Sodium-glucose cotransporter 1 promotes the biofunctions of perivascular preadipocytes mediated by Akt/mTOR/p70S6K signaling pathway.

The influence of SGLT-1 on perivascular preadipocytes (PVPACs) and vascular remodeling is not well understood. This study aimed to elucidate the role and mechanism of SGLT-1-mediated PVPACs bioactivity. PVPACs were cultured in vitro and applied ex vivo to the carotid arteries of mice using a lentivirus-based thermosensitive in situ gel (TISG). The groups were treated with Lv-SGLT1 (lentiviral vector, overexpression), Lv-siSGLT1 (RNA interference, knockdown), or specific signaling pathway inhibitors. Assays were conducted to assess changes in cell proliferation, apoptosis, glucose uptake, adipogenic differentiation, and vascular remodeling in the PVPACs. Protein expression was analyzed by Western blotting, immunocytochemistry, and/or immunohistochemistry. The methyl thiazolyl tetrazolium (MTT) assay and Hoechst 33342 staining indicated that SGLT-1 overexpression significantly promoted PVPACs proliferation and inhibited apoptosis in vitro. Conversely, SGLT-1 knockdown exerted the opposite effect. Oil Red O staining revealed that SGLT-1 overexpression facilitated adipogenic differentiation, while its inhibition mitigated these effects. 3H-labeled glucose uptake experiments demonstrated that SGLT-1 overexpression enhanced glucose uptake by PVPACs, whereas RNA interference-mediated SGLT-1 inhibition had no significant effect on glucose uptake. Moreover, RT-qPCR, Western blotting, and immunofluorescence analyses revealed that SGLT-1 overexpression upregulated FABP4 and VEGF-A levels and activated the Akt/mTOR/p70S6K signaling pathway, whereas SGLT-1 knockdown produced the opposite effects. In vivo studies corroborated these findings and indicated that SGLT-1 overexpression facilitated carotid artery remodeling. Our study demonstrates that SGLT-1 activation of the Akt/mTOR/p70S6K signaling pathway promotes PVPACs proliferation, adipogenesis, glucose uptake, glucolipid metabolism, and vascular remodeling.NEW & NOTEWORTHY SGLT-1 is expressed in PVPACs and can affect preadipocyte glucolipid metabolism and vascular remodeling. SGLT-1 promotes the biofunctions of PVPACs mediated by Akt/mTOR/p70S6K signaling pathway. Compared with caudal vein or intraperitoneal injection, the external application of lentivirus-based thermal gel around the carotid artery is an innovative attempt at vascular remodeling model, it may effectively avoid the transfection of lentiviral vector into the whole body of mice and the adverse effect on experimental results.

PFKFB2 Inhibits Ferroptosis in Myocardial Ischemia/Reperfusion Injury Through Adenosine Monophosphate-Activated Protein Kinase Activation.

ABSTRACT: Six-phosphofructo-2-kinase/fructose-2, 6-bisphosphatase 2 (PFKFB2) is a key regulator of glycolytic enzyme. This study identified whether PFKFB2 can regulate myocardial ferroptosis in ischemia/reperfusion (I/R) injury. Mice myocardial (I/R) injury and H9c2 cells oxygen-glucose deprivation/reperfusion (OGD/R) models were established. PFKFB2 expression was enhanced in I/R mice and OGD/R H9c2 cells. Overexpression of PFKFB2 improves heart function in I/R mice. Overexpression of PFKFB2 inhibits I/R and OGD/R-induced ferroptosis in mice and H9c2 cells. Mechanistically, overexpression of PFKFB2 activates the adenosine monophosphate-activated protein kinase (AMPK). AMPK inhibitor compound C reverses effect of PFKFB2 overexpression in reducing ferroptosis under OGD/R treatment. In conclusion, PFKFB2 protects hearts against I/R-induced ferroptosis through activation of the AMPK signaling pathway.

Association between triglyceride glucose index, coronary artery calcification and multivessel coronary disease in Chinese patients with acute coronary syndrome.

BACKGROUND: Multivessel coronary disease (MVCD) is the common type of coronary artery disease in acute coronary syndrome (ACS). Coronary artery calcification (CAC) has been confirmed the strong predictor of major adverse cardiovascular events (MACEs). Several studies have validated that triglyceride glucose (TyG) index can reflect the degree of coronary calcification or predict MACEs. However, no evidence to date has elucidated and compared the predictive intensity of TyG index or/and coronary artery calcification score (CACS) on multi-vascular disease and MACEs in ACS patients. METHODS: A total of 935 patients, diagnosed with ACS and experienced coronary computed tomography angiography (CCTA) from August 2015 to March 2022 in the Second Hospital of Shandong University, were selected for retrospective analysis. The subjects were divided into TyG index quartile 1-4 groups (Q1-Q4 groups), non-multivessel coronary disease (non-MVCD) and multivessel coronary disease (MVCD) groups, respectively. The general data, past medical or medication history, laboratory indicators, cardiac color Doppler ultrasound, CACS, and TyG indexes were respectively compared among these groups. The ROC curve preliminarily calculated and analyzed the diagnostic value of TyG index, CACS, and the combination of the two indicators for MVCD. Univariate and multivariate logistic regression analysis discriminated the independent hazard factors for forecasting MVCD. RESULTS: Compared with the lower TyG index and non-MVCD groups, the higher TyG index and MVCD groups had higher values of age, smoking history, waist circumference, systolic blood pressure, low-density lipoprotein cholesterol(LDL-C), fasting blood glucose and glycosylated hemoglobin, and CACS, but lower values of high-density lipoprotein cholesterol(HDL-C) (all P < 0.01). Coronary artery calcification is more common in the left anterior descending artery. Compared with non-MVCD, each unit increase in TyG index was associated with a 1.213-fold increased risk of MVCD. Logistic regression analysis adjusted for potential confounders indicated that TyG index is an independent risk factor for MVCD. With the increase of TyG index, the incidence of MACEs, apart from all-cause death, cardiac death, unexpected re-hospitalization of heart failure, recurrent ACS or unplanned revascularization, and non-fatal stroke in coronary artery increased (P log-rank < 0.001). CONCLUSION: TyG index could completely substitute for CACS as a reliable, practical, and independent indicator for predicting the severity and prognosis of MVCD in patients with ACS.

HIF-1/AKT Signaling-Activated PFKFB2 Alleviates Cardiac Dysfunction and Cardiomyocyte Apoptosis in Response to Hypoxia.

Myocardial infarction (MI) is the most prevalent disease with severe mortality, and hypoxia-induced cardiac injury and cardiomyocyte apoptosis are the significant and harmful consequences of this disease. The cross talk between hypoxia signaling and glycolysis energy flux plays a critical role in modulating MI-related heart disorder. However, the underlying mechanism remains unclear. Here, we aimed to explore the effect of a key glycolytic enzyme of 6-phosphofructo-2-kinase/fructose-2, 6-bisphosphatase 2 (PFKFB2) on cardiac dysfunction and apoptosis in response to hypoxia. Our data demonstrated that the mRNA and protein expression of PFKFB2 were significantly elevated in the MI mice. The MI treatment promoted the activation of PFKFB2 in vivo, as presented by the remarkably increased phosphorylation levels of PFKFB2. PFKFB2 depletion enhanced MI-induced cardiac dysfunction and cardiomyocyte apoptosis in the MI mouse model. Moreover, hypoxia treatment dramatically upregulated the expression and activation of PFKFB2 in a time-dependent manner in cardiomyocytes. Hypoxia-stimulated PFKFB2 relieved hypoxia-induced cardiomyocyte apoptosis in vitro. PFKFB2 activated the fructose-2, 6-bisphosphate (Fru-2, 6-p2) /PFK/anaerobic adenosine triphosphate (ATP) glycolysis energy flux in response to hypoxia in cardiomyocytes. Mechanically, hypoxia-activated PFKFB2 by stimulating the hypoxia-inducible factor 1 (HIF-1) /ATK signaling. Thus, we conclude that HIF-1/AKT axis-activated PFKFB2 alleviates cardiac dysfunction and cardiomyocyte apoptosis in response to hypoxia. Our finding presents a new insight into the mechanism by which HIF-1/AKT/PFKFB2 signaling modulates MI-related heart disorder under the hypoxia condition, providing potential therapeutic targets and strategy for hypoxia-related myocardial injury.

Chemerin-induced angiogenesis and adipogenesis in 3 T3-L1 preadipocytes is mediated by lncRNA Meg3 through regulating Dickkopf-3 by sponging miR-217.

PURPOSE: Obesity is often caused by the excess adipogenesis and regulated by long non-coding RNAs (lncRNAs) and microRNAs (miRNAs). We performed this study to investigate the influence of Meg3 expression on adipogenesis and also the Meg3/miR-217/Dkk3 axis-mediated molecular mechanism in adipogenesis and angiogenesis. METHODS: 3 T3-L1 preadipocytes were incubated with chemerin and transfected with Meg3-overexpressing (OE-Meg3) and Dkk3-overexpressing (OE-Dkk3) plasmids, siRNAs, and miR-217 mimics, inhibitor and scrambled sequences for 48 h or 72 h. The changes in cell proliferation, adipogenesis and angiogenesis ability in 3 T3-L1 preadipocytes was detected by using the corresponding assay. The expressions of related proteins were detected via western blot. RESULTS: Chemerin decreased miR-217 expression and increased Meg3 expression, meanwhile promoted the proliferation, adipogenesis and angiogenesis in 3 T3-L1 preadipocytes. Besides, OE-Meg3 exerted the synergistic effect on 3 T3-L1 preadipocytes when co-treated with chemerin. The target interactions between Meg3 and miR-217 as well as between miR-217 and Dkk3 were validated using dual-luciferase reporter system. SiMeg3 antagonized chemerin-induced changes, while the addition of miR-217 inhibitor attenuated siMeg3-induced changes in 3 T3-L1 preadipocytes. The proliferation, adipogenesis and angiogenesis in 3 T3-L1 preadipocytes were suppressed by miR-217 mimics, while promoted by the OE-Dkk3 Chemerin promoted the expression of fatty acid binding protein 4 and vascular endothelial growth factor (VEGF) proteins, and decreased the expression of cyclin D1, c-Myc, and ß-catenin proteins. Meanwhile, these effects were further enhanced by OE-Meg3 or OE-Dkk3. However, the transfection of siMeg3, or miR-217 mimics, or siDkk3 reversed the previous changes. CONCLUSIONS: Meg3/miR-217/Dkk3 induced adipogenesis and angiogenesis in 3 T3-L1 preadipocytes via activating VEGF signaling pathway and inhibiting Wnt/ß-catenin signaling pathway.

FSP1 promotes the biofunctions of adventitial fibroblast through the crosstalk among RAGE, JAK2/STAT3 and Wnt3a/ß-catenin signalling pathways.

Emerging evidence indicates that fibroblast-specific protein 1 (FSP1) provides vital effects in cell biofunctions. However, whether FSP1 influences the adventitial fibroblast (AF) and vascular remodelling remains unclear. Therefore, we investigated the potential role and action mechanism of FSP1-mediated AF bioactivity. AFs were cultured and stimulated with FSP1 and siRNA-FSP1 in vitro. Viability assays demonstrated that siRNA-FSP1 counteracted AFs proliferative, migratory and adherent abilities enhanced with FSP1. Flow cytometry revealed that FSP1 increased AFs number in S phase and decreased cellular apoptosis. Contrarily, siRNA-FSP1 displayed the contrary results. RT-PCR, Western blotting and immunocytochemistry showed that FSP1 synchronously up-regulated the expression of molecules in RAGE, JAK2/STAT3 and Wnt3a/ß-catenin pathways and induced a proinflammatory cytokine profile characterized by high levels of MCP-1, ICAM-1 and VCAM-1. Conversely, FSP1 knockdown reduced the expression of these molecules and cytokines. The increased number of autophagosomes in FSP1-stimulated group and fewer autophagic corpuscles in siRNA-FSP1 group was observed by transmission electron microscope (TEM). Autophagy-related proteins (LC3B, beclin-1 and Apg7) were higher in FSP1 group than those in other groups. Conversely, the expression of p62 protein was shown an opposite trend of variation. Therefore, these pathways can promote AFs bioactivity, facilitate autophagy and induce the expression of the proinflammatory cytokines. Contrarily, siRNA-FSP1 intercepts the crosstalk of these pathways, suppresses AF functions, restrains autophagy and attenuates the expression of the inflammatory factors. Our findings indicate that crosstalk among RAGE, STAT3/JAK2 and Wnt3a/ß-catenin signalling pathways may account for the mechanism of AF functions with the stimulation of FSP1.