EGR3 reduces podocyte inflammatory damage in obesity related glomerulopathy by inhibiting the PRMT1/p-STAT3 pathway. / EGR3通过抑制PRMT1/p-STAT3é€šè·¯å‡è½»è‚¥èƒ–ç›¸å ³æ€§è‚¾ç— è¶³ç»†èƒžç‚Žç—‡æŸä¼¤.

OBJECTIVES: Obesity related glomerulopathy (ORG) is induced by obesity, but the pathogenesis remains unclear. This study aims to investigate the expression of early growth response protein 3 (EGR3) in the renal cortex tissues of ORG patients and high-fat diet-induced obese mice, and to further explore the molecular mechanism of EGR3 in inhibiting palmitic acid (PA) induced human podocyte inflammatory damage. METHODS: Renal cortex tissues were collected from ORG patients (n=6) who have been excluded from kidney damage caused by other diseases and confirmed by histopathology, and from obese mice induced by high-fat diet (n=10). Human and mouse podocytes were intervened with 150 µmol/L PA for 48 hours. EGR3 was overexpressed or silenced in human podocytes. Enzyme linked immunosorbent assay (ELISA) was used to detcet the levels of interleukin-6 (IL-6) and interleukin-1ß (IL-1ß). Real-time RT-PCR was used to detect the mRNA expressions of EGR3, podocytes molecular markers nephrosis 1 (NPHS1), nephrosis 2 (NPHS2), podocalyxin (PODXL), and podoplanin (PDPN). RNA-seq was performed to detect differentially expressed genes (DEGs) after human podocytes overexpressing EGR3 and treated with 150 µmol/L PA compared with the control group. Co-immunoprecipitation (Co-IP) combined with liquid chromatography tandem mass spectrometry (LC-MS) was used to detect potential interacting proteins of EGR3 and the intersected with the RNA-seq results. Co-IP confirmed the interaction between EGR3 and protein arginine methyltransferases 1 (PRMT1), after silencing EGR3 and PRMT1 inhibitor intervention, the secretion of IL-6 and IL-1ß in PA-induced podocytes was detected. Western blotting was used to detect the expression of phosphorylated signal transducer and activator of transcription 3 (p-STAT3) after overexpression or silencing of EGR3. RESULTS: EGR3 was significantly upregulated in renal cortex tissues of ORG patients and high-fat diet-induced obese mice (both P<0.01). In addition, after treating with 150 µmol/L PA for 48 hours, the expression of EGR3 in human and mouse podocytes was significantly upregulated (both P<0.05). Overexpression or silencing of EGR3 in human podocytes inhibited or promoted the secretion of IL-6 and IL-1ß in the cell culture supernatant after PA intervention, respectively, and upregulated or downregulated the expression of NPHS1, PODXL, NPHS2,and PDPN (all P<0.05). RNA-seq showed a total of 988 DEGs, and Co-IP+LC-MS identified a total of 238 proteins that may interact with EGR3. Co-IP confirmed that PRMT1 was an interacting protein with EGR3. Furthermore, PRMT1 inhibitors could partially reduce PA-induced IL-6 and IL-1ß secretion after EGR3 silencing in human podocytes (both P<0.05). Overexpression or silencing of EGR3 negatively regulated the expression of PRMT1 and p-STAT3. CONCLUSIONS: EGR3 may reduce ORG podocyte inflammatory damage by inhibiting the PRMT1/p-STAT3 pathway.

Effects of furosemide, acetazolamide and amiloride on renal cortical and medullary tissue oxygenation in non-anaesthetised healthy sheep.

It has been proposed that diuretics can improve renal tissue oxygenation through inhibition of tubular sodium reabsorption and reduced metabolic demand. However, the impact of clinically used diuretic drugs on the renal cortical and medullary microcirculation is unclear. Therefore, we examined the effects of three commonly used diuretics, at clinically relevant doses, on renal cortical and medullary perfusion and oxygenation in non-anaesthetised healthy sheep. Merino ewes received acetazolamide (250 mg; n = 9), furosemide (20 mg; n = 10) or amiloride (10 mg; n = 7) intravenously. Systemic and renal haemodynamics, renal cortical and medullary tissue perfusion and P O 2 ${P_{{{\mathrm{O}}_{\mathrm{2}}}}}$ , and renal function were then monitored for up to 8 h post-treatment. The peak diuretic response occurred 2 h (99.4 ± 14.8 mL/h) after acetazolamide, at which stage cortical and medullary tissue perfusion and P O 2 ${P_{{{\mathrm{O}}_{\mathrm{2}}}}}$ were not significantly different from their baseline levels. The peak diuretic response to furosemide occurred at 1 h (196.5 ± 12.3 mL/h) post-treatment but there were no significant changes in cortical and medullary tissue oxygenation during this period. However, cortical tissue P O 2 ${P_{{{\mathrm{O}}_{\mathrm{2}}}}}$ fell from 40.1 ± 3.8 mmHg at baseline to 17.2 ± 4.4 mmHg at 3 h and to 20.5 ± 5.3 mmHg at 6 h after furosemide administration. Amiloride did not produce a diuretic response and was not associated with significant changes in cortical or medullary tissue oxygenation. In conclusion, clinically relevant doses of diuretic agents did not improve regional renal tissue oxygenation in healthy animals during the 8 h experimentation period. On the contrary, rebound renal cortical hypoxia may develop after dissipation of furosemide-induced diuresis.

The Ischemia and Reperfusion Injury Involves the Toll-Like Receptor-4 Participation Mainly in the Kidney Cortex.

BACKGROUND/AIMS: The renal inflammatory response and kidney regeneration in ischemia-reperfusion injury (IRI) are associated with Toll-like receptor 4 (TLR4). Here we study the role of TLR4 during IRI in the renal cortex and medulla separately, using wild-type (TLR4-WT) and Knockout (TLR4-KO) TLR4 mice. METHODS: We used 30 minutes of bilateral renal ischemia, followed by 48 hours of reperfusion in C57BL/6 mice. We measured the expression of elements associated with kidney injury, inflammation, macrophage polarization, mesenchymal transition, and proteostasis in the renal cortex and medulla by qRT-PCR and Western blot. In addition, we studied kidney morphology by H/E and PAS. RESULTS: Renal ischemia (30min) and reperfusion (48hrs) induced the mRNA and protein of TLR4 in the renal cortex. In addition, Serum Creatinine (SCr), blood urea nitrogen (BUN), Neutrophil gelatinase-associated lipocalin (NGAL), and acute tubular necrosis (ATN) were increased in TLR4-WT by IRI. Interestingly, the SCr and BUN had normal levels in TLR-KO during IRI. However, ATN and high levels of NGAL were present in the kidneys of TLR4-KO mice. The pro-inflammatory (IL-6 and TNF-α) and anti-inflammatory (Foxp3 and IL-10) markers increased by IRI only in the cortex of TLR4-WT but not in TLR4-KO mice. Furthermore, the M1 (CD38 and Frp2) and M2 (Arg-I, Erg-2, and c-Myc) macrophage markers increased by IRI only in the cortex of TLR4-WT. The TLR4-KO blunted the IRI-upregulation of M1 but not the M2 macrophage polarization. Vimentin increased in the renal cortex and medulla of TLR4-WT animals but not in the cortex of TLR4-KO mice. In addition, iNOS and clusterin were increased by IRI only in the cortex of TLR4-WT, and the absence of TLR4 inhibited only clusterin upregulation. Finally, Hsp27 and Hsp70 protein levels increased by IRI in the cortex and medulla of TLR4-WT and TRL4-KO lost the IRI-upregulation of Hsp70. In summary, TLR4 participates in renal ischemia and reperfusion through pro-inflammatory and anti-inflammatory responses inducing impaired kidney function (SCr and BUN). However, the IRI-upregulation of M2 macrophage markers (cortex), iNOS (cortex), IL-6 (medulla), vimentin (medulla), and Hsp27 (cortex and medulla) were independent of TLR4. CONCLUSION: The TLR4 inactivation during IRI prevented the loss of renal function due to the inactivation of inflammation response, avoiding M1 and preserving the M2 macrophage polarization in the renal cortex.

1400W Prevents Renal Injury in the Renal Cortex But Not in the Medulla in a Murine Model of Ischemia and Reperfusion Injury.

BACKGROUND/AIMS: Acute kidney injury (AKI) carries high morbidity and mortality, and the inducible nitric oxide synthase (iNOS) is a potential molecular target to prevent kidney dysfunction. In previous work, we reported that the pharmacological inhibitions of iNOS before ischemia/reperfusion (I/R) attenuate the I/R-induced AKI in mice. Here, we study the iNOS inhibitor 1400W [N-(3-(Aminomethyl)benzyl] acetamide, which has been described to be much more specific to iNOS inhibition than other compounds. METHODS: We used 30 minutes of bilateral renal ischemia, followed by 24 hours of reperfusion in Balb/c mice. 1400w (10 mg/kg i.p) was applied before I/R injury. We measured the expression of elements associated with kidney injury, inflammation, macrophage polarization, mesenchymal transition, and nephrogenic genes by qRT-PCR in the renal cortex and medulla. The Periodic Acid-Schiff (PAS) was used to study the kidney morphology. RESULTS: Remarkably, we found that 1400W affects the renal cortex and medulla in different ways. Thus, in the renal cortex, 1400W prevented the I/R-upregulation of 1. NGAL, Clusterin, and signs of morphological damage; 2. IL-6 and TNF-α; 3. TGF-ß; 4. M2(Arg1, Erg2, cMyc) and M1(CD38, Fpr2) macrophage polarization makers; and 5. Vimentin and FGF2 levels but not in the renal medulla. CONCLUSION: 1400W conferred protection in the kidney cortex compared to the kidney medulla. The present investigation provides relevant information to understand the opportunity to use 1400W as a therapeutic approach in AKI treatment.

Analysis of the Hypoxic Response in a Mouse Cortical Collecting Duct-Derived Cell Line Suggests That Esrra Is Partially Involved in Hif1α-Mediated Hypoxia-Inducible Gene Expression in mCCDcl1 Cells.

The kidney is strongly dependent on a continuous oxygen supply, and is conversely highly sensitive to hypoxia. Controlled oxygen gradients are essential for renal control of solutes and urine-concentrating mechanisms, which also depend on various hormones including aldosterone. The cortical collecting duct (CCD) is part of the aldosterone-sensitive distal nephron and possesses a key function in fine-tuned distal salt handling. It is well known that aldosterone is consistently decreased upon hypoxia. Furthermore, a recent study reported a hypoxia-dependent down-regulation of sodium currents within CCD cells. We thus investigated the possibility that cells from the cortical collecting duct are responsive to hypoxia, using the mouse cortical collecting duct cell line mCCDcl1 as a model. By analyzing the hypoxia-dependent transcriptome of mCCDcl1 cells, we found a large number of differentially-expressed genes (3086 in total logFC< −1 or >1) following 24 h of hypoxic conditions (0.2% O2). A gene ontology analysis of the differentially-regulated pathways revealed a strong decrease in oxygen-linked processes such as ATP metabolic functions, oxidative phosphorylation, and cellular and aerobic respiration, while pathways associated with hypoxic responses were robustly increased. The most pronounced regulated genes were confirmed by RT-qPCR. The low expression levels of Epas1 under both normoxic and hypoxic conditions suggest that Hif-1α, rather than Hif-2α, mediates the hypoxic response in mCCDcl1 cells. Accordingly, we generated shRNA-mediated Hif-1α knockdown cells and found Hif-1α to be responsible for the hypoxic induction of established hypoxically-induced genes. Interestingly, we could show that following shRNA-mediated knockdown of Esrra, Hif-1α protein levels were unaffected, but the gene expression levels of Egln3 and Serpine1 were significantly reduced, indicating that Esrra might contribute to the hypoxia-mediated expression of these and possibly other genes. Collectively, mCCDcl1 cells display a broad response to hypoxia and represent an adequate cellular model to study additional factors regulating the response to hypoxia.

Supplementation of amino acids and organic acids prevents the increase in blood pressure induced by high salt in Dahl salt-sensitive rats.

A high-salt (HS) diet leads to metabolic disorders in Dahl salt-sensitive (SS) rats, and promotes the development of hypertension. According to the changes in the metabolites of SS rats, a set of combined dietary supplements containing amino acids and organic acids (AO) were designed. The purpose of the present study was to evaluate the effect of AO supplementation on the blood pressure of SS rats after the HS diet and clarify the mechanism of AO by metabolomics and biochemical analyses. The results showed that AO supplementation avoided the elevation of blood pressure induced by the HS diet in SS rats, increased the renal antioxidant enzyme activities (catalase, superoxide dismutase, glutathione reductase, and glutathione S-transferase), reduced the H2O2 and MDA levels, and restored the normal antioxidant status of the serum and kidneys. AO also reversed the decrease in the nitric oxide (NO) levels and NO synthase activity induced by the HS feed, which involved the L-arginine/NO pathway. Metabolomics analysis showed that AO administration increased the levels of amino acids such as cysteine, glycine, hypotaurine, and lysine in the renal medulla and the levels of leucine, isoleucine, and serine in the renal cortex. Of note, lysine, hypotaurine and glycine had higher metabolic centrality in the metabolic correlation network of the renal medulla after AO administration. In conclusion, AO intervention could prevent HS diet-induced hypertension in SS rats by restoring the metabolic homeostasis of the kidneys. Hence, AO has the potential to become a functional food additive to improve salt-sensitive hypertension.

Detection of impaired renal allograft function in paediatric and young adult patients using arterial spin labelling MRI (ASL-MRI).

The study aimed to discriminate renal allografts with impaired function by measuring cortical renal blood flow (cRBF) using magnetic resonance imaging arterial spin labelling (ASL-MRI) in paediatric and young adult patients. We included 18 subjects and performed ASL-MRI on 1.5 T MRI to calculate cRBF on parameter maps. cRBF was correlated to calculated glomerular filtration rate (GFR) and compared between patient groups with good (GFR ≥ 60 mL/min/1.73 m2) and impaired allograft function (GFR < 60 mL/min/1.73 m2). Mean cRBF in patients with good allograft function was significantly higher than in patients with impaired allograft function (219.89 ± 57.24 mL/min/100 g vs. 146.22 ± 41.84 mL/min/100 g, p < 0.008), showing a highly significant correlation with GFR in all subjects (r = 0.75, p < 0.0001). Also, the diffusion-weighted imaging (DWI-MRI) apparent diffusion coefficient (ADC) and Doppler measurements of peak-systolic and end-diastolic velocities and the resistive index (PS, ED, RI) were performed and both methods showed no significant difference between groups. ADC implied no correlation with GFR (r = 0.198, p = 0.464), while PS indicated moderate correlation to GFR (r = 0.48, p < 0.05), and PS and ED moderate correlation to cRBF (r = 0.58, p < 0.05, r = 0.56, p < 0.05, respectively). Cortical perfusion as non-invasively measured by ASL-MRI differs between patients with good and impaired allograft function and correlates significantly with its function.

Identification of multinucleated cells in human kidney cortex: A way for tissue repairing?

The presence of multinucleated cells has never been demonstrated in renal tissue, although, polyploid cells were recently observed in the tubules of normal and pathological human kidney. Therefore, the aim of the present study is to identify and quantify, by electron microscopy, multinucleated cells in the cortical tissue of normal human kidney i.e., in the three compartments of renal tubule: the proximal tubule (PT), the distal tubule (DT), and the collecting duct (CD), as well as, in the glomerulus (podocytes). The percentage of the multinucleated cells observed was 5% (95%CI: 3.6%-6.7%) in renal cortical tubules with distribution in each tubular compartment of 6% in PT, 4% in DT and 3% in CD with no statistically significant difference in the distribution of multinucleated cells according to tubular compartments. Four percent of analysed podocytes (in total 149 podocytes) were multinucleated (95%CI: 1.5%-8.6%). In conclusion, multinucleated cells were identified and quantified in functionally normal kidneys, as previously demonstrated in other organs such as the liver.

Asymmetric Opening of Mitochondrial Permeability Transition Pore in Mouse Brain Hemispheres: A Link to the Mitochondrial Calcium Uniporter Complex

Abstract The prolonged entry of large amounts of calcium into the mitochondria through the mitochondrial calcium uniporter complex (MCUC) may cause the permeability transition pore (mPTP) to open, which contributes to the pathogenesis of several diseases. Tissue-specific differences in mPTP opening due to variable expression of MCUC components may contribute to disease outcomes. We designed this study to determine differential mPTP opening in mitochondria isolated from different regions of mouse brain and kidney and to compare it with the expression of MCUC components. mPTP opening was measured using mitochondria isolated from the left/right brain hemispheres (LH/RH, respectively) and from kidney cortex/medulla, while the expression level of MCUC components was assessed from total cellular RNA. Interestingly, LH mitochondria showed less calcium-induced mPTP opening as compared to RH mitochondria at two different calcium concentrations. Conversely, mPTP opening was similar in the renal cortex and renal medulla mitochondria. However, the kidney mitochondria demonstrated bigger and faster mPTP opening as compared to the brain mitochondria. Furthermore, asymmetric mPTP opening in the LH and RH mitochondria was not associated with the expression of MCUC components. In brief, this study demonstrates thus far unreported asymmetric mPTP opening in mouse brain hemispheres that is not associated with the mRNA levels of MCUC components.

Integrative transcriptomic profiling of a mouse model of hypertension-accelerated diabetic kidney disease.

The current understanding of molecular mechanisms driving diabetic kidney disease (DKD) is limited, partly due to the complex structure of the kidney. To identify genes and signalling pathways involved in the progression of DKD, we compared kidney cortical versus glomerular transcriptome profiles in uninephrectomized (UNx) db/db mouse models of early-stage (UNx only) and advanced [UNxplus adeno-associated virus-mediated renin-1 overexpression (UNx-Renin)] DKD using RNAseq. Compared to normoglycemic db/m mice, db/db UNx and db/db UNx-Renin mice showed marked changes in their kidney cortical and glomerular gene expression profiles. UNx-Renin mice displayed more marked perturbations in gene components associated with the activation of the immune system and enhanced extracellular matrix remodelling, supporting histological hallmarks of progressive DKD in this model. Single-nucleus RNAseq enabled the linking of transcriptome profiles to specific kidney cell types. In conclusion, integration of RNAseq at the cortical, glomerular and single-nucleus level provides an enhanced resolution of molecular signalling pathways associated with disease progression in preclinical models of DKD, and may thus be advantageous for identifying novel therapeutic targets in DKD.

A Higher Foci Density of Interstitial Fibrosis and Tubular Atrophy Predicts Progressive CKD after a Radical Nephrectomy for Tumor.

BACKGROUND: Chronic tubulointerstitial injury on kidney biopsy is usually quantified by the percentage of cortex with interstitial fibrosis/tubular atrophy (IF/TA). Whether other patterns of IF/TA or inflammation in the tubulointerstitium have prognostic importance beyond percentage IF/TA is unclear. METHODS: We obtained, stained, and digitally scanned full cortical thickness wedge sections of renal parenchyma from patients who underwent a radical nephrectomy for a tumor over 2000-2015, and morphometrically analyzed the tubulointerstitium of the cortex for percentage IF/TA, IF/TA density (foci per mm2 cortex), percentage subcapsular IF/TA, striped IF/TA, percentage inflammation (both within and outside IF/TA regions), and percentage subcapsular inflammation. Patients were followed with visits every 6-12 months. Progressive CKD was defined as dialysis, kidney transplantation, or 40% decline from the postnephrectomy eGFR. Cox models assessed the risk of CKD or noncancer mortality with morphometric measures of tubulointerstitial injury after adjustment for the percentage IF/TA and clinical characteristics. RESULTS: Among 936 patients (mean age, 64 years; postnephrectomy baseline eGFR, 48 ml/min per 1.73m2), 117 progressive CKD events and 183 noncancer deaths occurred over a median 6.4 years. Higher IF/TA density predicted both progressive CKD and noncancer mortality after adjustment for percentage IF/TA and predicted progressive CKD after further adjustment for clinical characteristics. Independent of percentage IF/TA, age, and sex, higher IF/TA density correlated with lower eGFR, smaller nonsclerosed glomeruli, more global glomerulosclerosis, and smaller total cortical volume. CONCLUSIONS: Higher density of IF/TA foci (a more scattered pattern with more and smaller foci) predicts higher risk of progressive CKD after radical nephrectomy compared with the same percentage of IF/TA but with fewer and larger foci.

Overexpressing STAMP2 attenuates diabetic renal injuries via upregulating autophagy in diabetic rats.

Diabetic nephropathy (DN) is one of the most serious and major renal complications of diabetes. Previously, Six-transmembrane Protein of Prostate 2 (STAMP2) was reported to contribute to nutritional stress. The purpose of this study is to investigate whether overexpression of STAMP2 attenuates diabetic renal injuries in DN rats. We induced the DN rat model by high-fat diet and low-dose streptozotocin and evaluated the metabolite and urine albumin/creatinine. Recombinant adeno-associated virus vectors were injected for overexpression of STAMP2. Pathophysiologic and ultrastructure features of DN by histochemical stain and transmission electron microscope, autophagy-related proteins and signaling pathway by western blotting were assessed. We found the expression of STAMP2 was decreased and autophagy was blunted in DN rat kidneys. Overexpressing STAMP2 significantly ameliorated metabolic disturbance, insulin resistance, and specifically restoring diabetic renal injury. Furthermore, overexpressing STAMP2 improved the autophagy deficiency in DN rats, as revealed by changes in the expressions of Beclin1, p62, and LC3. Furthermore, STAMP2 overexpressing promoted autophagy by inhibiting the mTOR and activating the AMPK/SIRT1 signaling pathway. Our results suggested that STAMP2 overexpression attenuated renal injuries via upregulating autophagy in DN rats. STAMP2 overexpressing promoted autophagy may been involved with inhibition of the mTOR/ULK1 and activation of the AMPK/SIRT1 signaling pathway.

Renal Cortical Perfusion Estimated in Color Doppler Dynamic Tissue Perfusion Measurement in Patients Treated with Levothyroxine Following Total Thyroidectomy for Resectable Thyroid Cancer Is Independently Associated with Free Thyroxine: A Single-Center Prospective Study.

BACKGROUND The thyroid state significantly influences renal function. However, a direct link between thyroid and kidney dysfunction has not been identified. Thyroid hormones affect cardiac output and vascular resistance, and thus can modify kidney perfusion. This prospective study aimed to test the association between renal cortical perfusion (RCP) estimated in color Doppler sonographic dynamic tissue perfusion measurement (DTPM) with thyroid hormones in 36 patients treated with levothyroxine following total thyroidectomy for resectable thyroid cancer. MATERIAL AND METHODS Blood tests, blood pressure monitoring, and DTPM of the renal cortex were performed. To exclude possible reading errors, the intrarater reliability of the ultrasound perfusion measurement method was estimated. RESULTS The absolute difference between the 2 ultrasound RCP measurements was 5.2±4.4%. RCP correlated significantly with free thyroxine (FT4) (r=0.46; p=0.006) but not with triiodothyronine and thyroid-stimulating hormone. In the adjusted to age backward stepwise multivariable regression analysis model, including estimated glomerular filtration rate, mean arterial pressure, and FT4, only FT4 was independently associated with RCP (R²=0.21; p=0.006). CONCLUSIONS Renal cortical perfusion is independently associated with free thyroxine, which can contribute to renal function abnormalities in the condition of impaired thyroid function. This small prospective study from a single center showed that the renal cortex's color Doppler sonographic dynamic tissue perfusion measurement had very good intraobserver reproducibility.

Quantitative Evaluation of Renal Cortex Perfusion Using Contrast-Enhanced Ultrasound Imaging Parameters in Ischemia-Reperfusion Injury in Rabbits.

The aim of this study was to evaluate blood perfusion of the renal cortex during ischemia-reperfusion (I/R) injury using quantitative contrast-enhanced ultrasound (CEUS) parameters. In this experiment, 24 rabbits were randomly divided into four groups (N = 6): sham-operated group, 24-h post-operation for I/R injury group (24-h I/R), 3-d post-operation for I/R injury group (3-d I/R) and 5 d post-operation for I/R injury group (5 d I/R). All quantitative CEUS parameters were monitored and included the gradient from the start frame to the peak frame (Grad), area under the curve (Area), time-to-peak (TTP), difference between B(intercept intensity at t=0) and A(the intensity attenuation t= 0) and arrival time (AT). Subsequently, we analyzed the changes in these parameters, as well as the correlation between changes in CEUS parameters and pathological parameters. AT and TTP values peaked 3 d after I/R surgery, which correlated with the most significant pathological changes at the same time point. These parameters (such as AT, TTP and Grad) may be useful in dynamically monitoring the severity of tissue damage at the early stage of I/R injury.

Astragaloside IV ameliorates diabetic nephropathy in db/db mice by inhibiting NLRP3 inflammasome­mediated inflammation.

Diabetic nephropathy (DN) is a primary cause of end­stage renal disease. Despite the beneficial effects of astragaloside IV (AS)­IV on renal disease, the underlying mechanism of its protective effects against DN has not been fully determined. The aims of the present study were to assess the effects of AS­IV against DN in db/db mice and to explore the mechanism of AS­IV involving the NLR family pyrin domain containing 3 (NLRP3), caspase­1 and interleukin (IL)­1ß pathways. The 8­week­old db/db mice received 40 mg/kg AS­IV once a day for 12 weeks via intragastric administration. Cultured mouse podocytes were used to further confirm the underlying mechanism in vitro. AS­IV effectively reduced weight gain, hyperglycemia and the serum triacylglycerol concentration in db/db mice. AS­IV also reduced urinary albumin excretion, urinary albumin­to­creatinine ratio and creatinine clearance rate, as well as improved renal structural changes, accompanied by the upregulation of the podocyte markers podocin and synaptopodin. AS­IV significantly inhibited the expression levels of NLRP3, caspase­1 and IL­1ß in the renal cortex, and reduced the serum levels of tumor necrosis factor (TNF)­α and monocyte chemoattractant protein­1. In high glucose­induced podocytes, AS­IV significantly improved the expression levels of NLRP3, pro­caspase­1 and caspase­1, and inhibited the cell viability decrease in a dose­dependent manner, while NLRP3 overexpression eliminated the effect of AS­IV on podocyte injury and the inhibition of the NLRP3 and caspase­1 pathways. The data obtained from in vivo and in vitro experiments demonstrated that AS­IV ameliorated renal functions and podocyte injury and delayed the development of DN in db/db mice via anti­NLRP3 inflammasome­mediated inflammation.

Inhibition of inducible nitric oxide synthase protects against the deleterious effects of sub-lethal sepsis and ethanol in the cardiorenal system.

We tested the hypothesis that ethanol would aggravate the deleterious effects of sub-lethal cecal ligation and puncture (SL-CLP) sepsis in the cardiorenal system and that inhibition of inducible nitric oxide synthase (iNOS) would prevent such response. Male C57BL/6 mice were treated with ethanol for 12 weeks. One hour before SL-CLP surgery, mice were treated with N6-(1-iminoethyl)-lysine (L-NIL, 5 mg/kg, i.p.), a selective inhibitor of iNOS. A second dose of L-NIL was administered 24 h after SL-CLP surgery. Mice were killed 48 h post surgery and the blood, the renal cortex, and the left ventricle (LV) were collected for biochemical analysis. L-NIL attenuated the increase in serum creatinine levels induced by ethanol, but not by SL-CLP. Ethanol, but not SL-CLP, increased creatine kinase (CK)-MB activity and L-NIL did not prevent this response. In the renal cortex, L-NIL prevented the redox imbalance induced by ethanol and SL-CLP. Inhibition of iNOS also decreased lipoperoxidation induced by ethanol and SL-CLP in the LV. L-NIL prevented the increase of pro-inflammatory cytokines and reactive oxygen species induced by ethanol and (or) SL-CLP in the cardiorenal system, suggesting that iNOS modulated some of the molecular mechanisms that underlie the deleterious effects of both conditions in the cardiorenal system.

Effects of Virgin Olive Oil on Blood Pressure and Renal Aminopeptidase Activities in Male Wistar Rats.

High saturated fat diets have been associated with the development of obesity and hypertension, along with other pathologies related to the metabolic syndrome. In contrast, the Mediterranean diet, characterized by its high content of monounsaturated fatty acids, has been proposed as a dietary factor capable of positively regulating cardiovascular function. These effects have been linked to changes in the local renal renin angiotensin system (RAS) and the activity of the sympathetic nervous system. The main goal of this study was to analyze the role of two dietary fat sources on aminopeptidases activities involved in local kidney RAS. Male Wistar rats (six months old) were fed during 24 weeks with three different diets: the standard diet (S), the standard diet supplemented with virgin olive oil (20%) (VOO), or the standard diet enriched with butter (20%) plus cholesterol (0.1%) (Bch). Kidney samples were separated in medulla and cortex for aminopeptidase activities (AP) assay. Urine samples were collected for routine analysis by chemical tests. Aminopeptidase activities were determined by fluorometric methods in soluble (sol) and membrane-bound (mb) fractions of renal tissue, using arylamide derivatives as substrates. After the experimental period, the systolic blood pressure (SBP) values were similar in standard and VOO animals, and significantly lower than in the Bch group. At the same time, a significant increase in GluAP and IRAP activities were found in renal medulla of Bch animals. However, in VOO group the increase of GluAP activity in renal medulla was lower, while AspAP activity decreased in the renal cortex. Furthermore, the VOO diet also affected other aminopeptidase activities, such as TyrAP and pGluAP, related to the regulation of the sympathetic nervous system and the metabolic rate. These results support the beneficial effect of VOO in the regulation of SBP through changes in local AP activities of the kidney.

Early critical cortical infarction by anti-angiotensin II type 1 receptor antibody: A case report and literature review.

RATIONALE: Anti-angiotensin II type 1 receptor antibodies (AT1R-Abs) have been demonstrated to increase the risk of antibody-mediated rejection. We report a case of AT1R-Ab mediated rejection which caused early critical cortical infarction. PATIENT CONCERNS: A 52-year-old man with end-stage kidney disease underwent preemptive kidney transplantation (KT) from his wife. He had no immunologic risk except ABO incompatibility. Proper desensitization treatment were applied prior to KT. On postoperative day 1, he showed stable clinical course with adequate urine output, but there was no decrease in serum creatinine level and imaging studies showed hypoperfusion in the transplanted kidney. DIAGNOSES: Allograft biopsy revealed total cortical infarction with severe necrotizing vasculitis, but the medullary area was preserved. Serum AT1R-Ab concentration was elevated from 10.9 U/mL before KT to 19.1 U/mL on 7 days after KT. INTERVENTIONS: He was treated with plasmapheresis, intravenous immunoglobulin, rituximab, high-dose methylprednisolone, and bortezomib. OUTCOMES: The treatment showed a partial response, and he was discharged with 7.3 mg/dL creatinine level. At 4 months, his creatinine plateaued at 5.5 mg/dL and AT1R-Ab decreased to 3.6 U/mL. LESSONS: This case highlights the risk of early active antibody-mediated rejection by preformed AT1R-Ab, suggesting its ability to exhibit atypical histopathologic findings, such as total cortical infarction.

Assessment of chronic allograft injury in renal transplantation using diffusional kurtosis imaging.

BACKGROUND: Chronic allograft injury (CAI) is a significant reason for which many grafts were lost. The study was conducted to assess the usefulness of diffusional kurtosis imaging (DKI) technology in the non-invasive assessment of CAI. METHODS: Between February 2019 and October 2019, 110 renal allograft recipients were included to analyze relevant DKI parameters. According to estimated glomerular filtration rate (eGFR) (mL/min/ 1.73 m2) level, they were divided to 3 groups: group 1, eGFR ≥ 60 (n = 10); group 2, eGFR 30-60 (n = 69); group 3, eGFR < 30 (n = 31). We performed DKI on a clinical 3T magnetic resonance imaging system. We measured the area of interest to determine the mean kurtosis (MK), mean diffusivity (MD), and apparent diffusion coefficient (ADC) of the renal cortex and medulla. We performed a Pearson correlation analysis to determine the relationship between eGFR and the DKI parameters. We used the receiver operating characteristic curve to estimate the predicted values of DKI parameters in the CAI evaluation. We randomly selected five patients from group 2 for biopsy to confirm CAI. RESULTS: With the increase of creatinine, ADC, and MD of the cortex and medulla decrease, MK of the cortex and medulla gradually increase. Among the three different eGFR groups, significant differences were found in cortical and medullary MK (P = 0.039, P < 0.001, P < 0.001, respectively). Cortical and medullary ADC and MD are negatively correlated with eGFR (r = - 0.49, - 0.44, - 0.57, - 0.57, respectively; P < 0.001), while cortical and medullary MK are positively correlated with eGFR (r = 0.42, 0.38; P < 0.001). When 0.491 was set as the cutoff value, MK's CAI assessment showed 87% sensitivity and 100% specificity. All five patients randomly selected for biopsy from the second group confirmed glomerulosclerosis and tubular atrophy/interstitial fibrosis. CONCLUSION: The DKI technique is related to eGFR as allograft injury progresses and is expected to become a potential non-invasive method for evaluating CAI.