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Background: G-protein-coupled receptor 43 (GPR43) is a posttranscriptional regulator involved in cholesterol metabolism. This study aimed to investigate the possible roles of GPR43 activation in podocyte lipotoxicity in diabetic nephropathy (DN) and explore the potential mechanisms. Methods: The experiments were conducted by using diabetic GPR43-knockout mice and a podocyte cell culture model. Lipid deposition and free cholesterol levels in kidney tissues were measured by BODIPY staining and quantitative cholesterol assays, respectively. The protein expression of GPR43, LC3II, p62, beclin1, low-density lipoprotein receptor (LDLR) and early growth response protein 1 (EGR1) in kidney tissues and podocytes was measured by real-time PCR, immunofluorescent staining and Western blotting. Results: There were increased LDL cholesterol levels in plasma and cholesterol accumulation in the kidneys of diabetic mice. However, GPR43 gene knockout inhibited these changes. An in vitro study further demonstrated that acetate treatment induced cholesterol accumulation in high glucose-stimulated podocytes, which was correlated with increased cholesterol uptake mediated by LDLR and reduced cholesterol autophagic degradation, as characterized by the inhibition of LC3 maturation, p62 degradation and autophagosome formation. Gene knockdown or pharmacological inhibition of GPR43 prevented these effects on podocytes. Furthermore, GPR43 activation increased extracellular regulated protein kinases 1/2 (ERK1/2) activity and EGR1 expression in podocytes, which resulted in an increase in cholesterol influx and autophagy inhibition. In contrast, after GPR43 deletion, these changes in podocytes were improved, as shown by the in vivo and in vitro results. Conclusion: GPR43 activation-mediated lipotoxicity contributes to podocyte injury in DN by modulating the ERK/EGR1 pathway.

Mesenchymal stem cells-derived exosomes (MSC-exos) have attracted great interest as a cell-free therapy for acute kidney injury (AKI). However, the in vivo biodistribution of MSC-exos in ischemic AKI has not been established. The potential of MSC-exos in promoting tubular repair and the underlying mechanisms remain largely unknown. Methods: Transmission electron microscopy, nanoparticle tracking analysis, and western blotting were used to characterize the properties of human umbilical cord mesenchymal stem cells (hucMSCs) derived exosomes. The biodistribution of MSC-exos in murine ischemia/reperfusion (I/R) induced AKI was imaged by the IVIS spectrum imaging system. The therapeutic efficacy of MSC-exos was investigated in renal I/R injury. The cell cycle arrest, proliferation and apoptosis of tubular epithelial cells (TECs) were evaluated in vivo and in HK-2 cells. The exosomal miRNAs of MSC-exos were profiled by high-throughput miRNA sequencing. One of the most enriched miRNA in MSC-exos was knockdown by transfecting miRNA inhibitor to hucMSCs. Then we investigated whether this candidate miRNA was involved in MSC-exos-mediated tubular repair. Results: Ex vivo imaging showed that MSC-exos was efficiently homing to the ischemic kidney and predominantly accumulated in proximal tubules by virtue of the VLA-4 and LFA-1 on MSC-exos surface. MSC-exos alleviated murine ischemic AKI and decreased the renal tubules injury in a dose-dependent manner. Furthermore, MSC-exos significantly attenuated the cell cycle arrest and apoptosis of TECs both in vivo and in vitro. Mechanistically, miR-125b-5p, which was highly enriched in MSC-exos, repressed the protein expression of p53 in TECs, leading to not only the up-regulation of CDK1 and Cyclin B1 to rescue G2/M arrest, but also the modulation of Bcl-2 and Bax to inhibit TEC apoptosis. Finally, inhibiting miR-125b-5p could mitigate the protective effects of MSC-exos in I/R mice. Conclusion: MSC-exos exhibit preferential tropism to injured kidney and localize to proximal tubules in ischemic AKI. We demonstrate that MSC-exos ameliorate ischemic AKI and promote tubular repair by targeting the cell cycle arrest and apoptosis of TECs through miR-125b-5p/p53 pathway. This study provides a novel insight into the role of MSC-exos in renal tubule repair and highlights the potential of MSC-exos as a promising therapeutic strategy for AKI.

Rationale: Albuminuria is an early clinical feature in the progression of diabetic nephropathy (DN). Podocyte insulin resistance is a main cause of podocyte injury, playing crucial roles by contributing to albuminuria in early DN. G protein-coupled receptor 43 (GPR43) is a metabolite sensor modulating the cell signalling pathways to maintain metabolic homeostasis. However, the roles of GPR43 in podocyte insulin resistance and its potential mechanisms in the development of DN are unclear. Methods: The experiments were conducted by using kidney tissues from biopsied DN patients, streptozotocin (STZ) induced diabetic mice with or without global GPR43 gene knockout, diabetic rats treated with broad-spectrum oral antibiotics or fecal microbiota transplantation, and cell culture model of podocytes. Renal pathological injuries were evaluated by periodic acid-schiff staining and transmission electron microscopy. The expression of GPR43 with other podocyte insulin resistance related molecules was checked by immunofluorescent staining, real-time PCR, and Western blotting. Serum acetate level was examined by gas chromatographic analysis. The distribution of gut microbiota was measured by 16S ribosomal DNA sequencing with faeces. Results: Our results demonstrated that GPR43 expression was increased in kidney samples of DN patients, diabetic animal models, and high glucose-stimulated podocytes. Interestingly, deletion of GPR43 alleviated albuminuria and renal injury in diabetic mice. Pharmacological inhibition and knockdown of GPR43 expression in podocytes increased insulin-induced Akt phosphorylation through the restoration of adenosine 5'-monophosphate-activated protein kinase α (AMPKα) activity. This effect was associated with the suppression of AMPKα activity through post-transcriptional phosphorylation via the protein kinase C-phospholipase C (PKC-PLC) pathway. Antibiotic treatment-mediated gut microbiota depletion, and faecal microbiota transplantation from the healthy donor controls substantially improved podocyte insulin sensitivity and attenuated glomerular injury in diabetic rats accompanied by the downregulation of the GPR43 expression and a decrease in the level of serum acetate. Conclusion: These findings suggested that dysbiosis of gut microbiota-modulated GPR43 activation contributed to albuminuria in DN, which could be mediated by podocyte insulin resistance through the inhibition of AMPKα activity.

Introduction: Hyperuricemia has been associated with increased cardiovascular events in the general population. However, the role of serum uric acid (SUA) level on the severity of coronary artery s...

Background: Our previous study demonstrated that the disruption of cholesterol homeostasis promotes tubulointerstitial injury in diabetic nephropathy (DN). This study aimed to further investigate the effects of gut microbiota dysbiosis on this process and explored its potential mechanism. Methods: Diabetic rats treated with broad-spectrum oral antibiotics or faecal microbiota transplantation (FMT) from the healthy donor group and human kidney 2 (HK-2) cells stimulated with sodium acetate were used to observe the effects of gut microbiota on cholesterol homeostasis. The gut microbiota distribution was measured by 16S rDNA sequencing with faeces. Serum acetate level was examined by gas chromatographic analysis. Protein expression of G protein coupled receptor 43 (GPR43) and molecules involved in cholesterol homeostasis were assessed by immunohistochemical staining, immunofluorescence staining, and Western Blotting. Results: Depletion of gut microbiota significantly attenuated albuminuria and tubulointerstitial injury. Interestingly, serum acetate levels were also markedly decreased in antibiotics-treated diabetic rats and positively correlated with the cholesterol contents in kidneys. An in vitro study demonstrated that acetate significantly increased cholesterol accumulation in HK-2 cells, which was caused by increased expression of proteins mainly modulating cholesterol synthesis and uptake. As expected, FMT effectively decreased serum acetate levels and alleviated tubulointerstitial injury in diabetic rats through overriding the disruption of cholesterol homeostasis. Furthermore, GPR43 siRNA treatment blocked acetate-mediated cholesterol homeostasis dysregulation in HK-2 cells through decreasing the expression of proteins governed cholesterol synthesis and uptake. Conclusion: Our studies for the first time demonstrated that the acetate produced from gut microbiota mediated the dysregulation of cholesterol homeostasis through the activation of GPR43, thereby contributing to the tubulointerstitial injury of DN, suggesting that gut microbiota reprogramming might be a new strategy for DN prevention and therapy.

Intestinal microbiota-derived membrane vesicles (MVs) play essential roles in immunomodulation and maintenance of the intestinal micro-ecosystem. The relationship between MVs and chronic kidney disease (CKD) has remained undefined. This review provides a survey of the structure and biological function of different vesicle types and summarizes the possible pathogenic mechanisms mediated by MVs, which may be of great clinical significance in the diagnosis and treatment of chronic kidney disease.

Diabetic nephropathy (DN) is the leading cause of end-stage renal disease in the developed world. Podocyte injury is a critical cellular event involved in the progression of DN. Our previous studies demonstrated that platelet-derived microparticles (PMPs) mediated endothelial injury in diabetic rats. This study aimed to investigate whether PMPs are deposited in podocytes and to assess their potential effects on podocyte injury in DN.The deposition of PMPs in podocytes was assessed by immunofluorescent staining and electron microscopy. The changes in renal pathology and ultra-microstructure were assessed by periodic acid-Schiff staining and electron microscopy, respectively. The expression of inflammatory cytokines and extracellular matrix proteins was measured by immuno-histochemical staining and western blot.PMPs were widely deposited in podocytes of glomeruli in diabetic patients and animal models and closely associated with DN progression. Interestingly, aspirin treatment significantly inhibited the accumulation of PMPs in the glomeruli of diabetic rats, alleviated mesangial matrix expansion and fusion of foot processes, and decreased the protein expression of inflammatory cytokines and extracellular matrix secretion. An in vitro study further confirmed the deposition of PMPs in podocytes. Moreover, PMP stimulation induced the phenotypic transition of podocytes through decreased podocin protein expression and increased protein expression of α-SMA and fibronectin, which was correlated with increased production of inflammatory cytokines.Our findings demonstrated for the first time that the deposition of PMPs in podocytes contributed to the development of DN.

Background and Aims The G-protein-coupled receptor 43 (GPR43) is a post-transcriptional regulator involved in cholesterol metabolism. Our previous studies demonstrated that intracellular cholesterol accumulation contributed to podocyte injury in diabetic nephropathy (DN). This study aimed to investigate possible roles of GPR43 activation in lipid nephrotoxicity in DN and to explore its potential mechanisms. Method The experiments were conducted by using diabetic GPR43 knockout mice and cell culture model of podocytes. Renal pathological changes were checked by periodic acid schiff staining, immunohistochemical staining and transmission electron microscopy (TEM). The lipid deposition and free cholesterol contents in kidney tissues were detected by Oil Red O staining, BODIPY staining, and cholesterol quantitative assay. The protein expressions of GPR43, LC3II, p62 and related molecules of LDLR pathway in kidney tissues and podocytes were detected by real-time PCR, immunofluorescent staining, and Western blotting. Results There were decreased plasma level of LDL-cholesterol and cholesterol accumulation in kidney tissues of diabetic GPR43 knockout mice. In vitro study demonstrated that acetate, a stimulator of GPR43, increased LDLR-mediated cholesterol uptake in podocytes, accompanied with reduced cholesterol autophagic degeneration, characterized by inhibited LC3 maturation, p62 degradation, and autophagosome formation. These may evoke synergistic effects attributing to cellular cholesterol accumulation in podocytes. While genetic knockdown or pharmacological inhibition of GPR43 inhibited this effect in podocytes. Furthermore, the activation of GPR43 led to the activation of ERK1/2/EGR1 pathway in podocytes, whereas blocking ERK1/2 activity or reducing EGR1 expression reversed cholesterol influx and the inhibition of autophagy mediated by GPR43 activation in podocytes. Meanwhile, deletion of the GPR43 improved autophagy and inhibited the ERK1/2-EGR1 pathway of podocyte in diabetic mice in vivo. Conclusion Activation of the GPR43 mediated lipid nephrotoxicity contributes to podocyte injury in DN, which was mainly through the activation of ERK/EGR1 pathways. These findings suggested that the GPR43 receptor could be a potential therapeutic target for the prevention of DN progression.

Background and Aims Diabetic nephropathy (DN) is the leading cause of end-stage renal disease in the developed world. Podocyte injury has been shown to be a critical cellular event in the progression of DN. Our previous studies demonstrated that platelet-derived microparticles (PMPs) mediated endothelial injury in diabetic rats. Therefore, this study aimed to investigate whether the PMPs deposit in podocytes and its potential effects on podocyte injury in DN. Method Deposition of PMPs in podocytes was checked by immunofluorescent staining and electron microscopy. Renal pathological change and ultra-microstructure were respectively checked by periodic acid-schiff staining and electron microscopy. The protein expression of inflammatory cytokines and extracellular matrix was measured by immunohistochemical staining and Western blot. Results The PMPs were found to be widely deposited in podocytes of glomeruli in diabetic patients and animal models and closely associated with progression of DN. Interestingly, aspirin treatment significantly inhibited accumulation of PMPs in glomeruli of diabetic rats, accompanied with alleviated mesangial matrix expansion, fusion of foot processes, decreased protein expression of inflammatory cytokines and extracellular matrix secretion. The in vitro study further confirmed the deposition of PMPs in podocytes. Moreover, PMPs stimulation induced phenotype transition of podocytes through decreased protein expression of podocin and increased protein expression of α-SMA and fibronectin, which was correlated with increased production of inflammatory cytokines. Conclusion Our findings for the first time demonstrated that deposition of PMPs in podocytes contributed to the development of DN.

The discovery of hypoxia-inducible factor (HIF)-prolyl hydroxylase inhibitor (PHI) has revolutionized the treatment strategy for renal anemia. However, the presence of multiple transcription targets of HIF raises safety concerns regarding HIF-PHI. Here, we explored the dose-dependent effect of MK-8617 (MK), a kind of HIF-PHI, on renal fibrosis. MK was administered by oral gavage to mice for 12 wk at doses of 1.5, 5, and 12.5 mg/kg. In vitro, the human proximal tubule epithelial cell line HK-2 was treated with increasing doses of MK administration. Transcriptome profiling was performed, and fibrogenesis was evaluated. The dose-dependent biphasic effects of MK on tubulointerstitial fibrosis (TIF) were observed in chronic kidney disease mice. Accordingly, high-dose MK treatment could significantly enhance TIF. Using RNA-sequencing, combined with in vivo and in vitro experiments, we found that Krüppel-like factor 5 (KLF5) expression level was significantly increased in the proximal tubular cells, which could be transcriptionally regulated by HIF-1α with high-dose MK treatment but not low-dose MK. Furthermore, our study clarified that HIF-1α–KLF5–TGF-β1 signaling activation is the potential mechanism of high-dose MK-induced TIF, as knockdown of KLF5 reduced TIF in vivo. Collectively, our study demonstrates that high-dose MK treatment initiates TIF by activating HIF-1α–KLF5–TGF-β1 signaling. These findings provide novel insights into TIF induction by high-dose MK (HIF-PHI), suggesting that the safety dosage window needs to be emphasized in future clinical applications.—Li, Z.-L., Lv, L.-L., Wang, B., Tang, T.-T., Feng, Y., Cao, J.-Y., Jiang, L.-Q., Sun, Y.-B., Liu, H., Zhang, X.-L., Ma, K.-L., Tang, R.-N., Liu, B.-C. The profibrotic effects of MK-8617 on tubulointerstitial fibrosis mediated by the KLF5 regulating pathway.

Aim: Idiopathic membranous nephropathy (IMN) is an immune-mediated inflammatory disease characterized by a high risk of thromboembolic complications. Microparticles (MPs), a type of extracellular vesicles, have procoagulant properties, especially when they display tissue factor (TF). This study aimed to investigate whether circulating TF-positive MPs contributed to the hypercoagulable state in patients with IMN. Methods: Twenty adult IMN patients and fourteen healthy subjects were included in the study. The basic indexes of a routine biochemical examination and coagulative function were determined. The plasma levels of MPs were detected by flow cytometry, and TF activity of MPs was examined using an assay kit. The plasma levels of lipopolysaccharide (LPS) were measured by an enzyme-linked immunosorbent assay. Results: Total circulating MPs were not increased in patients with IMN compared with healthy controls. Circulating CD14+/TF+MPs were significantly increased in IMN patients, but this achieved significance was not observed in CD41+/TF+MPs between the two groups. Interestingly, the circulating TF-positive MPs were increased significantly. Plasma MPs TF assays revealed high procoagulant activity, which was positively associated with the D-dimer level in IMN. In addition, circulating LPS in IMN patients were significantly higher than those in the controls. Furthermore, after two hours' incubation with healthy whole blood, LPS enhanced the release of circulating TF-positive MPs and the TF activity of MPs. Conclusion: Increased circulating LPS may mediate the release of monocyte-derived TF-positive MPs, which further contributes to the hypercoagulable state in IMN patients. These findings provide an additional mechanism by which patients with IMN have a higher risk of thromboembolic complication.

AIM Lipoprotein(a) [Lp(a)] is considered a risk factor for cardiovascular disease. However, the role of Lp(a) in chronic kidney disease (CKD) patients is not well understood. The aim of this study was to evaluate the association between Lp(a) levels and the presence of coronary artery disease (CAD) and the severity of coronary artery stenosis (CAS) in patients with stage 3-5 CKD. PATIENTS AND METHODS We retrospectively studied patients who were diagnosed with stage 3-5 CKD and underwent coronary artery angiography in Zhongda Hospital. The Gensini scoring system was used to assess the severity of coronary stenosis. RESULTS A total of 1003 patients were enrolled in this cross-sectional study, and 776 of these patients were diagnosed with CAD. The Lp(a) levels were significantly higher in the CAD group than the non-CAD group [271.50 (168.00-459.25) vs. 195.00 (131.00-347.00) mg/l, P

Background Platelet microparticles (PMPs) are closely associated with diabetic macrovascular complications. This study aimed to explore the underlying mechanisms of high glucose-induced PMPs generation. Methods Washed platelets, obtained from the plasma of healthy male Sprague-Dawley rats, were incubated with high glucose. PMPs were isolated using gradient centrifugation and counted by flow cytometry. Expression and activity of ROCK1 and caspase3 were evaluated by real-time PCR, Western blotting, and activity assay kit. Results High glucose enhanced PMPs shedding in the presence of collagen. The mRNA and protein levels of ROCK1, but not ROCK2, were increased in platelets incubated with high glucose. Y-27632, an inhibitor of ROCK, blocked the increased PMPs shedding induced by high glucose. Expression and activity of caspase3 were elevated in platelets under the high glucose conditions. Z-DVED-FMK, a caspase3 inhibitor, inhibited ROCK1 activity and decreased the PMPs generation under high glucose. Conclusion High glucose increased PMPs shedding via caspase3-ROCK1 signal pathway.

This study aimed to investigate the effects of aspirin on podocyte injury and its underlying mechanisms in diabetic nephropathy (DN). Eight-week-old male Sprague–Dawley rats were divided into three groups: non-diabetic rats (Control), streptozotocin-induced diabetic rats (DM), and diabetic rats treated with aspirin (DM + Aspirin) for 12 weeks. Intracellular lipid accumulation was evaluated by Oil Red O staining and quantitative free cholesterol assays. Podocyte injury and the levels of COX-2, inflammatory cytokines, and low-density lipoprotein receptor (LDLr) pathway-related proteins were evaluated by electron microscopy, immunohistochemical staining, and Western blotting, respectively. Lipid levels and urinary albumin–creatinine ratios were higher in the DM rats than in the Control rats. Periodic acid-Schiff staining showed glomerular hypertrophy and mild mesangial area widening in the DM rats. Electron microscopy showed that the podocyte foot processes were significantly flattened or absent in the DM rats. The protein expression levels of WT-1 and nephrin in the podocytes of DM rats were reduced. Interestingly, lipid accumulation in the kidneys of DM rats was significantly increased due to increased protein expression levels of LDLr, sterol regulatory element-binding protein (SREBP) cleavage-activating protein (SCAP), SREBP-2, cyclooxygenase-2 (COX-2), and inflammatory cytokines. Confocal immunofluorescent staining showed that COX-2 and WT-1 were co-expressed. Furthermore, COX-2 protein expression levels were positively correlated with LDLr protein expression levels. However, when COX-2 expression was inhibited by aspirin, these changes in the DM rats were significantly attenuated. Aspirin attenuates podocyte injury in DN, which may be through COX-2-mediated dysregulation of LDLr pathway.

Artemisinin (Art) is isolated from Artemisia annua L. and known as the most effective antimalaria drugs. Previous studies demonstrated that it could exert an immune-regulatory effect on autoimmune diseases. In this study, we first investigated its potential role in tubulointerstitial inflammation and fibrosis in rats with 5/6 nephrectomy. Subtotal nephrectomized (SNx) rats were orally administered Art (100 mg·kg -1 ·d - 1) for 16 weeks. Blood and urine samples were collected for biochemical examination. Kidney tissues were collected for immunohistochemistry and Western blot analyses. Ang II-induced injury of the human kidney 2 (HK-2) cells was used for in vitro study. It was shown that Art could significantly attenuate the renal function decline in SNx rats compared with control. More importantly, Art treatment significantly reduced the tubulointerstitial inflammation and fibrosis, as demonstrated by the evaluation of renal pathology. Furthermore, Art inhibited the activation of NLRP3 inflammasome and NF-κB in the kidneys. In in vitro study, Art pretreatment could significantly prevent the activation of NLRP3 inflammasome and NF-κB in Ang II-treated HK-2 cells, while BAY11-7082 (an inhibitor of NF-κB) significantly inhibited Ang II-induced NLRP3 inflammasome activation. This study suggested that Art could provide renoprotective role by attenuating the tubulointerstitial inflammation and fibrosis in SNx rats by downregulating the NF-κB/NLRP3 signaling pathway.

Background/Aims: Chronic kidney disease (CKD) is a worldwide public health problem. Regardless of the underlying primary disease, CKD tends to progress to end-stage kidney disease, resulting in unsatisfactory and costly treatment. Its common pathogenesis, however, remains unclear. The aim of this study was to provide an unbiased catalog of common gene-expression changes of CKD and reveal the underlying molecular mechanism using an integrative bioinformatics approach. Methods: We systematically collected over 250 Affymetrix microarray datasets from the glomerular and tubulointerstitial compartments of healthy renal tissues and those with various types of established CKD (diabetic kidney disease, hypertensive nephropathy, and glomerular nephropathy). Then, using stringent bioinformatics analysis, shared differentially expressed genes (DEGs) of CKD were obtained. These shared DEGs were further analyzed by the gene ontology (GO) and pathway enrichment analysis. Finally, the protein-protein interaction networks(PINs) were constructed to further refine our results. Results: Our analysis identified 176 and 50 shared DEGs in diseased glomeruli and tubules, respectively, including many transcripts that have not been previously reported to be involved in kidney disease. Enrichment analysis also showed that the glomerular and tubulointerstitial compartments underwent a wide range of unique pathological changes during chronic injury. As revealed by the GO enrichment analysis, shared DEGs in glomeruli were significantly enriched in exosomes. By constructing PINs, we identified several hub genes (e.g. OAS1, JUN, and FOS) and clusters that might play key roles in regulating the development of CKD. Conclusion: Our study not only further reveals the unifying molecular mechanism of CKD pathogenesis but also provides a valuable resource of potential biomarkers and therapeutic targets.

Glomerular sclerosis is characterized by mesangial cell proliferation and progressive extracellular matrix (ECM) accumulation. CCN3 belongs to the CCN family of matrix proteins; increasing evidence suggests that CCN3 is an endogenous negative regulator of the ECM and fibrosis. However, the exact role of CCN3 in the accumulation of ECM remains unknown. The aim of the present study was to investigate the effects of CCN3 on TGF-β1-induced production of ECM in human mesangial cells (HMCs) in vitro. Treatment with TGF-β1 (0.5–2.0 ng/mL) suppressed the mRNA and protein expression of CCN3 in HMCs in dose- and time-dependent manners. Furthermore, treatment with TGF-β1 significantly increased the expression of the two markers of renal fibrosis, fibronectin (FN) and type I collagen (COLI), in HMCs. Moreover, treatment with TGF-β1 significantly decreased the expression of metalloproteinase (MMP)-2 and MMP-9, and markedly increased the expression of tissue inhibitor of metalloproteinase (TIMP)-1 in HMCs. Pretreatment of HMCs with exogenous CCN3 (5–500 ng/mL) or overexpression of CCN3 significantly attenuated TGF-β1-induced changes in FN, COLI, MMP-2, MMP-9 and TIMP-1 in HMCs. These results suggest that CCN3 suppresses TGF-β1-induced accumulation of ECM in HMCs. CCN3 may have potential as a novel therapeutic target for alleviating glomerulosclerosis.

Microparticles (MPs) are a type of extracellular vesicles (EVs) shed from the outward budding of plasma membranes during cell apoptosis and/or activation. These microsized particles then release specific contents (e.g., lipids, proteins, microRNAs) which are active participants in a wide range of both physiological and pathological processes at the molecular level, e.g., coagulation and angiogenesis, inflammation, immune responses. Research limitations, such as confusing nomenclature and overlapping classification, have impeded our comprehension of these tiny molecules. Diabetic nephropathy (DN) is currently the greatest contributor to end-stage renal diseases (ESRD) worldwide, and its public health impact will continue to grow due to the persistent increase in the prevalence of diabetes mellitus (DM). MPs have recently been considered as potentially involved in DN onset and progression, and this review juxtaposes some of the research updates about the possible mechanisms from several relevant aspects and insights into the therapeutic perspectives of MPs in clinical management and pharmacological treatment of DN patients.

Background: Podocyte-derived microparticles (MPs) could be secreted from activated or apoptotic podocytes. An increased number of podocyte-derived MPs in the urine might reflect podocyte injury in renal diseases. This study aimed to observe the change of urinary podocyte-derived MP levels in patients with chronic kidney disease (CKD) and to further explore its correlation with the progression of CKD. Methods: A prospective, longitudinal study was conducted in eighty patients with biopsy-proven CKD. Podocyte-derived MPs (annexin V and podocalyxin positive) were detected by flow cytometry. The number of urinary podocyte-derived MPs was analyzed to evaluate the association with biochemical measurements and pathological glomerulosclerosis assessment. Patients with idiopathic membranous nephropathy (IMN) were followed up after the six-month treatment of prednisone combined with tacrolimus to evaluate the association of urinary podocyte-derived MP levels and the remission of IMN. Results: The CKD patients had higher urinary podocyte-derived MP levels compared with healthy controls (HCs). Baseline urinary levels of podocyte-derived MPs were positively correlated with 24-hour proteinuria, while were inversely correlated with the percentage of global glomerulosclerosis. The urinary podocyte-derived MPs levels had good discrimination for glomerulosclerosis [area under curve (AUC), 0.66]. The urinary podocyte-derived MPs levels in IMN patients were significantly decreased accompanied with the recovery of abnormal clinical parameters after six-month treatment. Conclusions: The urinary levels of podocyte-derived MPs were closely associated with podocyte injury and glomerulosclerosis, which could be useful for monitoring disease activity in CKD patients. Urinary podocyte-derived MPs might be a non-invasive biomarker for the evaluation of early CKD progression.

Background New non-invasive biomarkers are demanded to identify renal damage in various autoimmune-associated kidney diseases. Glomerular podocyte damage mediated by systemic lupus erythematosus (SLE) plays an important role in the pathogenesis and progression of lupus nephritis (LN). This study evaluated whether the podocyte-derived microparticles (MPs) were novel biomarkers of clinical and histological features in SLE patients with LN. Methods A cross-sectional study, including 34 SLE patients and 16 healthy controls, was designed. Urinary annexin V+ podocalyxin+ MPs of all participants were quantified by flow cytometry. The correlation of podocyte-derived MPs with clinical and histological parameters of SLE patients was analysed. Results The number of annexin V+ podocalyxin+ MPs from urine samples were markly increased in patients with SLE. Furthermore, the level of urinary podocyte-derived MPs was positively correlated with the SLE Disease Activity Index (SLEDAI) score, anti-dsDNA antibody titre, erythrocyte sedimentation rate, and proteinuria. Conversely, it was negatively correlated with the level of complement C3 and serum albumin. The number of urinary podocyte-derived MPs was significantly increased in SLE patients with high activity indices. Receiver operating characteristic (ROC) curves were calculated to assess the power for podocyte-derived MP levels in differentiating between SLE patients with and without LN. Podocyte-derived MP levels were able to differentiate between SLE patients with mild disease activity, as well as those with moderate and above disease activity. SLE patients showed increased podocyte-derived MP excretion into the urine. Conclusions These findings suggest that the change in urinary podocyte-derived MP levels could be useful for evaluating and monitoring SLE disease activity.

Some studies have shown that gut microbiota along with its metabolites is closely associated with diabetic mellitus (DM). In this study we explored the relationship between gut microbiota and kidney injuries of early diabetic nephropathy (DN) and its underlying mechanisms. Male SD rats were intraperitoneally injected with streptozotocin to induce DM. DM rats were orally administered compound broad-spectrum antibiotics for 8 weeks. After the rats were sacrificed, their blood, urine, feces, and renal tissues were harvested for analyses. We found that compared with the control rats, DM rats had abnormal intestinal microflora, increased plasma acetate levels, increased proteinuria, thickened glomerular basement membrane, and podocyte foot process effacement in the kidneys. Furthermore, the protein levels of angiotensin II, angiotensin-converting enzyme, and angiotensin II type 1 receptor in the kidneys of DM rats were significantly increased. Administration of broad-spectrum antibiotics in DM rats not only completely killed most intestinal microflora, but also significantly lowered the plasma acetate levels, inhibited intrarenal RAS activation, and attenuated kidney damage. Finally, we showed that plasma acetate levels were positively correlated with intrarenal angiotensin II protein expression (r = 0.969, P

Inflammation and lipid disorders play crucial roles in synergistically accelerating the progression of diabetic nephropathy (DN). In this study we investigated how inflammation and lipid disorders caused tubulointerstitial injury in DN in vivo and in vitro. Diabetic db/db mice were injected with 10% casein (0.5 mL, sc) every other day for 8 weeks to cause chronic inflammation. Compared with db/db mice, casein-injected db/db mice showed exacerbated tubulointerstitial injury, evidenced by increased secretion of extracellular matrix (ECM) and cholesterol accumulation in tubulointerstitium, which was accompanied by activation of the CXC chemokine ligand 16 (CXCL16) pathway. In the in vitro study, we treated HK-2 cells with IL-1β (5 ng/mL) and high glucose (30 mmol/L). IL-1β treatment increased cholesterol accumulation in HK-2 cells, leading to greatly increased ROS production, ECM protein expression levels, which was accompanied by the upregulated expression levels of proteins in the CXCL16 pathway. In contrast, after CXCL16 in HK-2 cells was knocked down by siRNA, the IL-1β-deteriorated changes were attenuated. In conclusion, inflammation accelerates renal tubulointerstitial lesions in mouse DN via increasing the activity of CXCL16 pathway.

Background Diabetic kidney disease (DKD) is the leading cause of end-stage kidney disease (ESKD) in the world. Emerging evidence has shown that urinary mRNAs may serve as early diagnostic and prognostic biomarkers of DKD. In this article, we aimed to first establish a novel bioinformatics-based methodology for analyzing the “urinary kidney-specific mRNAs” and verify their potential clinical utility in DKD. Methods To select candidate mRNAs, a total of 127 Affymetrix microarray datasets of diabetic kidney tissues and other tissues from humans were compiled and analyzed using an integrative bioinformatics approach. Then, the urinary expression of candidate mRNAs in stage 1 study (n = 82) was verified, and the one with best performance moved on to stage 2 study (n = 80) for validation. To avoid potential detection bias, a one-step Taqman PCR assay was developed for quantification of the interested mRNA in stage 2 study. Lastly, the in situ expression of the selected mRNA was further confirmed using fluorescent in situ hybridization (FISH) assay and bioinformatics analysis. Results Our bioinformatics analysis identified sixteen mRNAs as candidates, of which urinary BBOX1 (uBBOX1) levels were significantly upregulated in the urine of patients with DKD. The expression of uBBOX1 was also increased in normoalbuminuric diabetes subjects, while remained unchanged in patients with urinary tract infection or bladder cancer. Besides, uBBOX1 levels correlated with glycemic control, albuminuria and urinary tubular injury marker levels. Similar results were obtained in stage 2 study. FISH assay further demonstrated that BBOX1 mRNA was predominantly located in renal tubular epithelial cells, while its expression in podocytes and urothelium was weak. Further bioinformatics analysis also suggested that tubular BBOX1 mRNA expression was quite stable in various types of kidney diseases. Conclusions Our study provided a novel methodology to identify and analyze urinary kidney-specific mRNAs. uBBOX1 might serve as a promising biomarker of DKD. The performance of the selected urinary mRNAs in monitoring disease progression needs further validation. Electronic supplementary material The online version of this article (10.1186/s12967-019-1818-2) contains supplementary material, which is available to authorized users.

Finding new therapeutic targets of glomerulosclerosis treatment is an ongoing quest. Due to a living environment of various stresses and pathological stimuli, podocytes are prone to injuries; moreover, as a cell without proliferative potential, loss of podocytes is vital in the pathogenesis of glomerulosclerosis. Thus, sufficient understanding of factors and underlying mechanisms of podocyte injury facilitates the advancement of treating and prevention of glomerulosclerosis. The clinical symptom of podocyte injury is proteinuria, sometimes with loss of kidney functions progressing to glomerulosclerosis. Injury-induced changes in podocyte physiology and function are actually not a simple passive process, but a complex interaction of proteins that comprise the anatomical structure of podocytes at molecular levels. This chapter lists several aspects of podocyte injuries along with potential mechanisms, including glucose and lipid metabolism disorder, hypertension, RAS activation, micro-inflammation, immune disorder, and other factors. These aspects are not technically separated items, but intertwined with each other in the pathogenesis of podocyte injuries.

This study aimed to investigate possible synergistic effects of lipid disorder with renin-angiotensin-system (RAS) activation in non-alcoholic fatty liver disease (NAFLD). Apolipoprotein E gene-knockout mice, angiotensin II (Ang II) type 1 receptor (AT1) gene-knockout mice, and human hepatoblastoma cell line (HepG2) were used for experiments. Lipid accumulation was examined by Filipin staining and intracellular cholesterol quantitative assay. The gene and protein expression of molecules involved in RAS and low-density lipoprotein receptor (LDLr) pathway was examined by real-time PCR, immunofluorescent staining, and Western blot. There was significantly increased expression of RAS components and extracellular matrix (ECM) in livers of high-fat-diet-fed apolipoprotein E gene-knockout mice compared with controls. Upregulation of RAS components was positively associated with increased plasma levels of lipid profile. The in vitro study further confirmed that cholesterol loading increased supernatant renin activity and Ang II level of HepG2 cells, accompanied by increased ECM production that was positively associated with increased expression of intracellular RAS components. Interestingly, Ang II treatment increased lipid accumulation in livers of C57BL/6 mice and HepG2 cells. Furthermore, Ang II treatment increased gene and protein expression of sterol regulatory element-binding protein (SREBP) cleavage activating protein (SCAP), SREBP-2, and LDLr, which were mediated by enhanced SCAP/SREBP-2 complex translocation from endoplasmic reticulum to Golgi. However, LDLr pathway was accordingly downregulated in livers of AT1 gene-knockout C57BL/6 mice or in HepG2 cells treated by telmisartan. These findings demonstrate that lipid disorder and intrahepatic RAS activation synergistically accelerate NAFLD progression. This article is protected by copyright. All rights reserved.

Background: Chronic inflammation plays a critical role in the progression of atherosclerosis (AS). This study aimed to determine the effects of the CXC chemokine ligand 16 (CXCL16)/CXC chemokine receptor 6 (CXCR6) pathway on cholesterol accumulation in the radial arteries of end-stage renal disease (ESRD) patients with concomitant microinflammation and to further investigate the potential effects of the purinergic receptor P2X ligand-gated ion channel 7 (P2X7R). Methods: Forty-three ESRD patients were divided into the control group (n=17) and the inflamed group (n=26) based on plasma C-reactive protein (CRP) levels. Biochemical indexes and lipid profiles of the patients were determined. Surgically removed tissues from the radial arteries of patients receiving arteriovenostomy were used for preliminary evaluation of AS. Haematoxylin-eosin (HE) and Filipin staining were performed to assess foam cell formation. CXCL16/CXCR6 pathway-related protein expression, P2X7R protein expression and the expression of monocyte chemotactic protein-1 (MCP-1), tumour necrosis factor-α (TNF-α), and CD68 were detected by immunohistochemical and immunofluorescence staining. Results: Inflammation increased both MCP-1 and TNF-α expression and macrophage infiltration in radial arteries. Additionally, foam cell formation significantly increased in the radial arteries of the inflamed group compared to that of the controls. Further analysis showed that protein expression of CXCL16, CXCR6, disintegrin and metalloproteinase-10 (ADAM10) in the radial arteries of the inflamed group was significantly increased. Furthermore, CXCL16 expression was positively correlated with P2X7R expression in the radial arteries of ESRD patients. Conclusions: Inflammation contributed to foam cell formation in the radial arteries of ESRD patients via activation of the CXCL16/CXCR6 pathway, which may be regulated by P2X7R.

The low-density lipoprotein receptor (LDLR) pathway is a negative feedback system that plays important roles in the regulation of plasma and intracellular cholesterol homeostasis. To maintain a cholesterol homeostasis, LDLR expression is tightly regulated by sterol regulatory element-binding protein-2 (SREBP-2) and SREBP cleavage-activating protein (SCAP) in transcriptional level and by proprotein convertase subtilisin/kexin type 9 (PCSK9) in posttranscriptional level. The dysregulation of LDLR expression results in abnormal lipid accumulation in cells and tissues, such as vascular smooth muscle cells, hepatic cells, renal mesangial cells, renal tubular cells and podocytes. It has been demonstrated that inflammation, renin-angiotensin system (RAS) activation, and hyperglycemia induce the disruption of LDLR pathway, which might contribute to lipid disorder-mediated organ injury (atherosclerosis, non-alcoholic fatty liver disease, kidney fibrosis, etc). The mammalian target of rapamycin (mTOR) pathway is a critical mediator in the disruption of LDLR pathway caused by pathogenic factors. The mTOR complex1 activation upregulates LDLR expression at the transcriptional and posttranscriptional levels, consequently resulting in lipid deposition. This paper mainly reviews the mechanisms for the dysregulation of LDLR pathway and its roles in lipid disorder-mediated organ injury under various pathogenic conditions. Understanding these mechanisms leading to the abnormality of LDLR expression contributes to find potential new drug targets in lipid disorder-mediated diseases.

Platelet microparticles (PMPs) are closely associated with diabetic macrovascular complications. The present study aimed to investigate the effects of PMPs in diabetes on aortic vascular endothelial injury and to explore the underlying mechanisms. Peritoneal injection of streptozotocin was used to generate a diabetic rat model in vivo, and human umbilical vein endothelial cells (HUVECs) treated with PMPs were used in vitro. PMP levels in the circulation and aorta tissues were time-dependently increased in streptozotocin-induced diabetic rats at weeks 4, 8, and 12 (P

Background Increased plasma level of lipoprotein(a) (Lpa) is a risk factor of cardiovascular diseases. This study aimed to explore the role of Lpa in the progression of atherosclerosis in patients with end-stage renal disease (ESRD) and to investigate whether its potential mechanism is mediated by CXC chemokine ligand 16 (CXCL16) and low-density lipoprotein receptor (LDLr). Methods This is a retrospective clinical study. From January 2015 to April 2016, forty-six ESRD patients from Danyang First People’s Hospital were investigated. The patients were grouped according to their plasma Lpa levels: control group (Lpa

The mechanisms that underlie the profibrotic effect of interleukin (IL)-1β are complicated and not fully understood. Recent evidence has suggested the involvement of the calcium-sensing receptor (CaSR) in tubular injury. Therefore, the current study aimed to investigate whether CaSR mediates IL-1β-induced collagen expression in cultured mouse inner medullary collecting duct cells (mIMCD3) and to determine the possible downstream signaling effector. The results showed that IL-1β significantly upregulated the expression of type I and III collagens in a concentration- and time-dependent manner. Moreover, CaSR was expressed in mIMCD3 cells, and its expression was increased by increasing the concentrations and times of IL-1β treatment. Selective inhibitors (Calhex231 or NPS2143) or the siRNA of CaSR attenuated the enhanced expression of type I and III collagens. Furthermore, IL-1β increased nuclear β-catenin protein levels and decreased cytoplasmic β-catenin expression in cells. In contrast, blockage of CaSR by the pharmacological antagonists or siRNA could partially attenuate such changes in the IL-1β-induced nuclear translocation of β-catenin. DKK1, an inhibitor of β-catenin nuclear translocation, further inhibited the expression of type I and III collagens in cells treated with IL-1β plus CaSR antagonist. In summary, these data demonstrated that IL-1β-induced collagen I and III expressions in collecting duct cells might be partially mediated by CaSR and the downstream nuclear translocation of β-catenin.

Considerable interest nowadays has focused on gut microbiota owing to their pleiotropic roles in human health and diseases. This intestinal community can arouse a variety of activities in the host and function as "a microbial organ" by generating bioactive metabolites and participating in a series of metabolism-dependent pathways. Alternations in the composition of gut microbiota, referred to as intestinal dysbiosis, are reportedly associated with several diseases, especially diabetes mellitus and its complications. Here we focus on the relationship between gut microbiota and diabetic nephropathy (DN), as the latter is one of the major causes of chronic kidney diseases. The activation of renin angiotensin system (RAS) is a critical factor to the onset of DN, and emerging data has demonstrated a provoking and mediating role of gut microbiota for this system in the context of metabolic diseases. The purpose of the current review is to highlight some research updates about the underlying interplay between gut microbiota, their metabolites, and the development and progression of DN, along with exploring innovative approaches to targeting this intestinal community as a therapeutic perspective in clinical management of DN patients.

Non-alcoholic fatty liver disease (NAFLD), characterised by early lipid accumulation and subsequent inflammation in the liver, is becoming a worldwide challenge due to its increasing prevalence in developing and developed countries. This study aimed to investigate the role of CXC chemokine ligand 16 (CXCL16) and its receptor CXC chemokine receptor 6 (CXCR6) in NAFLD under inflammation. We used IL-1β stimulation in human hepatoblastoma cell line (HepG2) for in vitro studies and casein injection in apolipoprotein E knockout mice in vivo to induce inflammatory stress. The effects of inflammation on cholesterol accumulation were examined by histochemical staining and a quantitative intracellular cholesterol assay. The gene and protein expression of molecules involved in CXCL16/CXCR6 pathway and extracellular matrix (ECM) were examined by real-time polymerase chain reaction (PCR) and Western blotting. The fluorescence intensity of reactive oxygen species (ROS) was assessed by flow cytometry. Results showed that significantly elevated levels of serum amyloid protein A in casein-injected mice confirmed the successful induction of inflamed NAFLD model. Inflammation significantly increased lipid accumulation in livers compared with the high-fat diet group and the controls. Furthermore, inflammation increased the expression of CXCL16, CXCR6, and adisintegrin and metalloproteinase domain-containing protein 10 (ADAM10) in livers, accompanied with increased ECM expression and ROS production. These effects were further confirmed by in vitro studies. Interestingly, CXCL16 gene knockdown in HepG2 cells induced by CXCL16 siRNA resulted in decreased lipid accumulation, ECM excretion, and ROS production. These findings demonstrated that inflammation-mediated activation of CXCL16/CXCR6 is involved in the progression of NAFLD.

Ischemia/reperfusion (I/R) is a critical risk factor for acute kidney injury (AKI). Recent studies provided evidence that tubular epithelial cells (TEC)-associated inflammation aggravates kidney injury and impairs tissue repair after I/R injury. Here we demonstrated that the Nod-like receptor protein 3 (NLRP3) inflammasome is activated by mitochondrial reactive oxygen species (mROS) during I/R injury via direct interactions between the inflammasome and thioredoxin-interacting protein (TXNIP). Firstly, we found that NLRP3 inflammasome activation was induced by I/R injury, peaking at day 3 after reperfusion. Consistent with this observation, NLRP3 deletion significantly attenuated I/R-induced kidney damage and markers of inflammasome activation. Then, we observed mitochondrial dysfunction, characterized by ultrastructural changes and cytochrome C (Cyt c) redistribution. Mitochondria-targeted antioxidant MitoTEMPO prevented mROS overproduction and the decline in mitochondrial membrane potential (MMP) in vitro. MitoTEMPO treatment also inhibited NLRP3 inflammasome activation and co-localization of NLRP3 and TXNIP after simulated ischemia/reperfusion (SI/R) injury. Finally, we transfected HK-2 cells with TXNIP siRNA to explore the role of TXNIP in mROS-induced NLRP3 inflammasome activation. We found that TXNIP siRNA significantly inhibited NLRP3 inflammasome activation. These results demonstrate that NLRP3 inflammasome is activated through the mROS-TXNIP-NLRP3 pathway and provide a potential therapeutic target in ischemic AKI.

Dyslipidemia and chronic inflammation play crucial roles in the progression of diabetes. This study aimed to investigate the effects of inflammatory stress on lipid accumulation in multi-organs in diabetes. Eight-week-old male db/db mice were randomly assigned to inflamed group with alternating day subcutaneous injection of 10% casein or control group with daily injection of distilled water. The lipid profile and plasma levels of inflammatory cytokines were determined using a clinical biochemical assay and enzyme-linked immunosorbent assay. The effects of inflammation on lipid accumulation in target organs were evaluated by hematoxylin-eosin staining, Oil Red O staining, Filipin staining, and a quantitative intracellular cholesterol assay. The protein expressions of low-density lipoprotein receptor (LDLr), sterol regulatory element binding protein-2 (SREBP-2), and SREBP-cleavage-activating protein (SCAP) in tissues were assessed by immunohistochemical staining and western blotting. Results showed that the serum levels of inflammatory cytokines were significantly elevated in casein-injected mice, suggesting that an inflamed diabetic model was established. Furthermore, the protein expressions of inflammatory cytokines in aortas, livers, kidneys, and intestines were significantly increased in inflamed group compared with control. Whereas the serum levels of lipid moieties in inflamed mice were not different compared with the control, inflammatory stress significantly increased lipid accumulation in aortas, livers, kidneys, and intestines, which coincided with increased protein expressions of LDLr, SREBP-2, and SCAP in these organs of inflamed mice. In conclusion, inflammation induces lipid accumulation in multi-organs of db/db mice from the circulation to peripheral tissues, potentially due to lipid redistribution mediated by the disruption of LDLr feedback regulation.