Unspliced XBP1 Counteracts ß-Catenin to Inhibit Vascular Calcification.

BACKGROUND: Vascular calcification is a prevalent complication in chronic kidney disease and contributes to increased cardiovascular morbidity and mortality. XBP1 (X-box binding protein 1), existing as the XBP1u (unspliced XBP1) and XBP1s (spliced XBP1) forms, is a key component of the endoplasmic reticulum stress involved in vascular diseases. However, whether XBP1u participates in the development of vascular calcification remains unclear. METHODS: We aim to investigate the role of XBP1u in vascular calcification. XBP1u protein levels were reduced in high phosphate-induced calcified vascular smooth muscle cells, calcified aortas from mice with adenine diet-induced chronic renal failure, and calcified radial arteries from patients with chronic renal failure. RESULTS: Inhibition of XBP1u rather than XBP1s upregulated in the expression of the osteogenic markers Runx2 (runt-related transcription factor 2) and Msx2 (msh homeobox 2), and exacerbated high phosphate-induced vascular smooth muscle cell calcification, as verified by calcium deposition and Alizarin red S staining. In contrast, XBP1u overexpression in high phosphate-induced vascular smooth muscle cells significantly inhibited osteogenic differentiation and calcification. Consistently, smooth muscle cell-specific XBP1 deficiency in mice markedly aggravated the adenine diet- and 5/6 nephrectomy-induced vascular calcification compared with that in the control littermates. Further interactome analysis revealed that XBP1u is bound directly to ß-catenin, a key regulator of vascular calcification, via amino acid (aa) 205-230 in its C-terminal degradation domain. XBP1u interacted with ß-catenin to promote its ubiquitin-proteasomal degradation and thus inhibited ß-catenin/TCF (T-cell factor)-mediated Runx2 and Msx2 transcription. Knockdown of ß-catenin abolished the effect of XBP1u deficiency on vascular smooth muscle cell calcification, suggesting a ß-catenin-mediated mechanism. Moreover, the degradation of ß-catenin promoted by XBP1u was independent of GSK-3ß (glycogen synthase kinase 3ß)-involved destruction complex. CONCLUSIONS: Our study identified XBP1u as a novel endogenous inhibitor of vascular calcification by counteracting ß-catenin and promoting its ubiquitin-proteasomal degradation, which represents a new regulatory pathway of ß-catenin and a promising target for vascular calcification treatment.

Icariside II prevents kidney fibrosis development in chronic kidney disease by promoting fatty acid oxidation.

Renal interstitial fibrosis (RIF) is the main pathological basis for the progression of chronic kidney disease (CKD), however, effective interventions are limited. Here, we investigated the effect of Icariside II (ICA-II) on RIF and explored the underlying mechanisms. Rats receiving 5/6 ablation and infarction (A/I) surgery were gavaged with ICA-II (5 or 10 mg/kg) for 8 weeks. In vitro, TGF-ß1-stimulated NRK-52E cells were treated with ICA-II and (or) oleic acid, etomoxir, ranolazine, fenofibrate, and GW6471. The effects of ICA-II on RIF, fatty acid oxidation, lipid deposition, and mitochondrial function were determined by immunoblotting, Oil red O staining, colorimetric, and fluorometric assays. Using adeno-associated virus injection and co-culture methods, we further determined mechanisms of ICA-II anti-RIF. ICA-II ameliorated the fibrotic responses in vivo and in vitro. RNA-seq analysis indicated that ICA-II regulated fatty acid degradation and PPAR pathway in 5/6 (A/I) kidneys. ICA-II attenuated lipid accumulation and up-regulated expression of PPARα, CPT-1α, Acaa2, and Acadsb proteins in vivo and in vitro. Compared to ICA-II treatment, ICA-II combined with Etomoxir exacerbated mitochondrial dysfunction and fibrotic responses in TGF-ß-treated NRK-52E cells. Importantly, we determined that ICA-II improved lipid metabolism, fatty acid oxidation, mitochondrial function, and RIF by restoring PPARα. Co-culture revealed that ICA-II decreased the expression of Fibronectin, Collagen-I, α-SMA, and PCNA proteins in NRK-49F cells by restoring PPARα of renal tubular cells. ICA-II may serve as a promising therapeutic agent for RIF in 5/6 (A/I) rats, which may be important for the prevention and treatment of CKD.

Shenkang injection for the treatment of acute kidney injury: a systematic review and meta-analysis.

OBJECTIVE: Shenkang injection (SKI) has been widely used in China for many years for the treatment of kidney disease. The objective of this systematic review was to assess the efficacy of Shenkang injection for the treatment of acute kidney injury (AKI). METHODS: A search was conducted across seven databases, encompassing data from the inception of each database through October 8th, 2023. Randomized controlled trials comparing SKI-treated AKI patients with control subjects were extracted. The main outcome measure was serum creatinine (SCr) levels. Secondary outcomes included blood urea nitrogen (BUN), serum cystatin C (CysC), 24-h urine protein (24 h-Upro) levels, APACHE II score and adverse reactions. RESULTS: This meta-analysis included eleven studies, and the analysis indicated that, compared with the control group, SKI significantly decreased SCr [WMD = -23.31, 95% CI (-28.06, -18.57); p < 0.001]; BUN [WMD = -2.07, 95% CI (-2.56, -1.57); p < 0.001]; CysC [WMD = -0.55, 95% CI (-0.78, -0.32), p < 0.001]; 24-h urine protein [WMD = -0.43, 95% CI (-0.53, -0.34), p < 0.001]; and the APACHE II score [WMD = -3.07, 95% CI (-3.67, -2.48), p < 0.001]. There was no difference in adverse reactions between the SKI group and the control group [RR = 1.32, 95% CI (0.66, 2.63), p = 0.431]. CONCLUSION: The use of SKI in AKI patients may reduce SCr, BUN, CysC, 24-h Upro levels, and APACHE II scores in AKI patients. The incidence of adverse reactions did not differ from that in the control group. Additional rigorous clinical trials will be necessary in the future to thoroughly evaluate and establish the effectiveness of SKI in the treatment of AKI.

Shen Shuai II recipe improves renal hypoxia to attenuate renal injury in 5/6 renal ablation/infarction rats and effect evaluation using blood oxygenation level-dependent functional magnetic resonance imaging.

Background: Renal hypoxia plays a key role in the progression of chronic kidney disease (CKD). Shen Shuai II Recipe (SSR) has shown good results in the treatment of CKD as a common herbal formula. This study aimed to explore the effect of SSR on renal hypoxia and injury in CKD rats. Methods: Twenty-five Wistar rats underwent 5/6 renal ablation/infarction (A/I) surgery were randomly divided into three groups: 5/6 (A/I), 5/6 (A/I) + losartan (LOS), and 5/6 (A/I) + SSR groups. Another eight normal rats were used as the Sham group. After 8-week corresponding interventions, blood oxygenation level-dependent functional magnetic resonance imaging (BOLD-fMRI) was performed to evaluate renal oxygenation in all rats, and biochemical indicators were used to measure kidney and liver function, hemoglobin, and proteinuria. The expression of fibrosis and hypoxia-related proteins was analyzed using immunoblotting examination. Results: Renal oxygenation, evaluated by BOLD-fMRI as cortical and medullary T2* values (COT2* and MET2*), was decreased in 5/6 (A/I) rats, but increased after SSR treatment. SSR also downregulated the expression of hypoxia-inducible factor-1α (HIF-1α) in 5/6 (A/I) kidneys. With the improvement of renal hypoxia, renal function and fibrosis were improved in 5/6 (A/I) rats, accompanied by reduced proteinuria. Furthermore, the COT2* and MET2* were significantly positively correlated with the levels of creatinine clearance rate (Ccr) and hemoglobin, but negatively associated with the levels of serum creatinine (SCr), blood urea nitrogen (BUN), serum cystatin C (CysC), serum uric acid (UA), 24-h urinary protein (24-h Upr), and urinary albumin:creatinine ratio (UACR). Conclusion: The degree of renal oxygenation reduction is correlated with the severity of renal injury in CKD. SSR can improve renal hypoxia to attenuate renal injury in 5/6 (A/I) rats of CKD.

SDMA attenuates renal tubulointerstitial fibrosis through inhibition of STAT4.

BACKGROUND: Renal tubulointerstitial fibrosis is the hallmark of various chronic kidney diseases. Symmetric dimethylarginine (SDMA) is an independent cardiovascular risk factor in patients with chronic kidney diseases, which is mostly excreted through renal tubules. However, the effect of SDMA on kidneys in a pathological condition is currently unknown. In this study, we investigated the role of SDMA in renal tubulointerstitial fibrosis and explored its underlying mechanisms. METHODS: Mouse unilateral ureteral obstruction (UUO) and unilateral ischemia-reperfusion injury (UIRI) models were established to study renal tubulointerstitial fibrosis. SDMA was injected into kidneys through ureter retrogradely. TGF-ß stimulated human renal epithelial (HK2) cells were used as an in vitro model and treated with SDMA. Signal transducer and activator of transcription-4 (STAT4) was inhibited by berbamine dihydrochloride or siRNA or overexpressed by plasmids in vitro. Masson staining and Western blotting were performed to evaluate renal fibrosis. Quantitative PCR was performed to validate findings derived from RNA sequencing analysis. RESULTS: We observed that SDMA (from 0.01 to 10 µM) dose-dependently inhibited the expression of pro-fibrotic markers in TGF-ß stimulated HK2 cells. Intrarenal administration of SDMA (2.5 µmol/kg or 25 µmol/kg) dose-dependently attenuated renal fibrosis in UUO kidneys. A significant increase in SDMA concentration (from 19.5 to 117.7 nmol/g, p < 0.001) in mouse kidneys was observed after renal injection which was assessed by LC-MS/MS. We further showed that intrarenal administration of SDMA attenuated renal fibrosis in UIRI induced mouse fibrotic kidneys. Through RNA sequencing analysis, we found that the expression of STAT4 was reduced by SDMA in UUO kidneys, which was further confirmed by quantitative PCR and Western blotting analysis in mouse fibrotic kidneys and renal cells. Inhibition of STAT4 by berbamine dihydrochloride (0.3 mg/ml or 3.3 mg/ml) or siRNA reduced the expression of pro-fibrotic markers in TGF-ß stimulated HK2 cells. Furthermore, blockage of STAT4 attenuated the anti-fibrotic effect of SDMA in TGF-ß stimulated HK2 cells. Conversely, overexpression of STAT4 reversed the anti-fibrotic effect of SDMA in TGF-ß stimulated HK2 cells. CONCLUSION: Taken together, our study indicates that renal SDMA ameliorates renal tubulointerstitial fibrosis through inhibition of STAT4.

Correlation of Renal Oxygenation with Renal Function in Chronic Kidney Disease: A Preliminary Prospective Study.

INTRODUCTION: Chronic hypoxia is prevalent in chronic kidney disease (CKD), and blood oxygenation level-dependent magnetic resonance imaging (BOLD-MRI) provides noninvasive evaluation of renal oxygenation. This study aimed to explore the correlation of renal oxygenation evaluated by BOLD-MRI with renal function. METHODS: 97 non-dialysis patients with CKD stages 1-5 and healthy volunteers (HVs) were recruited in the study, all participants without diabetes. Based on their estimated glomerular filtration rate (eGFR), the patients were divided into two groups: CKD stages 1-3 (CKD 1-3) and CKD stages 4-5 (CKD 4-5). We measured cortical and medullary T2* (COT2* and MET2*) values in all participants by BOLD-MRI. Physiological indices were also recorded and compared among three groups. Correlation of T2* values with clinical characteristics was determined. RESULTS: The COT2* values were significantly higher than MET2* values in all participants. The COT2* and MET2* values of three groups were ranked as HV > CKD 1-3> CKD 4-5 (p < 0.0001). There were positive correlations between the COT2* values, MET2* values and eGFR, hemoglobin (r > 0.4, p < 0.01). The 24-h urinary protein (24-h Upr) showed weak correlation with the COT2* value (rs = -0.2301, p = 0.0265) and no correlation with the MET2* value (p > 0.05). Urinary microprotein, including urinary alpha1-microglobulin, urinary beta2-microglobulin (ß2-MG), and urinary retinol-binding protein (RBP), showed strong correlation with COT2* and MET2* values. According to the analysis of receiver operating characteristic curve, the optimal cut-points between HV and CKD 1-3 were "<61.17 ms" (sensitivity: 91.23%, specificity: 100%) for COT2* values and "<35.00 ms" (sensitivity: 77.19%, specificity: 100%) for MET2* values, whereas COT2* values ("<47.34 ms"; sensitivity: 90.00%, specificity: 92.98%) and MET2* values ("<25.09 ms"; sensitivity: 97.50%, specificity: 80.70%) between CKD 1-3 and CKD 4-5. CONCLUSION: The decline of renal oxygenation reflected on T2* values, especially in cortex, may be an effective diagnostic marker for early detection of CKD.

SENP1 protects cisplatin-induced AKI by attenuating apoptosis through regulation of HIF-1α.

BACKGROUND: Acute kidney injury is a clinical syndrome with both high morbidity and mortality. However, the underlying molecular mechanism of AKI is still largely unknown. The role of SENP1 in AKI is unclear, while one of its substrates, HIF-1α possesses nephroprotective effect in AKI. Herein, this study aimed to reveal the role of SENP1/HIF-1α axis in AKI by using both cell and animal models. METHODS: We investigated the effects of AKI on SENP1 expression using clinical samples, and cisplatin-induced AKI model based on mice or HK-2 cells. The influence of SENP1 knockdown or over-expression on cisplatin-induced AKI was studied in vitro and in vivo. Following the exploration of the change in HIF-1α expression brought by AKI, the synergistic effects of SENP1 knockdown and HIF-1α over-expression on AKI were examined. RESULTS: The results showed the up-regulation of SENP1 in clinical specimens, as well as cell and animal models. The knockdown or over-expression of SENP1 in HK-2 cells could promote or inhibit AKI through regulating cell apoptosis, respectively. Moreover, SENP1+/- mice suffered from much more serious AKI compared with mice in wild type group. Furthermore, we found that HIF-1α over-expression could attenuate the promoted cell apoptosis as well as AKI induced by SENP1 knockdown. CONCLUSIONS: we showed that SENP1 provided protection for kidney in AKI via regulating cell apoptosis and through the regulation of HIF-1α. This study could benefit for the understanding of the pathogenesis of AKI and provide potential therapeutic target for AKI treatment.

Randomized controlled trial of nalfurafine for refractory pruritus in hemodialysis patients.

Background: Chronic kidney disease-associated pruritus (CKD-aP) is very common and sometimes refractory to treatment in hemodialysis patients. In a trial conducted in Japan, nalfurafine, effectively reduced itching of treatment-resistant CKD-aP. Our present bridging study aimed to evaluate the efficacy and safety of nalfurafine in Chinese cohort with refractory CKD-aP.Methods: In this phase III, multicenter bridging study conducted at 22 sites in China, 141 Chinese cases with refractory CKD-aP were randomly (2:2:1) assigned to receive 5 µg, 2.5 µg of nalfurafine or a placebo orally for 14 days in a double-blind manner. The primary end point was the mean decrease in the mean visual analogue scale (VAS) from baseline.Results: A total of 141 patients were included. The primary endpoint analysis based on full analysis set (FAS), the difference of mean VAS decrease between 5 µg nalfurafine and placebo group was 11.37 mm (p = .041); the difference of mean VAS decrease between 2.5 µg and placebo group was 8.81 mm, but not statistically significantly different. Both differences were greater than 4.13 mm, which met its predefined success criterion of at least 50% efficacy of the key Japanese clinical trial. The per protocol set (PPS) analysis got similar results. The incidence of adverse drug reactions (ADRs) was 49.1% in 5µg, 38.6% in 2.5 µg and 33.3% in placebo group. The most common ADR was insomnia, seen in 21 of the 114 nalfurafine patients.Conclusions: Oral nalfurafine effectively reduced itching with few significant ADRs in Chinese hemodialysis patients with refractory pruritus.

Salvianolic acid B attenuates tubulointerstitial fibrosis by inhibiting EZH2 to regulate the PTEN/Akt pathway.

CONTEXT: Salvianolic acid B (SAB) can alleviate renal fibrosis and improve the renal function. OBJECTIVE: To investigate the effect of SAB on renal tubulointerstitial fibrosis and explore its underlying mechanisms. MATERIALS AND METHODS: Male C57 mice were subjected to unilateral ureteric obstruction (UUO) and aristolochic acid nephropathy (AAN) for renal fibrosis indication. Vehicle or SAB (10 mg/kg/d, i.p.) were given consecutively for 2 weeks in UUO mice while 4 weeks in AAN mice. The serum creatinine (Scr) and blood urine nitrogen (BUN) were measured. Masson's trichrome staining and the fibrotic markers (FN and α-SMA) were used to evaluate renal fibrosis. NRK-49F cells exposed to 2.5 ng/mL TGF-ß were treated with SAB in the presence or absence of 20 µM 3-DZNep, an inhibitor of EZH2. The protein expression of EZH2, H3k27me3 and PTEN/Akt signaling pathway in renal tissue and NRK-49F cells were measured by Western blots. RESULTS: SAB significantly improved the levels of Scr by 24.3% and BUN by 35.7% in AAN mice. SAB reduced renal interstitial collagen deposition by 34.7% in UUO mice and 72.8% in AAN mice. Both in vivo and in vitro studies demonstrated that SAB suppressed the expression of FN and α-SMA, increased PTEN and decreased the phosphorylation of Akt, which were correlated with the down-regulation of EZH2 and H3k27me3. The inhibition of EZH2 attenuated the anti-fibrotic effects of SAB in NRK-49Fs. CONCLUSION: SAB might have therapeutic potential on renal fibrosis of CKD through inhibiting EZH2, which encourages further clinical trials.

[Notoginsenoside R_1 interferes with renal oxidative stress and Nrf2/HO-1 signaling pathway to alleviate kidney injury in mice with IgA nephropathy and its mechanism].

The purpose of this study was to investigate the effect of notoginsenoside R_1(NGR_1) on alleviating kidney injury by regulating renal oxidative stress and the Nrf2/HO-1 signaling pathway in mice with IgA nephropathy(IgAN) and its mechanism. The mouse model of IgAN was established using a variety of techniques, including continuous bovine serum albumin(BSA) gavage, subcutaneous injections of carbon tetrachloride(CCl_4) castor oil, and tail vein injections of lipopolysaccharide(LPS). After successful modeling, mice with IgAN were randomly separated into a model group, low, medium, and high-dose NGR_1 groups, and a losartan group, and C57BL6 mice were utilized as normal controls. The model and normal groups were given phosphate buffered saline(PBS) by gavage, the NGR_1 groups were given varying dosages of NGR_1 by gavage, and the losartan group was given losartan by gavage for 4 weeks. The 24-hour urine of mice was collected after the last administration, and serum and kidney tissues of mice were taken at the end of the animal experiment. Then urine red blood cell count(URBCC), 24-hour urine protein(24 h protein), serum creatinine(Scr), and blood urea nitrogen(BUN) levels were measured. The enzyme-linked immunosorbent assay(ELISA) was used to detect the levels of galactose-deficient IgA1(Gd-IgA1), kidney injury molecule 1(Kim-1), and neutropil gelatinase-associated lipocalin(NGAL) in the mouse serum. The assay kits were used to detect the levels of malondialdehyde(MDA) and superoxide dismutase(SOD), and immunofluorescence(IF) was used to detect the expression level of glutathione peroxidase 4(GPX4) in the mesangial region. Western blot was used to detect the protein expression of nuclear transcription factor E2 related factor 2(Nrf2)/heme oxygenase 1(HO-1) signaling pathway in the renal tissue. Hematoxylin-eosin(HE) staining was used to observe pathological alterations in the glomerulus of mice. The results revealed that, as compared with the model group, the serum Gd-IgA1 level, URBCC, 24 h protein level, renal damage markers(Kim-1 and NGAL) in the high-dose NGR_1 group decreased obviously and renal function indicators(BUN, Scr) improved significantly. The activity of SOD activity and expression level of GPX4 increased significantly in the high-dose NGR_1 group, whereas the expression level of MDA reduced and protein expression levels of Nrf2 and HO-1 increased. Simultaneously, HE staining of the renal tissue indicated that glomerular damage was greatly decreased in the high-dose NGR_1 group. In conclusion, this study has clarified that NGR_1 may alleviate the kidney injury of mice with IgAN by activating the Nrf2/HO-1 signaling pathway, improving antioxidant capacity, and reducing the level of renal oxidative stress.

Xper computed tomography-guided translumbar inferior vena cava catheterization for long-term hemodialysis: A case report and literature review.

Hemodialysis is the most widely used renal replacement therapy for end-stage renal disease patients. Exhausted vascular access due to repeated indwelling central venous catheters is becoming a challenging clinical problem, which also contributes to reduced survival of the hemodialysis patients. Lack of conventional peripheral and central venous access mandates the use of alternative strategies. We present a case of translumbar dialysis catheter (TLDC) for long-term hemodialysis in a patient with central venous occlusion refractory to conventional endovascular techniques. After a careful literature review, totally 10 cohort studies including 216 cases through TLDC were reported. The incidence of procedure-related complications was very low. The catheter-related infection rate of TLDC was comparable with overall tunneled cuffed catheters (TCCs) reported by clinical practice guidelines for vascular access. Although the patency might be relatively low due to the catheter-related complications, TLDC could be rescued by multiple systemic and topical medications and interventional therapies. Percutaneous translumbar placement of a cuffed tunneled hemodialysis catheter directly into the inferior vena cava (IVC) can provide a relatively safe salvage when traditional central venous sites such as the internal jugular, femoral, subclavian veins are unavailable. Xper computed tomography together with real-time fluoroscopic guidance can reduce the intraoperative risks and complications.

The role of lung ultrasonography in the assessment of overhydration in maintenance hemodialysis patients.

PURPOSE: Existed methods like biochemical markers improve the accuracy of fluid evaluation for the maintenance hemodialysis patients, but none of them has become the gold standard. This study aimed to evaluate the potential of lung ultrasonography as a useful tool for monitoring the volume status of the patients. METHODS: A total of 88 patients undergoing maintenance hemodialytic were enrolled in this prospective observational study. Patients were divided into three groups: overhydration (OH), normohydration, and hypohydration according to bioimpedance spectroscopy. Lung ultrasonography parameters, echocardiography parameters, and clinical characteristics of three groups were analyzed. After an average follow-up of 433 days, all-cause mortality among groups was compared. RESULTS: The total number of lung comets was statistically reduced in patients after dialysis (Z= -6.891, p < 0.001). This reduction was related to ΔOH (OH - ΔW (the weight gain from dry weight)) and echocardiographic parameters, which proved the relationship among the comet-tail, hydration status of body and cardiac performance. The Kappa consistency test showed that lung ultrasonography and bioelectrical spectroscopy had moderate consistency. ROC analysis showed that the best cut-point of lung comet is 13. The pre-/post-dialysis lung comet-tail, cardiac function and total body impedance with all-cause mortality was investigated. Kaplan-Meier's analysis revealed that the all-cause mortality was higher in lung congestion patients. CONCLUSIONS: This study proposes a potentially reliable lung ultrasonography method for estimating fluids overload, which also has implication value of all-cause mortality.

Calycosin plays a protective role in diabetic kidney disease through the regulation of ferroptosis.

CONTEXT: Diabetic kidney disease (DKD) is a devastating complication of diabetes. Renal functional deterioration caused by tubular injury is the primary change associated with this disease. Calycosin shows protective roles in various diseases. OBJECTIVES: This study explored the function and underlying mechanism of calycosin in DKD. MATERIALS AND METHODS: HK-2 cells were treated with 25 mM high glucose (HG) to establish a renal tubule injury cell model. Then, the viability of cells treated with 0, 5, 10, 20, 40 and 80 µM of calycosin was measured using Cell Counting Kit-8. For the in vivo model, db/db mice were treated with 10 and 20 mg/kg/day of calycosin; db/m mice served as controls. The histomorphology was analyzed via haematoxylin and eosin staining. RESULTS: HG-induced decreased expression of glutathione (491.57 ± 33.56 to 122.6 ± 9.78 µmol/mL) and glutathione peroxidase 4 (inhibition rate 92.3%) and increased expression of lactate dehydrogenase (3.85 ± 0.89 to 16.84 ± 2.18 U/mL), malondialdehyde (3.72 ± 0.66 to 18.2 ± 1.58 nmol/mL), lipid ROS (4.31-fold increase) and NCOA4 (7.69-fold increase). The effects induced by HG could be blocked by calycosin. Moreover, calycosin alleviated the HG-induced decrease of cell viability and the increase of lipid ROS, but erastin could block the effects caused by calycosin. The in vivo model showed that calycosin alleviated the renal injury caused by diabetes. DISCUSSION AND CONCLUSION: Calycosin has a protective effect on diabetic kidney disease; ferroptosis may be involved in this process.

[Icariin improves renal interstitial fibrosis in a rat model of chronic renal failure by regulating mitochondrial dynamics].

This study aims to explore the effect of icariin(ICA) on mitochondrial dynamics in a rat model of chronic renal failure(CRF) and to investigate the molecular mechanism of ICA against renal interstitial fibrosis(RIF). CRF was induced in male Sprague-Dawley(SD) rats with 5/6(ablation and infarction, A/I) surgery(right kidney ablation and 2/3 infarction of the left kidney). Four weeks after surgery, the model rats were randomized into the following groups: 5/6(A/I) group, 5/6(A/I)+low-dose ICA group, and 5/6(A/I)+high-dose ICA group. Another 12 rats that received sham operation were randomly classified into 2 groups: sham group and sham+ICAH group. Eight weeks after treatment, the expression of collagen-Ⅰ(Col-Ⅰ), collagen-Ⅲ(Col-Ⅲ), mitochondrial dynamics-related proteins(p-Drp1 S616, p-Drp1 S637, Mfn1, Mfn2), and mitochondrial function-related proteins(TFAM, ATP6) in the remnant kidney tissues was detected by Western blot. The expression of α-smooth muscle actin(α-SMA) was examined by immunohistochemical(IHC) staining. The NRK-52 E cells, a rat proximal renal tubular epithelial cell line, were cultured in vitro and treated with ICA of different concentration. Cell viability was detected by CCK-8 assay. In NRK-52 E cells stimulated with 20 ng·mL~(-1) TGF-ß1 for 24 h, the effect of ICA on fibronectin(Fn), connective tissue growth factor(CTGF), p-Drp1 S616, p-Drp1 S637, Mfn1, Mfn2, TFAM, and ATP6 was detected by Western blot, and the ATP content and the mitochondrial morphology were determined. The 20 ng·mL~(-1) TGF-ß1-stimulated NRK-52 E cells were treated with or without 5 µmol·L~(-1) ICA+10 µmol·L~(-1) mitochondrial fusion promoter M1(MFP-M1) for 24 h and the expression of fibrosis markers Fn and CTGF was detected by Western blot. Western blot result showed that the levels of Col-Ⅰ, Col-Ⅲ, and p-Drp1 S616 were increased and the levels of p-Drp1 S637, Mfn1, Mfn2, TFAM, and ATP6 were decreased in 5/6(A/I) group compared with those in the sham group. The levels of Col-Ⅰ, Col-Ⅲ, and p-Drp1 S616 were significantly lower and the levels of p-Drp1 S637, Mfn1, Mfn2, TFAM, and ATP6 were significantly higher in ICA groups than that in 5/6(A/I) group. IHC staining demonstrated that for the expression of α-SMA in the renal interstitium was higher in the 5/6(A/I) group than in the sham group and that the expression in the ICA groups was significantly lower than that in the 5/6(A/I) group. Furthermore, the improvement in the fibrosis, mitochondrial dynamics, and mitochondrial function were particularly prominent in rats receiving the high dose of ICA. The in vitro experiment revealed that ICA dose-dependently inhibited the increase of Fn, CTGF, and p-Drp1 S616, increased p-Drp1 S637, Mfn1, Mfn2, TFAM, and ATP6, elevated ATP content, and improved mitochondrial morphology of NRK-52 E cells stimulated by TGF-ß1. ICA combined with MFP-M1 further down-regulated the expression of Fn and CTGF in NRK-52 E cells stimulated by TGF-ß1 compared with ICA alone. In conclusion, ICA attenuated RIF of CRF by improving mitochondrial dynamics.

Emodin ameliorates tubulointerstitial fibrosis in obstructed kidneys by inhibiting EZH2.

Emodin, a major component of Chinese herbal rhubarb, delays the progression of chronic renal failure. However, the effect and working mechanisms of Emodin on renal tubulointerstitial fibrosis remains elusive. We hypothesized that emodin inhibits renal tubulointerstitial fibrosis through EZH2, a histone methyltransferase. Our in vivo and in vitro studies demonstrate that emodin reduced extracellular collagen deposition and inhibited Smad3 and CTGF pro-fibrotic signaling pathways, which were correlated with the down-regulation of EZH2 and reduced trimethylation of histone H3 on lysine 27 (H3k27me3) in NRK-49F fibrotic cells and UUO kidneys. Inhibition of EZH2 by 3-DZNeP blocked or attenuated the anti-fibrotic effect of emodin in UUO kidneys and NRK-49F cells. These data indicate that emodin inhibits renal tubulointerstitial fibrosis in obstructed kidneys and this effect is mediated through EZH2.

Icariin attenuates renal fibrosis in chronic kidney disease by inhibiting interleukin-1ß/transforming growth factor-ß-mediated activation of renal fibroblasts.

Icariin (ICA) is a bioactive flavonoid extracted from Epimedium brevicornum Maxim and exhibits a variety of pharmacological activities including antiinflammatory and antioxidant effects. Recently, icariin has shown renoprotective role by inhibiting pathological matrix. However, the underlying mechanisms of the efficacy remain unknown. This study aimed to determine the effects of icariin on renal fibrosis and explore its molecular mechanisms. Chronic kidney disease (CKD) was induced in rats with 5/6 ablation and infarction (A/I) operation. Four weeks later, rats were treated with vehicle or 20 mg/kg (low dose) or 40 mg/kg (high dose) of icariin by daily gavage. Furthermore, to further elucidate the effect mechanisms of icariin, in vitro, NRK-49F cells stimulated by 8 ng/ml IL-1ß were treated with icariin in the presence or absence of SB431542 or the neutralizing antibody of transforming growth factor-ß (TGF-ß) for 24 h. We showed that icariin treatment for 8 weeks dose-dependently improved 5/6 (A/I)-induced kidney injury and fibrosis, and blocked the release of inflammatory cytokine IL-1ß. In vitro, icariin inhibited IL-1ß/TGF-ß-mediated activation of renal fibroblasts. In summary, anti-fibrotic effects of icariin were interconnected with the inhibition of renal fibroblast activation caused by IL-1ß/TGF-ß signaling.

Meclofenamic acid promotes cisplatin-induced acute kidney injury by inhibiting fat mass and obesity-associated protein-mediated m6A abrogation in RNA.

The role of RNA methylation on the sixth N atom of adenylate (m6A) in acute kidney injury (AKI) is unknown. FTO (fat mass and obesity-associated protein) reverses the m6A modification in cisplatin-induced AKI. Here, we aimed to determine FTO's role in AKI. We induced AKI in c57BL/6 mice by intraperitoneal cisplatin injection and treated the animal with vehicle or an FTO inhibitor meclofenamic acid (MA) for 3 days. Moreover, as an in vitro model, human kidney proximal tubular cells (HK2 cells) were treated with cisplatin. We found that the cisplatin treatment reduces FTO expression and increases m6A levels in vivo and in vitro MA aggravated renal damage and increased apoptosis in cisplatin-treated kidneys, phenotypes that were correlated with reduced FTO expression and increased m6A levels. Moreover, MA promoted apoptosis in cisplatin-treated HK2 cells, which was correlated with the reduced FTO expression and increased m6A in HK2 cells. FTO protein overexpression reduced m6A levels and inhibited apoptosis in cisplatin-treated HK2 cells and also blocked the MA-induced increase in m6A levels and apoptosis rates. In agreement, overexpression of the m6A-generating methyltransferase-like 3 and 14 (METTL3 and METTL14) or siRNA-mediated FTO knockdown promoted apoptosis and enhanced m6A levels in cisplatin-treated HK2 cells. MA increased p53 mRNA and protein levels in AKI both in vitro and in vivo, and FTO overexpression reduced p53 expression and reversed the MA-induced p53 increase in AKI. In conclusion, reduced renal FTO expression in cisplatin-induced AKI increases RNA m6A levels and aggravates renal damages.

Reduced asymmetric dimethylarginine accumulation through inhibition of the type I protein arginine methyltransferases promotes renal fibrosis in obstructed kidneys.

The role of asymmetric dimethylarginine (ADMA) in chronic kidney disease (CKD) is unclear. Through inhibition of type I protein arginine methyltransferases (PRMTs), a novel strategy, we aimed to determine the effect of ADMA on renal fibrosis and explore its underlying working mechanisms. After sham or unilateral ureter ligation (UUO) operation, 20-25 g male c57 mice were treated with vehicle or PT1001B, an inhibitor of type I PRMTs, for 13 d. Moreover, human kidney 2 (HK2) and normal rat kidney 49F (NRK-49F) cells were treated with various concentrations of PT1001B or ADMA in the presence of 2.5 ng/ml TGF-ß. We found that treatment with PT1001B increased the deposition of extracellular matrix proteins, the expression of α smooth muscle actin, and connective tissue growth factor in UUO-induced fibrotic kidneys, which is correlated with reduced expression of PRMT1, reduced the production of ADMA, and increased expression of uromodulin. In TGF-ß-stimulated HK2 and NRK-49F cells, PT1001B dose-dependently inhibited ADMA production, increased NO concentrations, and enhanced the expression of profibrotic proteins. Exogenous addition of ADMA inhibited the expression of profibrotic proteins dose-dependently and attenuated the profibrotic effect of PT1001B. Moreover, ADMA reduced the NO concentration in PT1001B-treated HK2 cells. Finally, we conclude that ADMA has an antifibrotic effect in obstructed kidneys, and future application of type I PRMT inhibitor should be done cautiously for patients with CKD.-Wu, M., Lin, P., Li, L., Chen, D., Yang, X., Xu, L., Zhou, B., Wang, C., Zhang, Y., Luo, C., Ye, C. Reduced asymmetric dimethylarginine accumulation through inhibition of the type I protein arginine methyltransferases promotes renal fibrosis in obstructed kidneys.

Magnesium lithospermate B improves renal hemodynamics and reduces renal oxygen consumption in 5/6th renal ablation/infarction rats.

BACKGROUND: Magnesium lithospermate B (MLB) can promote renal microcirculation. The aim of the current project was to study whether MLB improves renal hemodynamics, oxygen consumption and subsequently attenuates hypoxia in rats induced by 5/6th renal Ablation/Infarction(A/I). METHODS: Chronic renal failure (CRF) was induced in male SD rats by the 5/6 (A/I) surgery. 30 rats were randomly divided into three groups: sham group, 5/6 (A/I) + vehicle group (CRF group) and 5/6 (A/I) + MLB (CRF + MLB) group. 28 days after the surgery, rats were given with saline or 100 mg/kg MLB by i.p. injection for 8 weeks. The 24-h urinary protein (24hUp), serum creatinine (Scr), blood urine nitrogen (BUN), systolic blood pressure (SBP) and diastolic blood pressure (DBP) were measured. The protein expression of Fibronectin (FN), Collagen-I (Col-I), Connective Tissue Growth Factor(CTGF) and Interleukin-6 (IL-6) were measured by Western blot. Renal blood flow (RBF) and renal O2 consumption (QO2) indicated as sodium reabsorption (QO2/TNa) were detected before sacrifice. Renal hypoxia was assessed by measuring the protein expression of nNOS, HIF-1α and VEGF. RESULTS: MLB significantly reduced 24hUp, Scr, BUN, SBP and DBP levels in rats with CRF. The expression of FN, Col-I, CTGF and IL-6 were down-regulated by MLB treatment in rats with CRF. In comparison to sham operated rats, 5/6 (A/I) rats had significantly lower RBF, and MLB significantly increased RBF in rats with CRF. Moreover, QO2/TNa was higher in the CRF group as compared to that in the sham group, and it was significantly attenuated in the CRF + MLB group. MLB reversed the expression of nNOS (neuronal nitric oxide synthase), HIF-1α (hypoxia inducible factor-1) and VEGF in rats with CRF. CONCLUSIONS: MLB improves renal function, fibrosis and inflammation in CRF rats induced by 5/6 (A/I), which is probably related to the increase in RBF, reduction of oxygen consumption and attenuation of renal hypoxia in the remnant kidney with CRF.