Mesangial Cells Research Articles

The IRE1/JNK signaling pathway regulates inflammation cytokines and production of glomerular extracellular matrix in the acute kidney injury to chronic kidney disease transition.

The transition from acute kidney injury (AKI) to chronic kidney disease (CKD) is extremely complex. Incomplete renal tubule repair, inflammation, and endoplasmic reticulum (ER) stress all play major roles. AKI activates ER stress, and the sensor protein inositol-requiring kinase-1 (IRE1) mediates inflammation by promoting the phosphorylation of C-jun NH2-terminal kinase (JNK). The interleukin-6/signal transducer and activator of transcription 3 (IL-6/STAT3) signaling pathway is associated with the secretion of renal extracellular matrix (ECM) and fibrosis. It remains unclear whether these signaling pathways play a role in the AKI-CKD transition. In this study, a mouse model of ischemia-reperfusion (I/R) with bilateral renal artery clipping was used. IRE1 or JNK inhibitors were also injected to confirm their roles in the AKI-CKD transition. The renal function of the mice was determined by observing the pathology of the renal tubules and glomeruli through electron microscopy, immunohistochemistry, western blotting and quantitative real-time PCR. I/R stimulates ER stress and the IRE1/JNK pathway in the renal tubules in a short period of time, leading to continuous inflammation. Long-term I/R injury activates the STAT3 pathway in the glomeruli, activates mesangial cells proliferation, causes secretion of large amounts of glomerular ECM, and promotes glomerular sclerosis. This damage to the renal tubules and glomeruli is significantly reduced in I/R model mice pretreated with IRE1 or JNK inhibitors. These findings suggested that the IRE1/JNK pathway regulates the inflammatory cytokines caused by AKI and continues to activate the STAT3 pathway and production of ECM in the glomeruli at late stages, suggesting the feasibility of targeted therapy for the AKI-CKD transition.

MicroRNA-339-5p inhibits lipopolysaccharide-induced rat mesangial cells by regulating the Syk/Ras/c-Fos pathway.

Chronic glomerulonephritis (CGN) is a disease occurred in glomeruli. The mechanism of CGN is regarded to be involved in a range of inflammatory responses. MicroRNA-339-5p (miR-339-5p) has been reported to be involved in inflammatory responses in many diseases. However, the role of miR-339-5p in CGN remains unclear. The purpose of this study was to investigate the role of miR-339-5p in lipopolysaccharide (LPS)-induced nephritis injury in vitro. The real-time reverse transcriptase-polymerase chain reaction (RT-qPCR) and western blot (WB) were used to detect the expression of miR-339-5p and Syk/Ras/c-Fos pathway. Double luciferase was performed to identify targeted binding of miR-339-5p to Syk. Cell counting kit-8 (CCK-8) and flow cytometry were used to observe cell viability and cell cycle. Enzyme-linked immunosorbent assay (ELISA) was performed to measure the concentrations of inflammatory cytokines IL-1β, IL-10, IL-6, and TNF-α. Lipopolysaccharide (LPS) could increase HBZY-1 (rat mesangial cells) cell viability, decrease the G2 phase, and promote cell proliferation and accelerate inflammatory cytokine. However, overexpression of miR-339-5p could inhibit LPS-induced HBZY-1 cell viability, decrease the expression of Syk/Ras/c-Fos signaling pathway, downregulate the expression level of inflammatory cytokines, increase the G2 phase, and inhibit cell proliferation. miR-339-5p could inhibit the proliferation and inflammation of the rat mesangial cells through regulating Syk/Ras/c-Fos signaling pathway.

Axl Expression in Renal Mesangial Cells Is Regulated by Sp1, Ap1, MZF1, and Ep300, and the IL-6/miR-34a Pathway.

Axl receptor tyrosine kinase expression in the kidney contributes to a variety of inflammatory renal disease by promoting glomerular proliferation. Axl expression in the kidney is negligible in healthy individuals but upregulated under inflammatory conditions. Little is known about Axl transcriptional regulation. We analyzed the 4.4 kb mouse Axl promoter region and found that many transcription factor (TF)-binding sites and regulatory elements are located within a 600 bp fragment proximal to the translation start site. Among four TFs (Sp1, Ap1, MZF1, and Ep300) identified, Sp1 was the most potent TF that promotes Axl expression. Luciferase assays confirmed the siRNA results and revealed additional mechanisms that regulate Axl expression, including sequences encoding a 5′-UTR mini-intron and potential G-quadruplex forming regions. Deletion of the Axl 5′-UTR mini-intron resulted in a 3.2-fold increases in luciferase activity over the full-length UTR (4.4 kb Axl construct). The addition of TMPyP4, a G-quadruplex stabilizer, resulted in a significantly decreased luciferase activity. Further analysis of the mouse Axl 3′-UTR revealed a miRNA-34a binding site, which inversely regulates Axl expression. The inhibitory role of miRNA-34a in Axl expression was demonstrated in mesangial cells using miRNA-34a mimicry and in primary kidney cells with IL-6 stimulated STAT3 activation. Taken together, Axl expression in mouse kidney is synergistically regulated by multiple factors, including TFs and secondary structures, such as mini-intron and G-quadruplex. A unique IL6/STAT3/miRNA-34a pathway was revealed to be critical in inflammatory renal Axl expression.

Circ_0000064 knockdown attenuates high glucose-induced proliferation, inflammation and extracellular matrix deposition of mesangial cells through miR-424-5p-mediated WNT2B inhibition in cell models of diabetic nephropathy.

Circular RNA (circRNA) is widely shown to be associated with the development of diabetic nephropathy (DN). Our study aimed to further explore the role of circ_0000064 and provide a new mechanism for its action in DN. Cell models of DN in vitro were constructed by treating human renal mesangial cells (HRMCs) with high glucose (HG). The expression of circ_0000064, microRNA-424-5p (miR-424-5p) and Wnt family member 2B (WNT2B) mRNA was detected by quantitative real-time PCR (qPCR). Cell proliferation was assessed by CCK-8 assay and EdU assay. Cell cycle was characterized by DNA content using flow cytometry. The releases of pro-inflammatory factors were checked using commercial ELISA kits. The expression of cell cycle- and fibrosis-associated proteins was detected by western blot. The interplays between miR-424-5p and circ_0000064 or WNT2B were verified by dual-luciferase reporter assay and RIP assay. Circ_0000064 and WNT2B were upregulated, while miR-424-5p was downregulated in HG-treated HRMCs. Circ_0000064 knockdown largely attenuated HG-induced proliferation, inflammatory responses and extracellular matrix (ECM) accumulation in HRMCs, and miR-424-5p deficiency reversed the role of circ_0000064 knockdown. MiR-424-5p was a target of circ_0000064, and miR-424-5p directly bound to WNT2B. MiR-424-5p restoration alleviated HG-induced proliferation, inflammatory responses and ECM accumulation in HRMCs, and WNT2B overexpression partially abolished the effects of miR-424-5p. Circ_0000064 knockdown ameliorated HG-induced HRMC dysfunctions through miR-424-5p enrichment-mediated WNT2B inhibition, hinting that circ_0000064 contributed to DN development.

LncRNA NEAT1/microRNA-124 regulates cell viability, inflammation and fibrosis in high-glucose-treated mesangial cells.

Long non-coding RNA (lncRNA) nuclear enriched abundant transcript 1 (NEAT1) has been frequently found to be dysregulated, which contributes to diabetes-related complications. The present study aimed to explore the effect of knockdown on mouse mesangial cell (MMC) viability, apoptosis, inflammation and fibrosis in an in vitro model of diabetic nephropathy (DN). The SV40 MES13 MMC cell line was first cultured with high glucose to establish an in vitro MMC DN cell model. Lnc-NEAT1 shRNA or the negative control shRNA were transfected into MMC DN cells, followed by the measurement of cell viability, apoptosis, inflammation, fibrosis and microRNA (miR)-124 expression, a known target of lnc-NEAT1, using Cell Counting Kit-8, flow cytometry, ELISA, western blotting [Capain1 (capn1), β-catenin (CTNNB1), cleaved caspase 3, cleaved poly-(ADP ribose) polymerase, fibronectin and Collagen] and reverse transcription-quantitative PCR (Capn1, CTNNB1, lnc-NEAT1, fibronectin, collagen and miR-124), respectively. In rescue experiments, the miR-124 and negative control inhibitor were co-transfected into lnc-NEAT1-downregulated cells, following which cell viability, apoptosis, inflammation, fibrosis, capn1 and CTNNB1 expression were measured. Lnc-NEAT1 expression was increased in high glucose-treated cells compared with that in normal glucose-treated cells and osmotic control cells, suggesting that lnc-NEAT1 is overexpressed in the MMC DN cell model. In the MMC DN cell model, lncRNA-NEAT1 knockdown enhanced cell apoptosis but reduced cell viability and the secretion of inflammatory cytokines in the supernatant (IL-1β, IL-8, monocyte chemotactic protein 1 and TNF-α), in addition to reducing the expression of fibrosis markers fibronectin and collagen I in the lysates. Lnc-NEAT1 knockdown increased miR-124 expression. Furthermore, transfection with the miR-124 inhibitor reduced cell apoptosis but increased cell viability, inflammation and fibrosis in lnc-NEAT1-downregulated MMC DN cells. miR-124 inhibitor transfection also increased the expression levels of Capn1 and CTNNB1. Taken together, the findings of the present study demonstrated that lnc-NEAT1 knockdown was able to attenuate MMC viability, inflammation and fibrosis by regulating miR-124 expression and the Capn1/β-catenin signaling pathway downstream. Therefore, Lnc-NEAT1 may serve as a potential therapeutic target for DN.

Procyanidin B2 suppresses hyperglycemia-induced renal mesangial cell dysfunction by modulating CAV-1-dependent signaling.

The dysfunction of renal mesangial cells (MCs) is a hallmark of diabetic kidney disease (DKD), which triggers glomerulosclerosis leading to end-stage renal disease. Procyanidin B2 (PB2), the main component of proanthocyanidin, is well known for its antioxidant and anti-inflammatory effects; however, it remains unclear as to whether it has protective effects on DKD. The present study investigated the protective effect of PB2 against hyperglycemia-induced renal MC dysfunction in mouse SV40-Mes13 (Mes13) cells. The Mes13 cells were treated with or without PB2 under HG conditions. Cell proliferation was assessed using an MTT assay and oxidative stress was assessed by examining intracellular ROS generation and H2O2 production. The changes in extracellular matrix accumulation- and cellular inflammation-related proteins were measured by western blot analysis, ELISA and immunofluorescence analysis. The results showed that PB2 treatment markedly attenuated hyperglycemia-induced cell proliferation, oxidative stress, extracellular matrix accumulation and cellular inflammation in Mes13 cells, which was accompanied by an inactivation of redoxosomes, TGF-β1/SMAD and IL-1β/TNF-α/NF-κB signaling pathways. The present study also demonstrated that hyperglycemia upregulated and activated caveolin-1 (CAV-1), whereas PB2 treatment potently reversed this effect. In accordance, CAV-1 overexpression abolished the protective effects of PB2 against hyperglycemia in Mes13 cells, indicating that the cytoprotective effect of PB2 was CAV-1-dependent. These findings form the basis of the potential clinical applications of PB2 in the treatment of DKD.

Circular ribonucleic acid nucleoporin 98 knockdown alleviates high glucose-induced proliferation, fibrosis, inflammation and oxidative stress in human glomerular mesangial cells by regulating the microribonucleic acid-151-3p-high mobility group AT-hook 2 axis.

Aims/IntroductionThis study aimed to investigate the role and mechanism of circular ribonucleic acid nucleoporin 98 (circNUP98) in diabetic nephropathy (DN).Materials and MethodsHuman glomerular mesangial cells (HMCs) were stimulated with high glucose (HG) to imitate the growth environment of cells under the DN condition. Levels of genes and proteins were tested by quantitative reverse transcription polymerase chain reaction and western blot. Cell proliferation, apoptosis and inflammatory response were analyzed by using cell counting kit‐8, flow cytometry and enzyme‐linked immunosorbent assay analysis, respectively. Oxidative stress and fibrosis were evaluated by detecting the activity of reactive oxygen species, malondialdehyde, superoxide dismutase, fibronectin and collagen IV. The binding interaction between microribonucleic acid (miR)‐151‐3p and high mobility group AT‐hook 2 (HMGA2) or circNUP98 was confirmed using dual‐luciferase reporter, pull‐down and ribonucleic acid immunoprecipitation assays. Exosomes were isolated by ultracentrifugation, and qualified by transmission electron microscopy, nanoparticle tracking analysis and western blot.ResultsCircNUP98 expression was higher in the serum of DN patients and HG‐stimulated HMCs. Functionally, circNUP98 knockdown alleviated HG‐induced proliferation, fibrosis, inflammatory response and oxidative stress in HMCs. Mechanistically, circNUP98 directly sponged miR‐151‐3p, which targeted HMGA2. Rescue experiments showed that miR‐151‐3p reversed the inhibitory effects of circNUP98 knockdown on HG‐induced HMC dysfunction. Furthermore, miR‐151‐3p re‐expression also led to an inhibition of the aforementioned biological behaviors, which was attenuated by HMGA2 upregulation. Besides that, CircNUP98 was found to be packaged into exosomes of DN, and exosomal circNUP98 possessed diagnostic value for DN patients.ConclusionCircNUP98 knockdown alleviates HG‐induced proliferation, fibrosis inflammation and oxidative stress in HMCs by regulating the miR‐151‐3p–HMGA2 axis, which might provide a potential approach for DN therapeutics.

The extract of Polygala fallax Hemsl. slows the progression of diabetic nephropathy by targeting TLR4 anti-inflammation and MMP-2/9-mediated anti-fibrosis in vitro

BackgroundPolygala fallax Hemsl. is a plant that is commonly used as a folk medicine by Guangxi ethnic minorities, and it is also widely used in the clinical treatment of chronic diseases in China. The extract of P. fallax (EPF) contains key biologically active components from the roots and stems. However, the role of P. fallax or EPF in diabetic nephropathy (DN) is unclear. PurposeThis study aimed to investigate the effects and mechanisms of EPF on high glucose (HG)-induced human glomerular mesangial cell (HMC) injury, inflammation, fibrosis, and apoptosis in vitro. MethodsFor the in vitro study, MTT and ELISA assays were performed with HG-treated HMCs, as well as MMP, Hoechst, flow cytometry, qRT-PCR, and western blot analyses. The expression of the TLR4/NF-κB pathway, along with its downstream inflammatory, apoptosis, and fibrosis factors, was measured. The expression of the TLR4/NF-κB pathway and its downstream inflammatory factors were also measured after the addition of TLR4 inhibitors. ResultsOur results suggest that EPF can reverse the hyperproliferation and apoptosis of HMCs induced by HG. In addition, the extract inhibited the increase in inflammatory factors IL-6, TNF-α, IL-1β, MCP-1, and IL-18 in cells treated with HG. The mRNA and protein expression of TLR4, MyD88, NF-κB, Col IV, FN, MMP-9, and MMP-2 were downregulated by EPF. In addition, EPF significantly reduced the loss of MMP and the upregulation of Bcl-2/Bax mRNA and protein levels after HG treatment. ConclusionThese results demonstrated that EPF protects against diabetes-induced renal injury in vitro. EPF protected against HG-induced HMCs proliferation, apoptosis, fibrosis, and inflammation likely via inhibition of TLR4-dependent NF-κB signaling. This herbal extract may also be a novel treatment for DN.

Quercetin prevents chronic kidney disease on mesangial cells model by regulating inflammation, oxidative stress, and TGF-β1/SMADs pathway.

BackgroundChronic kidney disease (CKD) happens due to decreasing kidney function. Inflammation and oxidative stress have been shown to result in the progression of CKD. Quercetin is widely known to have various bioactivities including antioxidant, anticancer, and anti-inflammatory activities.ObjectiveTo evaluate the activity of quercetin to inhibit inflammation, stress oxidative, and fibrosis on CKD cells model (mouse mesangial cells induced by glucose).Methods and MaterialThe SV40 MES 13 cells were plated in a 6-well plate with cell density at 5,000 cells/well. The medium had been substituted for 3 days with a glucose-induced medium with a concentration of 20 mM. Quercetin was added with 50, 10, and 5 µg/mL concentrations. The negative control was the untreated cell. The levels of TGF-β1, TNF-α, and MDA were determined using ELISA KIT. The gene expressions of the SMAD7, SMAD3, SMAD2, and SMAD4 were analyzed using qRT-PCR.ResultsGlucose can lead to an increase in inflammatory cytokines TNF-α, TGF-β1, MDA as well as the expressions of the SMAD2, SMAD3, SMAD4, and a decrease in SMAD7. Quercetin caused the reduction of TNF-α, TGF-β1, MDA as well as the expression of the SMAD2, SMAD3, SMAD4, and increased SMAD7.ConclusionQuercetin has anti-inflammation, antioxidant, antifibrosis activity in the CKD cells model. Thus, quercetin is a promising substance for CKD therapy and further research is needed to prove this in CKD animal model.

Suppressing the activity of CXCR4 down-regulates the expression of renal fibrosis related genes in primary glomerular cells.

BackgroundC-X-C chemokine receptor type 4 (CXCR4) has a certain effect on renal fibrosis, and there are few specific studies in cells. We want to investigate the impact of suppressing CXCR4 activity on the expression of renal fibrosis-related genes in primary glomerular endothelial cells, mesangial cells, and podocytes.MethodsImmunofluorescence assays were used to determine the purity of isolated glomerular endothelial cells, mesangial cells, and podocytes. CXCR4 knockdown cell lines were established by transfecting the short hairpin (sh)RNA against CXCR4. T140 and AMD3100 were used to inhibit the activity of CXCR4. LY294002 was used to inhibit the activity of phosphoinositide 3-kinase (PI3K). The mRNA expression of CXCR4 was determined by real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR). The protein expression of CXCR4, collagen IV, matrix metallopeptidase (MMP)-9, PI3K, Rac1, and vascular cell adhesion protein 1 (VCAM-1) was evaluated by Western blot analysis.ResultsHigh purity was observed on isolated primary glomerular endothelial cells and podocytes. However, the purity of isolated mesangial cells was relatively low. The mRNA expression of CXCR4 was significantly suppressed by the transfection of shRNA. Compared to control cells, the expression of CXCR4, collagen IV, MMP-9, PI3K, Rac1, and VCAM-1 were dramatically downregulated in cell lines transfected with shRNA against CXCR4. Furthermore, cell lines treated with T140, AMD3100, or LY294002 also showed downregulated expression of these proteins compared to untreated cells. No significant differences were observed in the protein expression of these proteins between control cells and cells transfected with the shRNA negative control (NC).ConclusionsSuppressing the activity of CXCR4 downregulated the expression of renal fibrosis-related genes in primary glomerular cells, even under a non-inflammatory state.

Endothelins and liver cirrhosis

AbstractEndothelins are a family of 21‐amino acid oligopeptides, called endothelin‐1 (ET‐1), endothelin‐2 (ET‐2), and endothelin 3 (ET‐3). Endothelins act on hepatocytes, liver endothelial cells, and Kupffer cells in a paracrine or autocrine manner through two G protein‐coated receptors, called endothelin A and B receptors, which are mainly located in interlobular veins, interlobular artery endothelial cells and hepatic stellate cells (HSCs). ET B receptor (ETBR)‐1 is responsible for the induction of endothelial nitric oxide synthase, resulting in nitric oxide release and vasodilatation, whereas ETBR‐2 is located on HSCs and is responsible for vasoconstriction. Endothelins are not stored in the organs. Approximately 20% of endothelins are secreted into the circulation system, and are rapidly cleared by the lungs, liver, heart, and kidneys. As a potent vasoconstrictor, endothelins may have a key role for the treatment of hypertensive vascular diseases, inflammation, fibrosis, and metabolic diseases. By clearing ET‐1 from the circulation, endothelin A receptor (ETAR) antagonists can reduce intraportal vascular resistance by dilatating the portal vein, reducing the contraction of HSCs, increasing the diameter of sinusoids, facilitating the regression of liver fibrosis, and restoring liver parenchyma. ET‐1 induces nitric oxide synthase and upregulates cyclooxygenase 2 mRNA levels, making it a key factor during the onset of fibrosis and in the prognosis of patient outcomes. Endothelins can be used to treat porto‐pulmonary hypertension, portal hypertension, angiogenesis, and liver fibrosis and have been approved for the treatment of porto‐pulmonary hypertension. Endothelins are produced on mesangial cells, podocytes, tubular epithelium, and renal collecting tubes in high amounts. Activation of ETAR supports the progression of kidney diseases, whereas activation of ETBR has protective effects on the kidneys. ET‐1 plays an important role in normal cardiovascular homeostasis. Endothelins are closely associated with severe systemic hypertension, congestive heart failure, atherosclerosis, and pulmonary hypertension. Dual ET receptor antagonists reduce blood pressure, but have several side effects, including liver toxicity, acute liver failure, accumulation of salt and water, testicular toxicity, headache, and teratogenicity. Liver enzymes should be checked in all patients who have received ET receptor antagonists, and women of childbearing years should use contraception.

Deconvolution of gene expression for microenvironmental cell types in thymomas.

e20623 Background: Thymomas are rare neoplasms in the anterior mediastinum that account for less than 1% of tumors. The presence of immune cells within the tumor is one of the cardinal features of thymomas. However, the nature of the immune infiltration is not well characterized. Methods: Deconvolution of the gene expression of thymomas in the TCGA-THYM cohort was performed using the CIBERSORT and xCELL programs. Pairwise correlative analyses were performed to study the relationships of cell types and thymoma histological subtypes. Results: The differences in presence of immune cells was better highlighted using xCELL, which identifies 67 categories of cells rather than in the 23 categories identified by CIBERSORT. As expected, there were dramatic differences in the presence of T-cells (almost all subtypes) and B-cells between A/AB and type B (-1, -2, -3) thymomas. A significant difference (P < 0.0001) in different type B thymomas was observed for the presence of monocytes, macrophages, and M1, but not in M2 macrophages. Significant differences in dendritic cells were observed within subtypes of B thymoma. Similarly, statistically significant differences were observed between most subtypes in Immunoscore (except AB/B3) and MicroenvironmentScore (except A/B3; AB/B2 and AB/B3). However, StromalScore differences were observed only in comparisons of type A/AB with B type thymomas (except AB/B3). Interestingly, there also differences in postulated pre-adipocytes, hepatocytes, mesangial cells and osteoblasts in types A vs B, and melanocytes between subtypes of B thymomas. Conclusions: The differences in lymphoid content of the tumor microenvironment is associated with dramatic changes in cell type distribution in subtypes of thymoma. These differences in composition underpin their clinicopathological behavior including incidence of myasthenia and likelihood of recurrence.

386-P: ROCK1 Regulation of AMPK in the Development of Diabetic Nephropathy

Rho-associated, coiled-coil-containing protein kinase (ROCK) is activated in the diabetic kidney. Studies over the past decade have illuminated ROCK signaling as an essential pathway in the pathogenesis of diabetic nephropathy. Accumulating evidence shows that renal dysfunction among patient with diabetes is associated with abnormal fatty acid oxidation in the kidney. However, the interaction of ROCK and fatty acid oxidation in diabetic kidneys remains unclear. Extracellular flux analyzer demonstrated that deficiency of ROCK1, one of the ROCK isoforms, improves TGF-β-induced mitochondrial dysfunction in primary cultured mesangial cells. ROCK1 deficiency rescued TGF-β-induced downregulation of fatty acid oxidation mediators such as peroxisome proliferator-activated receptor gamma coactivator 1-α (PGC-1α) , and carnitine palmitoyltransferase 1A (CPT1A) . An investigation of mechanisms underlying this observation revealed activated ROCK1 axis through the phosphorylation of AMPK and increased expression of PGC-1α. Fluorescence histochemistry showed that ROCK1 deficiency suppresses ROS production induced by mitochondrial damage due to abnormal fatty acid metabolism. Consistent with this result, compared to wild-type mice, ROCK1 knockout mice exhibited reduced albuminuria and histological abnormalities along with the recovery of impaired fatty acid utilization and mitochondrial fragmentation. Furthermore, RNA-seq showed that ROCK1 deficiency in mesangial cells significantly altered the expression of genes involved in metabolic pathway. These observations indicate that ROCK1 is a key player in the development of diabetic renal injury. ROCK1 may be a potential therapeutic target for the treatment of diabetic nephropathy. Disclosure Y.Nagai: None. K.Matoba: None. K.Sekiguchi: None. Y.Takeda: None. Y.Kanazawa: None. D.Kawanami: None. K.Utsunomiya: None. R.Nishimura: Speaker's Bureau; Abbott Diabetes, Astellas Pharma Inc., AstraZeneca, Boehringer Ingelheim International GmbH, Eli Lilly and Company, Medtronic, Merck & Co., Inc., Novo Nordisk, Ono Pharmaceutical Co., Ltd., Sanofi, Taisho Pharmaceutical Holdings Co., Ltd., Takeda Pharmaceutical Company Limited, Teijin Pharma Limited, Terumo Corporation.

CLINICAL AND MORPHOLOGICAL FEATURES OF KIDNEY DAMAGE IN PATIENTS WITH ARTERIAL HYPERTENSION DIED OF SARS-COV-2 INFECTION

Objective: To assess clinical and pathomorphological features of kidney damage in patients with arterial hypertension (AH) who died of the new coronavirus infection COVID-19. Design and method: A complex analysis of 268 kidney autopsies was carried out, including the study of macro- and microscopic changes reflected in the protocols of pathological and anatomical autopsies and identified during the histological examination. In 224 patients (83.6%) with AH, the diagnosis was confirmed by isolating the SARS-CoV-2 RNA using the polymerase chain reaction; in 44 (16.4%) – through computed tomography of the lungs. The causes of deaths were the following: in 31 patients (11.6%) acute myocardial infarction; in 40 (14.9%) cerebrovascular accident; in 11 (4.1%) pulmonary embolism; 222 patients (83%) had acute respiratory distress syndrome. The analysis included 130 men aged 36 to 92 (72.6 years old on average) and 138 women aged 40 to 106 (77.1 years old on average). Results: In the kidneys we detected ischemic changes caused by disturbances in the microvasculature. These are stases, sludges, erythrocyte and fibrin thrombi predominantly in the medulla. In the glomeruli diapedesis hemorrhages, mesangial cells proliferation, basement membrane thickening and fibrinoid necrosis of the capillary wall were observed. In the epithelium of the convoluted tubules, a granular, hyaline-drop dystrophy and a necrosis as the extreme degree of the damage were noted. In the kidneys, a pronounced lymphoid and leukocyte infiltration was detected. These changes were accompanied by inflammation and renal failure symptoms. In particular, the level of C-reactive protein was 140.6 ± 7.42 mg/l; blood ferritin 1258.0 ± 110.1 mcg/l; blood leukocytes 15.0 ± 0.67 10^9/l; blood lymphocytes 1.54 ± 0.29 10^9/l; procalcitonin 1.350 ± 0.51 ng/ml. The development of hemorrhagic syndrome was observed in 93 (35%) cases. The level of D-dimer was 4326.4 ± 696.1 ng/ml; fibrinogen 18.2 ± 13.04 g/l; platelets 179.0 ± 8.7 10^9/l. Blood creatinine in patients aged 36 to 59 (27 people) was 120.6 ± 12.2 mmol/l; aged 60 to 80 (136 people) 136.6 ± 10.12 mmol/l; aged over 80 (105 people) 139.0 ± 11.7 mmol/l. Conclusions: The study of the kidneys in patients who died from the new coronavirus infection revealed microvasculature vessels damage and development of coagulation disorders amid the renal failure.

Correlation of Body Mass Index with Clinicopathologic Parameters in Patients with Idiopathic Membranous Nephropathy.

ObjectiveTo investigate the association between body mass index (BMI) and clinicopathologic parameters in patients with idiopathic membranous nephropathy (IMN).MethodsThis study was retrospective and included patients with biopsy-proven IMN from 2018 to 2021 in Hebei General Hospital. Patients were categorized into two groups based on BMI. Clinical and histopathologic data were analyzed at the time of renal biopsy. Pathological data included immunofluorescence staining, glomerulosclerosis (GS, 0–2), mesangial cell proliferation (MCP, 0–1), tubular atrophy (TA, 0–1), interstitial fibrosis (IF, 0–1), vascular wall thickness (VWT, 0–1) and a combination score (GMTIV) graded from 0 to 5.ResultsOur study revealed that the obese group had a higher prevalence of hypertension and diabetes than the overweight/normal weight group (P=0.001, P=0.002). Systolic blood pressure (P=0.005), diastolic blood pressure (P<0.001), haemoglobin (P=0.006), triglycerides (P<0.001), serum uric acid (P=0.05), 24 h urine proteinuria concentration (UP) (P=0.012), MCP (P=0.042), IF (P=0.033), and GMITV (P=0.033) score were higher in obese group compared to the other group, while the high-density lipoprotein-cholesterol (P=0.034) and immunoglobulin A deposition score (P=0.005) were lower. Factors significantly associated with UP were the ratio of lymphocyte count to white blood cell count, serum pre-albumin, immunoglobulin G, microscopic hematuria, anti-phospholipase A2 receptor (anti-PLA2R), C3 deposit on multivariable analysis (adjusted R2=0.343). Binary logistic regression analysis illustrated that MCP was correlated to BMI (OR=2.528, P=0.036). Ordinal logistic regression analysis demonstrated that GMTIV was associated with BMI (OR=1.114, P=0.010) and C3 deposit (OR=1.655, P=0.001).ConclusionHigh BMI was associated with MCP and GMTIV score in IMN patients. Obesity may play an essential role in mesangial lesions of IMN. This study emphasized the relation between BMI and histological parameters under the universal usage of anti-PLA2R antibodies for diagnosis and prognosis in IMN.

Curcumin Blocks High Glucose-Induced Podocyte Injury via RIPK3-Dependent Pathway.

Podocyte loss is well known to play a critical role in the early progression of diabetic nephropathy. A growing number of studies are paying attention to necroptosis, a programmed form of cell necrosis as a mechanism of podocyte loss. Although necroptosis is a recently established concept, the significance of receptor interacting serine/threonine kinase 3 (RIPK3), a gene that encodes for the homonymous enzyme RIPK3 responsible for the progression of necroptosis, is well studied. Curcumin, a natural hydrophobic polyphenol compound responsible for the yellow color of Curcuma longa, has drawn attention due to its antioxidant and anti-inflammatory effects on cells prone to necroptosis. Nonetheless, effects of curcumin on high glucose-induced podocyte necroptosis have not been reported yet. Therefore, this study investigated RIPK3 expression in high glucose-treated podocytes to identify the involvement of necroptosis via the RIPK3 pathway and the effects of curcumin treatment on RIPK3-dependent podocytopathy in a hyperglycemic environment. The study discovered that increased reactive oxygen species (ROS) in renal podocytes induced by high glucose was improved after curcumin treatment. Curcumin treatment also significantly restored the upregulated levels of VEGF, TGF-β, and CCL2 mRNAs and the downregulated level of nephrin mRNA in cultured podocytes exposed to a high glucose environment. High glucose-induced changes in protein expression of TGF-β, nephrin, and CCL2 were considerably reverted to their original levels after curcumin treatment. Increased expression of RIPK3 in high glucose-stimulated podocytes was alleviated by curcumin treatment as well as N-acetyl cysteine (NAC, an antioxidant) or GSK′872 (a RIPK3 inhibitor). Consistent with this, the increased necroptosis-associated molecules, such as RIPK3, pRIPK3, and pMLKL, were also restored by curcumin in high glucose-treated mesangial cells. DCF-DA assay confirmed that such a result was attributed to the reduction of RIPK3 through the antioxidant effect of curcumin. Further observations of DCF-DA-sensitive intracellular ROS in NAC-treated and GSK′872-treated podocyte groups showed a reciprocal regulatory relationship between ROS and RIPK3. The treatment of curcumin and GSK′872 in podocytes incubated with high glucose protected from excessive intracellular superoxide anion production. Taken together, these results indicate that curcumin treatment can protect against high glucose-induced podocyte injuries by suppressing the abnormal expression of ROS and RIPK3. Thus, curcumin might be a potential therapeutic agent for diabetic nephropathy as an inhibitor of RIPK3.

The Effects of The Bongardia Chrysogonum (L) In A Rat Model of Diabetic Nephropathy

Diabetic nephropathy is increasing in frequency and is one of the most important causes of end-stage renal disease. In this study, the effect of Bongardia chrysogonum on kidneys by induced diabetic nephropathy in a diabetic rat model induced by streptozotocin (STZ) was investigated. Forty male Wistar Albino rats were used in the study. Rats; They were divided into 4 groups as the control group (K), the group given B. chrysogonum (B), the group with diabetes (DM) and the group given B. chrysogonum for treatment after diabetes was established (DMB). 45 mg/kg STZ was given intravenously to the DM and DMB groups. One day later, the blood sugar levels of the rats were checked with a glucometer, and values above 300 mg/dl were accepted as diabetes. B. chrysogonum infusion prepared by weighing 3 grams every day for 5 weeks was given to the B and DMB groups by gavage. Their weights and blood sugar levels were measured and recorded every week. At the end of 5 weeks, it was observed that the blood sugar level of the DMB group decreased statistically significantly compared to the DM group. Then, the rats were sacrificed under deep anesthesia and their kidneys were removed. As a result of histopathological examination, glomerular basement membrane thickening and mesangial cell increase were observed in the kidneys of the DM group. Similar histopathological findings were observed in the kidneys of DMB group rats and K group kidneys. These results were supported by Periodic acid Schiff and reticular staining. As a result of this study, it was observed that B. chrysogonum lowered blood sugar level after diabetes and reduced kidney damage. The mechanism by which B. chrysogonum regulates blood sugar in diabetic rats and affects commonly occurring secondary injuries should be investigated in detail.

Therapeutic Potential of Phlorotannin-Rich Ecklonia cava Extract on Methylglyoxal-Induced Diabetic Nephropathy in In Vitro Model.

Advanced glycation end-products (AGEs) play a vital role in the pathogenesis of diabetic complications. Methylglyoxal (MGO), one of the major precursors of AGEs, is a highly reactive dicarbonyl compound that plays an important role in the pathogenesis of diabetic nephropathy. This study was designed to evaluate the therapeutic potential of phlorotannin-rich Ecklonia cava extract (ECE) on MGO-induced diabetic nephropathy in in vitro models using mouse glomerular mesangial cells. ECE showed anti-glycation activity via breaking of AGEs-collagen cross-links and inhibition of AGEs formation and AGE-collagen cross-linking formation. The renoprotective effects were determined by assessing intracellular reactive oxygen species (ROS) and MGO accumulation, cell apoptosis, and the Nrf-2/ARE signaling pathway. MGO-induced renal damage, intracellular ROS production level, and MGO-protein adduct accumulation were significantly decreased by pretreating ECE. Moreover, ECE pretreatment exhibited preventive properties against MGO-induced dicarbonyl stress via activation of the Nrf2/ARE signaling pathway and reduction of RAGE protein expression in mouse glomerular mesangial cells. Collectively, these results indicated potential anti-glycation properties and prominent preventive effects of ECE against MGO-induced renal damage. Additionally, ECE may be utilized for the management of AGE-related diabetic nephropathy.

EOS789, pan-phosphate transporter inhibitor, ameliorates the progression of kidney injury in anti-GBM-induced glomerulonephritis rats.

Hyperphosphatemia associated with chronic kidney disease (CKD) not only dysregulates mineral metabolism and bone diseases, but also strongly contributes to the progression of kidney disease itself. We have identified a novel drug for hyperphosphatemia, EOS789, that interacts with several sodium‐dependent phosphate transporters (NaPi‐IIb, PiT‐1, and PiT‐2) known to contribute to intestinal phosphate absorption. In this study, we investigated whether EOS789 could ameliorate kidney disease progression in glomerulonephritis rats. Anti‐glomerular basement membrane (GBM) nephritis was induced in rats by intravenously administering two types of anti‐rat GBM antibodies. We evaluated the effect of EOS789 administered in food admixture on hyperphosphatemia and kidney disease progression. In an anti‐GBM nephritis rats, which exhibit a significant increase in serum phosphate and a decline in renal function, EOS789 dose‐dependently improved hyperphosphatemia and EOS789 at 0.3% food admixture significantly ameliorated kidney dysfunction as shown in the decline of serum creatinine and BUN. Renal histopathology analysis showed that EOS789 significantly decreased crescent formation in glomeruli. To elucidate the mechanism underlying glomerular disease progression, human mesangial cells were used. High phosphate concentration in media significantly increased the expression of Collagen 1A1, 3A1, and αSMA mRNA in human mesangial cells and EOS789 dose‐dependently suppressed these fibrotic markers. These results indicate that EOS789 prevented glomerular crescent formation caused by mesangial fibrosis by ameliorating hyperphosphatemia. In conclusion, EOS789 would not only be useful against hyperphosphatemia but may also have the potential to relieve mesangial proliferative glomerulonephritis with crescent formation.

Bruceine A protects against diabetic kidney disease via inhibiting galectin-1

Bruceine A is a natural quassinoid compound extracted from the fruit of the Traditional Chinese Medicine Brucea javanica (L.) Merr. that has various types of various biological activities. However, whether the compound has a protective effect on diabetic kidney disease remains unknown. Galectin-1 is actively involved in a variety of chronic inflammation-relevant human diseases including diabetic kidney disease. Here, we identified Bruceine A as a kidney protective molecule against a model of diabetic kidney disease in db/db mice with potent anti-inflammatory activity both invitro and invivo. Mechanistically, by selectively binding to the conserved carbohydrate-recognition domain of galectin-1 and disrupting the interaction between galectin-1 and the receptor for activated protein C kinase 1, Bruceine A was found to inhibit galectin-1-mediated inflammatory signal transduction under high glucose stress in rat mesangial HBZY-1 cells. Thus, our findings reveal Bruceine A as an unidentified galectin-1 inhibitor affording significant protection against diabetic kidney disease and may provide novel pharmacological therapeutics for the disease.