Mesangial Cell Activation Research Articles

Sialylation of IgG inhibits the formation of galactose-deficient IgA1-containing immune complexes and protects mesangial cells from injury in IgA nephropathy

BackgroundThe addition of sialic acid alters IgG from a pro-inflammatory state to an anti-inflammatory state. However, there is a lack of research on the changes of IgG sialylation in IgA nephropathy (IgAN).MethodsThis study included a total of 184 IgAN patients. The sialylated IgG (SA-IgG), IgG-galactose-deficient IgA1 complex (IgG-Gd-IgA1-IC), IL-6, TNF-α, and TGF-β were detected using commercial ELISA kits. SA-IgG, non-sialylated IgG (NSA-IgG), sialylated IgG-IgA1 complex (SA-IgG-IgA1), and non-sialylated IgG-IgA1 complex (NSA-IgG-IgA1) were purified from IgAN patients and healthy controls (HCs).ResultsThe mean SA-IgG levels in plasma and B lymphocytes in IgAN patients were significantly higher than those of healthy controls. A positive correlation was found between SA-IgG levels in plasma and B lymphocytes. In vitro, the results showed that the release of IgG-Gd-IgA1-IC was significantly decreased in peripheral blood mononuclear cells (PBMCs) cultured with SA-IgG from both IgAN patients and healthy controls. The proliferation ability and the release of IL-6, TNF-α, and TGF-β in human mesangial cells (HMCs) were measured after stimulating with SA-IgG-IgA1-IC and NSA-IgG-IgA1-IC. The mesangial cell proliferation levels induced by NSA-IgG-IgA1-IC derived from IgAN patients were significantly higher than those caused by SA-IgG-IgA1-IC derived from IgAN patients and healthy controls. Compared with NSA-IgG-IgA1 from healthy controls, IgAN-NSA-IgG-IgA1 could significantly upregulate the expression of IL-6 and TNF-α in mesangial cells. The data showed that there weren’t any significant differences in the levels of IL-6, TNF-α, and TGF-β when treated with IgAN-SA-IgG-IgA1 and HC-NSA-IgG-IgA1.ConclusionsThe present study demonstrated that the sialylation of IgG increased in patients with IgA nephropathy. It exerted an inhibitory effect on the formation of Gd-IgA1-containing immune complexes in PBMCs and the proliferation and inflammation activation in mesangial cells.

Targeting Mechanosensitive Piezo1 Alleviated Renal Fibrosis Through p38MAPK-YAP Pathway.

Renal fibrosis is the most common pathological manifestation of a wide variety of chronic kidney disease. Increased extracellular matrix (ECM) secretion and enhanced microenvironment stiffening aggravate the progression of renal fibrosis. However, the related mechanisms remain unclear. Here, we evaluated the mechanism by which ECM stiffness aggravates renal fibrosis. In the present study, renal mesangial cells (MCs) were cultured on polyacrylamide hydrogels with different stiffness accurately detected by atomic force microscope (AFM), simulating the in vivo growth microenvironment of MCs in normal kidney and renal fibrosis. A series of in vitro knockdown and activation experiments were performed to establish the signaling pathway responsible for mechanics-induced MCs activation. In addition, an animal model of renal fibrosis was established in mice induced by unilateral ureteral obstruction (UUO). Lentiviral particles containing short hairpin RNA (sh RNA) targeting Piezo1 were used to explore the effect of Piezo1 knockdown on matrix stiffness-induced MCs activation and UUO-induced renal fibrosis. An in vitro experiment demonstrated that elevated ECM stiffness triggered the activation of Piezo1, which increased YAP nuclear translocation through the p38MAPK, and consequently led to increased ECM secretion. Furthermore, these consequences have been verified in the animal model of renal fibrosis induced by UUO and Piezo1 knockdown could alleviate UUO-induced fibrosis and improve renal function in vivo. Collectively, our results for the first time demonstrate enhanced matrix stiffness aggravates the progression of renal fibrosis through the Piezo1-p38MAPK-YAP pathway. Targeting mechanosensitive Piezo1 might be a potential therapeutic strategy for delaying the progression of renal fibrosis.

Mineralocorticoid receptor antagonists in diabetic kidney disease - mechanistic and therapeutic effects.

Chronic kidney disease (CKD) is the leading complication in type 2 diabetes (T2D) and current therapies that limit CKD progression and the development of cardiovascular disease (CVD) include angiotensin-converting enzyme inhibitors, angiotensin II receptor blockers and sodium-glucose co-transporter 2 (SGLT2) inhibitors. Despite the introduction of these therapeutics, an important residual risk of CKD progression and cardiovascular death remains in patients with T2D. Mineralocorticoid receptor antagonists (MRAs) are a promising therapeutic option in diabetic kidney disease (DKD) owing to the reported effects of mineralocorticoid receptor activation in inflammatory cells, podocytes, fibroblasts, mesangial cells and vascular cells. In preclinical studies, MRAs consistently reduce albuminuria, CKD progression, and activation of fibrotic and inflammatory pathways. DKD clinical studies have similarly demonstrated that steroidal MRAs lead to albuminuria reduction compared with placebo, although hyperkalaemia is a major secondary effect. Non-steroidal MRAs carry a lower risk of hyperkalaemia than steroidal MRAs, and the large FIDELIO-DKD clinical trial showed that the non-steroidal MRA finerenone also slowed CKD progression and reduced the risk of adverse cardiovascular outcomes compared with placebo in patients with T2D. Encouragingly, other non-steroidal MRAs have anti-albuminuric properties in DKD. Whether or not combining MRAs with other renoprotective drugs such as SGLT2 inhibitors might provide additive protective effects warrants further investigation.

Mesangial-Cell Activation by Circulating Immune Complexes Consisting of Galactose-Deficient IgA1 and IgG Autoantibodies from Patients with IgA Nephropathy

Mesangial-Cell Activation by Circulating Immune Complexes Consisting of Galactose-Deficient IgA1 and IgG Autoantibodies from Patients with IgA Nephropathy

An Interplay of Glucose, IL-1β, and PDGF-B Trigger cPLA2 Activation, Prostaglandin Secretion, and Proliferation in Human Mesangial Cells

An Interplay of Glucose, IL-1β, and PDGF-B Trigger cPLA2 Activation, Prostaglandin Secretion, and Proliferation in Human Mesangial Cells

Calcium‐Dependent Dual Oxidase 2 is a Novel Source of Reactive Oxygen Species Implicated in Glomerular Mesangial Cell Fibrotic Response to Angiotensin II

The vasoactive peptide angiotensin II (Ang II) contributes to the initiation and the progression of glomerular fibrosis via activation of glomerular mesangial cells (MCs) and subsequent extracellular matrix expansion. We have previously shown that oxidative stress is critical for MC fibrotic response to Ang II. Here, we demonstrate that Dual oxidase 2 (Duox2), a member of the Nox/Duox family of NADPH oxidases, is present in MCs and that its protein expression is upregulated by Ang II. Small interfering RNA (siRNA)-mediated downregulation of Duox2 significantly reduces Ang II-induced increase in reactive oxygen (ROS) generation and prevents the stimulatory effect of Ang II on MC fibrotic injury (as assessed by measuring a-smooth muscle actin and fibronectin expression). To demonstrate that Duox2 activation is Ca2+-dependent, we show that the extracellular Ca2+ chelator BAPTA prevented Ang II-induced ROS generation and the stimulation of MC fibrotic injury by Ang II. Moreover, treatment of MCs with ionomycin resulted in increased ROS production and enhanced MC fibrotic injury. These effects were abrogated by siRNA targeting Duox2. Fura-2 fluorescence was utilized as well to show calcium mobilization in response to Ang II and this effect was abrogated with siDuox2 transfection. These data indicate that Ang II-stimulated Duox2 activation and ROS generation subsequently lead to MC fibrotic injury. In summary, we have identified a novel role for Duox2 as a major source of ROS in response to Ang II and established the significance of Duox2 in Ang II-mediated MC activation and fibrotic injury. Therapeutic targeting of this pathway may prevent or reverse pathophysiologic manifestations of renal fibrotic diseases.

POS-370 HUMAN RENAL MESANGIAL CELL ACTIVATION INDUCED BY ENDOTHELIN-1 OR IGA NEPHROPATHY PATIENT-DERIVED IMMUNE COMPLEXES IS BLOCKED BY SELECTIVE ETA ANTAGONIST ATRASENTAN

Mesangial cell activation in response to deposition of pathogenic IgA-containing immune complexes is characterized by cellular proliferation and inflammatory cytokine secretion, and is considered the initiating intra-renal event in the pathogenesis of IgA nephropathy (IgAN). The molecular mechanisms have not been well defined. Endothelin (ET) pathway activation has been observed in kidney biopsies of IgAN patients and may be an important driver of disease progression by promoting proteinuria along with kidney inflammation and fibrosis via ETA receptor activation.

Activin A and Cell-Surface GRP78 Are Novel Targetable RhoA Activators for Diabetic Kidney Disease.

Diabetic kidney disease (DKD) is the leading cause of kidney failure. RhoA/Rho-associated protein kinase (ROCK) signaling is a recognized mediator of its pathogenesis, largely through mediating the profibrotic response. While RhoA activation is not feasible due to the central role it plays in normal physiology, ROCK inhibition has been found to be effective in attenuating DKD in preclinical models. However, this has not been evaluated in clinical studies as of yet. Alternate means of inhibiting RhoA/ROCK signaling involve the identification of disease-specific activators. This report presents evidence showing the activation of RhoA/ROCK signaling both in vitro in glomerular mesangial cells and in vivo in diabetic kidneys by two recently described novel pathogenic mediators of fibrosis in DKD, activins and cell-surface GRP78. Neither are present in normal kidneys. Activin inhibition with follistatin and neutralization of cell-surface GRP78 using a specific antibody blocked RhoA activation in mesangial cells and in diabetic kidneys. These data identify two novel RhoA/ROCK activators in diabetic kidneys that can be evaluated for their efficacy in inhibiting the progression of DKD.

Glomerular Endothelial Cells as Instigators of Glomerular Sclerotic Diseases.

Glomerular endothelial cell (GEnC) dysfunction is important in the pathogenesis of glomerular sclerotic diseases, including Focal Segmental Glomerulosclerosis (FSGS) and overt diabetic nephropathy (DN). GEnCs form the first cellular barrier in direct contact with cells and factors circulating in the blood. Disturbances in these circulating factors can induce GEnC dysfunction. GEnC dysfunction occurs in early stages of FSGS and DN, and is characterized by a compromised endothelial glycocalyx, an inflammatory phenotype, mitochondrial damage and oxidative stress, aberrant cell signaling, and endothelial-to-mesenchymal transition (EndMT). GEnCs are in an interdependent relationship with podocytes and mesangial cells, which involves bidirectional cross-talk via intercellular signaling. Given that GEnC behavior directly influences podocyte function, it is conceivable that GEnC dysfunction may culminate in podocyte damage, proteinuria, subsequent mesangial activation, and ultimately glomerulosclerosis. Indeed, GEnC dysfunction is sufficient to cause podocyte injury, proteinuria and activation of mesangial cells. Aberrant gene expression patterns largely contribute to GEnC dysfunction and epigenetic changes seem to be involved in causing aberrant transcription. This review summarizes literature that uncovers the importance of cross-talk between GEnCs and podocytes, and GEnCs and mesangial cells in the context of the development of FSGS and DN, and the potential use of GEnCs as efficacious cellular target to pharmacologically halt development and progression of DN and FSGS.

ICAT acts as a Coactivator in Regulating PPARγ Transcriptional Activity in Mesangial Cells.

Our study aims to explore the role of β-catenin interaction protein-1(ICAT) in regulating peroxisome proliferator-activated receptor γ (PPARγ) transcriptional activity in mesangial cells. The abnormal ICAT expression in mesangial cells under high glucose(HG) contributes to the development of diabetes and its complications such as diabetic nephropathy (DN). Human mesangial cells (HMCs) were cultured in either 5.5 (normal control) or 30 (high glucose) mmol/L glucose medium. Overexpression and knock-down of ICAT or β-catenin were carried out by transient transfection. PPARγ transcriptional activity was evaluated by luciferase assay. Protein-protein interactions were tested by Coimmunoprecipitation and GST-pull down assay. Cell phenotype transition of HMCs was detected by the expression level of α-SMA and fibronectin, as well as MTT assay. High β-catenin protein expression but low ICAT was accompanied by low PPARγ transcriptional activity in HMCs cultured in HG. By using bioinformatics prediction, protein-protein and protein-DNA interaction experimental methods, ICAT and β-catenin were confirmed to act as coactivators in regulating PPARγ transcriptional activity. Overexpression of ICAT could mitigate the decrease of PPARγ transcriptional activity and partly relieve cell phenotype transition in HMCs. β-catenin and ICAT interact as coactivator to modulate PPARγ transcriptional activation. In HMCs cultured in HG, the low expression of ICAT leads to low PPARγ transcriptional activation.

Icariin Prevents Extracellular Matrix Accumulation and Ameliorates Experimental Diabetic Kidney Disease by Inhibiting Oxidative Stress via GPER Mediated p62-Dependent Keap1 Degradation and Nrf2 Activation.

The present study aimed to determine whether icariin could attenuate type 1 diabetic nephropathy (T1DN) induced by streptozotocin (STZ) after 4 weeks or not. Therefore, its therapeutic effect on diabetic kidney disease was investigated in view of reactive oxygen (ROS) and extracellular matrix (ECM) generation in human glomerular mesangial cells under high glucose. To establish the participation and the key role of GPER and Nrf2 in ECM deposition, a combination of G15 (antagonist of GPER) or siGPER and siNrf2 were performed, respectively. The results showed that T1DN can be significantly inhibited by oral icariin, evidenced by improvement of 24 h urinary volume, 24 h proteinuria, microalbuminuria, and histopathological changes of kidney. Icariin decreased the levels of intracellular superoxide anion, impeded the generation of fibronectin and increased the expression and activity of antioxidant enzymes in the human glomerular mesangial cells treated with high glucose. It acted as a GPER activator, increased dissociation of Nrf2/Keap1 complexes, combination of Keap1/p62 complexes, Nrf2 translocation to nuclear, Nrf2/ARE DNA binding activity, and ARE luciferase reporter gene activity in glomerular mesangial cells. The Nrf2 inhibitor ML385 or siNrf2 obviously abolished extracellular matrix (ECM) generation inhibited by icariin. Furthermore, icariin-induced Nrf2 activation was mainly dependent on p62-mediated Keap1 degradation, which functions as an adaptor protein during autophagy. The GPER antagonist G15 and siGPER obviously abolished the above effects by icariin. Taken together, the present study demonstrated that the therapeutic effects of icariin on type 1 diabetic nephropathy in rats via GPER mediated p62-dependent Keap1 degradation and Nrf2 activation.

P0174NEPHROPROTECTIVE ACTIVITY OF GOSSYPETIN THROUGH INHIBITORY EFFECT ON XANTHINE OXIDASE, NUCLEAR FACTOR KAPPA B (NF-KB) AND SOLUBLE EPOXIDE HYDROLASE (SEH): IN-VITRO AND IN-SILICO EXPERIMENT

Abstract Background and Aims Oxidative stress and inflammation is the major contributor of kidney injury and the drugs which have antioxidant and anti-inflammatory activity could protect kidney against renal damage. Mechanisms involved in renal failure include oxidative stress, inflammation and apoptosis which lead myoglobinemia, myoglobinuria and cast formation. Inflammatory mediators such as IL-1, ICAM-1 and TNFα also play important role in renal failure. Excess production of uric acid can cause serious consequence in hyperuricemia and xanthine oxidase (XO) catalyzes the oxidation of xanthine to uric acid. Protective effects of gossypetin in the management of kidney injury and related disorders have been investigated in the present work through inhibitory potential of gossypetin on nuclear factor kappa B (NF-κB), soluble epoxide hydrolase (sEH) and XO. Method Present work described the medicinal importance of gossypetin with their beneficial effect on kidney disorder. In-silico molecular docking and dynamic experiments were carried out with gossypetin against nuclear factor kappa B (NF-κB) and soluble epoxide hydrolase (sEH). Further docking was also performed to investigate how gossypetin and the active site of XO fit together. Results From the analysis of the available data’s in the present work, it was found that gossypetin have protective effect against nephrotoxicity. Gossypetin also showed potent anti-inflammatory activity in kidney mesangial cells which further support application of natural compounds on nephritis treatment. Importance of gossypetin for preventing renal damage has been also emphasized due to its antioxidants potential. In-silico studies showed that, gossypetin exhibited a higher docking score against NF-κB and sEH. Docking studies revealed gossypetin surrounds the active sites of XO and reduces conversion of xanthine to uric acid. Conclusion Study revealed their antioxidant, anti-mutagenic, anti-atherosclerotic, anti-microbial and cytoprotective properties. The protective effect of gossypetin in kidney could be due to its antioxidant and anti-inflammatory activity through inhibition of NF-κB and sEH upregulation.

β-Caryophyllene inhibits high glucose-induced oxidative stress, inflammation and extracellular matrix accumulation in mesangial cells

β-Caryophyllene (BCP) is a bicyclic sesquiterpene compound that has anti-diabetic activity. However, the effect of BCP on diabetic nephropathy (DN) remains unclear. Here, we aimed to evaluate the potential role of BCP in high glucose (HG)-induced glomerular mesangial cells (MCs). MCs were maintained under HG condition to simulate DN in vitro. Our results showed that BCP inhibited HG-induced cell proliferation, ROS production and NADPH oxidase (NOX) 2/4 expression. BCP exhibited anti-inflammatory activity with decreased levels of TNF-α, IL-1β, IL-6 in HG-induced MCs. Moreover, BCP treatment suppressed the HG-induced secretion of fibronectin (FN) and collagen IV (Col IV) in MCs. Furthermore, BCP suppressed the NF-κB activation and enhanced the Nrf2 activation in HG-induced MCs. However, inhibition of Nrf2 attenuated the protective effects of BCP on HG-induced MCs, while inhibition of NF-κB enhanced the nephro-protective effects of BCP on MCs. In conclusion, these findings demonstrated that BCP executed protective effects on HG-induced MCs via regulating NF-κB and Nrf2 signaling pathways.

Calcium‐Dependent Dual Oxidase 2 is a Novel Source of Reactive Oxygen Species Implicated in Glomerular Mesangial Cell Fibrotic Response to Angiotensin II

The vasoactive peptide angiotensin II (Ang II) contributes to the initiation and the progression of glomerular fibrosis via activation of glomerular mesangial cells (MCs) and subsequent extracellular matrix expansion. We have previously shown that oxidative stress is critical for MC fibrotic response to Ang II. Here, we demonstrate that Dual oxidase 2 (Duox2), a member of the Nox/Duox family of NADPH oxidases, is present in MCs and that its protein expression is upregulated by Ang II.Small interfering RNA (siRNA)‐mediated downregulation of Duox2 significantly reduces Ang II‐induced increase in reactive oxygen (ROS) generation and prevents the stimulatory effect of Ang II on MC fibrotic injury (as assessed by measuring α‐smooth muscle actin and fibronectin expression).To demonstrate that Duox2 activation is Ca2+‐dependent, we show that the extracellular Ca2+ chelator BAPTA prevented Ang II‐induced ROS generation and the stimulation of MC fibrotic injury by Ang II. Moreover, treatment of MCs with ionomycin resulted in increased ROS production and enhanced MC fibrotic injury. These effects were abrogated by siRNA targeting Duox2. Fura‐2 fluorescence was utilized as well to show calcium mobilization in response to Ang II and this effect was abrogated with siDuox2 transfection. These data indicate that Ang II‐stimulated Duox2 activation and ROS generation subsequently lead to MC fibrotic injury.In summary, we have identified a novel role for Duox2 as a major source of ROS in response to Ang II and established the significance of Duox2 in Ang II‐mediated MC activation and fibrotic injury. Therapeutic targeting of this pathway may prevent or reverse pathophysiologic manifestations of renal fibrotic diseases.Support or Funding InformationThis project was funded by the University of the Incarnate Word, Office of Research and Graduate Studies and the following grants: NRSA 5T32HL007446‐32

Ampelopsin inhibits high glucose-induced extracellular matrix accumulation and oxidative stress in mesangial cells through activating the Nrf2/HO-1 pathway.

Oxidative stress plays an important role in diabetic nephropathy (DN), which is a diabetic complication. Ampelopsin (AMP) is a natural flavonoid that has been found to possess antidiabetic and antioxidative activities. However, the effect of AMP on DN remains unclear. In this study, we aimed to evaluate the protective effect of AMP on glomerular mesangial cells (MCs) exposed to high glucose (HG). We found that AMP improved HG-caused cell viability reduction in MCs. AMP significantly suppressed the intracellular ROS production and expression levels of ROS producing enzymes NADPH oxidase 2 (NOX2) and NOX4. Increased of NOX activity in HG-stimulated MCs was suppressed by AMP. Pretreatment with AMP inhibited extracellular matrix (ECM) accumulation in HG-stimulated MCs with decreased expression levels of fibronectin (FN) and collagen type IV (Col IV). Furthermore, AMP elevated the expression levels of nuclear Nrf2 and heme oxygenase-1 (HO-1), as well as increased the mRNA levels of Nrf2-driven genes NAD(P)H dehydrogenase quinone-1 (NQO-1) and HO-1 in HG-treated MCs. Knockdown of Nrf2 reversed the protective effects of AMP against HG-induced oxidative stress and EMC accumulation in MCs. In conclusion, these findings indicated that AMP protected MCs from HG-induced oxidative damage and ECM accumulation, which might be mediated by Nrf2/HO-1 pathway.

SUN-164 CXCR4-AngII axis plays an important role in glomerular injury through promoting the crosstalk between podocytes and mesangial cells

Glomerular disease has a high incidence worldwide. Podocyte injury and mesangial cell activation play pivotal roles in the progression of proteinuria and glomerulosclerosis. However, the linkage and underlying mechanisms are rarely reported. Herein, we investigated the role of chemokine receptor (CXCR4) and angiotensin II (AngII) in the pathogenesis of podocyte and mesangial cell injury. The association of CXCR4-AngII axis in crosstalk between podocytes and mesangial cells was also testified. 5/6 nephrectomized mice was established. To identify the role of CXCR4, CXCR4 plasmid was injected once a week by tail injection. Some mice were injected with adriamycin (ADR), advanced oxidation protein products (AOPPs) or Losartan. In vitro, MPC5 cells, an immortalized mouse podocyte cell line, were incubated with AOPPs or transfected with CXCR4 expression plasmid or siRNA to CXCR4. RMC cells, an immortalized mesangial cell line, were also adopted. RMC were treated with the conditional medium from AOPPs-treated MPC5 cells to assess the crosstalk between podocytes and mesangial cells. In 5/6NX and ADR mice model, CXCR4 was upregulated predominantly in podocytes. Whereas, SDF-1α, the ligand of CXCR4, was highly expressed in mesangial cells. CXCR4 colocalized with AngII in podocytes. Administration of AOPPs significantly induced the upregulation of CXCR4 and accelerated the progression of proteinuria. AOPPs induced podocyte injury and mesangial cell proliferation. These glomerular lesions were accompanied by upregulation of CXCR4, SDF-1α and activation of ACE, Renin and AT1. In ADR model, ectopic expression of CXCR4 aggravated proteinuria, podocyte injury, mesangial cell proliferation and glomerulosclerosis. Treatment with losartan significantly ameliorated these pathological changes. In cultured MPC5 cells, incubation of AOPPs or overexpression of CXCR4 promoted the activation of ACE, Renin, and AT1. However, blockade of CXCR4 reduced these effects. In RMC cells, administration of AngII stimulated fibrogenesis and cell proliferation. The conditional medium from AOPPs-treated MPC5 cells induced upregulation of fibronectin, PCNA and angiotensin II receptor AT1, but blocked by incubation with losartan. Our studies indicate that CXCR4 could upregulate the expression of RAS system in podocyte. AngII may subsequently bind to its receptor AT1 in mesangial cells to induce matrix deposition. The activation of mesangial cells would lead to the secretion of SDF-1α, the ligand of CXCR4, which further triggers podocyte injury. These studies demonstrate that CXCR4-AngII axis plays a vital role in glomerular injury via mediating pathologic crosstalk between podocytes and mesangial cells. Our findings uncover a new pathogenic mechanism by which CXCR4-AngII axis promotes glomerulosclerosis.

Farrerol alleviates high glucose-induced renal mesangial cell injury through the ROS/Nox4/ERK1/2 pathway

Hyperproliferation and oxidative stress induced by hyperglycemia in mesangial cells plays crucial roles in the pathological process of diabetic nephropathy. Farrerol, isolated from rhododendron leaves, possesses broad anti-oxidative and anti-inflammatory properties towards several diseases, but its role in diabetic neuropathy remains unclear. The aim of this study was to evaluate the effects of farrerol in high glucose induced mesangial cell injury, and to explore underlying molecular mechanisms. Our results showed that high glucose in vitro conditions significantly stimulated cell proliferation, inflammatory cytokine secretion, extracellular matrix deposition, excessive oxidative stress, and NADPH oxidase activity in mesangial cells. Levels of NADPH oxidase 4 (Nox4) expression, ERK1/2 phosphorylation, and TGF-β1/Smad2 activation were significantly induced by high glucose conditions in mesangial cells. Inversely, farrerol treatments at 40, 60, and 80 μM concentrations, dose-dependently alleviated this molecular damage by high glucose in mesangial cells. We also found that restoration of Nox4 expression abolished the protective effects of farrerol on high glucose-induced proliferation and reactive oxygen species generation. Furthermore, pretreatment with the Nox4 inhibitor diphenyliodonium or the ERK1/2 pathway inhibitor PD98059, displayed similar ameliorated effects of farrerol on high glucose-induced mesangial cell damage. Taken together, these data suggest that farrerol displays protective effects on high glucose induced mesangial cell injury, partly through the Nox4-mediated ROS/ERK1/2 signaling pathway. These observations may provide novel insights into the application of farrerol as a diabetic neuropathy treatment.

Sonic hedgehog connects podocyte injury to mesangial activation and glomerulosclerosis.

Glomerular disease is characterized by proteinuria and glomerulosclerosis, two pathologic features caused by podocyte injury and mesangial cell activation, respectively. However, whether these two events are linked remains elusive. Here, we report that sonic hedgehog (Shh) is the mediator that connects podocyte damage to mesangial activation and glomerulosclerosis. Shh was induced in glomerular podocytes in various models of proteinuric chronic kidney diseases (CKD). However, mesangial cells in the glomeruli, but not podocytes, responded to hedgehog ligand. In vitro, Shh was induced in podocytes after injury and selectively promoted mesangial cell activation and proliferation. In a miniorgan culture of isolated glomeruli, Shh promoted mesangial activation but did not affect the integrity of podocytes. Podocyte-specific ablation of Shh in vivo exhibited no effect on proteinuria after adriamycin injection but hampered mesangial activation and glomerulosclerosis. Consistently, pharmacologic blockade of Shh signaling decoupled proteinuria from glomerulosclerosis. In humans, Shh was upregulated in glomerular podocytes in patients with CKD and its circulating level was associated with glomerulosclerosis but not proteinuria. These studies demonstrate that Shh mechanistically links podocyte injury to mesangial activation in the pathogenesis of glomerular diseases. Our findings also illustrate a crucial role for podocyte-mesangial communication in connecting proteinuria to glomerulosclerosis.

Microcystin exposure worsens nonalcoholic fatty liver disease associated ectopic glomerular toxicity via NOX-2-MIR21 axis

NAFLD often results in cardiovascular, intestinal and renal complications. Previous reports from our laboratory highlighted NAFLD induced ectopic inflammatory manifestations in the kidney that gave rise to glomerular inflammation. Extending our studies, we hypothesized that existing inflammatory conditions in NAFLD could make the kidneys more susceptible to environmental toxicity. Our results showed that exposure of Microcystin-LR (MC) in NAFLD mice caused a marked increase in cellular scarring with a concomitant increase in mesangial cell activation as observed by increased α-SMA in the extracellular matrix surrounding the glomeruli. Renal tissue surrounding the glomeruli also showed increased NOX2 activation as shown by greater co-localization of p47 Phox and its membrane component gp91Phox both in the mesangial cell and surrounding tissue. Mechanistically, mesangial cells incubated with apocynin, nitrone spin trap DMPO and miR21 inhibitor showed significantly decreased α-SMA, miR21 levels and proinflammatory cytokine release in the supernatant. In parallel, mice lacking miR21, known to be activated by NOX2, when exposed to MC in NAFLD showed decreased mesangial cell activation. Strikingly, phenyl boronic acid incubated cells that were exposed to MC showed significantly decreased mesangial cell activation showing that peroxynitrite might be the major reactive species involved in mediation of the activation process, release of proinflammatory micro RNAs and cytokines that are crucial for renal toxicity. Thus, in conclusion, MC exposure causes NOX2 activation that leads to mesangial cell activation and toxicity via release of peroxynitrite that also represses PTEN by the upregulation of miR21 thus amplifying the toxicity.

MiR-29b expression is altered in crescent formation of HSPN and accelerates Ang II-induced mesangial cell activation

MicroRNA-29b (miR-29b) has been suggested to possess pro-inflammatory activity, which can partially be explained by the repression of tumor necrosis factor alpha protein three antibody (TNFAIP3). Meanwhile, it also promotes thyroid cell proliferation via Smad signaling pathways. The present study aimed to elucidate the role of miR-29b in Henoch Schönlein purpura nephritis (HSPN) and its underlying molecular mechanism in angiotensin II (Ang II)-induced human glomerular mesangial cell (HGMC) activation. We evaluated miR-29b expression in 35 HSPN renal tissues based on crescent formation, glomerular sclerosis, interstitial fibrosis, thrombosis formation and capillary loop necrosis. Meanwhile, HGMCs were cultured, treated with Ang II and then transfected with LV-hsa-miR-29b-1 to induce miR-29b overexpression or LV-hsa-miR-29b-3p-inhibition to inhibit miR-29b expression. Finally, we examined the effects of miR-29b on cell proliferation and release of inflammatory mediators. We observed that miR-29b expression was significantly higher in the crescent group than in the no crescent group. MiR-29b overexpression induced the release of intercellular adhesion molecule-1, interleukin-1β (IL-1β), IL-6, IL-8, the increase of CyclinA2, CyclinD1, and cell proliferation. It also could inhibit the expressions of TNFAIP3 and NF-kappa-B-repressing factor (NKRF). Correspondingly, miR-29b inhibition produced the opposite effects and increased the expression of TNFAIP3 and NKRF. MiR-29b expression is altered in crescent formation of HSPN and accelerates Ang II-induced mesangial cell proliferation and release of inflammatory mediators.