Poricoic acid A suppresses renal fibroblast activation and interstitial fibrosis in UUO rats via upregulating Sirt3 and promoting ß-catenin K49 deacetylation.

Renal interstitial fibrosis is the common pathological process of various chronic kidney diseases to end-stage renal disease. Inhibition of fibroblast activation attenuates renal interstitial fibrosis. Our previous studies show that poricoic acid A (PAA) isolated from Poria cocos is a potent anti-fibrotic agent. In the present study we investigated the effects of PAA on renal fibroblast activation and interstitial fibrosis and the underlying mechanisms. Renal interstitial fibrosis was induced in rats or mice by unilateral ureteral obstruction (UUO). UUO rats were administered PAA (10 mg·kg-1·d-1, i.g.) for 1 or 2 weeks. An in vitro model of renal fibrosis was established in normal renal kidney fibroblasts (NRK-49F cells) treated with TGF-ß1. We showed that PAA treatment rescued Sirt3 expression, and significantly attenuated renal fibroblast activation and interstitial fibrosis in both the in vivo and in vitro models. In TGF-ß1-treated NRK-49F cells, we demonstrated that Sirt3 deacetylated ß-catenin (a key transcription factor of fibroblast activation) and then accelerated its ubiquitin-dependent degradation, thus suppressing the protein expression and promoter activity of pro-fibrotic downstream target genes (twist, snail1, MMP-7 and PAI-1) to alleviate fibroblast activation; the lysine-49 (K49) of ß-catenin was responsible for Sirt3-mediated ß-catenin deacetylation. In molecular docking analysis, we found the potential interaction of Sirt3 and PAA. In both in vivo and in vitro models, pharmacological activation of Sirt3 by PAA significantly suppressed renal fibroblast activation via facilitating ß-catenin K49 deacetylation. In UUO mice and NRK-49F cells, Sirt3 overexpression enhanced the anti-fibrotic effect of PAA, whereas Sirt3 knockdown weakened the effect. Taken together, PAA attenuates renal fibroblast activation and interstitial fibrosis by upregulating Sirt3 and inducing ß-catenin K49 deacetylation, highlighting Sirt3 functions as a promising therapeutic target of renal fibroblast activation and interstitial fibrosis.

Association of mineral metabolism biomarkers with chronic kidney disease in Chinese adults.

BACKGROUND: We aimed to examine the association of three mineral metabolism markers, including serum calcium, inorganic phosphorus, and intact parathyroid hormone with the risk of chronic kidney disease (CKD) at all stages. METHODS: This retrospective cohort study involved 3563 participants, including 3274 CKD patients and 289 healthy controls. CKD is diagnosed according to clinical guidelines from the 2012 KDIGO. Effect sizes are expressed odds ratio (OR) and 95 confidence interval (CI). RESULTS: After propensity score matching, per 0.5 mg/dL increment of inorganic phosphorus was significantly associated with 1.33-, 1.61-, and 2.85-fold increased risk of CKD at stages 1-2, 4, and 5, respectively. Regarding per 8 pg/mL increment of intact parathyroid hormone, significance was only noted for stage 5. In subsidiary analyses, the risk prediction of mineral metabolism markers under study was more evident in males and hypertensive subjects. A nomogram prediction model was constructed based on age, sex, and three mineral metabolism markers for CKD, with decent accuracy. CONCLUSIONS: Our findings indicate that serum calcium was associated with all-stage CKD risk, whereas the association for inorganic phosphorus and intact parathyroid hormone was significant at advanced stages.

Association of Serum Thyroid Hormones with the Risk and Severity of Chronic Kidney Disease Among 3563 Chinese Adults.

BACKGROUND Chronic kidney disease (CKD) is a global health problem with an increasing prevalence. We explored the association of serum thyroid hormones with the risk and severity of CKD among Chinese adults. MATERIAL AND METHODS This retrospective study involved 3563 participants. CKD was diagnosed according to the clinical practice guidelines of the 2012 Kidney Disease Improving Global Outcomes guidelines. Effect-size estimates are expressed as odds ratio (OR) and 95% confidence interval (CI). RESULTS Given the strong magnitude of correlation, only 3 thyroid hormones were analyzed: free triiodothyronine (FT3), free thyroxin (FT4), and thyroid-stimulating hormone (TSH). After propensity score matching on age, sex, diabetes, and hypertension, per 0.2 pg/mL increase in FT3 was significantly associated with 35-38% reduced risk of CKD at stage 1-4, and per 0.3 ng/dL increase in FT4 was only significantly associated with 21% reduced risk of CKD at stage 5 (OR, 95% CI: 0.79, 0.69-0.89), and per 0.5 µIU/mL increment in TSH increased the risk of CKD stage 5 by 8% (1.08, 1.02-1.14). Importantly, 3 thyroid hormones acted interactively, particularly for the interaction between FT3 and FT4 in predicting CKD at stage 5 (OR, 95% CI: 1.81, 1.30-2.55 for high FT3-low FT4, 17.72, 7.18-43.74 for low FT3-high FT4, and 22.28, 9.68-51.30 for low FT3-low FT4). CONCLUSIONS Our findings indicate that serum FT3 can be used as an early-stage biomarker for CKD, and FT4 and TSH can be used as advanced-stage biomarkers among Chinese adults.

The effectiveness of PC6 acupuncture in the prevention of postoperative nausea and vomiting in children: A systematic review and meta-analysis.

BACKGROUND: A growing number of studies have demonstrated the effectiveness of acupuncture in preventing and treating postoperative nausea and vomiting. Here, we used meta-analysis to confirm these benefits in children and to determine the optimal time to perform this treatment. METHODS: Four databases (MEDLINE, EMBASE, CENTRAL, and Chinese Database of Biology and Medicine) were searched from inception until January 16, 2019. We included randomized controlled trials for evaluating the effectiveness of acupuncture in the prevention and treatment of postoperative nausea and vomiting during the early stage (0-4 hours) and within 24 hours postoperatively in pediatrics. Control groups received standardized care control or standardized care combined with sham control. RESULTS: Sixteen literatures and 1773 patients undergoing general anesthesia were included in the study. The results indicated that acupuncture was effective in reducing postoperative vomiting, both during the first 4 hours (RR = 0.47, 95% CI 0.26, 0.84; low quality) and within 24 hours postoperatively (RR = 0.74, 95% CI 0.60, 0.91; low quality). Stratifying by the timing of acupuncture, acupuncture was effective in reducing the first 4 hours (RR = 0.34, 95% CI 0.18, 0.64; moderate quality), and 0-24 hours postoperative vomiting (RR = 0.81, 95% CI 0.70, 0.93; moderate quality) when performed before and during anesthesia, respectively. Further, the RR value was more robust when acupuncture was performed before anesthesia. Acupuncture was also effective in treating 0-24 hours postoperative nausea (RR = 0.73, 95% CI 0.60, 0.88; moderate quality) and in reducing the utilization of remedies during the first 4 hours (RR = 0.64, 95% CI 0.45, 0.89; moderate quality). CONCLUSION: Acupuncture reduces the incidence of postoperative nausea and vomiting as well as the utilization of antiemetic remedies, particularly during the first 4 hours following the operation. Acupuncture performed before anesthesia was demonstrated to be the most ideal intervention time for children.

Chinese medicine for diabetic kidney disease in China.

Diabetic kidney disease (DKD) is a major global public health concern due to its high prevalence and mortality, and high healthcare cost. Conventional Western therapies for DKD fail to provide favourable efficacy. Chinese medicine (CM) has been widely used to combat DKD in China and other Asian countries. The clinical effects and mechanism of CM in treating DKD have not been fully elucidated. This review summarizes current knowledge of CM therapeutic modalities for DKD, especially Chinese herbal medicine (CHM), and potential mechanisms.

Effects of Tangshen Formula on urinary and plasma liver-type fatty acid binding protein levels in patients with type 2 diabetic kidney disease: post-hoc findings from a multi-center, randomized, double-blind, placebo-controlled trial investigating the efficacy and safety of Tangshen Formula in patients with type 2 diabetic kidney disease.

BACKGROUND: Tangshen Formula (TSF) is a traditional Chinese medicine for the treatment of diabetic kidney disease (DKD). Liver-type fatty acid binding protein (L-FABP) is expressed in various tissues, including the kidney, where it is known as urinary L-FABP. Other studies demonstrated that urinary L-FABP may be a useful biomarker for monitoring DKD. This post-hoc analysis and cross-sectional study evaluated the changes in urinary L-FABP in DKD patients treated with TSF and conventional medicine. METHODS: Post-hoc analysis was conducted on a multicenter, randomized, double-blind, placebo-controlled trial. A total of 180 participants with DKD including 98 with microalbuminuria and 82 with macroalbuminuria were enrolled in the original study. In addition to conventional treatment, 122 participants were randomly assigned to receive TSF and 58 to receive placebo. After 24-weeks of treatment, the intention-to-treat population in microalbuminuria stage was 56 in the TSF group and 25 in the placebo group, and in the macroalbuminuria stage 42 and 19, respectively. The primary outcome in the original trial was urinary protein level. In the current study, urinary and plasma L-FABP levels were measured in 30 microalbuminuria patients (15 in the TSF group and 15 in the placebo group) and 30 macroalbuminuria patients (15 in the TSF group and 15 in the placebo group). In addition, another 30 patients with normoalbuminuria (urinary albumin excretion rate (UAER) < 20 µg/min) were recruited for the cross-sectional study. RESULTS: (1) In microalbuminuria patients, UAER in the TSF group displayed a significant decrease after 24 weeks of treatment (P = 0.045). Levels of urinary L-FABP in the TSF group were markedly lower than in the placebo group after 12 and 24 weeks (P = 0.004 and P = 0.047, respectively). (2) In macroalbuminuria patients, 24-h urinary protein levels decreased significantly compared with baseline in the TSF group at week 12 (P = 0.042) and week 24 (P = 0.041). The TSF group showed a significant decrease in urinary L-FABP after 12 and 24 weeks (P = 0.036 and P = 0.046, respectively). (3) Levels of urinary L-FABP increased markedly, correlating with severity of DKD. L-FABP in patients with normoalbuminuria, microalbuminuria, and macroalbuminuria were 5.9 (5.2, 7.8) µg/ml, 11.4 (6.8, 13.4) µg/ml and 18.5 (10.9, 23.4) µg/ml, respectively (P = 0.000). CONCLUSIONS: TSF combined with conventional therapy appeared to be effective in reducing urinary protein and urinary L-FABP. Urinary L-FABP levels appear to be associated with the severity of DKD. TRIAL REGISTRATION: Chinese Clinical Trial Registry ChiCTR-TRC-10000843 . Registered 15 April 2010.

[Experimental study of Tangshen formulain improved lipid metabolism and phenotypic switch of macrophage in db/db mice].

Obesity and its associated metabolic disorders, including non-alcoholic fatty liver disease (NAFLD), have become major chronic diseases threatening public health. NAFLD is a chronic liver disorder that is strongly associated with type 2 diabetes and obesity. In this study, we investigated the effects and mechanism of Tangshen formula (TSF) on hepatic dyslipidemia and phenotypic switch of macrophage in db/db mice. Eight-week-old male C57BLKS/J db/m control and db/db mice were divided into 3 groups (namely db/m, db/db, db/db+TSF), and fed with TSF or distilled water for 12 weeks. It was found that after treatment with TSF, the triglycerides accumulation in db/db mice was decreased on the basis of oil red O staining with cryosections of liver tissues. And protein expressions of macrophage activation markers CD68 and F4/80 were decreased according to immunohistochemical analysis of hepatic sections. The mRNA level of TNF-α (M1 marker) was significantly decreased by TSF in db/db mice, but with no significant difference in Mrc1 and Arg1 (M2 marker). According to the results, TSF attenuated hepatic steatosis and relieved dyslipidemia, its mechanism may be correlated with the regulation of macrophage activation and phenotypic switch.

Renoprotective effect of berberine on type 2 diabetic nephropathy in rats.

Inflammation, fibrosis, and lipid disorder are essential promoters in the pathogenesis of diabetic kidney injury in diabetes mellitus type 2. Berberine (BBR) has been reported to have beneficial effects on diabetic nephropathy, but its action mechanism is still unclear. The present study was designed to elucidate the therapeutic mechanism of BBR in a type 2 diabetic nephropathy rat model induced by a high-fat diet and low-dose streptozotocin injection. The diabetic rats were treated with or without BBR by gavage for 20 weeks and examined by serology, 24-h albuminuria, histology, immunohistochemistry, and molecular analyses. Results showed that treatment with BBR significantly reduced serum levels of blood glucose and lipids, inhibited urinary excretion of albumin, and attenuated renal histological injuries in diabetic rats. Berberine treatment also inhibited renal inflammation, which was associated with inactivation of nuclear factor kappa-light-chain-enhancer of activated B-cell signalling. As a result, the upregulation of pro-inflammatory cytokines (interleukin-1ß, tumour necrosis factor-α) and chemokine (monocyte chemotactic protein-1) was blocked. In addition, BBR treatment also inactivated transforming growth factor-ß/Smad3 signalling and suppressed renal fibrosis, including expression of fibronectin, collagen I, and collagen IV. The present study reveals that BBR is a therapeutic agent for attenuating type 2 diabetic nephropathy by inhibiting nuclear factor kappa-light-chain-enhancer of activated B cell-driven renal inflammation and transforming growth factor-ß/Smad3 signalling pathway.

Irbesartan ameliorates diabetic kidney injury in db/db mice by restoring circadian rhythm and cell cycle.

Background and Objectives: Irbesartan has been widely used in the clinical treatment of diabetic kidney disease (DKD). However, the molecular mechanism of its delay of DKD disease progression has not been fully elucidated. The aim of the present study was to investigate the mechanism of irbesartan in the treatment of DKD. Materials and Methods: C57BL/KsJ db/db mice were randomly divided into the model group and irbesartan-treated group. After treatment with irbesartan for 12 weeks, the effects on blood glucose, body weight, 24-h urinary albumin, and renal injuries were evaluated. Microarray was used to determine the differentially expressed genes (DEGs) in the renal cortex of mice. |Log FC| <0.5 and false discovery rate (FDR) <0.25 were set as the screening criteria. Kyoto Encyclopedia of Genes and Genomes (KEGG), gene ontology (GO), protein-protein interaction (PPI) network and modules, and microRNA (miRNA)-DEGs network analysis were applied to analyze the DEGs. Furthermore, quantitative real-time polymerase chain reaction (qRT-PCR) was used to validate the results of microarray. Results: The present study demonstrated irbesartan could significantly improve the renal function in db/db mice through decreasing 24-h urinary albumin and alleviating the pathological injury of kidney. Irbesartan may affect the expression of numerous kidney genes involved in circadian rhythm, cell cycle, micoRNAs in cancer, and PI3K-AKT signaling pathway. In the miRNA-DEGs network, miR-1970, miR-703, miR-466f, miR-5135, and miR-132-3p were the potential targets for irbesartan treatment. The validation test confirmed that key genes regulating circadian rhythm (Arntl, Per3, and Dbp) and cell cycle (Prc1, Ccna2, and Ccnb2) were restored in db/db mice on treatment with Irbesartan. Conclusion: Generally, irbesartan can effectively treat DKD by regulating the circadian rhythm and cell cycle. The DEGs and pathways identified in the study will provide new insights into the potential mechanisms of irbesartan in the treatment of DKD.

Irbesartan ameliorates diabetic kidney injury in db/db mice by restoring circadian rhythm and cell cycle.

Background and Objectives: Irbesartan has been widely used in the clinical treatment of diabetic kidney disease (DKD). However, the molecular mechanism of its delay of DKD disease progression has not been fully elucidated. The aim of the present study was to investigate the mechanism of irbesartan in the treatment of DKD. Materials and Methods: C57BL/KsJ db/db mice were randomly divided into the model group and irbesartan-treated group. After treatment with irbesartan for 12 weeks, the effects on blood glucose, body weight, 24-h urinary albumin, and renal injuries were evaluated. Microarray was used to determine the differentially expressed genes (DEGs) in the renal cortex of mice. |Log FC| <0.5 and false discovery rate (FDR) <0.25 were set as the screening criteria. Kyoto Encyclopedia of Genes and Genomes (KEGG), gene ontology (GO), protein-protein interaction (PPI) network and modules, and microRNA (miRNA)-DEGs network analysis were applied to analyze the DEGs. Furthermore, quantitative real-time polymerase chain reaction (qRT-PCR) was used to validate the results of microarray. Results: The present study demonstrated irbesartan could significantly improve the renal function in db/db mice through decreasing 24-h urinary albumin and alleviating the pathological injury of kidney. Irbesartan may affect the expression of numerous kidney genes involved in circadian rhythm, cell cycle, micoRNAs in cancer, and PI3K-AKT signaling pathway. In the miRNA-DEGs network, miR-1970, miR-703, miR-466f, miR-5135, and miR-132-3p were the potential targets for irbesartan treatment. The validation test confirmed that key genes regulating circadian rhythm (Arntl, Per3, and Dbp) and cell cycle (Prc1, Ccna2, and Ccnb2) were restored in db/db mice on treatment with Irbesartan. Conclusion: Generally, irbesartan can effectively treat DKD by regulating the circadian rhythm and cell cycle. The DEGs and pathways identified in the study will provide new insights into the potential mechanisms of irbesartan in the treatment of DKD.

Tangshen formula targets the gut microbiota to treat non-alcoholic fatty liver disease in HFD mice: A 16S rRNA and non-targeted metabolomics analyses.

BACKGROUND: Tangshen formula (TSF) has an ameliorative effect on hepatic lipid metabolism in non-alcoholic fatty liver disease (NAFLD), but the role played by the gut microbiota in this process is unknown. METHOD: We conducted three batches of experiments to explore the role played by the gut microbiota: TSF administration, antibiotic treatment, and fecal microbial transplantation. NAFLD mice were induced with a high-fat diet to investigate the ameliorative effects of TSF on NAFLD features and intestinal barrier function. 16S rRNA sequencing and serum untargeted metabolomics were performed to further investigate the modulatory effects of TSF on the gut microbiota and metabolic dysregulation in the body. RESULTS: TSF ameliorated insulin resistance, hypercholesterolemia, lipid metabolism disorders, inflammation, and impairment of intestinal barrier function. 16S rRNA sequencing analysis revealed that TSF regulated the composition of the gut microbiota and increased the abundance of beneficial bacteria. Antibiotic treatment and fecal microbiota transplantation confirmed the importance of the gut microbiota in the treatment of NAFLD with TSF. Subsequently, untargeted metabolomics identified 172 differential metabolites due to the treatment of TSF. Functional predictions suggest that metabolisms of choline, glycerophospholipid, linoleic acid, alpha-linolenic acid, and arachidonic acid are the key metabolic pathways by which TSF ameliorates NAFLD and this may be influenced by the gut microbiota. CONCLUSION: TSF treats the NAFLD phenotype by remodeling the gut microbiota and improving metabolic profile, suggesting that TSF is a functional gut microbial and metabolic modulator for the treatment of NAFLD.

Characteristics of Serum Metabolites and Gut Microbiota in Diabetic Kidney Disease.

Disturbance of circulating metabolites and disorders of the gut microbiota are involved in the progression of diabetic kidney disease (DKD). However, there is limited research on the relationship between serum metabolites and gut microbiota, and their involvement in DKD. In this study, using an experimental DKD rat model induced by combining streptozotocin injection and unilateral nephrectomy, we employed untargeted metabolomics and 16S rRNA gene sequencing to explore the relationship between the metabolic profile and the structure and function of gut microbiota. Striking alterations took place in 140 serum metabolites, as well as in the composition and function of rat gut microbiota. These changes were mainly associated with carbohydrate, lipid, and amino acid metabolism. In these pathways, isomaltose, D-mannose, galactonic acid, citramalic acid, and prostaglandin B2 were significantly upregulated. 3-(2-Hydroxyethyl)indole, 3-methylindole, and indoleacrylic acid were downregulated and were the critical metabolites in the DKD model. Furthermore, the levels of these three indoles were restored after treatment with the traditional Chinese herbal medicine Tangshen Formula. At the genera level, g_Eubacterium_nodatum_group, g_Lactobacillus, and g_Faecalibaculum were most involved in metabolic disorders in the progression of DKD. Notably, the circulating lipid metabolites had a strong relationship with DKD-related parameters and were especially negatively related to the mesangial matrix area. Serum lipid indices (TG and TC) and UACR were directly associated with certain microbial genera. In conclusion, the present research verified the anomalous circulating metabolites and gut microbiota in DKD progression. We also identified the potential metabolic and microbial targets for the treatment of DKD.

Subproteomic profiling from renal cortices in OLETF rats reveals mutations of multiple novel genes in diabetic nephropathy.

BACKGROUND: Diabetic nephropathy (DN) is a serious threat to human health, but its pathogenesis is not fully understood. Otsuka Long-Evans Tokushima Fatty (OLETF) rats are very similar to human DN in many aspects such as pathological changes and processes, and are deemed to be an ideal rodent model. OBJECTIVE: This study was aimed to explore the pathogenesis of DN by analyzing the protein expression profile from renal cortices in OLETF rats. METHODS: Thirty-six-week-old diabetic OLETF rats and normal control Long-Evans Tokushima Otsuka (LETO) rats were nephrectomized, and the renal cortices were isolated. The proteins were separated by soluble and insoluble high-resolution subproteomics methods for the analysis and identification of differential proteins. RESULTS: Thirty-six differentially expressed proteins were found. Among them, 11 proteins had different isoelectric points and molecular weights between OLETF and LETO rats. Further sequencing identified point mutations in genes encoding eight of these proteins, which are involved in many biological processes closely related to DN, including oxidative stress and inflammation. Five of these eight proteins have not been reported in DN. CONCLUSION: This study reveals mutations of multiple novel genes in diabetic OLETF rats, providing some new potential targets for the pathogenesis of DN and helping to better understand the pathogenesis of DN.

Association between PNPLA2 Gene Polymorphisms and the Risk of Diabetic Kidney Disease in a Chinese Han Population with Type 2 Diabetes.

Diabetic kidney disease (DKD) is one of the most common complications of diabetes and the leading cause of end-stage renal disease. Here, we investigated the association of PNPLA2 gene variations with DKD susceptibility in a Chinese Han population. A total of 818 participants with type 2 diabetes were recruited in the case-control study, including 379 patients diagnosed with DKD. We observed that 2 tagSNPs, PNPLA2 rs28633403 (A>G) and rs1138714 (A>G), were associated with DKD (rs28633403: genotype, P = 0.017; allele, P = 0.015; rs1138714: genotype, P = 0.029; allele, P = 0.018). PNPLA2 rs1138693 (T>C), a missense SNP, showed no association with DKD (genotype, P = 0.966; allele, P = 0.845). Genetic model analysis revealed that minor allele G of PNPLA2 rs28633403 was a protective factor of DKD in a dominant model adjusted by confounders (AG+GG vs. AA: adjusted odds ratio (aOR), 0.619; 95% CI 0.447-0.857; P = 0.004) and in an additive model (AG vs. AA: aOR, 0.633; 95% CI 0.447-0.895; P = 0.010; GG vs. AA: aOR, 0.588; 95% CI 0.385-0.897; P = 0.014). Minor allele G of PNPLA2 rs1138714 was associated with a higher risk of DKD in a dominant model adjusted by confounders (AG+GG vs. AA: adjusted odds ratio (aOR), 1.531; 95% CI 1.134-2.067; P = 0.005) and in an additive model (AG vs. AA: aOR, 1.529; 95% CI 1.118-2.091; P = 0.008). The combined effect of PNPLA2 rs28633403 AA+rs1138714 AG or GG genotype showed an association with DKD, adjusted by confounders (aOR, 2.194; 95% CI 1.378-3.492; P = 0.001), which was considered statistically significant with a markedly increased risk of DKD after a Holm-Bonferroni correction for multiple tests (P < 0.00125). Our results suggest that PNPLA2 rs28633403 and rs1138714 are significantly associated with the risk of DKD in a Chinese Han population with type 2 diabetes.

Association between LXR-α and ABCA1 Gene Polymorphisms and the Risk of Diabetic Kidney Disease in Patients with Type 2 Diabetes Mellitus in a Chinese Han Population.

We designed a case-control study and selected LXR-α rs7120118 C>T and ABCA1 rs2230806 A>G polymorphisms to determine the correlation between these polymorphisms and diabetic kidney disease (DKD) susceptibility in a Chinese Han population. Three hundred DKD patients and 346 type 2 diabetes mellitus (DM) patients without kidney disease were recruited. Our results showed that rs7120118 was associated with DKD (genotype, P = .027; allele, P < .011). rs7120118 was associated with a higher risk of DKD under a dominant model adjustment by age and sex (P = .015) and an additive model (P = .040); rs2230806 was associated with a higher risk of DKD under an recessive model (P < .03); the combined effect of rs7120118 CC+rs2230806 GG genotype showed an association of DKD adjustment for age and sex (P = .009). In subgroup analysis of patients without hypercholesterolemia, the rs2230806 genotype frequencies were different between the two groups (P = .042). rs2230806 was associated with increased risk of DKD under a recessive model adjustment for age and sex (P = .013) and an additive model (P = .031). Our results suggest that LXR-α rs7120118 is significantly associated with a higher risk of DKD, and ABCA1 rs2230806 is significantly associated with a higher risk of DKD without hypercholesterolemia in Chinese Han individuals.

Tangshen formula modulates gut Microbiota and reduces gut-derived toxins in diabetic nephropathy rats.

Growing evidence shows that diabetic kidney disease (DKD) is linked with intestinal dysbiosis from gut-derived toxins. Tangshen Formula (TSF) is a traditional Chinese herbal medicine that has been used to treat DKD. In this study, streptozotocin injection and uninephrectomy-induced diabetic nephropathy (DN) rat model was established to explore the impact of TSF on gut microbiota composition, gut-derived toxins, and the downstream inflammatory pathway of urotoxins in the kidney. TSF treatment for 12 weeks showed significant attenuation of both renal histologic injuries and urinary excretion of albumin compared with DN rats without treatment. TSF treatment also reconstructed gut dysbiosis and reduced levels of indoxyl sulfate and metabolic endotoxemia/lipopolysaccharide. MCP-1 and TNF-α were decreased by TSF both in the serum and kidney. In addition, we revealed that the inhibitory effect of TSF on renal inflammation was associated with the inhibition of aryl hydrocarbon, a receptor of indoxyl sulfate, and TLR4, thereby inhibiting JNK and NF-κB signaling in the kidney. Spearman correlation analysis found that a cluster of gut bacterial phyla and genera were significantly correlated with renal pathology, renal function, and systemic inflammation. In conclusion, orally administered TSF significantly inhibited diabetic renal injury, and modulated gut microbiota, which decreased levels of lipopolysaccharide and indoxyl sulfate, and attenuated renal inflammation. Our results indicate that TSF may be used as an agent in the prevention of gut dysbiosis and elimination of intestinal toxins in DN individuals.

Tangshen Formula Attenuates Diabetic Kidney Injury by Imparting Anti-pyroptotic Effects via the TXNIP-NLRP3-GSDMD Axis.

We previously reported that Tangshen formula (TSF), a Chinese herbal medicine for diabetic kidney disease (DKD) therapy, imparts renal protective effects. However, the underlying mechanisms of these effects remain unclear. Pyroptosis is a form of programmed cell death that can be triggered by the NLRP3 inflammasome. Recently, the association between the pyroptosis of renal resident cells and DKD was established, but with limited evidence. This study aimed to investigate whether the renal protective effects of TSF are related to its anti-pyroptotic effect. DKD rats established by right uninephrectomy plus a single intraperitoneal injection of STZ and HK-2 cells stimulated by AGEs were used. In vivo, TSF reduced the 24 h urine protein (24 h UP) of DKD rats and alleviated renal pathological changes. Meanwhile, the increased expression of pyroptotic executor (GSDMD) and NLRP3 inflammasome pathway molecules (NLRP3, caspase-1, and IL-1ß) located in the tubules of DKD rats were downregulated with TSF treatment. In vitro, we confirmed the existence of pyroptosis in AGE-stimulated HK-2 cells and the activation of the NLRP3 inflammasome. TSF reduced pyroptosis and NLRP3 inflammasome activation in a dosage-dependent manner. Then, we used nigericin to determine that TSF imparts anti-pyroptotic effects by inhibiting the NLRP3 inflammasome. Finally, we found that TSF reduces reactive oxygen species (ROS) production and thioredoxin-interacting protein (TXNIP) expression in AGE-stimulated HK-2 cells. More importantly, TSF decreased the colocalization of TXNIP and NLRP3, indicating that ROS-TXNIP may be the target of TSF. In summary, the anti-pyroptotic effect via the TXNIP-NLRP3-GSDMD axis may be an important mechanism of TSF for DKD therapy.

Association of Epoxide Hydrolase 2 Gene Arg287Gln with the Risk for Primary Hypertension in Chinese.

BACKGROUND: Epoxide hydrolase 2 (EPHX2) gene coding for soluble epoxide hydrolase is a potential candidate in the pathogenesis of hypertension. OBJECTIVES: We aimed to assess the association of a missense mutation, R287Q, in EPHX2) gene coding for soluble epoxide hydrolase is a potential candidate in the pathogenesis of hypertension. METHODS: This study involved 782 patients with primary hypertension and 458 healthy controls. Genotyping was done using TaqMan technique. Activity of soluble epoxide hydrolase fusion proteins was evaluated by the conversion of 11,12-EET to corresponding 11,12-DHET using ELISA kit. RESULTS: After taking carriers of R287Q variant GG genotype as a reference, those with GA genotype had a significantly reduced risk of hypertension (adjusted odds ratio: 0.72, 95% confidence interval: 0.56 to 0.93, P = 0.013). Five significant risk factors were identified, including age, body mass index, total cholesterol, homocysteine, and R287Q variant. These five risk factors for hypertension were represented in a nomogram, with a descent prediction accuracy (C-index: 0.833, P = 0.013). Five significant risk factors were identified, including age, body mass index, total cholesterol, homocysteine, and R287Q variant. These five risk factors for hypertension were represented in a nomogram, with a descent prediction accuracy (C-index: 0.833. CONCLUSIONS: We provide evidence that R287Q mutation in EPHX2 gene was associated with reduced risk of primary hypertension and low activity of soluble epoxide hydrolase.EPHX2) gene coding for soluble epoxide hydrolase is a potential candidate in the pathogenesis of hypertension.

Inhibiting calpain 1 and 2 in cyclin G associated kinase-knockout mice mitigates podocyte injury.

Evidence for reduced expression of cyclin G associated kinase (GAK) in glomeruli of patients with chronic kidney disease was observed in the Nephroseq human database, and GAK was found to be associated with the decline in kidney function. To examine the role of GAK, a protein that functions to uncoat clathrin during endocytosis, we generated podocyte-specific Gak-knockout mice (Gak-KO), which developed progressive proteinuria and kidney failure with global glomerulosclerosis. We isolated glomeruli from the mice carrying the mutation to perform messenger RNA profiling and unearthed evidence for dysregulated podocyte calpain protease activity as an important contributor to progressive podocyte damage. Treatment with calpain inhibitor III specifically inhibited calpain-1/-2 activities, mitigated the degree of proteinuria and glomerulosclerosis, and led to a striking increase in survival in the Gak-KO mice. Podocyte-specific deletion of Capns1, essential for calpain-1 and calpain-2 activities, also improved proteinuria and glomerulosclerosis in Gak-KO mice. Increased podocyte calpain activity-mediated proteolysis of IκBα resulted in increased NF-κB p65-induced expression of growth arrest and DNA-damage-inducible 45 beta in the Gak-KO mice. Our results suggest that loss of podocyte-associated Gak induces glomerular injury secondary to calcium dysregulation and aberrant calpain activation, which when inhibited, can provide a protective role.

Formononetin alleviates hepatic steatosis by facilitating TFEB-mediated lysosome biogenesis and lipophagy.

Formononetin has been reported to ameliorate hyperlipidemia and obesity, but its effect and mechanism of action in anti-non-alcoholic fatty liver disease (NAFLD) remain unclear. Lipophagy is a critical protective mechanism during steatosis development that results in the decomposition of lipid droplets through autophagy and the prevention of cellular lipid accumulation. This study aimed to investigate the beneficial role of formononetin in treating NAFLD and explore the mechanism of lipophagy in formononetin anti-hepatic steatosis effects. Formononetin treatment significantly ameliorated hepatic steatosis in HFD mice. Consistently, formononetin also reduced FFAs-stimulated lipid accumulation in HepG2 cells and primary mouse hepatocytes. Further analysis revealed that steatosis increased LC3B-II, a marker of autophagy, but caused blockade of autophagic flux associated with a lack of lysosomes. Treatment with formononetin promoted lysosome biogenesis and autophagosome-lysosome fusion, relieving the blockade in autophagic flux and further induced lipophagy. Mechanistically, formononetin activated adenosine monophosphate activated protein kinase (AMPK) and promoted subsequent nuclear translocation of transcription factor EB (TFEB), a key regulator of lysosome biogenesis. TFEB inhibition markedly abolished formononetin-induced lysosome biogenesis, autophagosome-lysosome fusion and lipophagy and concomitantly alleviated lipid accumulation. Formononetin improved hepatic steatosis via TFEB-mediated lysosome biogenesis, which provides new evidence regarding formononetin's anti-NAFLD effects.