Glomerular Damage Research Articles

Supplying rats with halfa-bar and liquorice extracts ameliorate doxorubicin-induced nephrotic syndrome

In the current work, we aimed to evaluate the protective effects of liquorice and halfa-bar extract against doxorubicin (DOX)-induced nephritic syndrome (NS) in rats. Twenty albino male rats were intraperitoneally injected with 50 mg/kg of DOX. The injected rats were supplied daily with 400 mg/kg of liquorice, halfa-bar extract, or their combination for 2 weeks. Our findings confirmed the induction of NS in rats indicated by alteration in Bowman’s space, damaged in glomerular capsules, and tubules. Moreover, the levels of produced tumour necrosis factor-alpha (TNF-α) and interleukin-8 (IL-8) were increased, accompanied by decreasing levels of IL-4 and IL-10. Supplement NS-rats with liquorice and halfa-bar extracts restored the glomerular and tubules damage and adjusted the level of produced TNF-α and IL-8. Interestingly, both extracts can stimulate the expression profile of small proline-rich protein 2 F (sprr2f) and metalloproteinase-10 (MMP-10), which are responsible for repairing and regeneration mechanisms of renal syndromes.

Adhesion G Protein-Coupled Receptor Gpr126 (Adgrg6) Expression Profiling in Diseased Mouse, Rat, and Human Kidneys.

Uncovering the function of understudied G protein-coupled receptors (GPCRs) provides a wealth of untapped therapeutic potential. The poorly understood adhesion GPCR Gpr126 (Adgrg6) is widely expressed in developing kidneys. In adulthood, Gpr126 expression is enriched in parietal epithelial cells (PECs) and epithelial cells of the collecting duct and urothelium. Whether Gpr126 plays a role in kidney disease remains unclear. Here, we characterized Gpr126 expression in diseased kidneys in mice, rats, and humans. RT-PCR data show that Gpr126 expression is altered in kidney disease. A quantitative RNAscope® analysis utilizing cell type-specific markers revealed that Gpr126 expression upon tubular damage is mainly increased in cell types expressing Gpr126 under healthy conditions as well as in cells of the distal and proximal tubules. Upon glomerular damage, an increase was mainly detected in PECs. Notably, Gpr126 expression was upregulated in an ischemia/reperfusion model within hours, while upregulation in a glomerular damage model was only detected after weeks. An analysis of kidney microarray data from patients with lupus nephritis, IgA nephropathy, focal segmental glomerulosclerosis (FSGS), hypertension, and diabetes as well as single-cell RNA-seq data from kidneys of patients with acute kidney injury and chronic kidney disease indicates that GPR126 expression is also altered in human kidney disease. In patients with FSGS, an RNAscope® analysis showed that GPR126 mRNA is upregulated in PECs belonging to FSGS lesions and proximal tubules. Collectively, we provide detailed insights into Gpr126 expression in kidney disease, indicating that GPR126 is a potential therapeutic target.

Glomerulonefritis focal y segmentaria post COVID-19 en una paciente lúpica

We present the case of a 29-year-old woman, with a diagnosis of chronic kidney disease, systemic lupus erythematosus without lupus nephritis criteria, who developed SARS-COV-2 infection, and had acute kidney injury, requiring dialysis. Renal biopsy showed focal segmental glomerulosclerosis, perihilar variant, and evidence of advanced sclerosis. However, renal function recovered several months later, without immunosuppression, so the glomerular damage and acute kidney injury were considered to be secondary to the COVID-19 infection. Since focal and segmental glomerulosclerosis occurred in patients with COVID-19, but its association was infrequent in patients who also had systemic lupus erythematosus, this case is reported.

New concepts in the physiopathology of hypertension. Purinergic receptors

Hypertension is a major risk of morbidity and mortality in patients when it is uncontrolled. In spite of improved therapies currently available for blood pressure control, their complications are far away from being accomplished. Therefore, chronic renal failure is frequently observed in hypertensive patients. Thus, insights on mechanisms that may contribute to arterial pressure control should be studied to prevent life-threatening cardiovascular disorders. Purinergic receptors have been recognized in the physiopathology of hypertension; this review summarizes their participation in the renal abnormalities of the kidney in hypertension. Several studies have suggested the activation of renal purinergic receptors under an elevated interstitial ATP milieu as a fundamental pathway that leads to generation and maintained hypertension. Elevated ATP concentration alters fundamental mechanisms involved in the long-term control of blood pressure such as pressure natriuresis, autoregulation of glomerular filtration rate and renal blood flow, as well as increased tubule-glomerular feedback responses, overall, these alterations decrease sodium excretion; in addition, the expression of ATP receptors is modified. Under a genetical background, ATP induces the production of vasoactive compounds, decreases renal function and induces tubulointerstitial injury before glomerular damage. Simultaneously, a deleterious interaction between angiotensin II and purinergic receptors lead to the progression of renal damage.

Tribbles pseudokinase 3 promoted renal fibrosis by regulating the expression of DNA damage-inducible transcript 3 in diabetic nephropathy.

Diabetic nephropathy (DN) is a severe complication of prolonged diabetes, impacting millions worldwide with an increasing incidence. This study investigates the role of tribbles pseudokinase 3 (TRIB3), a protein implicated in the progression of DN, focusing on its mechanisms underlying glomerular damage. Through analysis of the Gene Expression Omnibus (GEO) database, we identified TRIB3 among differentially expressed genes in streptozotocin (STZ)-treated C57BL/6J mice. Both in vitro and in vivo experiments were conducted to examine the effects of TRIB3 inhibition on high glucose (HG)-induced damage in podocytes and DN mouse models. The results demonstrated that TRIB3 inhibition reduced inflammatory responses and extracellular matrix (ECM) production in MPC5 cells, mediated by the downregulation of DNA damage-inducible transcript 3 (DDIT3) - a critical regulator of proinflammatory cytokine secretion and ECM synthesis. Inhibiting TRIB3 decreased inflammatory factors and ECM deposition in diabetic mice in vivo, confirming its pivotal role in DN pathogenesis. These findings indicate that TRIB3 and its interaction with DDIT3 contribute significantly to DN by promoting inflammatory cascades and ECM accumulation, presenting potential therapeutic targets for managing the disease.

Assessment of single and combined administration of ubiquinone and lactoferrin on histopathology, ultrastructure, oxidative stress, and WNT4 expression gene induced by thioacetamide on hepatorenal system of adult male rats

BackgroundHepatorenal syndrome is a life-threatening medical complication of liver cirrhosis. Hepatic cirrhosis is commonly accompanied by rapid failure of renal functions. Thioacetamide (TAA) is a potent hepatotoxin and a class 2-type carcinogen. Ubiquinone (Coq10) and lactoferrin (LF) are potent antioxidants with antifibrotic and antiinflammatory effects. However, whether Coq10 and LF reduce the hepatorenal injury induced by TAA remains unclear. Here, we investigated the potential protective effect of both/or Coq10 and LF in ameliorating TAA-induced hepatorenal injury and the role of WNT4 gene expression in detecting TAA-induced renal injury in rats. Seventy healthy and mature male Sprague Dawley rats, weighting (200 g ± 20 g) and aging (4–6) weeks were randomly divided into seven groups (n = 10): control, Coq10, LF, TAA, TAA + Coq10, TAA + LF, and TAA + Coq10 + LF. The hepatorenal injury was induced through intraperitoneal (i.p.) injection of TAA (150 mg/kg/twice/weekly) for nine weeks. Coq10 (10 mg/kg/day) and LF (200 mg/kg/day) were orally administered for nine weeks.ResultsTAA induced marked hepatorenal damage, evident by the significant increase in the alanine aminotransferase (ALT), aspartate transaminase (AST), serum creatinine (SCr) activities, and the blood urea nitrogen (BUN) level. Besides, the significant increases in concentrations of malondialdehyde (MDA) and nitric oxide (NOx) together with significant decreases in the activities of catalase (CAT) and superoxide dismutase (SOD). The histopathological analysis of the TAA group showed obvious fibrosis, steatosis, and inflammation of the hepatic parenchyma as well as severe glomerular and tubular damage of the renal parenchyma. In addition, TAA induced marked ultrastructural alterations and up-regulation in the expression of the WNT4 gene in the kidney. Meanwhile, the biochemical, histopathological, and ultrastructural alterations were significantly decreased with significant down-regulation in the expression of WNT4 in the groups exposed to TAA and treated with Coq10 and LF.ConclusionOur data suggested that Coq10 and LF could have protective effects on TAA hepatorenal damage, through improving the hepatic and renal functions, reduction of oxidative stress, structural and ultrastructural alterations, besides down-regulation in the expression of WNT4.

Preventive effects of vasohibin-2-targeting peptide vaccine for diabetic nephropathy.

Diabetic nephropathy remains the leading cause of end-stage kidney disease in many countries, and additional therapeutic targets are needed to prevent its development and progression. Some angiogenic factors are involved in the pathogenesis of diabetic nephropathy. Vasohibin-2 (VASH2) is a novel proangiogenic factor, and our previous study showed that glomerular damage is inhibited in diabetic Vash2 homozygous knockout mice. Therefore, we established a VASH2-targeting peptide vaccine as a tool for anti-VASH2 therapy in diabetic nephropathy. In this study, the preventive effects of the VASH2-targeting peptide vaccine against glomerular injury were examined in a streptozotocin (STZ)-induced diabetic mouse model. The mice were subcutaneously injected with the vaccine at two doses 2 wk apart and then intraperitoneally injected with 50 mg/kg STZ for 5 consecutive days. Glomerular injury was evaluated 20 wk after the first vaccination. Treatment with the VASH2-targeting peptide vaccine successfully induced circulating anti-VASH2 antibody without inflammation in major organs. Although the vaccination did not affect blood glucose levels, it significantly prevented hyperglycemia-induced increases in urinary albumin excretion and glomerular volume. The vaccination did not affect increased VASH2 expression but significantly inhibited renal angiopoietin-2 (Angpt2) expression in the diabetic mice. Furthermore, it significantly prevented glomerular macrophage infiltration. The preventive effects of vaccination on glomerular injury were also confirmed in db/db mice. Taken together, the results of this study suggest that the VASH2-targeting peptide vaccine may prevent diabetic glomerular injury in mice by inhibiting Angpt2-mediated microinflammation.NEW & NOTEWORTHY This study demonstrated preventive effects of VASH2-targeting peptide vaccine therapy on albuminuria and glomerular microinflammation in STZ-induced diabetic mouse model by inhibiting renal Angpt2 expression. The vaccination was also effective in db/db mice. The results highlight the importance of VASH2 in the pathogenesis of early-stage diabetic nephropathy and the practicability of anti-VASH2 strategy as a vaccine therapy.

Juxtaglomerular (JG) cell-specific deletion of the NADPH oxidase accessory protein, NOXA1, decreases pro-renin and proteinuria in vivo in a mouse model of diabetes

A large fraction of patients with diabetes develops diabetic nephropathy. Plasma pro-renin levels are elevated in diabetic patients and thought to contribute to hypertension, renal damage and inflammation. Studying the mechanisms by which diabetes reprograms JG cells to increase pro-renin release could provide new targets for kidney disease. We previously found that JG cells from diabetic mice had higher reactive oxygen species production (superoxide and H2O2), pro-renin expression and pro-renin and renin release. We hypothesized that increased ROS production by specific NADPH oxidases (NOXs) in JG cells is responsible for higher pro-renin release and contributes to glomerular damage in diabetes. Western Blots showed that the NADPH oxidase isoform NOX1 and NOXA1 are upregulated in JG cells from diabetic mice (C57=100%; Akita=147±10%; n=4; p<0.05), whereas NOX2 or NOX4 were not different. Confocal microscopy showed that NOX1 and NOXA1 are present in JG cells and co-localize with renin-containing granules (n=3). Silencing NOX1 in primary cultures of JG cells from diabetic mice decreased renin release to C57 control levels; (n=6; p<0.01) whereas silencing NOX4 had no effect (n=6). To study the role of NOXA1 and NOX1 in JG cells in vivo, we used Cas9 mediated silencing in kidneys of control mice. Adenoviruses expressing Cas9 under control of the Ren1 promoter (AV-Ren1-Cas9) and AAV-gRNA-NOXA1 were loaded into 28-day minipumps and delivered into the renal subcapsular space in C57 mice. After 5 weeks, we obtained control (non-transduced) and Ren-Cas9-NOXA1-edited kidneys and isolated JG cells. NOXA1 expression was reduced by 60±22% in isolated JG cells and pro-renin expression was decreased by 46±19% compared to non-transduced kidneys (n=3). We then specifically deleted NOXA1 from JG cells in diabetic mice by subcapsular delivery of AV-Ren1-Cre in the kidney cortex of transgenic NOXA1 floxed diabetic Akita mice. JG cell-specific deletion of NOXA1 in diabetic mice decreased urinary ROS (8-Isoprostane pg/24hrs: Akita=4091±307; NOXA1 JG/KO=3086 ±137; n=4; p<0.05), and glomerular damage as indicated by decrease proteinuria (24h Ualbumin Akita= 98.6± 9.4; NOXA1 JG/KO= 57.8± 16.4 μg/24h; n=4; p<0.05). We conclude that enhanced NOX1/NOXA1 induced ROS in JG cells drive enhanced pro-renin release from JG cells which contribute to glomerular damage in diabetes. Thus, NOXA1/NOX1 in JG cells are a potential new target for diabetic nephropathy. DK105300-RO3 Henry Ford Hospital Institutional Funds. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Nicotine Exposure Causes Nitrosative and Oxidative Stress in Glomerular Podocytes, Resulting in Renal Damage

Introduction: Nicotine consumption is the leading cause of preventable disease and death. Patients with hypertension or kidney disease are more likely to exhibit renal pathologies associated with nicotine, such as glomerulopathy. In endothelial cells, nicotine was linked to mitochondrial oxidative stress and nitric oxide synthase (NOS) remodeling. However, there is a gap in knowledge regarding the effects of nicotine in glomerular epithelial cells, podocytes. We hypothesized that nicotine promotes nitrosative stress in the podocytes by producing peroxynitrite (ONOO-) and mitochondrial damage, thereby reducing glomerular function. Methods: We performed confocal imaging on human cultured podocytes to detect ONOO− (HPF), Ca2+ (Fluo-8) and nitric oxide NO (DAF-FM) in response to nicotine. Seahorse assay (Agilent XFe24) was used to test mitochondrial respiration. Then, nicotine was chronically infused in Dahl SS rats (0.2 mg/kg/day s.c., 0.4% NaCl, 21 days), and blood pressure was recorded with telemetry. Glomerular damage was evaluated, and electron microscopy was employed to assess mitochondrial ultrastructure. Electron paramagnetic resonance (EPR) spectroscopy was used to access the NO levels in cultured podocytes and in vivo. OriginPro was used for statistical analysis. Results: Acute application of nicotine promoted intracellular Ca2+ and ONOO− transients in podocytes. The ONOO− response was blocked in the presence of SOD, indicating that ONOO− production requires superoxide. The application of specific α7 or α4β2, α2β4, α4β4, and α3β4 nicotinic acetylcholine receptor (nAChR) agonists elicited Ca2+ transients but did not produce ONOO−, suggesting that nitrosative stress occurs independently from Ca2+ influx or nAChR activation. Incubation with nicotine (12 hrs) resulted in a decrease in podocytes’ mitochondrial respiration (two-sample t-test, p<0.05). In vivo, nicotine infusion did not affect blood pressure but promoted mitochondrial damage in podocytes, which exhibited swelling, loss of cristae, and mitophagy (t-test, p<0.05). Histopathological assessment showed higher glomerular damage in nicotine-exposed rats (t-test, p<0.05). Both EPR and confocal approaches demonstrated nicotine-mediated changes in NO bioavailability and an increase in NOS2 activity (t-test, p<0.05) Conclusion: Nicotine elicits superoxide-mediated nitrosative stress and peroxynitrite formation in podocytes, promoting mitochondrial damage and glomerular dysfunction. Revealing the mechanisms of nicotine-mediated damage in the podocytes impacts our progress towards new treatments for smoking and vaping-associated renal pathologies. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

The Effect of a High-Fat Western Diet on Diabetic Nephropathy and the Pathogenesis of Type 2 Diabetes

The high-fat Western diet is a key contributor to the growing rate of obesity, which has become a global health concern. Additionally, obesity increases the risk of developing metabolic syndrome which leads to a range of chronic illnesses like diabetes, heart disease, and stroke. Diabetic Nephropathy (DN) is a complication prevalent in one-third of people with Type 2 Diabetes and leads to End Stage Renal Disease. The aim of this study was to assess how a high-fat Western diet impacts diabetic severity and renal, cardiovascular, and hepatic injury, in a rodent model of type 2 Diabetes. We used aged (>38 weeks old, with severe renal disease) and young (12 weeks old, prior to major kidney damage) male and female Type 2 Diabetic Nephropathy (T2DN) rats, a non-obese diabetic animal model, and placed them on either normal (NFD) or high fat (HFD) calorically balanced diets for 12 weeks. Urine sampling and glucose tolerance tests (GTT) were assessed every 3 weeks. All rats on the HFD had significantly increased total body weights (TBW). However, average caloric intake over the 12 weeks was not significantly different in young males and females or in old males, while the aged HFD females had a significant increase (52.18 ± 1.536 vs 67.43 ± 0.4682 kcal/g, p=0.0001). The HFD had no effect on female GTT curves or fasting blood glucose in either age group. In contrast, the HFD males’ glucose intolerance and fasting blood glucose levels increased progressively and became significantly different between diets by week 9 (old: 204.5 ± 10.8 vs 284.4 ± 15.8 mg/dL, p=0.003, young: 193.3 ± 9.497 vs 257.5 ± 17.19 mg/dL, p=0.02). The HFD did not affect urine output or 2-kidney/TBW ratios in both sexes, while heart weight/TBW ratios in females were significantly reduced on the HFD in both age groups. Studies are ongoing in young animals; however, old animals revealed a significant increase in lipid deposits in the liver of HFD group for both males and females. There were no significant changes in circulating cholesterol and triglyceride levels between diets; however, there was a trend to increased triglycerides in females on HFD (p=0.051). The liver function panel showed a significant increase in ALP liver enzyme in the HFD group of both sexes. Picrosirius red-stained heart tissue showed increased fibrosis in the HFD aged males. Microbiome analysis revealed a significant decrease in gut microbiota alpha diversity in males on HFD, while no significant changes were observed in females between diets. Ongoing studies will quantify renal damage, including cortical fibrosis, medullary protein casts, glomerular damage, albuminuria, and KIM-1 staining, as well as lipidemia and heart fibrosis resulting from the HFD for both old and young animals. In conclusion, the HFD appears to have a more deleterious effect on glucose handling in both old and young T2DN males than in female rats; however, further studies are warranted to validate this effect. R01 DK129227, R01 DK135644, R00 HL153686. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Unraveling a Novel Mechanism of Altered Glomerular Hemodynamics in Pre-diabetic Obesity

Background: In pre-diabetic obesity, disrupted glomerular autoregulation leads to increased glomerular capillary pressure and renal blood flow, causing potential barotrauma-induced glomerular damage. A decrease in Afferent arteriole ( Af-Art) resistance causes higher glomerular capillary pressure but the underlying mechanism behind this, specifically in pre-diabetic obesity, remains unclear. Hyperinsulinemia is common in pre-diabetic obesity, but its role in reducing Af-Art resistance in the kidneys is unknown. Kidney blood flow is auto-regulated by vasoconstriction via tubuloglomerular feedback (TGF) and vasodilation through the Distal convoluted tubule-2/connecting tubule (DCT-2/CNT) mechanism. Our earlier studies indicate increased Af-Art dilation in pre-diabetic obese rats through the DCT-2/CNT mechanism without affecting TGF. This DCT-2/CNT induced Af-Art dilation requires initiation from epithelial sodium channels (ENaC) present at this tubular segment and interestingly insulin is a known ENaC activator. Hypothesis: We hypothesize that in the context of pre-diabetic obesity, hyperinsulinemia contributes to an increase in renal cortical blood flow (CBF) by increasing DCT-2/CNT-induced Af-Art dilation, subsequently leading to glomerular injury. Methods: We assessed the effects of insulin on Af-Art dilation by perfusing juxtaposed DCT-2/CNT segments with insulin in normal rabbits. Contribution of DCT-2/CNT induced Af-Art dilation was measured by co-administration of the ENaC blocker benzamil. To study hyperinsulinemia's role in an in-vivo setting without altering blood glucose levels, we used a hyperinsulinemic-euglycemic (HI-EUG) clamp in obese rats with intravenous (IV) glucose and insulin infusions. Renal CBF and blood pressure were monitored using a laser Doppler probe and femoral artery catheter, respectively, both with and without benzamil administration (400 μg/kg, IV). To evaluate insulin's potential role in causation of renal damage, we infused insulin directly into one kidney of lean rats for 6 weeks via a renal sub-capsular catheter, with the contralateral kidney serving as control. Glomerular basement membrane (GBM) thickness, a marker of renal damage, was measured via electron microscopy. Untargeted lipidomic analysis of the renal cortex was performed to identify potential renal damage pathways in insulin infused kidneys. Results: Insulin perfusion in rabbits' DCT-2/CNT potentiated the dilation of adherent Af-Art compared to control infusions with artificial tubular fluid. ENaC inhibition reversed the insulin-induced Af-Art dilation, confirming that insulin increases Af-Art dilation through the ENaC-initiated DCT-2/CNT mechanism. In obese rats, insulin delivery using a HI-EUG clamp resulted in a significant 20.8±4.9% increase in CBF, which was negated by the administration of the ENaC blocker benzamil, indicating the potential involvement of DCT-2/CNT-mediated vasodilation. Neither the HI-EUG clamp nor benzamil infusion affected blood pressure. Additionally, continuous direct insulin infusion into the kidneys of lean rats led to an increase in glomerular basement membrane (GBM) thickness and ceramide accumulation in the renal cortex, both indicative of renal damage. Conclusion: Our study demonstrates that insulin directly augments DCT-2/CNT-mediated Af-Art dilation and increases renal cortical blood flow in obese rats. Chronic renal insulin infusion led to structural changes and damage markers in the kidney, independent of blood glucose and blood pressure levels. Henry Ford Health System Internal Funding. Pilot and Feasibility grant from southeast center for integrated metabolomics. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Abstract 1078: A Cross-species Platform And Phosphoproteomics Analysis To Mitigate Vegfri-induced Hypertension And Endothelial Dysfunction

Introduction: Vascular endothelial growth factor receptor inhibitors (VEGFRis) are a class of tyrosine kinase inhibitors (TKIs) used to treat human and canine cancers, but these drugs also cause hypertension (HTN) and endothelial cell (EC) dysfunction. Purpose: We combined phosphoproteomic analysis in ECs with a cross-species in-vitro and in-vivo approach to identify mitigating therapies for VEGFRi-induced EC dysfunction and HTN. Methods and Results: We developed a sorafenib-induced HTN mouse model showing increased blood pressure, mesenteric resistance vessel EC dysfunction (decreased phosphorylation of Ser1177 on endothelial nitric oxide synthase [p-eNOS], increased endothelin-1 [ET-1]), and renal glomerular endotheliosis. Human umbilical vein ECs were treated with VEGFR TKIs, non-VEGFR TKIs, and cardiovascular drugs. Phosphoproteomic mass spectrometry and a marker selection algorithm identified a 14-phosphopeptide signature differentiating VEGFR TKIs from non-VEGFR TKIs. The signature peptide RBM17 pS222 increased with VEGFRi treatment in human, canine, and mouse aortic ECs, correlating with reduced p-eNOS. By screening for cardiovascular drugs that reverse the VEGFRi phosphoproteomic signature, we found alpha-adrenergic antagonists, particularly doxazosin, had the most anti-correlated signatures. Doxazosin prevented VEGFRi-induced EC dysfunction in primary aortic ECs across species. In mice, doxazosin reversed sorafenib-induced EC dysfunction in resistance vessels without affecting blood pressure or renal glomerular endotheliosis. Conversely, lisinopril reduced HTN and glomerular endotheliosis but not EC dysfunction. Preliminary results from our clinical trial in canine cancer patients confirmed that toceranib induced HTN and showed that both doxazosin and lisinopril significantly lowered blood pressure in dogs with cancer and VEGFRi-induced HTN. Conclusion: Our study demonstrates the utility of phosphoproteomics and cross-species analysis in identifying EC-protective effects of doxazosin in VEGFRi-induced HTN. Our mouse data suggest distinct mechanisms for VEGFRi-induced EC dysfunction and HTN, potentially involving renal glomerular damage.

Divergent cardiac and renal effects of miR-181c-5p inhibition in a rodent heart failuremodel.

MiR-181c-5p overexpression associates with heart failure (HF) and cardiac damage, but the underlying pathophysiology remains unclear. This study investigated the effect of miR-181c-5p inhibition on cardiac function and fibrosis in a rodent model of diastolic dysfunction, and evaluated additional effects on kidney as relevant comorbid organ. Diastolic dysfunction was induced in male C57/BL6J mice (n = 20) by combining high-fat diet, L-NG-nitroarginine methyl ester, and angiotensin II administration, and was compared to sham controls (n = 18). Mice were randomized to subcutaneous miR-181c-5p antagomiR (INH) or scrambled antagomiR injections (40 mg/kg/week). HF mice demonstrated diastolic dysfunction and increased fibrosis, which was attenuated by INH treatment. Remarkably, HF + INH animals had a threefold higher mortality rate (60%) compared to HF controls (20%). Histological examination revealed increased glomerular damage in all INH treated mice, and signs of thrombotic microangiopathy (TMA) in mice who died prematurely. Quantitative polymerase chain reaction demonstrated a miR-181c-5p-related downregulation of cardiac but not renal Tgfbr1 in HF + INH mice, while INH treatment reduced renal but not cardiac Vegfa expression in all mice. This study demonstrates cardiac anti-fibrotic effects of miR-181c-5p inhibition in a rodent HF model through targeting of Tgfbr1 in the heart. Despite improved diastolic function, HF + INH mice had higher mortality due to increased predisposition for TMA, increased renal fibrosis and glomerular damage, associated with Vegfa downregulation in kidneys.

Set7 Methyltransferase and Phenotypic Switch in Diabetic Glomerular Endothelial Cells.

Hyperglycaemia influences the development of glomerular endothelial cell damage and nowhere is this more evident than in the progression of diabetic kidney disease (DKD). While the Set7 lysine methyltransferase is a known hyperglycaemic sensor, its role in endothelial cell function in the context of DKD remains poorly understood. Single-cell transcriptomics was used to investigate Set7 regulation in a mouse model of DKD, followed by validation of findings using pharmacological and shRNA inhibition of Set7. Set7 knockout (Set7KO) improved glomerular structure and albuminuria in a mouse model of diabetes. Analysis of single cell RNA-seq (scRNA-seq) data showed dynamic transcriptional changes in diabetic renal cells. Set7KO controls phenotype switching of GEN cell populations through transcriptional regulation of IGFBP5 (Insulin growth factor binding protein 5). Chromatin immunoprecipitation assays confirmed the expression of the IGFBP5 gene was associated with mono- and di-methylation of histone H3 lysine 4 (H3K4me1/2). The generalisability was investigated in human renal and circulating hyperglycaemic cells exposed to TGFβ1. We show that the highly selective Set7 inhibitor, PFI-2, attenuated indices associated with renal cell damage and mesenchymal transition; specifically (i) reactive oxygen species production, (ii) IGFBP5 gene regulation, and (iii) expression of mesenchymal markers. Furthermore, renal benefit observed in Set7KO diabetic mice closely correspond in human GEN cells with PFI-2 inhibition or Set7 shRNA silencing. Set7 regulates the phenotypic endothelial-mesenchymal transition (EDMT) switch and suggest that targeting the lysine methyltransferase could protect glomerular cell injury in DKD.

Dissociation of Hypertension and Renal Damage After Cessation of High-Salt Diet in Dahl Rats.

Every year, thousands of patients with hypertension reduce salt consumption in an effort to control their blood pressure. However, hypertension has a self-sustaining character in a significant part of the population. We hypothesized that chronic hypertension leads to irreversible renal damage that remains after removing the trigger, causing an elevation of the initial blood pressure. Dahl salt-sensitive rat model was used for chronic, continuous observation of blood pressure. Rats were fed a high salt diet to induce hypertension, and then the diet was switched back to normal sodium content. We found that developed hypertension was irreversible by salt cessation: after a short period of reduction, blood pressure grew even higher than in the high-salt phase. Notably, the self-sustaining phase of hypertension was sensitive to benzamil treatment due to sustaining epithelial sodium channel hyperactivity, as shown with patch-clamp analysis. Glomerular damage and proteinuria were also irreversible. In contrast, some mechanisms, contributing to the development of salt-sensitive hypertension, normalized after salt restriction. Thus, flow cytometry demonstrated that dietary salt reduction in hypertensive animals decreased the number of total CD45+, CD3+CD4+, and CD3+CD8+ cells in renal tissues. Also, we found tubular recovery and improvement of glomerular filtration rate in the postsalt period versus a high-salt diet. Based on earlier publications and current data, poor response to salt restriction is due to the differential contribution of the factors recognized in the developmental phase of hypertension. We suggest that proteinuria or electrolyte transport can be prioritized over therapeutic targets of inflammatory response.

Activation of the IL-6/STAT3 pathway contributes to the pathogenesis of membranous nephropathy and is a target for Mahuang Fuzi and Shenzhuo Decoction (MFSD) to repair podocyte damage

BackgroundPrimary membranous nephropathy (PMN) is an autoimmune glomerular disease. IL-6 is a potential therapeutic target for PMN. Previous clinical studies have demonstrated the effectiveness of Mahuang Fuzi and Shenzhuo Decoction (MFSD) in treating membranous nephropathy. However, the mechanism of action of MFSD remains unclear. MethodsSerum IL-6 levels were measured in patients with PMN and healthy subjects. The passive Heymann nephritis (PHN) rat model was established, and high and low doses of MFSD were used for intervention to observe the repair effect of MFSD on renal pathological changes and podocyte injury. RNA-seq was used to screen the possible targets of MFSD, and the effect of MFSD targeting IL-6/STAT3 was further verified by combining the experimental results. Finally, the efficacy of tocilizumab in PHN rats was observed. ResultsSerum IL-6 levels were significantly higher in PMN patients than in healthy subjects. These levels significantly decreased in patients in remission after MFSD treatment. MFSD treatment improved laboratory indicators in PHN rats, as well as glomerular filtration barrier damage and podocyte marker protein expression. Renal transcriptome changes showed that MFSD-targeted differential genes were enriched in JAK/STAT and cytokine-related pathways. MFSD inhibits the IL6/STAT3 pathway in podocytes. Additionally, MFSD significantly reduced serum levels of IL-6 and other cytokines in PHN rats. However, treatment of PHN with tocilizumab did not achieve the expected effect. ConclusionThe IL-6/STAT3 signaling pathway is activated in podocytes of experimental membranous nephropathy. MFSD alleviates podocyte damage by inhibiting the IL-6/STAT3 pathway.

Combination of Fushengong decoction with Western medicine on patients with chronic renal failure: An observational study.

Chronic renal failure (CRF) causes a reduction in glomerular filtration rate and damage to renal parenchyma. Fushengong decoction (FSGD) showed improvement in renal function in CRF rats. This study aims to analyze the differentially expressed proteins in CRF patients treated with Western medicine alone or in combination with FSGD. Sixty patients with CRF recruited from Yongchuan Traditional Chinese Medicine Hospital affiliated to Chongqing Medical University were randomly assigned into control (treated with Western medicine alone) and observation groups (received additional FSGD treatment thrice daily for 8 weeks). The clinical efficacy and changes in serum Bun, serum creatinine, Cystatin C, and transforming growth factor beta 1 (TGF-β1) before and after treatment were observed. We employed isotope relative labeling absolute quantification labeling and liquid chromatography-mass spectrometry to identify differentially expressed proteins and carried out bioinformatics Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses. Patients in the observation group showed greater clinical improvement and lower levels of serum Bun, serum creatinine, Cyc-c, and TGF-β1 than the control group. We identified 32 differentially up-regulated and 52 down-regulated proteins in the observation group. These proteins are involved in the blood coagulation system, protein serine/threonine kinase activity, and TGF-β, which are closely related to the pathogenesis of CRF. Protein-protein-interaction network analysis indicated that candidate proteins fibronectin 1, fibrinogen alpha chain, vitronectin, and Serpin Family C Member 1 were in the key nodes. This study provided an experimental basis suggesting that FSGD combined with Western medicine could significantly improve renal function and renal fibrosis of CRF patients, which may be through the regulation of fibronectin 1, fibrinogen alpha chain, vitronectin, Serpin Family C Member 1, TGF-β, and the complement coagulation pathway (see Graphical abstract S1, Supplemental Digital Content, http://links.lww.com/MD/L947).

Phloretamide Protects against Diabetic Kidney Damage and Dysfunction in Diabetic Rats by Attenuating Hyperglycemia and Hyperlipidemia, Suppressing NF-κβ, and Upregulating Nrf2.

Potent hypoglycemic and antioxidant effects were recently reported for the apple-derived phenolic compound phloretamide (PLTM). The renoprotective effects of this compound are yet to be shown. This study aimed to examine the potential of PLTM to prevent diabetic nephropathy in streptozotocin-induced diabetic rats and to examine the possible mechanisms of protection. Non-diabetic and STZ-diabetic male rats were treated orally by gavage with either the vehicle or with PTLM (200 mg/kg; twice/week) for 12 weeks. PTLM significantly increased urine volume and prevented glomerular and tubular damage and vacuolization in STZ-diabetic rats. It also increased creatinine excretion and reduced urinary albumin levels and the renal levels of kidney injury molecule-1 (KIM-1), 8-hydroxy-2'-deoxyguanosine (8-OHdG), neutrophil gelatinase-associated lipocalin (NGAL), and nephrin in the diabetic rats. PTLM also prevented an increase in the nuclear levels of NF-κβ, as well as the total levels of tumor necrosis factor-alpha (TNF-α), interleukin 6 (IL-6), caspase-3, and Bax in the kidneys of diabetic rats. These effects were associated with reduced serum levels of triglycerides, cholesterol, and low-density lipoprotein cholesterol. In both the control and diabetic rats, PTLM significantly reduced fasting plasma glucose and enhanced the renal mRNA and cytoplasmic levels of Nrf2, as well as the levels of Bcl2, superoxide dismutase (SOD), and glutathione (GSH). However, PTLM failed to alter the cytoplasmic levels of keap1 in diabetic rats. In conclusion, PTLM prevents renal damage and dysfunction in STZ-diabetic rats through its hypoglycemic and hypolipidemic activities, as well as through its antioxidant potential, which is mediated by activating the Nrf2/antioxidant axis.

Nlrp2 deletion ameliorates kidney damage in a mouse model of cystinosis.

Cystinosis is a rare autosomal recessive disorder caused by mutations in the CTNS gene that encodes cystinosin, a ubiquitous lysosomal cystine/H+ antiporter. The hallmark of the disease is progressive accumulation of cystine and cystine crystals in virtually all tissues. At the kidney level, human cystinosis is characterized by the development of renal Fanconi syndrome and progressive glomerular and interstitial damage leading to end-stage kidney disease in the second or third decade of life. The exact molecular mechanisms involved in the pathogenesis of renal disease in cystinosis are incompletely elucidated. We have previously shown upregulation of NLRP2 in human cystinotic proximal tubular epithelial cells and its role in promoting inflammatory and profibrotic responses. Herein, we have investigated the role of NLRP2 in vivo using a mouse model of cystinosis in which we have confirmed upregulation of Nlrp2 in the renal parenchyma. Our studies show that double knock out Ctns-/- Nlrp2-/- animals exhibit delayed development of Fanconi syndrome and kidney tissue damage. Specifically, we observed at 4-6 months of age that animals had less glucosuria and calciuria and markedly preserved renal tissue, as assessed by significantly lower levels of inflammatory cell infiltration, tubular atrophy, and interstitial fibrosis. Also, the mRNA expression of some inflammatory mediators (Cxcl1 and Saa1) and the rate of apoptosis were significantly decreased in 4-6-month old kidneys harvested from Ctns-/- Nlrp2-/- mice compared to those obtained from Ctns-/-mice. At 12-14 months of age, renal histological was markedly altered in both genetic models, although double KO animals had lower degree of polyuria and low molecular weight proteinuria and decreased mRNA expression levels of Il6 and Mcp1. Altogether, these data indicate that Nlrp2 is a potential pharmacological target for delaying progression of kidney disease in cystinosis.

Application of atmospheric cold plasma for zearalenone detoxification in cereals: Kinetics, mechanisms, and cytotoxicity analysis

IntroductionZearalenone (ZEN) is one of the most widely contaminated mycotoxins in world, posing a severe threat to human and animal health. Atmospheric cold plasma (ACP) holds great penitential in mycotoxin degradation. ObjectivesThis study aimed to investigate the degradation efficiency and mechanisms of ACP on ZEN as well as the cytotoxicity of ZEN degradation products by ACP. Additionally, this study also investigated the degradation efficiency of ACP on ZEN in cereals and its effect on cereal quality. MethodsThe degradation efficiency and products of ZEN by ACP was analyzed by HPLC and LC-MS/MS. The human normal liver cells and mice were employed to assess the cytotoxicity of ZEN degradation products. The ZEN artificially contaminated cereals were used to evaluate the feasibility of ACP detoxification in cereals. ResultsThe results showed that the degradation rate of ZEN was 96.18 % after 30-W ACP treatment for 180 s. The degradation rate was dependent on the discharge power, and treatment time and distance. Four major ZEN degradation products were produced after ACP treatment due to the oxidative destruction of CC double bond, namely C18H22O7 (m/z = 351.19), C18H22O8 (m/z = 367.14), C18H22O6 (m/z = 335.14), and C17H20O6 (m/z = 321.19). L02 cell viability was increased from 52.4 % to 99.76 % with ACP treatment time ranging from 0 to 180 s. Mice results showed significant recovery of body weight and depth of colonic crypts as well as mitigation of glomerular and liver damage. Additionally, ACP removed up to 50.55 % and 58.07 % of ZEN from wheat and corn. ConclusionsThis study demonstrates that ACP could efficiently degrade ZEN in cereals and its cytotoxicity was significantly reduced. Therefore, ACP is a promising effective method for ZEN detoxification in cereals to ensure human and animal health. Future study needs to develop large-scale ACP device with high degradation efficiency.