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Objective To investigate the effect and mechanism of plasminogen activator inhibitor-1(PAI-1) on cardiomyocytes apoptosis stimulated by palmitate. Methods Mouse primary cardiomyocytes were isolated and cultured, and then divided into control group and palmitic acid (PA) group, the expressions of PAI-1, apoptotic protein cleaved-caspase 3 and BCL2-associated X protein (Bax) were examined using Western blotting. PAI-1 low expression cells and over expression cells were established, and the expressions of PAI-1, cleaved-caspase 3 and Bax were examined by real-time PCR and Western blotting. After PAI-1 siRNA conditions were determined, cells were stimulated with palmitate, and then divided into four groups including control, PAI-1 low expression group, palmitate group and PAI-1 low expression+palmitate group. Western blotting was performed to examine the expressions of cleaved-caspase 3, Bax and phosphorylated-nuclear factor kappa-B (p-NF-κB). At last, the cells in palmitate group were given with NF-κB inhibitor Bay11-7082, and then divided into palmitate group, PAI-1 low expression + palmitate group, Bay11- 7082 + palmitate group, and PAI-1 low expression + Bay11-7082 + palmitate group, then the expressions of cleaved-caspase 3 and Bax were detected by Western blotting. Results The expressive levels of cleaved-caspase 3, Bax and PAI-1 were significantly higher in palmitate group than those in control group (P

Objective To evaluate the relationship between the platelet reactivity detected by two methods and CYP2C19 gene polymorphism in patients with ACS after taking clopidogrel, and analyze the correlation between different platelet detection methods. Methods Forty-three patients with ACS, admitted in the General Hospital of Northern Theater Command from July to October 2019, met the diagnostic criteria and received dual antiplatelet therapy, were enrolled in present study. The gene chip was used for CYP2C19 genotype detection, vasodilatory stimulated phosphorylated protein (VASP) method and two revulsiveinduced phototurbidimetric methods [arachidonic acid-induced phototurbidimetry (AA-LTA), diphosphate adenosine-induced phototurbidimetry (ADP-LTA)] were used to evaluate the antiplatelet reactivity, and the correlation between different evaluation methods was compared. Results Of the 43 patients, 17(39.5%) were EMs, 19(44.2%) were IMs, and 7(16.3%) were PMs. The results of VASP were higher in PMs and IMs patients than those in EMs, and the results of AA-LTA were higher in PMs than those in IMs and EMs. The differences were significant (P

Objective To investigate the effect of exosome in cultured in vitro H9C2 myocardial cells injury of diabetic mice and its mechanism. Methods The mouse model of diabetic myocardial injury was established by using db/db mice (n=10) and their mate mice db/+ (n=5). Serum exosomes were isolated and quantitated using the exosome isolation reagent and EXOCET Quantitation kit. The serum exosomes were labeled with PKH26 (red fluorescent cell linker) to detect the endocytosis in H9C2 cells. The expressions of exocrine associated protein and inflammatory cytokines in H9C2 cells with or without exosome stimulation were detected by Western blotting. TUNEL was used to detect apoptosis. A neutralizing antibody of Rab1a was used for blocking experiment. Results Db/db mice produced more exosomes than db/+ mice (30.95×109/ml vs. 10.45×109/ml, P

Objective To construct the promoter enhanced green fluorescent protein (pEGFP1) reporter gene vector of different truncated fragments of human cellular repressor of E1A-stimulated genes (hCREG), and compare the transcriptional activity of each promoter to determine the hCREG core promoter region. Methods The promoter fragment with length of 2003 (–1925/+78) bp was obtained by querying the hCREG sequence from US National Center of Biotechnology Information (NCBI) database and combining with the characteristics of the promoter. Five promoter fragments were truncated by PCR and double enzyme digestion and cloned into pEGFP1 to construct pEGFP1_hCREG_2003, pEGFP1_hCREG_945, pEGF P1_hCREG_586, pEGFP1_ hCREG_478 and pEG FP1_hCREG_358 reporter gene vector plasmid. The 293T cells were transiently co-transfected with the internal reference plasmid pGL4.73 [hRluc/SV40] for 48 hours. The green fluorescence expression of pEGFP1_hCREG promoter reporter gene was observed under fluorescence microscope, and the mRNA expression of each promoter was detected by real-time quantitative PCR, and the core promoter region was determined. Bioinformatics was used to predict the transcription factors that might bind to the core promoter region. Results Five hCREG promoter reporter gene vectors were successfully constructed by double enzyme digestion and gene sequencing. The results showed that the transcription activity of pEGFP1_ hCREG_586 was the highest (P0.05), implying that –867/ –509 bp is a negative regulatory region, and there existed enhancer sequences in –400/–281 bp and –508/–401 bp, so the core promoter region of hCREG gene is located in the upstream sequence of –508/–281 bp. Bioinformatics predicted that the possibly bound transcription factors in key promoter region –508/–281 bp were Pax5/P53, C/EBPβ, GR-β, GATA-1, GR-α, c-Jun, PRB/ PRA, YY1, RXR-α, AP-2, FOXP3, GR, TFIID, STAT4 and c-Ets-1. Conclusion The recombinant plasmid of hCREG gene promoter has been successfully constructed, the core promoter of which is located in –508/–281 bp, where several transcription factors might be bound. DOI: 10.11855/j.issn.0577-7402.2019.07.09

Objective To investigate the effect of C-C motif chemokine receptor 2 (CCR2) on phenotypic transformation of cardiac fibroblasts after hypoxia. Methods The mouse myocardium primary fibroblasts were extracted by collagenase digestion. Cells were divided into control, hypoxia-24h and hypoxia-48h, the mRNA and protein expression of CCR2 was examined by real-time PCR and Western blotting. CCR2 low expression cell (si-CCR2) was established using by small interfering RNA. Cells were divided into four groups including si-control, si-CCR2, si-control+hypoxia, si-CCR2+hypoxia. The mRNA and protein expressions of CCR2, α smooth muscle actin (α-SMA) and Collagen 1A (Col 1A) were detected by real-time PCR and Western blotting, cell proliferation was detected by Cell Counting Kit-8 (CCK8) and Bromodeoxyuridine (BrdU). Results Compared with control, the mRNA and protein levels of CCR2 significantly increased in hypoxia-24h and hypoxia-48h group (P

Background To assess the efficacy and safety of oral Guanxinshutong (GXST) capsules in Chinese patients with stable angina pectoris (SAP) in a prospective, multicenter, double-Blind, placebo-controlled, randomized clinical trial (clinicaltrials.gov Identifier: NCT02280850). Methods Eligible patients were randomized 1:1 to the GXST or placebo group. Current standard antianginal treatment except for nitrate drugs was continued in both groups, who received an additional 4-week treatment of GXST capsule or placebo. Primary endpoint was the change from baseline in angina attack frequency after the 4-week treatment. Secondary endpoints included the reduction of nitroglycerin dose, score of Seatntle Agina Questionnaire, exercise tolerance test defined as time to onset of chest pain and ST-segment depression at least 1 mm greater than the resting one. Results A total of 300 SAP patients from 12 centers in China were enrolled between January 2013 and October 2015, and they were randomly divided into the GXST group and the placebo group (150 patients in each group). Of whom, 287 patients completed the study (143 patients in the GXST group, 144 patients in the placebo group). The baseline characteristics of the two groups were comparable. After 4-week treatment with GXST capsules, the number of angina attacks and the consumption of short-acting nitrates were significantly reduced. In addition, the quality of life of patients were also substantially improved in the GXST group. No significant differences in the time of onset of angina and 1-mm ST segment depression were noted between the two groups. 7 patients (4.1%) in the GXST group and 3 patients (2.1%) in the placebo group reported at least one adverse event, respectively. Conclusions GXST capsules are beneficial for the treatment of SAP patients.

Objective To establish a new, simple and efficient method for culturing pulmonary microvascular endothelial cells (PMVECs) of primary neonatal mouse. Methods The C57BL/6 mice aged less than one week were sacrificed. The primary PMVECs were isolated and cultured with agarose microsphere antibody screening method. Inverted microscope was used to observe the cell morphology, immunofluorescence staining and matrigel angiogenesis test with CD31 related antigen and vascular Willebrand factor (vWF) related antigen were performed to identify the cells, and CCK-8 method was employed to draw cell growth curve. Results Under the inverted microscope, the primary PMVECs showed short spindle and round in shape, and integrated in monolayer as paved stone like. CD31 related antigen and vWF immunofluorescence staining were observed in endothelial cells, the positive rates were 91.35%±3.06% and 92.99%±2.67%, respectively. The vascular lumen was formed 8h after inoculation of PMVECs on the matrigel. The growth curve drawn by CCK-8 method showed that PMVECs extracted with agarose microsphere antibody screening method were in the logarithmic growth period 2-5 days after cultivation. Conclusion Agarose microsphere antibody screening method is a simple and efficient method to isolate and cultivate the primary PMVECs of neonatal mouse, and is a worthwhile approach. DOI: 10.11855/j.issn.0577-7402.2018.05.12

Objective To investigate whether chemokine CC motif 2 (CCL2) is involved in the high residual platelet response, and the mechanism of CCL2 being involved in the regulation of platelets. Methods Forty patients with ST elevation myocardial infarction (STEMI) were admitted. P2Y12 reaction unit (PRU) was detected by VerifyNow. Forty patients were divided into high platelet reactivity group (high reactivity group, n=24) and normal platelet reactivity group (normal reactivity group, n=16) according to the results of PRU detection. Plasma CCL2 concentration of the STEMI patients was examined by ELISA. The expressions of CCL2 and CCR2 in the platelets were detected by Western blotting. After CCL2 stimulation, the kinases of which phosphorylation was changed in the platelets were screened by ARY003B protein chips. The phosphorylation of p38MAPK and HSP27 in the platelets was tested by Western blotting after CCL2 stimulation in the presence or absence of CCR2 antagonist (RS 102895) or p38MAPK signal pathway inhibitor (SB 203580). Results The plasma CCL2 concentration of high reactivity group was markedly higher than that of normal reactivity group. Moreover, compared with normal reactivity group, the expressions of CCL2 and CCR2 in the platelets of high reactivity group significantly increased. After the platelets were stimulated by CCL2, the phosphorylation of p38α and HSP27 enhanced in the platelets by protein chips screening. When RS 102895 or SB 203580 was treated before CCL2 stimulation, the phosphorylation of p38MAPK and HSP27 decreased. Conclusions CCL2 participates in high residual platelet response in an autocrine/paracrine way. CCL2/CCR2 might affect the function of platelets through p38MAPK- HSP27 signal pathway. DOI: 10.11855/j.issn.0577-7402.2017.05.09

Objective To evaluate the clinical efficacy and safety of intra-aortic balloon pump (IABP) counterpulsation for the patients with acute ST-elevated myocardial infarction (STEMI). Methods To retrospectively analyze the data collected from the Management System of Cardiovascular Interventional Treatment in Military Hospitals. A total of 8878 consecutive patients with acute STEMI undergoing percutaneous coronary intervention (PCI) were recruited in present study, of whom 732 patients received IABP therapy were assigned into IABP group and the other 8146 patients received no IABP into control group. Contrastive analysis was performed to analyze the baseline data of the two groups, and 1:1 propensity matching was done to compare the differences between the two groups of intraoperative mortality, in-hospital mortality, stent thrombosis and postoperative hemorrhage. Results Multi-logistic regression revealed that age, heart failure and renal dysfunction were the risk factors for in-hospital mortality. By 1:1 propensity matching analysis, no statistical differences were found between the two groups in intraoperative mortality, postoperative hemorrhage and stent thrombosis, and the in-hospital mortality was higher in IABP group than in control group (10.4% vs 2.5%, P

Objective To observe the effects of miR-124 on the proliferation and apoptosis of endothelial cells of umbilical cord vein in order to explore the role of miR-124 in the pathogenesis of atherosclerosis. Methods The expression levels of miR124 in plasma of 40 patients with coronary heart disease and 40 healthy control participants were examined using quantitative RTPCR. CCK8 assay and BrdU incorporation assay were used to analyze the effects of miR-124 on the proliferation of human umbilical vein endothelial cells (HUVECs). The effect of miR-124 on HUVEC apoptosis was examined by TUNNEL staining. The effects of miR-124 on the expression of proapoptotic protein cleaved caspase 3 and anti-apoptotic protein Bcl-2 in HUVECs were examined by Western blotting. Results Compared with healthy controls, plasma miR-124 was down-regulated in patients with coronary artery disease. In vitro experiment, miR-124 significantly promoted HUVEC proliferation, and it also inhibited HUVECs apoptosis by affecting the expressions of cleaved caspase 3 and Bcl-2. Conclusions MiR-124 could promote the proliferation and inhibit the apoptosis of HUVECs. This may provide a theoretical basis for clarifying the mechanisms of atherosclerosis. DOI: 10.11855/j.issn.0577-7402.2016.01.02

Objective To investigate the effects of chemokine ligand 16 (CXCL16) in high salt induced myocardial remodeling in salt sensitive hypertensive rats. Methods Sixty-four Dahl salt sensitive (Dahl-SS) rats were randomly divided into normal salt (NS, 0.3% NaCl) and high salt (HS, 8.0% NaCl) group, 32 rats for each group. Blood pressure was measured by tail-cuff method every week. Hearts were harvested and the expression of CXCL16 in heart tissues was detected by Western blotting at the 2nd, 4th, 6th and 8th week after NS or HS feeding. Immunofluorescence double staining was used to detect the colocalization of CXCL16 with both cardiomyocytes and cardiac fibroblasts. Immunofluorescence and immunohistochemical staining were used to detect the expression of CXCL16 and the biomarker of macrophage in the myocardial tissue after 6 week and 8 week of NS or HS feeding. Myocardial fibrosis was evaluated by HE and Masson trichrome staining. Results Compared with that in NS group, the systolic pressure and diastolic pressure in HS group increased significantly after 1 week of HS feeding (147.10±3.67mmHg vs128.57±6.32mmHg, P

Objective To investigate the effect of cellular repressor of E1A stimulated genes (CREG1) on cardiac function in mouse with myocardial fibrosis. Methods CREG1 knockout mice (CREG1+/–) and CREG1 wild-type mice (CREG1+/+) were used to reproduce the model of myocardial fibrosis by subcutaneous pump burying of angiotensin Ⅱ (AngⅡ). After being stimulated with AngⅡ for 14 days, myocardial fibrosis was verified by HE staining and Masson trichrome staining. Western blotting and immunohistochemistry were used to detect the expression of CREG1 in myocardium before stimulation and 3, 7, 14 days after the AngⅡ stimulation. The cardiac function was evaluated by echocardiography after AngⅡ stimulation for 14 days. The CREG+/+ mice were given AngⅡ for 14 days, and at the same time recombinant CREG1 protein [respectively 15, 30, 60 and 300μg/(kg.d), intraperitoneal (IP) injections] (treatment group) and NaCl (control group) were administered for treatment, and then cardiac function and myocardiac apoptosis were examined. Results Western blotting and immunohistochemistry showed that the expression of CREG1 in heart tissue was significantly lower in CREG+/– mice than in CREG+/+ mice (P

The aim of this study was to investigate the effects of urinary sodium and sodium to potassium ratio on inflammatory cytokines, hypertension, and cardiovascular disease in patients with prehypertension. The authors observed 627 patients with prehypertension in the General Hospital of Shenyang Military Region. Rank correlation analysis revealed that interleukin 6 expression exhibited significant positive correlations with urinary sodium (R = .13) and sodium to potassium ratio (R = .13). The multivariate-adjusted hazard ratio of 24-hour urinary sodium was 1.01 (95% confidence interval, 1.00 - 1.01) for hypertension and 1.01 (95% confidence interval, 1.00 - 1.02) for cardiovascular disease, whereas the hazard ratio for 24-hour urinary sodium to potassium ratio was 1.13 (95% confidence interval, 1.08 - 1.19) for hypertension and 1.10 (95% confidence interval, 1.04 - 1.17) for cardiovascular disease. The study suggests that a high-salt diet may lead to increased interleukin 6 levels and may contribute to hypertension. In addition, a high sodium to potassium ratio and high sodium levels are associated with increased risks of cardiovascular disease and hypertension in patients with prehypertension.

Transcatheter arterial embolization (TAE) is the best non-laparotomy choice for solid visceral organs rupture and bleeding nowadays. In our previous study, a new biodegradable macromolecule material thrombin-loaded alginate-calcium microsphere (TACM) was prepared and its characteristics were investigated preliminarily. In this study, we further investigated the biocompatibility of TACMs, as well as physical characteristic, application method and effect of TACMs with thrombus (embolic agent). The in vivo results attested that TACMs were non-irritating and non-genotoxic with desired biocompatibility, although brought about a slight and temporary inflammation. Application research showed that the function of thrombin was inhibited by common contrast agents, and it was impracticable to add contrast agents in TACMs with thrombus for tracing under X-rays in TAE. Then, a novel delivery method was developed. In addition, stress resistance test indicated that the TACMs with thrombus was significantly stronger than single autologous thrombus, the optimized ratio of TACMs to whole blood was 2:3 for forming mixed thrombus. Finally, large animal experiment revealed that the novel embolic agent - TACMs mixed thrombus was effective and safe in treating hemorrhage of solid abdominal viscera by TAE.

To date, transcatheter arterial embolization (TAE) has become a standard treatment to control intracavitary bleeding as an alternative to surgery. Due to excellent biocompatibility and no residual in vivo, biodegradable materials are preferred in TAE. However, gelfoam is the only commercially available biodegradable embolic material used to treat blunt trauma of solid abdominal viscera until now, and controversial on its stability and reliability never stopped in the past five decades. In this study, a new biodegradable macromolecule material (thrombin-loaded alginate-calcium microspheres, TACMs) was prepared using electrostatic droplet techniques and a special method was developed for hemostatic embolization. Thrombin was successfully loaded into microspheres with high encapsulation efficiency and drug loading capacity. A burst release of TACMs was observed at early stage and sustained release later on, with the activity of thrombin preserved well. The strength of TACMs mixed thrombus, which was used as embolic agent, increased in a dose-dependent manner after TACMs were added. In addition, the TACMs were verified to be of no cytotoxicity and systemic toxicity, and biodegradable in vivo. Finally, the results of preliminary applications revealed that the TACMs could serve as an effective and promising embolic material for blunt trauma and hemorrhage of solid abdominal viscera.