Associations between cardiometabolic indices and the risk of diabetic kidney disease in patients with type 2 diabetes.

BACKGROUND: This study was designed to assess the associations between emerging cardiometabolic indices-the atherogenic index of plasma (AIP), the stress hyperglycemia ratio (SHR), the triglyceride-glucose (TyG) index, and the homeostasis model assessment of insulin resistance (HOMA-IR)-and the incidence of diabetic kidney disease (DKD) in type 2 diabetes (T2D) patients. METHODS: We consecutively enrolled 4351 T2D patients. The AIP, SHR, TyG index, and HOMA-IR were calculated from baseline parameters. DKD was defined as a urine albumin/creatinine ratio > 30 mg/g or an eGFR < 60 mL/min per 1.73 m. All participants were categorized into tertiles based on the cardiometabolic indices. Multivariate logistic regression models, restricted cubic splines, and receiver operating characteristic (ROC) curves were used for analysis. RESULTS: A total of 1371 (31.5%) patients were diagnosed with DKD. A restricted cubic spline showed a J-shaped association of the AIP and TyG index with DKD, a log-shaped association between HOMA-IR and DKD, and a U-shaped association between the SHR and DKD incidence. Multivariate logistic regression revealed that individuals in the highest tertile of the four cardiometabolic indices had a significantly greater risk of DKD than did those in the lowest tertile (AIP: OR = 1.08, 95% CI = 1.02-1.14, P = 0.005; SHR: OR = 1.42, 95% CI = 1.12-1.81, P = 0.004; TyG index: OR = 1.86, 95% CI = 1.42-2.45, P < 0.001; HOMA-IR: OR = 2.24, 95% CI = 1.52-3.30, P < 0.001). The receiver operating characteristic curves showed that the HOMA-IR score was better than other indices at predicting the risk of DKD, with an optimal cutoff of 3.532. CONCLUSIONS: Elevated AIP, SHR, TyG index and HOMA-IR are associated with a greater risk of DKD in patients with T2D. Among these indices, the HOMA-IR score demonstrated the strongest association with and predictive value for DKD incidence.

Novel Angiogenesis Role of GLP-1(32-36) to Rescue Diabetic Ischemic Lower Limbs via GLP-1R-Dependent Glycolysis in Mice.

BACKGROUND: Restoring the capacity of endothelial progenitor cells (EPCs) to promote angiogenesis is the major therapeutic strategy of diabetic peripheral artery disease. The aim of this study was to investigate the effects of GLP-1 (glucagon-like peptide 1; 32-36)-an end product of GLP-1-on angiogenesis of EPCs and T1DM (type 1 diabetes) mice, as well as its interaction with the classical GLP-1R (GLP-1 receptor) pathway and its effect on mitochondrial metabolism. METHODS: In in vivo experiments, we conducted streptozocin-induced type 1 diabetic mice as a murine model of unilateral hind limb ischemia to examine the therapeutic potential of GLP-1(32-36) on angiogenesis. We also generated Glp1r-/- mice to detect whether GLP-1R is required for angiogenic function of GLP-1(32-36). In in vitro experiments, EPCs isolated from the mouse bone marrow and human umbilical cord blood samples were used to detect GLP-1(32-36)-mediated angiogenic capability under high glucose treatment. RESULTS: We demonstrated that GLP-1(32-36) did not affect insulin secretion but could significantly rescue angiogenic function and blood perfusion in ischemic limb of streptozocin-induced T1DM mice, a function similar to its parental GLP-1. We also found that GLP-1(32-36) promotes angiogenesis in EPCs exposed to high glucose. Specifically, GLP-1(32-36) has a causal role in improving fragile mitochondrial function and metabolism via the GLP-1R-mediated pathway. We further demonstrated that GLP-1(32-36) rescued diabetic ischemic lower limbs by activating the GLP-1R-dependent eNOS (endothelial NO synthase)/cGMP/PKG (protein kinase G) pathway. CONCLUSIONS: Our study provides a novel mechanism with which GLP-1(32-36) acts in modulating metabolic reprogramming toward glycolytic flux in partnership with GLP-1R for improved angiogenesis in high glucose-exposed EPCs and T1DM murine models. We propose that GLP-1(32-36) could be used as a monotherapy or add-on therapy with existing treatments for peripheral artery disease. REGISTRATION: URL: www.ebi.ac.uk/metabolights/; Unique identifier: MTBLS9543.

Transferrin receptor levels and its rare variant are associated with human obesity.

AIM: Iron homeostasis is critical for functional respiratory chain complex of mitochondrial, thus potentially contributing to fat biology and energy homeostasis. Transferrin receptor (Tfrc) binds to transferrin for extracellular iron uptake and is recently reported to be involved in brown fat development and functionality. However, whether TFRC levels and variants are associated with human obesity is unknown. METHODS: To investigate the association of TFRC levels and variants with human obesity, fat biopsies were obtained from surgery. Exon-sequencing and genetic assessments were conducted of a case-control study. For TFRC levels assessment in fat biopsy, 9 overweight and 12 lean subjects were involved. For genetic study, obese (n = 1271) and lean subjects (n = 1455) were involved. TFRC levels were compared in abdominal mesenteric fat of pheochromocytoma patients versus control subjects, and overweight versus lean subjects. For genetic study, whole-exome sequencing of obese and matched control subjects were conducted and analyzed. In addition, the possible disruption in protein stability of TFRC variant was assessed by structural and molecular analysis. RESULTS: TFRC levels are increased in human browning adipose tissue and decreased in fat of overweight patients. Besides, TFRC levels are negatively correlated with body mass index and positively correlated with uncoupling protein 1 levels. Furthermore, a rare heterozygous missense variant p.I337V in TFRC shows a tendency to enrich in obese subjects. Structural and functional study reveals impaired protein stability of the TFRC variant compared to wild-type. CONCLUSIONS: Reduced TFRC levels and its rare variant p.I337V with protein instability are associated with human obesity.

Lipidomics Profiling Reveals Serum Phospholipids Associated with Albuminuria in Early Type 2 Diabetic Kidney Disease.

Early screening and administration of DKD are beneficial for renal outcomes of type 2 diabetic patients. However, the current early diagnosis using the albuminuria/creatine ratio (ACR) contains limitations. This study aimed to compare serum lipidome variation between type 2 diabetes and early DKD patients with increased albuminuria through an untargeted lipidomics method to explore the potential lipid biomarkers for DKD identification. 92 type 2 diabetic patients were enrolled and divided into two groups: DM group (ACR < 3 mg/mmol, n = 49) and early DKD group (3 mg/mmol ≤ ACR < 30 mg/mmol, n = 43). Fasting serum was analyzed through an ultraperformance liquid mass spectrometry tandem chromatography system (LC-MS). Orthogonal partial least-squares discriminant analysis (OPLS-DA) and univariate and multivariate analysis were performed to filter differentially depressed lipids. Receiver operating characteristic (ROC) curves were used to estimate the diagnostic capability of potential lipid biomarkers. We found that serum phospholipids including phosphatidylserine (PS), sphingomyelin (SM), and phosphatidylcholine (PC) were significantly upregulated in the DKD group and were highly correlated with the ACR. In addition, a panel of two phospholipids including PS(27:0)-H and PS(30:2e)-H showed good performance to help clinical lipids in early DKD identification, which increased the area under the curve (AUC) from 0.568 to 0.954. The study exhibited the serum lipidome variation in early DKD patients, and the increased phospholipids might participate in the development of albuminuria. The panel of PS(27:0)-H and PS(30:2e)-H could be a potential biomarker for DKD diagnosis.

ELABELA/APJ Axis Prevents Diabetic Glomerular Endothelial Injury by Regulating AMPK/NLRP3 Pathway.

ELABELA (ELA), a recently discovered peptide, is highly expressed in adult kidneys and the endothelium system. It has been identified as a novel endogenous ligand for the apelin receptor (APJ). This study aims to investigate the role of ELA in diabetic glomerular endothelial pyroptosis and its underlying mechanism. Initially, a significant decrease in ELA mRNA levels was observed in the renal cortex of db/db mice and high glucose-treated glomerular endothelial cells (GECs). It was also found that ELA deficiency in ELA+/- mice significantly accelerated diabetic glomerular injury, as shown by exacerbated glomerular morphological damage, increased serum creatine and blood urea nitrogen, and elevated 24-h urinary albumin excretion. In addition, in vivo overexpression of ELA prevented diabetic glomerular injury, reduced von Willebrand factor expression, restored endothelial marker CD31 expression, and attenuated the production of adhesive molecules such as intercellular adhesion molecule-1 and vascular cell adhesion molecule-1. Furthermore, in vitro studies confirmed that treatment with ELA inhibited GEC injury by regulating the NOD-like receptor protein 3 (NLRP3) inflammasome, as indicated by blocking NLRP3 inflammasome formation, decreasing cleaved Caspase-1 production, and inhibiting interleukin-1ß and interleukin-18 production. Moreover, in vitro experiments demonstrated that the protective effects of ELA in GECs during hyperglycemia were diminished by inhibiting adenosine monophosphate-activated protein kinase (AMPK) using Compound C or by APJ deficiency. Taken together, this study provides the first evidence that ELA treatment could prevent diabetic glomerular endothelial injury, which is partly mediated by the regulation of the AMPK/NLRP3 signaling pathway. Therefore, pharmacologically targeting ELA may serve as a novel therapeutic strategy for diabetic kidney disease.

Nomogram-Based Prediction of the Futile Recanalization Risk Among Acute Ischemic Stroke Patients Before and After Endovascular Therapy: A Retrospective Study.

Background and Purpose: Futile recanalization (FRC) is common among large artery occlusion (LAO) patients after endovascular therapy (EVT). We developed nomogram models to identify LAO patients at a high risk of FRC pre- and post-EVT to help neurologists select the optimal candidates for EVT. Methods: From April 2020 to July 2022, EVT and mTICI score ≥2b LAO patients were recruited. Nomogram models was developed by two-step approach for predicting the outcomes of LAO patients. First, the least absolute shrinkage and selection operator (LASSO) regression analysis was to optimize variable selection. Then, a multivariable analysis was to construct an estimation model with significant indicators from the LASSO. The accuracy of the model was verified using receiver operating characteristic (ROC), calibration curve, and decision curve analyses (DCA), along with validation cohort (VC). Results: Using LASSO, age, sex, hypertension history, baseline NIHSS, ASPECTS and baseline SBP upon admission were identified from the pre-EVT variables. Model 1 (pre-EVT) showed good predictive performance, with an area under the ROC curve (AUC) of 0.815 in the training cohort (TrC) and 0.904 in VC. Under the DCA, the generated nomogram was clinically applicable where risk cut-off was between 15%-85% in the TrC and 5%-100% in the VC. Moreover, age, ASPECTS upon admission, onset duration, puncture-to-recanalization (PTR) duration, and lymphocyte-to-monocyte ratio (LMR) were screened by LASSO. Model 2 (post-EVT) also demonstrated good predictive performance with AUCs of 0.888 and 0.814 for TrC and VC, respectively. Under the DCA, the generated nomogram was clinically applicable if the risk cut-off was between 13-100% in the TrC and 22-85% of VC. Conclusion: In this study, two nomogram models were generated that showed good discriminative performance, improved calibration, and clinical benefits. These nomograms can potentially accurately predict the risk of FRC in LAO patients pre- and post-EVT and help to select appropriate candidates for EVT.

Selective PPARγ modulator diosmin improves insulin sensitivity and promotes browning of white fat.

Peroxisome proliferator-activated receptor γ (PPARγ) is a master regulator of adipocyte differentiation, glucolipid metabolism, and inflammation. Thiazolidinediones are PPARγ full agonists with potent insulin-sensitizing effects, whereas their oral usage is restricted because of unwanted side effects, including obesity and cardiovascular risks. Here, via virtual screening, microscale thermophoresis analysis, and molecular confirmation, we demonstrate that diosmin, a natural compound of wide and long-term clinical use, is a selective PPARγ modulator that binds to PPARγ and blocks PPARγ phosphorylation with weak transcriptional activity. Local diosmin administration in subcutaneous fat (inguinal white adipose tissue [iWAT]) improved insulin sensitivity and attenuated obesity via enhancing browning of white fat and energy expenditure. Besides, diosmin ameliorated inflammation in WAT and liver and reduced hepatic steatosis. Of note, we determined that iWAT local administration of diosmin did not exhibit obvious side effects. Taken together, the present study demonstrated that iWAT local delivery of diosmin protected mice from diet-induced insulin resistance, obesity, and fatty liver by blocking PPARγ phosphorylation, without apparent side effects, making it a potential therapeutic agent for the treatment of metabolic diseases.

Sappanone a prevents diabetic kidney disease by inhibiting kidney inflammation and fibrosis via the NF-κB signaling pathway.

Background: Low grade of sterile inflammation plays detrimental roles in the progression of diabetic kidney disease (DKD). Sappanone A (SA), a kind of homoisoflavanone isolated from the heartwood of Caesalpinia sappan, exerts anti-inflammatory effects in acute kidney injury. However, whether SA has beneficial effects on diabetic kidney disease remains further exploration. Methods and Results: In the present study, uninephrectomized male mice were treated with Streptozotocin (STZ, 50 mg/kg) for five consecutive days to induce diabetes. Next, the diabetic mice were administered orally with SA (10, 20, or 30 mg/kg) or vehicle once per day. Our results showed that STZ treatment significantly enhanced damage in the kidney, as indicated by an increased ratio of kidney weight/body weight, elevated serum creatinine and blood urea nitrogen (BUN), as well as increased 24-h urinary protein excretion, whereas SA-treated mice exhibited a markedly amelioration in these kidney damages. Furthermore, SA attenuated the pathological changes, alleviated fibrotic molecules transforming growth factor-ß1 (TGF-ß1) and Collagen-IV (Col-IV) production, decreased inflammatory cytokines interleukin-1ß (IL-1ß) and tumor necrosis factor-α (TNF-α) expression in STZ-treated mice. Similarly, in glomerular mesangial cells, SA pretreatment decreased high glucose (HG)-induced proliferation, inflammatory cytokines excretion, and fibrotic molecules expression. Mechanistically, SA decreased the expression of nuclear factor kappa B (NF-κB) and restored the expression of total NF-κB inhibitor alpha (IκBα) both in vivo and in vitro. Conclusion: Our data suggest that SA may prevent diabetes-induced kidney inflammation and fibrosis by inhibiting the NF-κB pathway. Hence, SA can be potential and specific therapeutic value in DKD.

Antibiotic Azithromycin inhibits brown/beige fat functionality and promotes obesity in human and rodents.

Obesity, a metabolic disease caused by multiple factors, has become a global health problem. In addition to nutrient intake and sedentary lifestyle, environmental pollutants exposure has been shown to be involved in obesity epidemics. Antibiotics, a new type of environmental pollutant, have been widely used in animal husbandry, aquaculture and microorganism. However, the effects of antibiotics exposure on fat metabolism and metabolic diseases are largely unknown. Methods: We screened major types of antibiotics to examine their effects on the differentiation capacity and thermogenic functionality of brown and beige adipocytes, and found that azithromycin, one major kind of macrolide antibiotics suppressed brown and beige adipocyte functionality. We thus examined azithromycin accretion in adipose tissues of obese patients that correlates with BMI by high performance liquid chromatography-tandem mass spectrometry and systematically explore the influences of azithromycin on adiposity and metabolic performance in mice under high diet. Results: Azithromycin (macrolides) inhibits the mitochondrial and thermogenic gene programs of brown and beige adipocytes, thus disrupting their mitochondrial function and thermogenic response. Consistently, azithromycin treatment are more prone to diet-induced obesity in mice, and this was associated with impaired energy expenditure. Importantly, azithromycin is more accumulated in adipose tissue of obese patients and correlates with BMI and body weight. Mechanistically, we found that azithromycin inhibits mitochondria respiratory complex I protein levels and increases reactive oxidative species (ROS) levels, which causes damage of mitochondrial function in brown and beige adipocytes. The deleterious effects of azithromycin can be ameliorated by antioxidant N-acetyl-L-cysteine. Conclusions: Taken together, this work highlights the possible role of azithromycin in obesity epidemic and presents strategies for safe applications of antibiotics in the future.

Combination Usage of AdipoCount and Image-Pro Plus/ImageJ Software for Quantification of Adipocyte Sizes.

In recent decades, the prevalence of obesity has been rising. One of the major characteristics of obesity is fat accumulation, including hyperplasia (increase in number) and hypertrophy (increase in size). After histological staining, it is critical to accurately measure the number and size of adipocytes for assessing the severity of obesity in a timely fashion. Manual measurement is accurate but time-consuming. Although commercially available adipocyte counting tools, including AdipoCount, Image-Pro Plus, and ImageJ were helpful, limitations still exist in accuracy and time consuming. In the present study, we introduced the protocol of combined usage of these tools and illustrated the process with histological staining slides from adipose tissues of lean and obese mice. We found that the adipocyte sizes quantified by the tool combination were comparable as manual measurement, whereas the combined methods were more efficient. Besides, the recognition effect of monochrome segmentation image is better than that of color segmentation image. Overall, we developed a combination method to measure adipocyte sizes accurately and efficiently, which may be helpful for experimental process in laboratory and also for clinic diagnosis.

Neutrophil-to-Lymphocyte Ratio and Platelet-to-Lymphocyte Ratio Predict Mortality in Patients with Diabetic Foot Ulcers Undergoing Amputations.

PURPOSE: Elevated platelet-to-lymphocyte ratio (PLR) and neutrophil-to-lymphocyte ratio (NLR) are associated with poor outcomes in various diseases. The objectives of this study were to explore the utility of PLR and NLR in predicting all-cause mortality in patients with diabetic foot ulcers (DFU) undergoing amputations. PATIENTS AND METHODS: A retrospective observational study was performed that included a total of 348 DFU patients undergoing amputations. The primary end-point was all-cause death. According to the PLR and NLR cut-off values, patients were divided into two groups and Kaplan-Meier survival curves were constructed. Multivariable Cox regression was conducted to test the independent predictors of mortality in the study cohort. RESULTS: All-cause mortality was significantly higher in patients with a high PLR/NLR compared to those with a low PLR/NLR. In the low NLR group, the overall survival (OS) rates at 1, 3, and 5 years after amputation were 96.8%, 84% and 80.1%, respectively (p=0.001). In the high NLR group the corresponding OS rates at 1, 3, and 5 years were 85.2%, 58.6% and 23.9% (p<0.001). According to the multivariate analysis, age (HR 1.074, 95% CI 1.045-1.104, p<0.001), Wagner classification (HR 2.274, 95% CI 1.351-3.828, p=0.002), PLR (HR 1.794, 95% CI 1.014-3.174, p=0.045), NLR (HR 2.029, 95% CI 1.177-3.499, p=0.011), creatinine (HR 1.003, 95% CI 1.001-1.004, p<0.001) and direct bilirubin (HR 1.154, 95% CI 1.081-1.232, p<0.001) were independent predictors of mortality following amputation. CONCLUSION: Postoperative PLR and NLR values may be reliable predictive biomarkers of mortality in patients following amputation for DFU. Considering the high mortality in those patients, the patients with elevated PLR/NLR should be given more intensive in clinical practice.

Autonomous climbing: An effective exercise mode with beneficial outcomes of aerobic exercise and resistance training.

AIMS: To assess the effects of three specific exercise training modes, aerobic exercise (A), resistance training (R) and autonomous climbing (AC), aimed at proposing a cross-training method, on improving the physical, molecular and metabolic characteristics of mice without many side effects. MATERIALS AND METHODS: Seven-week-old male mice were randomly divided into four groups: control (C), aerobic exercise (A), resistance training (R), and autonomous climbing (AC) groups. Physical changes in mice were tracked and analysed to explore the similarities and differences of these three exercise modes. Histochemistry, quantitative real-time PCR (RT-PCR), western blot (WB) and metabolomics analysis were performed to identify the underlying relationships among the three training modes. KEY FINDINGS: Mice in the AC group showed better body weight control, glucose and energy homeostasis. Molecular markers of myogenesis, hypertrophy, antidegradation and mitochondrial function were highly expressed in the muscle of mice after autonomous climbing. The serum metabolomics landscape and enriched pathway comparison indicated that the aerobic oxidation pathway (pentose phosphate pathway, galactose metabolism and fatty acid degradation) and amino acid metabolism pathway (tyrosine, arginine and proline metabolism) were significantly enriched in group AC, suggesting an increased muscle mitochondrial function and protein balance ability of mice after autonomous climbing. SIGNIFICANCE: We propose a new exercise mode, autonomous climbing, as a convenient but effective training method that combines the beneficial effects of aerobic exercise and resistance training.

Transferrin Receptor Functionally Marks Thermogenic Adipocytes.

BACKGROUND: Thermogenic adipocytes, including beige and brown adipocytes, are critical for thermogenesis and energy homeostasis. Identification of functional cell surface markers of thermogenic adipocytes is of significance for potential application in biological and clinical practices. METHODS: With a combination of RNA-sequencing of in vivo and in vitro models, we identified transferrin receptor (Tfr1), a receptor specialized for cellular iron uptake, as a previously unappreciated cell surface molecule for thermogenic adipocytes compared to white adipocytes. The alternation of Tfr1 levels under physiological and pathological stimuli was assessed, and the mitochondria functionality, browning capacity, and iron metabolism of mature adipocytes were examined with Tfr1 knockdown. RESULTS: Tfr1 was expressed predominantly in thermogenic adipocytes versus white adipocyte, and its expression levels were tightly correlated with the activation or inhibition status of thermogenic adipocytes under external stimuli. Besides, Tfr1 gene deficiency in thermogenic adipocytes led to reduced thermogenic gene programs and mitochondrial integrity. CONCLUSION: Tfr1 functionally marks thermogenic adipocytes and could serve as a potential thermogenic adipocyte surface marker.

Ginsenoside Rb2 improves insulin resistance by inhibiting adipocyte pyroptosis.

Pyroptosis plays a critical role in the development of obesity-associated inflammation and insulin resistantance (IR). Ginsenoside Rb2 (Rb2), the main component of ginsenosides has drawn appreciable interest in the context of glucose metabolism. In the present study, we investigated Rb2-mediated protection against obesity-induced IR and the related mechanisms. Rb2 could significantly reduce high-fat diet (HFD)-induced body weight changes, fat accumulation and IR. In addition, Rb2 treatment inhibited pyroptosis-related genes and proteins, such as caspase-1, ASC, NLRP3, IL-1ß and GSDMD in HFD-fed mice. The above results were recapitulated in 3T3-L1 adipocytes and demonstrated that Rb2 improved TNF-α induced IR and pyroptosis in 3T3-L1 adipocytes. Furthermore, Rb2 reduced the phosphorylation levels of p65 and IκBα both in vitro and in vivo. The present study showed that Rb2, which could serve as a promising agent for the treatment of IR and obesity, ameliorated IR by inhibiting pyroptosis in adipocytes in vivo and in vitro through the NF-κB pathway.

Comprehensive analysis of circular RNA profiles in skeletal muscles of aging mice and after aerobic exercise intervention.

Aging induces gradual accumulation of damages in cells and tissues, which leads to physiological dysfunctions. Aging-associated muscle dysfunction is commonly seen in aged population and severely affects their physical activity and life quality, against which aerobic training has been shown to exert antagonizing or alleviating effects. Circular RNAs (circRNAs) play important roles in various physiological processes, yet their involvement in aging-associated muscle dysfunction is not well understood. In this study, we performed comprehensive analysis of circRNAs profiles in quadriceps muscles in sedentary young and aging mice, as well as aging mice with aerobic exercise using RNA sequencing. Our results identified circRNAs altered by factors of aging and aerobic exercise. Their host genes were then predicted and analyzed by gene ontology enrichment analysis. Importantly, we found that circBBS9 featured decreased levels in aging compared to young mice and elevated expression in exercise versus sedentary aging mice. Besides, we performed GO and KEGG analysis on circBBS9 target genes, as well as established the circBBS9-miRNA-mRNAs interaction network. Our results indicate that circBBS9 may play active roles in muscle aging by mediating the benefits of aerobic training intervention, thus may serve as a novel therapeutic target combating aging-associated muscle dysfunction.