/eNOS in vitro. When TPCA-1 was used, high glucose induced high PTX3 expression, and the expression of p-eNOS/eNOS increased. After PTX3 gene silencing, the expression of p-eNOS/eNOS also increased. In vivo, CaD reduced the expression of PTX3 and p-IKBa/IKBa in the kidneys of db/db mice and increased the expression of p-eNOS/eNOS. After PTX3 gene silencing, the urine protein and renal function of db/db mice were ameliorated, the glomerular extracellular matrix was decreased, and the expression of p-eNOS/eNOS was increased. CONCLUSIONS: Our results suggested that CaD may inhibit the expression of PTX3 by altering the IKK/IKB/NF-κB pathway, thereby improving endothelial dysfunction in HUVECs. PTX3 may be a potential therapeutic target for DKD.

NLRP3 inflammasome inhibition attenuates cisplatin-induced renal fibrosis by decreasing oxidative stress and inflammation.

BACKGROUND/AIMS: The NOD-like receptor, pyrin domain containing-3 (NLRP3) inflammasome is involved in the progression of chronic kidney disease in several rodent models. Here, we investigated whether a specific inhibitor of NLRP3 inflammasome, MCC950, can attenuate cisplatin-induced renal fibrosis. MATERIALS: Renal fibrosis was induced via a series of three injections of cisplatin to male C57BL/6 mice (7.5 mg/kg body weight). Activation of NLRP3 inflammasome was detected by immunoblotting, real-time PCR, and immunofluorescence. To validate the protective effect of NLRP3 inflammasome inhibition, MCC950(20 mg/kg body weight) was daily injected into multiple-cisplatin-treated mice intraperitoneally for 14 days, starting from 4 weeks after the first dose of cisplatin. NLRP3-/- mice were used to confirm the role of NLRP3 inflammasome in cisplatin-induced renal fibrosis. RESULTS: Mice were euthanized at 6 weeks after the first dose of cisplatin treatment. In multiple-cisplatin-induced murine model, renal fibrosis was accompanied by the activation of NLRP3 inflammasome. MCC950, the specific inhibitor of NLRP3 inflammasome, reduced cisplatin-induced renal dysfunction, tubular damage, interstitial collagen deposit, and the expression of profibrotic parameters. NLRP3 inhibition might protect against cisplatin-induced renal fibrosis through the alleviation of oxidative stress and inflammation. Furthermore, inhibition of NLRP3 inflammasome activation by deleting NLRP3 gene halted the progression of cisplatin-induced renal fibrosis. CONCLUSION: Inhibition of NLRP3 inflammasome attenuates renal fibrosis due to repeated cisplatin injections, and might be identified as a potential target for attenuating cisplatin-induced chronic kidney disease.

Leflunomide versus cyclophosphamide in the induction treatment of proliferative lupus nephritis in Chinese patients: a randomized trial.

OBJECTIVES: A prospective, multi-center, randomized controlled study was conducted to evaluate the efficacy and safety of a 24-week course low-dose leflunomide combined with prednisone in the induction treatment of proliferative lupus nephritis in Chinese patients. METHOD: Patients (n = 100) with biopsy-proved proliferative lupus nephritis were enrolled in this study. They were randomized into two groups and received either leflunomide or cyclophosphamide in conjunction with prednisone for 24 weeks. Leflunomide was given orally with a loading dose of 40 mg/day for 3 days followed by 20 mg/day. Intravenous cyclophosphamide was administered monthly at a dosage of 0.8-1.0 g. The primary efficacy outcome was the frequency of complete remission and partial remission at week 24. The secondary outcomes included changes of urinary protein excretion, serum albumin, complement 3, anti-dsDNA antibody level, and systemic lupus erythematosus disease activity index (SLEDAI) after 24-week therapy. RESULTS: Of 100 patients, 48 received leflunomide combined with prednisone and other 52 received cyclophosphamide with concomitant prednisone. There were no statistically significant differences between groups in complete remission rate and partial remission rate. At week 24, 23% of patients in the leflunomide group and 27% of patients in the cyclophosphamide group achieved complete remission (P = 0.64), while 56% of patients in the leflunomide group and 42% of patients in the cyclophosphamide group achieved partial remission at week 24 (P = 0.16). SLEDAI, serum albumin, complement 3, anti-dsDNA antibody level, and urinary protein excretion improved significantly in both groups. No significant difference was seen in the changes of clinical parameters after therapy between the two groups. There was no significant difference in side effects in both groups. CONCLUSIONS: Compared with cyclophosphamide, low-dose leflunomide in combination with prednisone showed both effectiveness and safety in the induction therapy of proliferative lupus nephritis in Chinese patients.

Diabetic Nephropathy Can Be Treated with Calcium Dobesilate by Alleviating the Chronic Inflammatory State and Improving Endothelial Cell Function.

BACKGROUND/AIMS: Diabetic kidney disease (DKD) is a leading cause of end-stage renal disease. However, no effective treatments for this disease are available. Calcium dobesilate (CaD) is widely used to treat diabetic retinopathy. DKD and retinopathy often co-exist and have similar mechanisms of pathogenesis. The aim of the present study was to elucidate the safety and efficacy of CaD in the treatment of DKD. METHODS: In the prospective randomised controlled study, 100 DKD from Type 2 diabetes mellitus (DM) patients with a urinary albumin/ creatinine ratio (ACR) ≥30 mg/g and urinary protein level between 150 mg/24 h and 2 g/24 h with GFR> 90ml/min were enrolled. The patients were randomly divided into the treatment group (500 mg of CaD, administered orally, 3 times per day) and the control group. DKD patients were treated for 3 months. In the case control study, DM patients without proteinuria and healthy individuals were also enrolled. Clinical data and related biochemical parameters were collected. Endothelial function markers (VEGF, ET-1, eNOS, NO) and inflammatory markers (MCP-1, ICAM, PTX3) were detected by ELISA. RESULTS: In the prospective randomised controlled study, the 24 h urinary albumin and 24 h urinary protein levels significantly decreased after three months of treatment with CaD in the patients with DKD, but the cystatin C-based glomerular filtration rate (GFR) remained unchanged. In addition, the levels of inflammatory markers (PTX3, MCP-1, hsCRP, ICAM) and endothelial dysfunction markers (VEGF, ET-1) were significantly reduced compared to pre-treatment levels, NO was signifcantly increased post treatment. In the case control study, we found that PTX3, MCP-1, ICAM, VEGF and ET-1 levels were positively correlated with urinary albumin in DKD patients, while the NO level was negatively correlated. Logistic regression analysis showed that PTX3, NO and HbAlc were influential factors in DKD. After patients with DKD were treated with CaD for three months, the 24 h urinary albumin and 24 h urinary protein levels significantly decreased, but the cystatin C-based glomerular filtration rate (GFR) remained unchanged. In addition, the levels of inflammatory markers (PTX3, MCP-1, hsCRP, ICAM) and endothelial dysfunction markers (VEGF, NO, ET-1) were significantly reduced compared to pre-treatment levels. CONCLUSION: CaD can be safely and effectively used to treatdiabetic nephropathy.

Calcium dobesilate may alleviate diabetes­induced endothelial dysfunction and inflammation.

Diabetic kidney disease (DKD) is a leading cause of end­stage renal disease. However, the pathogenesis of DKD remains unclear, and no effective treatments for the disease are available. Thus, there is an urgent need to elucidate the pathogenic mechanisms of DKD and to develop more effective therapies for this disease. Human umbilical vein endothelial cells (HUVECs) were cultured using different D­glucose concentrations to determine the effect of high glucose (HG) on the cells. Alternatively, HUVECs were incubated with 100 µmol/l calcium dobesilate (CaD) to detect its effects. The authors subsequently measured HUVEC proliferation via cell counting kit­8 assays. In addition, HUVEC angiogenesis was investigated via migration assays and fluorescein isothiocyanate (FITC)­labelled bovine serum albumin (BSA) permeability assays. The content or distribution of markers of endothelial dysfunction [vascular endothelial growth factor (VEGF), VEGF receptor (R) and endocan) or inflammation [intercellular adhesion molecule (ICAM)­1, monocyte chemotactic protein (MCP)­1 and pentraxin­related protein (PTX3)] was evaluated via reverse transcription­quantitative polymerase chain reaction and western blotting. HG treatment induced increased in VEGF, VEGFR, endocan, ICAM­1, MCP­1 and PTX3 mRNA and protein expression in HUVECs. HG treatment for 24 to 48 h increased cell proliferation in a time­dependent manner, but the cell proliferation rate was decreased at 72 h of HG treatment. Conversely, CaD inhibited abnormal cell proliferation. HG treatment also significantly enhanced HVUEC migration compared to the control treatment. In contrast, CaD treatment partially inhibited HUVEC migration compared to HG exposure. HG­treated HUVECs exhibited increased FITC­BSA permeability compared to control cells cultured in medium alone; however, CaD application prevented the HG­induced increase in FITC­BSA permeability and suppressed HG­induced overexpression of endothelial markers (VEGF, VEGFR­2, endocan) and inflammation markers (ICAM­1, MCP­1, PTX3) in HUVECs. CaD has angioprotective properties and protects endothelial cells partly by ameliorating HG­induced inflammation. The current results demonstrated the potential applicability of CaD to the treatment of diabetic nephropathy, particularly during the early stages of this disease.