Hedgehog signaling is a potential therapeutic target for vascular calcification.

BACKGROUND: Patients with chronic kidney disease (CKD) suffered from vascular calcification (VC), one major contributor for their increased mortality rate. Hedgehog (Hh) signaling plays a crucial role in physiological bone mineralization and is associated with several cardiovascular diseases. However, the molecular changes underlying VC is ill defined and it remains unclear whether Hh signaling intervention affects VC. METHODS: We constructed human primary vascular smooth muscle cell (VSMC) calcification model and performed RNA sequencing. Alizarin red staining and calcium content assay were conducted to identify the occurrence of VC. Three different R packages were applied to determine differentially expressed genes (DEGs). Enrichment analysis and protein-protein interaction (PPI) network analysis were carried out to explore the biological roles of DEGs. qRT-PCR assay was then applied to validate the expression of key genes. By using Connectivity Map (CMAP) analysis, several small molecular drugs targeting these key genes were obtained, including SAG (Hedgehog signaling activator) and cyclopamine (CPN) (Hedgehog signaling inhibitor), which were subsequently used to treat VSMC. RESULTS: Obvious Alizarin red staining and increased calcium content identified the occurrence of VC. By integrating results from three R packages, we totally obtained 166 DEGs (86 up-regulated and 80 down-regulated), which were significantly enriched in ossification, osteoblast differentiation, and Hh signaling. PPI network analysis identified 10 key genes and CMAP analysis predicted several small molecular drugs targeting these key genes including chlorphenamine, isoeugenol, CPN and phenazopyridine. Notably, our in vitro experiment showed that SAG markedly alleviated VSMC calcification, whereas CPN significantly exacerbated VC. CONCLUSIONS: Our research provided deeper insight to the pathogenesis of VC and indicated that targeting Hh signaling pathway may represent a potential and effective therapy for VC.

Exosomal Circsafb2 Reshaping Tumor Environment to Promote Renal Cell Carcinoma Progression by Mediating M2 Macrophage Polarization.

Background: Macrophages are the most abundant infiltrating immune-related stromal cells present in and around tumors, showing different phenotypes and functions. M2 macrophages mainly exert immunosuppressive functions and promote tumor growth. Exosomes are emerging as important mediators of cross-talk between tumor cells and the microenvironment. CircRNAs are novel members of non-coding RNAs that regulate cancer proliferation and progression. However, the mechanism by which exosomal circRNA regulates macrophage polarization in renal cell carcinoma (RCC) is still largely unknown. Methods: RCC-derived exosomes were characterized using transmission electron microscopy and nanoparticle tracking analysis (NTA). CCK-8, wound healing, and Transwell assays were performed to assess whether exosomes would affect the proliferation, migration, and invasion of RCC. Furthermore, we performed a bioinformatics analysis to identify circRNAs in RCC serum-derived exosomes from the GEO database. The fluorescence in situ hybridization (FISH) assay was used to detect the cellular distribution of circSAFB2. Bioinformatics analyses (StarBase 2.0) were used to pool the miRNA targets of circSAFB2. Luciferase assays were performed to verify the direct interactions. Western blotting was used to detect markers of macrophage M2 polarization. Lastly, mouse xenograft and bioluminescence imaging were used to examine the clinical relevance of exosomal circSAFB2 in vivo. Results: We report the circRNA derived from SAFB2 and evaluate its biological function in promoting the immune escape of RCC. We found that circSAFB2 was highly expressed in RCC tissues and RCC-derived exosomes. Furthermore, we demonstrated that exosomal circSAFB2 mediates the polarization of M2 macrophages through the miR-620/JAK1/STAT3 axis to promote RCC metastasis. Conclusions: Our data first demonstrated that circSAFB2 leads to immune escape from RCC by mediating M2 macrophage polarization via the miR-620/JAK1/STAT3 axis. These findings indicate a novel molecular mechanism of exosomal circSAFB2 in the progression of RCC and implicate circSAFB2 as a target for exosome-mediated tumor immune evasion.

Machine Learning Reveals Ets2 as a Novel Target for Membranous Nephropathy Treatment and Its Role in Immune Infiltration.

Background: Membranous nephropathy (MN) is a common pathological phenotype for adult nephrotic syndrome (NS). The occurrence of MN is increasing across China, but diagnostic methods for MN still rely on kidney biopsy and PLA2R and THSD7A detection in plasma and kidney tissue, and there has been no new biomarker for MN discovered since 2014. Immune infiltration status in MN patients suffers from the dearth of associated studies. In the present study, we aimed to find new bio-markers for MN and evaluate the role of immune cells infiltration in MN pathology. Methods: We downloaded MN expression profile from the Gene Expression Omnibus database and used R-project to screen differentially expressed genes (DEGs) and performed functional correlation analysis. Least absolute shrinkage and selection operator (LASSO) logistic regression and Radom Forest algorithms were used to screen and verify the bio-markers of MN. Finally, CIBERSORT was used to evaluate the infiltration of immune cells in MN tissues. Results: A total of 463 DEGs were screened from the MN tissue in this study. ETS2 was identified as bio-marker for MN. The CIBERSORT results showed that there were statistical differences in monocytes, plasma cells, regulatory T cells, and memory B cells. In addition, ETS2 was positively related to monocytes, M1 phase macrophages, and neutrophils and negatively correlated to plasma cells, CD4+ T memory cells, M2 macrophages, CD8+ T cells, memory B cells, and resting mast cells. Conclusion: (1) Machine learning algorithms reveals Ets2 as a novel target for membranous nephropathy patients. (2) Immune infiltration plays an important part in membranous nephropathy. (3) Ets2 expression is related to immune cells infiltration.

Effect of Magnesium Supplementation on Chronic Kidney Disease-Mineral and Bone Disorder in Hemodialysis Patients: A Meta-Analysis of Randomized Controlled Trials.

OBJECTIVES: Research about the effects of magnesium (Mg) supplementation on chronic kidney disease-mineral bone disorder (CKD-MBD) among hemodialysis (HD) patients is controversial. Thus, we conducted a meta-analysis to examine Mg supplementation's effects on CKD-MBD in patients requiring dialysis. METHODS: The PubMed and EMBASE databases were searched for English language studies up to September 2020. The main indicators of our study were changes in serum Mg, calcium (Ca), phosphate, parathyroid hormone (PTH), and C-reactive protein levels, and carotid intima-media thickness (CIMT) after Mg supplementation. Mg efficacy was evaluated by weighted mean difference (WMD) and confidence intervals (CIs), and subgroup analyses of intervention type and intervention duration were also performed. RESULTS: Eight eligible studies comprising 309 HD patients were included in our meta-analysis. Mg supplementation alone produced a negative effect on serum PTH levels (WMD = -236.56; 95% CI -349.71 to -123.41) and CIMT (WMD = -0.18; 95% CI -0.34 to -0.01). A subgroup analysis based on intervention type showed a significant improvement in serum Mg (WMD = 1.08; 95% CI 0.51-1.64) and Ca (WMD = -0.50; 95% CI -0.77 to -0.23) levels when Mg was administered via dialysate and oral medication, respectively. Different intervention durations had no effect on serum Mg levels. Mg supplementation had no significant effect on serum phosphate (WMD = -0.25; 95% CI -0.64 to 0.14) and C-reactive protein levels (WMD = -0.02; 95% CI -2.80 to 2,76). CONCLUSIONS: Our results showed that Mg supplementation alone could improve CKD-MBD by regulating serum Ca and PTH metabolism and decreasing CIMT among HD patients.

Multifactorial analysis of renal anemia-associated substandard hemoglobin levels and prevalence of anemia in patients on maintenance hemodialysis in Liaoning Province: a cross-sectional study.

BACKGROUND: In hemodialysis (HD) patients, anemia is greatly improved due to regular weekly use of iron and erythropoietin (EPO), but a large number of patients still show persistent anemia. We do a survey to elucidate the influencing factors that contribute to the failure of hemoglobin (Hb) to meet the standard and provide epidemiological data reference for promoting the recognition of renal anemia and improving the treatment effect of renal anemia. METHODS: The clinical data of End-Stage Renal Disease (ESRD) HD patients in 22 tertiary hospital HD centers in Liaoning Province from September 2021 to June 2022 were collected by convenient sampling. According to the 2012 Kidney Disease: Improving Global Outcomes (KDIGO) anemia diagnostic criteria. The standard of Hb compliance: Hb ≥110 g/L is considered as Hb compliant, and Hb <110 g/L as Hb non-compliant. The factors influencing Hb up-to-standard in ESRD HD patients and their correlations were analyzed by comparison between the two groups. RESULTS: The results of this study showed that among the 1,652 ESRD patients investigated in Liaoning Province, the prevalence rate of anemia was 89.29% (1,475/1,652), and the Hb compliance rate was 46.25% (764/1,652). The Hb compliance rate in maintenance hemodialysis (MHD) patients with different primary diseases was statistically significant (P<0.05). Compared with the Hb non-standard group, the gender, dialysis access, HD frequency, concurrent infection, primary disease of ESRD patients, red blood cell (RBC) count, hematocrit (HCT), mean RBC Hb concentration, mean RBC Hb content, platelet (PLT), albumin (ALB), total protein (TP), serum creatinine (Cr), serum calcium (Ca), serum potassium (K), ferritin (Fer), serum iron (SI), and transferrin (TRE) saturation were significantly different between both groups (P<0.05). Adrenaline was an independent risk factor affecting Hb failure in ESRD patients (OR =1.001, 95% CI: 1.000-1.002); dialysis frequency (OR =0.726, 95% CI: 0.601-0.878), ALB (OR =0.959, 95% CI: 0.929-0.990), TP (OR =0.982, 95% CI: 0.968-0.996), serum Cr (OR =0.959, 95% CI: 0.929-0.999), and SI (OR =0.961, 95% CI: 0.940-0.982) were protective factors affecting Hb failure in ESRD patients (P<0.05). Pearson correlation analysis showed that ALB, TP, serum Cr, serum Ca, serum K, SI, and TRE saturation were positively correlated with Hb (P<0.05). CONCLUSIONS: The anemia rate of ESRD patients treated with MHD in Liaoning Province is high. Based on the results, increasing the frequency of dialysis can improve anemia. Parathyroid hormone levels need to be controlled.

Prognostic Value of Soluble Suppression of Tumorigenicity 2 in Chronic Kidney Disease Patients: A Meta-Analysis.

OBJECTIVE: Previous studies have controversial results about the prognostic role of soluble suppression of tumorigenicity 2 (sST2) in chronic kidney disease (CKD). Therefore, we conduct this meta-analysis to access the association between sST2 and all-cause mortality, cardiovascular disease (CVD) mortality, and CVD events in patients with CKD. METHODS: The publication studies on the association of sST2 with all-cause mortality, CVD mortality, and CVD events from PubMed and Embase were searched through August 2020. We pooled the hazard ratio (HR) comparing high versus low levels of sST2 and subgroup analysis based on treatment, continent, and diabetes mellitus (DM) proportion, and sample size was also performed. RESULTS: There were 15 eligible studies with 11,063 CKD patients that were included in our meta-analysis. Elevated level of sST2 was associated with increased risk of all-cause mortality (HR 2.05; 95% confidence interval (CI), 1.51-2.78), CVD mortality (HR 1.68; 95% CI, 1.35-2.09), total CVD events (HR 1.88; 95% CI, 1.26-2.80), and HF (HR 1.35; 95% CI, 1.11-1.64). Subgroup analysis based on continent, DM percentage, and sample size showed that these factors did not influence the prognostic role of sST2 levels to all-cause mortality. CONCLUSIONS: Our results show that high levels of sST2 could predict the all-cause mortality, CVD mortality, and CVD events in CKD patients.

Effect of local anti-vascular endothelial growth factor therapy to prevent the formation of stenosis in outflow vein in arteriovenous fistula.

BACKGROUND AND OBJECTIVES: Vascular stenosis and angiogenesis are the major causes of short expectancy of arteriovenous fistula (AVF). Increased expression of vascular endothelial growth factor-A (VEGF-A) has been suggested to play an important role in the pathophysiologic process. Anti-VEGF has been proved to be effective on anti-angiogenesis and applied in clinical practice, but its effect on anti-stenosis remains to be verified before it could be applied to prevent stenosis of AVF. This study was aimed to evaluate the effect of local anti-VEGF therapy to prevent the formation of stenosis in the outflow vein in AVF and its mechanism. METHODS: Bioinformatics of VEGF-A and its downstream-regulated molecules from the STRING PPI database were analyzed in this study. The biopsy samples from outflow veins of AVF in patients and C57BL/6 mouse models were analyzed to examine the mechanisms of pathologic vascular stenosis associated with VEGF pathways and their potential therapeutic targets. RESULTS: We found that the reduction of VEGF-A could downregulate downstream molecules and subsequently reduce the intimal hyperplasia and abnormal vascular remodeling by analyzing the STRING PPI database. Venous wall thickening, intimal neointima formation, and apoptosis of vascular endothelial cells in the proliferative outflow vein of the AVF were significantly more obvious, and upregulation of expression of VEGF was observed in dysfunctional AVF in patients. In mouse models, the expression of VEGF, Ephrin receptor B4 (EphB4), matrix metalloproteinase (MMP)2, MMP9, tissue inhibitor of metalloproteinase (TIMP)1, TIMP2, and caspase 3 in the control-shRNA surgical group was significantly higher than in the sham group (P < 0.05), and all of these indicators were significantly lower in lentiviral transfection group and Avastin group than in control-shRNA surgical group (P < 0.05) on the 14th day after AVF operation. CONCLUSION: VEGF expression is significantly increased in vascular endothelial cells in stenosed or occluded outflow veins of dysfunctional AVF. Local injection of Avastin into the adventitia of the proximal outflow vein in autologous AVF procedure has an excellent potential to prevent the subsequent local stenosis of the proximal outflow vein.

The roles of collagen in chronic kidney disease and vascular calcification.

The extracellular matrix component collagen is widely expressed in human tissues and participates in various cellular biological processes. The collagen amount generally remains stable due to intricate regulatory networks, but abnormalities can lead to several diseases. During the development of renal fibrosis and vascular calcification, the expression of collagen is significantly increased, which promotes phenotypic changes in intrinsic renal cells and vascular smooth muscle cells, thereby exacerbating disease progression. Reversing the overexpression of collagen substantially prevents or slows renal fibrosis and vascular calcification in a wide range of animal models, suggesting a novel target for treating patients with these diseases. Stem cell therapy seems to be an effective strategy to alleviate these two conditions. However, recent findings indicate that the natural pore structure of collagen fibers is sufficient to induce the inappropriate differentiation of stem cells and thereby exacerbate renal fibrosis and vascular calcification. A comprehensive understanding of the role of collagen in these diseases and its effect on stem cell biology will assist in improving the unmet requirements for treating patients with kidney disease.

Efficacy of transanal drainage tube and self-expanding metallic stent in acute left malignant colorectal obstruction.

BACKGROUND: The self-expanding metal stent (SEMS) or transanal drainage tube (TDT) methods can be used as a palliative treatment before tumor resection surgery. Studies systematically comparing the efficacy and characteristics between SEMS with TDT are limited, especially in a large-scale Chinese population. This retrospective study aimed to compare the outcomes of these treatment approaches. METHODS: This was a retrospective comparative study of patients with an acute malignant left colorectal obstruction who underwent a preoperative decompressive procedure with SEMS or TDT intervention between December 2014 and October 2017. The indicators after endoscopic treatment and tumor resection surgery between the SEMS and TDT groups were compared. RESULTS: 206 patients underwent endoscopic intervention to relieve obstruction, including 139 patients treated with SEMS and 67 patients treated with TDT. The technical success rates of the SEMS group and TDT group were 97.1% and 95.6%, respectively, and the rates of obstruction relief were 92.8% and 86.6%, respectively. TDT was more easily translocated than SEMS (P=0.02), and there was no significant difference in the incidence of other complications. However, SEMS had a lower complication rate than TDT (P=0.02), and could alleviate the obstruction faster (P<0.01). There were 72 patients and 44 patients who took resection surgery in the SEMS group and TDT group, respectively. The direct anastomosis rates were 73.6% and 63.6% (P=0.26), respectively, and only 1 case in the TDT group had anastomotic leakage. The surgery time of the SEMS group was significantly shorter than that of the TDT group (P=0.01). There was no significant difference between the 2 groups in terms of postoperative hospital stay (P=1.00) or total treatment costs for patients undergoing surgery (P=0.26). CONCLUSIONS: Both TDT and SEMS could effectively relieve acute left malignant colorectal obstruction with safe and reliable results, and they could both reduce the stomas rate compared with traditional surgery. SEMS could alleviate obstruction faster than TDT and had fewer complications.

circHLA-C Plays an Important Role in Lupus Nephritis by Sponging miR-150.

Circular RNAs (circRNAs) participate in the pathogenesis of various diseases by sponging microRNAs (miRs). However, the roles of circRNAs remain unreported in glomerular diseases. We previously reported that miR-150 positively correlated with renal chronicity index in patients with lupus nephritis (LN). We aimed to investigate renal circRNA profiling and the interaction between circRNAs and miR-150 in LN patients. Six renal biopsies from untreated female patients with LN class IV and five normal kidney tissues from urology patients were used for circRNA sequencing. 171 circRNAs with 2-fold differential expression were identified in LN compared with normal control. Ten selected circRNAs were validated by real-time qPCR, and seven circRNAs showed the same significant increases as the sequencing results. circHLA-C positively correlated with proteinuria (R = 0.92, p < 0.01), serum creatinine (R = 0.76, p = 0.08), renal activity index (R = 0.88, p < 0.05), and crescentic glomeruli (R = 0.93, p < 0.01). Renal circHLA-C increased 2.72-fold, and miR-150 decreased 66% in LN compared with normal control (p < 0.05). Bio-informatic analysis predicted miR-150 was regulated by circHLA-C and displayed one perfect match seed between circHLA-C and miR-150. The renal miR-150 showed a tendency of negative correlation with circHLA-C in LN patients. In conclusion, circHLA-C may play an important role in the pathogenesis of lupus nephritis by sponging miR-150.

Nrf2 deficiency promotes the progression from acute tubular damage to chronic renal fibrosis following unilateral ureteral obstruction.

Background: Nuclear factor erythroid 2-related factor 2 (Nrf2) is a central mediator of cellular responses to oxidative stress. We hypothesized that Nrf2 modulates progression from acute tubular damage to renal fibrosis. We asked whether Nrf2 deletion increases renal injury in mice following unilateral ureteral obstruction (UUO). Methods: We explored the time course of renal injury and Nrf2 expression in Nrf2+/+ mice following UUO. We compared Nrf2+/+ and Nrf2-/- mice following UUO in tubular damage, transdifferentiation [vimentin, proliferating cell nuclear antigen (PCNA)], fibrosis [fibronectin, α-smooth muscle actin (SMA)], antioxidative and inflammatory responses. We studied Nrf2 in renal biopsies of patients with acute, subacute and chronic tubulointerstitial nephritis (TIN). Results: In Nrf2+/+ mice, renal Nrf2 expression and Nrf2-regulated glutamate-cysteine ligase catalytic (Gclc) and heme oxygenase-1 (Ho-1) were elevated, and renal injury occurred between 2 and 14 days after UUO. On Day 2 following UUO, in Nrf2-/- mice compared with Nrf2+/+ mice, tubular damage, apoptotic cell numbers, cleaved caspase3 and cleaved-poly ADP-ribose polymerase were increased. On Day 5, protein levels of vimentin and PCNA and the co-expressed cells of both proteins were increased. On Day 14, fibronectin and α-SMA protein levels were increased. Nrf2 deletion decreased expression of antioxidative genes (Gclc and Ho-1) and increased expression of inflammatory response genes (Tgfß, Tnf, IL-6, IL-1ß and F4/80). Finally, Nrf2 expression was upregulated in renal biopsies of patients with TIN. Conclusions: Following UUO, Nrf2 deficiency increased tubular damage, transdifferentiation, fibrosis and inflammatory response while decreasing antioxidative responses. The renal protective role of Nrf2 in the development of tubulointerstitial fibrosis in UUO may be mediated by antioxidative and anti-inflammatory pathways.