Serpina3c deficiency induced necroptosis promotes non-alcoholic fatty liver disease through ß-catenin/Foxo1/TLR4 signaling.

Non-alcoholic fatty liver disease (NAFLD) is a public health challenge and an increasing cause of chronic liver disease worldwide. However, the underlying molecular mechanism remains unclear. The aim of this study was to determine the precise role of serpina3c in the process of NAFLD. Male Apoe-/- /serpina3c-/- double knockout (DKO) and Apoe-/- mice were fed a high-fat diet (HFD) for 12 weeks. Several markers of steatosis and inflammation were evaluated. In vitro cell models induced by palmitic acid (PA) treatment were used to evaluate the beneficial effect of serpina3c on necroptosis and the underlying molecular mechanism. Compared with Apoe-/- mice, DKO mice exhibited a significantly exacerbated hepatic steatosis, increased hepatic triglyceride content and expression of genes involved in lipid metabolism (SREBP1c and SCD1), promoted hepatic inflammation and fibrosis, promoted necroptosis by increasing expression of receptor-interacting protein 3 (RIP3), phosphorylated mixed lineage kinase domain-like (MLKL) and high mobility group box 1 (HMGB1). Notably, serpina3c deficiency increased ß-catenin, Foxo1, and Toll-like receptor 4 (TLR4) protein expression. In vitro , serpina3c knockdown promoted necroptosis and lipid droplet formation under condition of lipotoxicity. However, these phenomena were reversed by the overexpression of serpina3c. Mechanistically, downregulation of serpina3c expression promoted Foxo1 and ß-catenin colocalized in the nucleus under condition of lipotoxicity, consequently upregulating the expression of TLR4. Conversely, disruption of Foxo1-ß/catenin by Foxo1 and ß-catenin inhibitors decreased TLR4 expression and ameliorated hepatic necroptosis in vitro. This study highlights a novel mechanism that serpina3c modulates NAFLD development by inhibiting necroptosis via ß-catenin/Foxo1/TLR4.

Maintained P2Y12 inhibitor monotherapy after shorter-duration of dual antiplatelet therapy in patients undergoing coronary drug-eluting stents implantation: An updated meta-analysis of randomized trials.

WHAT IS KNOWN AND OBJECTIVE: It is well known that high in-stent thrombotic risk due to the superimposition of a platelet-rich thrombus was considered as the main origin of major adverse cardiac events after stent implantation. The clinical management of antiplatelet therapy strategy after percutaneous coronary intervention (PCI) remains controversial. This study is sought to explore the efficacy and safety of a maintained P2Y12 inhibitor monotherapy after shorter-duration of dual antiplatelet therapy (DAPT) in these patients. METHODS: Medline, Google Scholar, Web of Science, and the Cochrane Controlled Trials Registry were searched online for retrieving eligible citations. A composite of all-cause death, myocardial infarction (MI) and stroke was defined as major adverse cardio- and cerebro-vascular events (MACCE), which is analysed as the primary efficacy endpoint. The risk of bleeding events was chosen as safety endpoint. RESULTS: Five randomized clinical trials (RCT) with 32,143 patients were finally analysed. A maintained P2Y12 inhibitor monotherapy after shorter-duration of DAPT cloud not only reduce the incidence of MACCE [odds ratios (OR): 0.89, 95% confidence intervals (CI): 0.79-0.99, p = 0.037], but also the bleeding risk (OR 0.61, 95% CI: 0.44-0.85, p = 0.003). No higher incidence of any ischaemic events, including MI, stroke or definite stent thrombosis (ST) was observed with respect to this new antiplatelet therapy option. CONCLUSIONS: A maintained P2Y12 inhibitor monotherapy after shorter-duration of DAPT was suggested as a more preferable antiplatelet therapy option in patients undergoing coronary drug-eluting stents (DES) placement. Larger and more powerful randomized trials with precise sub-analyses are still necessary for further confirming these relevant benefits.

Protective role of serpina3c as a novel thrombin inhibitor against atherosclerosis in mice.

Abnormal vascular smooth muscle cell (VSMC) proliferation is a critical step in the development of atherosclerosis. Serpina3c is a serine protease inhibitor (serpin) that plays a key role in metabolic diseases. The present study aimed to investigate the role of serpina3c in atherosclerosis and regulation of VSMC proliferation and possible mechanisms. Serpina3c is down-regulated during high-fat diet (HFD)-induced atherosclerosis. An Apoe-/-/serpina3c-/--double-knockout mouse model was used to determine the role of serpina3c in atherosclerosis after HFD for 12 weeks. Compared with Apoe-/- mice, the Apoe-/-/serpina3c-/- mice developed more severe atherosclerosis, and the number of VSMCs and macrophages in aortic plaques was significantly increased. The present study revealed serpina3c as a novel thrombin inhibitor that suppressed thrombin activity. In circulating plasma, thrombin activity was high in the Apoe-/-/serpina3c-/- mice, compared with Apoe-/- mice. Immunofluorescence staining showed thrombin and serpina3c colocalization in the liver and aortic cusp. In addition, inhibition of thrombin by dabigatran in serpina3c-/- mice reduced neointima lesion formation due to partial carotid artery ligation. Moreover, an in vitro study confirmed that thrombin activity was also decreased by serpina3c protein, supernatant and cell lysate that overexpressed serpina3c. The results of experiments showed that serpina3c negatively regulated VSMC proliferation in culture. The possible mechanism may involve serpina3c inhibition of ERK1/2 and JNK signaling in thrombin/PAR-1 system-mediated VSMC proliferation. Our results highlight a protective role for serpina3c as a novel thrombin inhibitor in the development of atherosclerosis, with serpina3c conferring protection through the thrombin/PAR-1 system to negatively regulate VSMC proliferation through ERK1/2 and JNK signaling.

Erythropoietin Attenuates Cardiac Dysfunction in Rats by Inhibiting Endoplasmic Reticulum Stress-Induced Diabetic Cardiomyopathy.

PURPOSE: Enhanced endoplasmic reticulum (ER) stress and down-regulated SERCA2a expression play crucial roles in diabetes. We aimed to verify whether erythropoietin (EPO) attenuates cardiac dysfunction by suppressing ER stress in diabetic rats. METHODS: Forty male SD rats were randomly divided into four groups: control, EPO-treated control, vehicle-treated diabetic, and EPO-treated diabetic groups. The animals in the EPO-treated control and diabetic groups were administered recombinant human EPO (1000 U/kg body weight) once per week for 12 weeks. RT-PCR and Western blotting assays were performed to detect the expression of 78-kDa glucose-regulated protein precursor (GRP78) and sarcoplasmic/endoplasmic reticulum Ca2+-ATPase (SERCA2a). We cultured neonatal rat cardiomyocytes and investigated the protective effects of EPO against high glucose (HG)-induced apoptosis. Intracellular calcium levels were measured through confocal microscopy. RESULTS: We observed increased myocardial GRP78 expression and decreased myocardial SERCA2a expression in diabetic rats. EPO prevented the changes in GRP78, SERCA2a expression and cardiac dysfunction in diabetic rats. The levels of GRP78 protein were significantly reduced in EPO-treated diabetic rats compared with vehicle-treated diabetic rats (GRP78 protein 0.09 ± 0.03 vs. 0.54 ± 0.04, P < 0.01). The levels of the SERCA2a proteins were significantly increased in EPO-treated diabetic rats compared with vehicle-treated diabetic rats (SERCA2a protein 0.60 ± 0.05 vs. 0.13 ± 0.04, P < 0.01). A reduction in apoptosis was observed in the cardiomyocytes treated with 20 U/mL EPO compared with the cardiomyocytes cultured under HG conditions (apoptosis rate 18.9 ± 1.94 vs. 37.9 ± 1.59%, P < 0.01). CONCLUSIONS: This study demonstrates that EPO treatment improved the parameters of cardiac function following HG-induced injury by suppressing ER stress and inducing SERCA2a expression.

Left ventricular subclinical dysfunction associated with myocardial deformation changes in obstructive sleep apnea patients estimated by real-time 3D speckle-tracking echocardiography.

BACKGROUND: Previous studies have demonstrated that patients with obstructive sleep apnea (OSA) may develop left ventricular (LV) diastolic dysfunction. We aimed to study whether OSA patients have LV regional systolic dysfunction with myocardial deformation changes, despite a normal LV ejection fraction, using real-time 3D speckle-tracking echocardiography (Rt3D-STE). METHODS: Seventy-eight patients with OSA and no comorbidities were studied. They were divided into the following three groups according to the apnea-hypopnea index (AHI): 5~15/h as group I (mild OSA, 26 cases), 15~30/h as group II (moderate OSA, 29 cases), and ≥30/h as group III (severe OSA, 23 cases). Thirty gender-age-matched normal subjects were included as controls. The parameters of LV diastolic function were acquired with traditional echocardiography. The LV myocardial deformation parameters were obtained, including the longitudinal (LS), circumferential (CS), radial (RS), and area (AS) strains, with Rt3D-STE. RESULTS: LV global systolic function was normal in all patients, but diastolic function was impaired in groups II and III (E/E' was 9.6 ± 2.8 and 10.4 ± 2.5, respectively, p < 0.0001). The global LS and AS were significantly reduced in groups II and III compared with the controls and group I (LS 15.9 ± 1.4 % and 14.8 ± 1.5 % vs 18.2 ± 1.7 % and 17.8 ± 1.5 %; AS 27.4 ± 1.8 % and 24.9 ± 2.3 % vs 33.4 ± 2.2 % and 32.7 ± 2.9 %, respectively, p < 0.0001), but the global CS and RS were significantly reduced only in group III (17.3 ± 1.4 % and 43.1 ± 6.5 % vs 19.6 ± 1.6 % and 55.4 ± 4.0 %, respectively, <0.0001). The severity of OSA was significantly associated with the LV global AS value (r = -0.80, p < 0.0001), LS (r = -0.64, p < 0.0001), CS (r = -0.51, p < 0.0001), and RS (r = -0.62, p < 0.0001). CONCLUSIONS: Patients with moderate and severe OSA tended to have both LV diastolic dysfunction and abnormalities in regional systolic function with myocardial deformation changes, in spite of the normal LV ejection fraction. Myocardial strains of the LV were negatively correlated with the AHI. Rt-3DST had important clinical significance in the early evaluation of cardiac dysfunction in OSA patients.

Prognostic significance of serum dynamin­related protein 1 in patients with heart failure: Findings from a prospective observational study.

Mitochondrial dysfunction plays a critical role in the development and exacerbation of heart failure (HF). Dynamin-related protein 1 (Drp1), a key regulator of mitochondrial fission, influences cardiac energy metabolism. The present study investigated the relationship between serum Drp1 levels and the prognosis of patients with HF across a broad spectrum. Serum Drp1 concentrations were measured using ELISA. The primary outcome was the risk of composite major adverse cardiac events (MACEs), which included instances of cardiac death and HF-related readmissions. To assess the prognostic significance of serum Drp1, a receiver operating characteristic curve was constructed to predict MACE-free survival. Additionally, an optimal threshold value for Drp1 was determined and was used to stratify patients into different risk categories. A total of 256 HF patients were finally included and categorized into two groups based on their serum Drp1 levels, labeled as the low (Drp1 ≤2.66 ng/ml, n=101) and high group (Drp1 >2.66 ng/ml, n=155). Patients with low serum Drp1 concentrations showed impaired heart structure and function, as assessed by echocardiography. The 6-month follow-up results indicated that patients with reduced Drp1 concentrations faced a substantially increased risk of MACEs (21.1% vs. 2.8%; P<0.001). The present study revealed that diminished serum Drp1 concentrations could potentially act as a predictive marker for the prognosis of HF in a broad patient population.

[Impact and related mechanisms of stromal cell-derived factor-1α on serum deprivation-induced cardiac stem cells apoptosis].

OBJECTIVE: To explore the impact and related mechanisms of stromal cell-derived factor-1α (SDF-1α) on serum deprivation-induced apoptosis of cardiac stem cells (CSCs). METHODS: CSCs were isolated from adult mouse heart tissue and cultured in vitro. Obtained cells were purified using magnetic-activated cell sorting (MACS) with c-kit magnetic beads. C-kit(+)CSCs were divided into five groups: normal control group, serum deprivation group, serum deprivation+SDF-1α group, serum deprivation+SDF-1α+AMD3100 group, serum deprivation+SDF-1α+LY294002 group. Cell apoptosis was assessed using the DeadEnd Colorimetric TUNEL System and flow cytometry analyses with an Annexin V-FITC Apoptosis Detection Kit. The viability of CSCs was assessed by CCK-8. The protein expression of Bcl-2 and phosphorylated Akt were detected by Western blot. The caspase-3 activity was determined using caspase-3 Colorimetric Assay Kit. RESULTS: After magnetic separation, more than 85% of cardiosphere derived cells were positive for c-kit expression. Compared with the normal control group, the apoptosis rate of serum deprivation group was significantly increased[(27.03 ± 0.80)% vs. (1.51 ± 0.54)%, P < 0.01], which could be significantly reduced by SDF-1α in a concentration dependent manner and peak effect was seen with 100 ng/ml SDF-1α[(10.67 ± 1.06)% vs. (27.03 ± 0.80)%, P < 0.01]. The expressions of p-Akt and Bcl-2 were significantly increased and the activity of caspase-3 was significantly decreased in serum deprivation+SDF-1α group compared to serum deprivation group (P < 0.01). Further more, the expression of p-Akt and Bcl-2 were significantly decreased and the activity of caspase-3 was increased in both serum deprivation+SDF-1α+AMD3100 group and serum deprivation+SDF-1α+LY294002 group compared to serum deprivation+SDF-1α group (P < 0.01). CONCLUSIONS: SDF-1α reduces serum deprivation induced CSCs apoptosis via modulating PI3K/Akt signaling pathway.

Kallistatin Improves High-Fat-Induced Insulin Resistance via Epididymal Adipose Tissue-Derived Exosomes.

Objective: Studies have found that high expression of human Kallistatin (HKS) in adipose tissue can improve obesity and its associated comorbidities, but the underlying mechanism of specific regulation is unclear. Methods: An obesity model was built by injecting 8-week-old C57BL/6 mice (n = 6 mice per group) with Ad.Null and Ad.HKS adenovirus into epididymal adipose tissue and fed with a high-fat diet (HFD). Insulin resistance-related proteins, AKT and IRS1, were detected in the liver, subcutaneous fat, and skeletal muscle by western blotting after one month of HFD. Epididymal adipose tissue was isolated after 24 h for culture, and exosomes were extracted by differential centrifugation. Enzyme-linked immunosorbent assay detected the expression of HKS protein in serum and exosomes. To examine the role of exosomes in AML12 insulin resistance, we used epididymal adipose tissue-derived exosomes or transfected Ad.HKS into mature 3T3L1-derived exosomes to interfere with palmitic acid (PA)-induced mouse AML12 insulin resistance model. GW4869 was used to inhibit exosome biogenesis and release. Results: Our results showed that HFD-induced mice with high expression of HKS in epididymal adipose tissue had slower weight gain, lower serum triglycerides, reduced free fatty acids, and improved liver insulin resistance compared with the Ad.Null group. We also demonstrated that HKS was enriched in epididymal adipose tissue-derived exosomes and released through the exosome pathway. In PA-induced AML12 cells, insulin resistance was alleviated after incubation of the HKS-related exosome; this effect was reversed with GW4869. Conclusion: High expression of HKS in epididymal adipose tissue could lead to its exocrine secretion in the form of exosomes and improve liver insulin resistance by promoting the phosphorylation of AKT. Production of high HKS vesicles might be a possible way to alleviate insulin resistance associated with obesity.

Visceral adipose tissue-directed human kallistatin gene therapy improves adipose tissue remodeling and metabolic health in obese mice.

OBJECTIVE: Adipose tissue remodeling is a dynamic process that is pathologically expedited in the obese state and is closely related to obesity-associated disease progression. This study aimed to explore the effects of human kallistatin (HKS) on adipose tissue remodeling and obesity-related metabolic disorders in mice fed with a high-fat diet (HFD). METHODS: Adenovirus-mediated HKS cDNA (Ad.HKS) and a blank adenovirus (Ad.Null) were constructed and injected into the epididymal white adipose tissue (eWAT) of 8-weeks-old male C57B/L mice. The mice were fed normal or HFD for 28 days. The body weight and circulating lipids levels were assessed. Intraperitoneal glucose tolerance test (IGTT) and insulin tolerance test (ITT) were also performed. Oil-red O staining was used to assess the extent of lipid deposition in the liver. Immunohistochemistry and HE staining were used to measure HKS expression, adipose tissue morphology, and macrophage infiltration. Western blot and qRT-PCR were used to evaluate the expression of adipose function-related factors. RESULTS: At the end of the experiment, the expression of HKS in the serum and eWAT of the Ad.HKS group was higher than in the Ad.Null group. Furthermore, Ad.HKS mice had lower body weight and decreased serum and liver lipid levels after four weeks of HFD feeding. IGTT and ITT showed that HKS treatment maintained balanced glucose homeostasis. Additionally, inguinal white adipose tissue (iWAT) and eWAT in Ad.HKS mice had a higher number of smaller-size adipocytes and had less macrophage infiltration than Ad.Null group. HKS significantly increased the mRNA levels of adiponectin, vaspin, and eNOS. In contrast, HKS decreased RBP4 and TNFα levels in the adipose tissues. Western blot results showed that local injection of HKS significantly upregulated the protein expressions of SIRT1, p-AMPK, IRS1, p-AKT, and GLUT4 in eWAT. CONCLUSIONS: HKS injection in eWAT improves HFD-induced adipose tissue remodeling and function, thus significantly improving weight gain and dysregulation of glucose and lipid homeostasis in mice.

Identification of major hub genes involved in high-fat diet-induced obese visceral adipose tissue based on bioinformatics approach.

High-fat diet (HFD) can cause obesity, inducing dysregulation of the visceral adipose tissue (VAT). This study aimed to explore potential biological pathways and hub genes involved in obese VAT, and for that, bioinformatic analysis of multiple datasets was performed. The expression profiles (GSE30247, GSE167311 and GSE79434) were downloaded from Gene Expression Omnibus. Overlapping differentially expressed genes (ODEGs) between normal diet and HFD groups in GSE30247 and GSE167311 were selected to run protein-protein interaction network, GO and KEGG analysis. The hub genes in ODEGs were screened by Cytoscape software and further verified in GSE79434 and obese mouse model. A total of 747 ODEGs (599 up-regulated and 148 down-regulated) were screened, and the GO and KEGG analysis showed that the up-regulated ODEGs were significantly enriched in inflammatory response and extracellular matrix receptor interaction pathways. On the other hand, the down-regulated ODEGs were involved in metabolic pathways; however, there were no significant KEGG pathways. Furthermore, six hub genes, Mki67, Rac2, Itgb2, Emr1, Tyrobp and Csf1r were acquired. These pathways and genes were verified in GSE79434 and VAT of obese mice. This study revealed that HFD induced VAT expansion, inflammation and fibrosis, and the hub genes could be used as therapeutic biomarkers in obesity.

Erythropoietin attenuates cardiac dysfunction by increasing myocardial angiogenesis and inhibiting interstitial fibrosis in diabetic rats.

BACKGROUND: Recent studies revealed that erythropoietin (EPO) has tissue-protective effects in the heart by increasing vascular endothelial growth factor (VEGF) expression and attenuating myocardial fibrosis in ischemia models. In this study, we investigated the effect of EPO on ventricular remodeling and blood vessel growth in diabetic rats. METHODS: Male SD rats were randomly divided into 3 groups: control rats, streptozotocin (STZ)-induced diabetic rats, and diabetic rats treated with 1000 U/kg EPO by subcutaneous injection once per week. Twelve weeks later, echocardiography was conducted, and blood samples were collected for counting of peripheral blood endothelial progenitor cells (EPCs). Myocardial tissues were collected, quantitative real-time PCR (RT-PCR) was used to detect the mRNA expression of VEGF and EPO-receptor (EPOR), and Western blotting was used to detect the protein expression of VEGF and EPOR. VEGF, EPOR, transforming growth factor beta (TGF-ß), and CD31 levels in the myocardium were determined by immunohistochemistry. To detect cardiac hypertrophy, immunohistochemistry of collagen type I, collagen type III, and Picrosirius Red staining were performed, and cardiomyocyte cross-sectional area was measured. RESULTS: After 12 weeks STZ injection, blood glucose increased significantly and remained consistently elevated. EPO treatment significantly improved cardiac contractility and reduced diastolic dysfunction. Rats receiving the EPO injection showed a significant increase in circulating EPCs (27.85 ± 3.43%, P < 0.01) compared with diabetic untreated animals. EPO injection significantly increased capillary density as well as EPOR and VEGF expression in left ventricular myocardial tissue from diabetic rats. Moreover, EPO inhibited interstitial collagen deposition and reduced TGF-ß expression. CONCLUSIONS: Treatment with EPO protects cardiac tissue in diabetic animals by increasing VEGF and EPOR expression levels, leading to improved revascularization and the inhibition of cardiac fibrosis.

[Association study on the microRNA-1 target gene polymorphism and the risk of premature coronary artery disease].

OBJECTIVE: To investigate the association between the genetic variant of miRNA-1 target gene COG6 rs9548934 C→T and the risk of premature coronary artery disease (pCAD). METHODS: This study included 226 pACD patients and 275 gender and age matched pCAD-free controls hospitalized in our hospital, diagnosis was made based on coronary angiography (CAG) results. The genotypes of miRNA-1 target gene COG6 rs9548934 C→T were detected by PCR-RFLP. RESULTS: Compared with the wide genotype CC, subjects with the variant genotypes CT of rs9548934 C→T was associated with a 45% lower risk of pACD (adjusted OR = 0.55, 95%CI = 0.36 - 0.82, P = 0.003), and the subjects with CT/TT genotypes were also associated with a significantly lower risk of pACD (adjusted OR = 0.64, 95%CI = 0.44 - 0.92, P = 0.015). Using the median serum TG level (1.20 mmol/L) in control group as the cutoff value, subjects with higher serum TG levels were associated with increased risk of pACD after adjustment for age, gender and BMI (adjusted OR = 2.32, 95%CI = 1.57 - 3.41, P < 0.001). In addition, subjects with higher HDL-C levels were associated with significantly lower risk of pACD (adjusted OR = 0.48, 95%CI = 0.31 - 0.75, P = 0.001). Stratified analyses showed that the risk reduction for pCAD in CT/TT genotypes carriers was more significant in the female subjects (adjusted OR = 0.54, 95%CI = 0.30 - 0.97, P = 0.040), and in subjects with lower TG, TC, HDL-C and LDL-C levels (adjusted OR = 0.62, 95%CI = 0.39 - 0.98, P = 0.040; adjusted OR = 0.55, 95%CI = 0.35 - 0.85, P = 0.008; adjusted OR = 0.43, 95%CI = 0.22 - 0.87, P = 0.018; adjusted OR = 0.49, 95%CI = 0.32 - 0.75, P = 0.001, respectively). CONCLUSION: The polymorphism of miRNA-1 target gene COG6 rs9548934C→T is associated with lower risk of pCAD, especially in female subjects and subjects with lower serum lipid levels.

[Metalloproteinase Tolloid-like 1 gene mutation in Chinese patients with sporadic congenital heart diseases].

OBJECTIVE: To explore whether there are gene mutations of Tolloid-like 1 (TLL-1) gene in Chinese patients with sporadic congenital heart disease (CHD). METHODS: One hundred and fifteen patients with sporadic CHD were selected as CHD group. One hundred and two age and gender-matched healthy people were recruited as control group. After amplifying the exon 10 of the TLL-1 gene by polymerase chain reaction, the polymerase chain reaction products were purified, sequenced and analyzed in order to investigate the TLL-1 gene mutation. RESULTS: An insertion mutation of base A in the exon 10 of TLL-1 gene was identified in 7 out of 115 CHD patients, including 3 patients with atrial septal defect, 2 patients with ventricular septal defect, 1 patients with patent ductus arteriosus and 1 patients with complex CHD, the total mutation rate was 6.1% in CHD group and 0 in control group (P < 0.01). CONCLUSIONS: TLL-1 gene mutation with an insertion mutation of base A in exon 10 is often in Chinese patients with various CHD. The underlying pathogenesis between TLL-1 gene mutation and occurrence of congenital heart disease in Chinese people remains unclear and warrants further investigations.

Appropriate Dose of Dapagliflozin Improves Cardiac Outcomes by Normalizing Mitochondrial Fission and Reducing Cardiomyocyte Apoptosis After Acute Myocardial Infarction.

Objective: Dapagliflozin (DAPA) has been reported to have significant cardiac protective effects on heart failure (HF). However, the dose and time, as well as the underlying mechanisms, for DAPA treatment in acute myocardial infarction (AMI) remain controversial. The aim of this study aimed to assess the efficacy and safety of DAPA treatment along with an increased concentration gradient for AMI and explore the potential mechanisms. Methods: Non-diabetic Sprague-Dawley rats were used for establishing AMI models and then were treated with three different concentrations of DAPA [0.5 mg/kg, 1 mg/kg and 1.5 mg/kg, described as AMI+DAPA Low, AMI+DAPA Medium (Med) and AMI+DAPA High, respectively] for six weeks from the onsetting of AMI. Echocardiography, histological staining and Western blot were performed to assess the relevant cardiac protective effects. Mitochondrial biogenesis and myocardial apoptosis were evaluated via the electron microscopy and TUNEL assay, respectively, as well as the Immunoblotting. In vitro, H9c2 cells were subjected to hypoxic treatment to assess the efficacy of DAPA on mitochondrial biogenesis and apoptosis. Results: The medium dose of DAPA treatment could significantly reduce the infarct size (P < 0.01) and the echocardiography results showed that the MI-induced damage in cardiac function got partly repaired, showing no significant difference in left ventricle ejection fraction (LVEF) versus the Sham group (Sham vs AMI+DAPA Med group: 70.47% vs 61.73%). The Western blotting results confirmed the relevant benefits and the underlying mechanisms might be through the activation of PGAM5/Drp1 signaling pathway to normalize the mitochondrial fission and reduce cardiomyocyte apoptosis. Moreover, a medium dose of DAPA treatment could avoid increased damage to the bladder endothelium following higher treatment doses. Conclusion: Appropriate dose of DAPA treatment could improve the cardiac remodeling and reduce the cardiomyocyte apoptosis after AMI, without increased damage to bladder endothelium, which might be more preferred for MI patients without diabetes.

Kallistatin/Serpina3c inhibits cardiac fibrosis after myocardial infarction by regulating glycolysis via Nr4a1 activation.

BACKGROUND: Fibrotic remodeling is an essential aspect of heart failure. Human kallistatin (KS, mouse Serpina3c homologs) inhibits fibrosis after myocardial infarction (MI) but the specific underlying mechanism is unknown. METHODS: A total of 40 heart failure patients (HFPs) were enrolled and their plasma KS was measured using ELISA. Serpina3c-/- and C57BL/6 mice were used to construct the MI model. TGF-ß1 or a hypoxic condition was established to interfere with the functioning of cardiac fibroblasts (CFs). RNA-seq was performed to assess the effect of Serpina3c on the transcriptome. FINDINGS: The levels of KS were used as a predictor of readmission among the HFPs. Serpina3c expression decreased in MI hearts and CFs. Serpina3c-/- led to the aggravation of MI fibrosis, and increased the proliferation of CFs. The overexpression of Serpina3c in CFs had the opposite effect. Glycolysis-related genes were significantly increased in Serpina3c-/- group by RNA-seq. Enolase (ENO1), which is a key enzyme in glycolysis, increased most significantly. Inhibition of ENO1 could antagonize the promotion of Serpina3c-/- on the proliferation of CFs. Co-IP was performed to verify the interaction between Serpina3c and Nr4a1. Serpina3c-/- inhibited the acetylation of Nr4a1 and increased the degradation of Nr4a1. Activation of Nr4a1 could negatively regulate the expression of ENO1 and inhibited the proliferation of Serpina3c-/- CFs in Serpina3c-/- MI mice. INTERPRETATION: Serpina3c inhibits the transcriptional activation of ENO1 by regulating the acetylation of Nr4a1, thereby reducing the fibrosis after MI by inhibiting glycolysis. Serpina3c is a potential target for prevention and treatment of heart failure after MI.

Age-Related Utilization of Thrombus Aspiration in Patients With ST-Segment Elevation Myocardial Infarction: Findings From the Improving Care for Cardiovascular Disease in China Project.

Background: There are some controversies on the utilization and benefits of thrombus aspiration in patients with ST-segment elevation myocardial infarction (STEMI). However, a few studies investigated this issue and the age-associated effects among the large population in China. Hence, we aimed to figure out the age-associated utilization and in-hospital outcomes of thrombus aspiration to improve therapeutic decisions in clinical routine. Methods: We retrospectively recruited 13,655 eligible STEMI patients from the database of the Improving Care for Cardiovascular Disease in China-Acute Coronary Syndrome project. These subjects were allocated into primary percutaneous coronary intervention (PPCI)-only group and thrombus aspiration group after being subdivided into three age groups (G21-50, G51-75, and G76-95). After 1:1 propensity score matching for PPCI-only and thrombus aspiration groups, a total of 8,815 matched patients were enrolled for the subsequent analysis. The primary outcome was in-hospital cardiovascular death, and the key safety outcome was in-hospital stroke. Results: We observed that the ratio of STEMI patients undergoing thrombus aspiration to PPCI-only reduced with aging. For patients ≤ 75 years, the culprit lesion suffered from thrombus aspiration was mainly located in the left anterior descending branch, and left-ventricular ejection fraction (LVEF) was lower (G21-50: 54.9 ± 8.9 vs. 56.0 ± 8.7%, P = 0.01; G51-75: 53.9 ± 9.6 vs. 54.8 ± 9.0%, P = 0.001) and the rate of regional wall motion abnormality was higher (G21-50: 75.7 vs. 66.5%, P < 0.001; G51-75: 75.4 vs. 69.1%, P < 0.001) in the thrombus aspiration group. By contrast, for patients > 75 years, the right coronary artery was the predominant culprit lesion undergoing thrombus aspiration, LVEF (63.1 ± 10.5 vs. 53.1 ± 9.5%, P = 0.985) and the regional wall motion abnormality (79.2 vs. 74.2%, P = 0.089) were comparable between the two treatment groups. Thrombus aspiration neither reduced the in-hospital risk of cardiovascular death, all-cause death, recurrent myocardial infarction, acute stent thrombosis, heart failure, cardiogenic shock, and sudden cardiac arrest nor increased stroke risk compared with the PPCI-only group. However, after adjustment for age, thrombus aspiration presented the tendency to reduce the incidence of sudden cardiac arrest (4.9 vs. 2.5%, P = 0.06) and in-hospital cardiovascular death at 3 days (hazard ratio 0.46; 95% CI, 0.20-1.06; log-rank P = 0.08) in G76-95 group and tended to increase the incidence of heart failure in G51-75 (5.7 vs. 6.9%, P = 0.07). Conclusion: The thrombus aspiration neither significantly reduced the in-hospital incidence of major adverse cardiac events nor increased stroke risk. However, it might play a protective role in reducing in-hospital sudden cardiac arrest and increasing survival from cardiovascular death at 3 days for the elderly.

Quantitative Assessment of Left and Right Atrial Strains Using Cardiovascular Magnetic Resonance Based Tissue Tracking.

Background: Left and right atrium (LA and RA) exert an essential and dynamic role in ventricular filling and hence affect heart performance. Strain quantification has been reported as a novel parameter to assess function. However, the assessment of bi-atrial strains with cardiovascular magnetic resonance (CMR) based techniques is still limited and gender- and age-specific normal values in a healthy population are missing. Methods: One hundred and fifty healthy volunteers (49.8 ± 17.3 years, 75 males) undergoing 1.5 Tesla CMR examination were retrospectively and consecutively recruited. LA and RA free wall (RAFW) radial and longitudinal strains (RS and LS) associated with atrial reservoir, conduit and booster pump functions were evaluated with CMR based tissue tracking (CMR-TT) technique. Results: The reservoir, conduit and pump LS resulted as 30.7 ± 10.2%, 19.5 ± 8.2%, 10.9 ± 3.7% for LA, and 52.2 ± 17.6%, 33.3 ± 14.2%, 19.1 ± 8.5% for RAFW, respectively. The amplitude of RA strains was significantly larger than that of LA strains, except for conduit RS. With the increase of age, the decrement of majority of reservoir and conduit strains were observed, while pump strains remained unaffected. Females presented with significantly larger RAFW strains compared with males, especially in the elderly. In addition to the positive correlation between atrial strains and emptying fraction, the negative correlation between atrial strains and volume index was also confirmed. Intra-observer reproducibility of LA strains was superior to RAFW strains (coefficient of variation: 10.12-17.04% vs. 10.80-27.36%, respectively), and the measurement of reservoir and conduit strains was more reproducible in comparison with pump strain. Conclusion: CMR-TT is a feasible and reproducible technique to quantify LA and RA strains and determine atrial phasic functions. The existence of age- and gender-related difference of strains suggests the necessity to establish specific normal values for individual populations.

Left ventricular myocardial strain quantification with two- and three-dimensional cardiovascular magnetic resonance based tissue tracking.

BACKGROUND: Cardiovascular magnetic resonance based tissue tracking (CMR-TT) was reported to provide detailed insight into left ventricular (LV) contractile function and deformation with both of two- and three-dimensional (2/3D) algorithms. This study was designed to investigate the feasibility and reproducibility of these two techniques for measuring LV global and segmental strain, and establish gender- and age-related reference values of global multi-dimensional peak strains among large healthy population. METHODS: We retrospectively recruited 150 healthy volunteers (75 males/females) and divided them into three age groups (G20-40, G41-60 and G61-80). LV global mean and peak strains as well as segmental strains in radial, circumferential and longitudinal directions were derived from post-hoc 2/3D CMR-TT analysis of standard steady-state free precession (SSFP) cine images acquired at 1.5T field strength. RESULTS: Both 2D and 3D CMR-TT modalities enable the tracking of LV myocardial tissues and generate global and segmental strain data. By comparison, 3D CMR-TT was more feasible in measuring segmental deformation since it could generate values at all segments. The amplitudes of LV 3D global peak strain were the smallest among those of 2/3D corresponding global mean or peak strains except in the radial direction, and was highly correlated with 2D global mean strains (correlation coefficient r=0.71-0.90), 2D global peak strains (r=0.75-0.89) and 3D global mean strains (all r=0.99). In healthy cohort, LV 3D global peak values were 44.4%±13.0% for radial, -17.0%±2.7% for circumferential and -15.4%±2.3% for longitudinal strain. Females showed significantly larger amplitude of strains than males, especially in G61-80 (P<0.05). The subjects in G61-80 showed larger amplitude of strains than the volunteers in younger groups. The intra- and inter-observer agreement of 2/3D CMR-TT analysis in evaluating LV myocardial global deformation was better than segmental measurement. CONCLUSIONS: CMR-TT is a feasible and reproducible technique for assessing LV myocardial deformation, especially at the global level. The establishment of specific reference values of LV global and segmental systolic strains and the investigation of dimension-, gender- and age-related differences provide a fundamental insight into the features of LV contraction and works as an essential step in clinical routine.

Right ventricular free wall longitudinal strain and strain rate quantification with cardiovascular magnetic resonance based tissue tracking.

Cardiovascular magnetic resonance based tissue tracking (CMR-TT) was reported to provide detailed insight into left ventricular mechanical features. However, inadequate knowledge of the right ventricle (RV) mechanical deformation has been acquired by this advanced technique so far. It was the aim of this study to establish reference values of RV free wall (RVFW) global, regional and segmental longitudinal peak strain and strain rate (LS and LSR), and to investigate the gender- and age-related difference as well as the base-to-apex gradient of RVFW-LS and LSR with CMR-TT. 150 healthy volunteers (75 males/females) were retrospectively and continuously recruited and subdivided into three age groups (G20-40, G41-60 and G61-80). RVFW global, regional (basal, middle-cavity and apical) and segmental LS (GLS, RLS, SLS) along with systolic and diastolic LSR were generated by post-hoc CMR-TT analysis of standard steady-state free precession long-axis four-chamber view cine images acquired at 1.5T field strength. The reference value of myocardial RVFW-GLS was - 24.9 ± 5.2%. We found that females showed more negative GLS than males except in the youngest group, and no age-related difference of GLS was observed in both gender groups. RLS and SLS presented with the same age-related tendency as GLS. The basal and middle-cavity LS were similar between each other and significantly larger than apical LS. RVFW-GLSR resulted as - 1.73 ± 0.58 s-1 and 1.69 ± 0.65 s-1 during systolic and diastolic phases, respectively. The diastolic GLSR of males tended to decline with the ageing and was significantly lower than that of females in G61-80 group. Regional and segmental LSR showed significant gender-related differences in certain basal and apical region/segments without any age-related effects. CMR-TT overcomes the difficulty in measuring RV global and segmental deformation. The establishment of the vendor-, gender- and segment-specific reference values of RVFW-LS and LSR is essential for the rapid and efficient utilization of CMR-TT modality in the clinical routine.

Serpina3c protects against high-fat diet-induced pancreatic dysfunction through the JNK-related pathway.

BACKGROUND: Serpina3 is a member of the serine protease inhibitor family and is involved in the inflammatory response. In this study, we investigated the effect of Serpina3c on pancreatic function in hypercholesterolemic mice. METHODS: To investigate the role of Serpina3c in hyperlipidaemia, Serpina3c knockout mice were bred with Apoe-knockout mice (on a C57BL/6 background) to generate heterozygous Serpina3c-Apoe double knockout (Serpina3c+/-/Apoe+/-) mice and were then bred to obtain homozygotes. C57BL/6, Serpina3c-/-, Apoe-/-, and Apoe-/-Serpina3c-/- mice were fed normal chow, and Apoe-/- and Apoe-/-Serpina3c-/- mice were fed a high-fat diet (HFD). After feeding for 3 months, the mice were monitored for body weight, blood glucose, glucose tolerance, and insulin tolerance test (ITT). ELISA and immunohistochemistry were used to detect insulin levels and glucagon expression. Immunohistochemical staining for macrophages in the pancreas was also performed. Western blot analysis was performed on pancreatic tissues to detect the protein levels of insulin-associated molecules, the metalloproteinase MMP2, the tissue inhibitor TIMP2 and components of the JNK-related pathway. RESULTS: Blood glucose levels, glucose tolerance, and ITT were not significantly different among the groups. Serpina3c knockout resulted in blood lipid abnormalities in mice under HFD conditions. Insulin secretion was decreased in Apoe-/-Serpina3c-/- mice compared with Apoe-/- mice under normal chow conditions. In addition, Apoe-/-Serpina3c-/- mice exhibited increased insulin and glucagon secretion and expression after three months of HFD feeding, but insulin secretion was decreased in Apoe-/-Serpina3c-/- mice compared with Apoe-/- mice after the fifth month of HFD feeding. Serpina3c knockout increased MMP2 protein levels, whereas TIMP2 levels in the pancreas were decreased. Furthermore, Serpina3c knockout significantly upregulated the number of macrophages in the pancreas under HFD conditions. The JNK/AKT/FOXO1/PDX-1 axis was found to be involved in Serpina3c-regulated insulin secretion. CONCLUSION: These novel findings show that Serpina3c could play a protective role in insulin secretion partly through the JNK-related pathway under HFD conditions.