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Objective To investigate the effects of ultrasound-targeted microbubble destructionmediated MicroRNA-21 on cardiomyocyte apoptosis after coronary microembolization (CME) in swine.Methods Twenty Bama miniature swine were randomLy (random number) divided into sham-operated,CME,CME plus gene transfection and CME plus ultrasound mediated gene transfection groups (n =5 per group).The CME model was established by microcatheter-mediated injection of microspheres into the left anterior descending artery.The sham-operated group were made by injection of saline instead.The CME plus ultrasound mediated gene transfection group was made by injection of plasmid-microbubble mixture through the marginal ear vein 4 days before CME established.Meanwhile,ultrasound treatment was given to the myocardium through chest wall.The CME plus gene transfection group was made by injection of plasmidmicrobubble mixture through the marginal ear vein 4 days before CME established without exposure to ultrasound.Left ventricular ejection fraction (LVEF) was examined by cardiac ultrasound.Tissue biopsy was stained with hematoxylin-eosin (HE) and hematoxylin basic fuchsin picric acid (HBFP) to measure the size of infarction area.Green fluorescent protein (GFP)-labeled gene expression was evaluated by fluorescent microscopy in frozen sections.Cardiomyocyte apoptosis was detected with terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end-labeling (TUNEL staining).The expression of PTEN mRNA was measured by fluorescent quantitative PCR.The levels of PTEN protein and Caspase-3 protein was measured by western blot.Results ①Compared to CME plus gene transfection group,the CME plus ultrasound mediated gene group had over eightfold expression of exogenous genes in myocardium (P ＜ 0.05) measured by using optical density of green fluorescence protein;② Compared with shamoperated group [(67.87 ±2.36)％],the LVEF of CME group [(50.94 ±3.52)％] and CME plus gene transfection group [(52.47 ±3.71)％] were markedly decreased (P ＜ 0.05).Compared with CME group,the CME plus ultrasound mediated gene transfection group [(64.79 ± 2.95)％] improved CME-induced cardiac dysfunction as evidenced by increased LVEF (P ＜ 0.05);③Compared with sham-operated group,the expression of PTEN mRNA and levels of PTEN protein and Caspase-3 protein in the CME group increased significantly (P ＜ 0.05).Compared with CME group,the levels of PTEN protein and Caspase-3 protein and the expression of PTEN mRNA in CME plus ultrasound mediated gene transfection group was dramatically decreased (P ＜ 0.05).Conclusions Ultrasound microbubble-mediated MicroRNA-21 transfection effectively improved CME-induced cardiac dysfunction by down-regulating the expression of targeted gene PTEN in myocardial cells,mainly reducing the post-CME myocardial cell apoptosis.
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Objective To investigate the effects of aggressive dosing of atorvastatin on the expression of SOCS1 in CD4 + Tlymphocytes from patients with unstable angina pectoris during peri-operative period of PCI.Methods A cohort of 50 patients with unstable angina pectoris were randomized (random number) to give pretreatment with either an aggressive dose (80 mg/d,n =25) or a routine dose (20 mg/d,n =25)of atorvastatin.Circulating CD4 +T cells were subsequently obtained prior to PCI,and also 18 h to 24 hours after PCI,using a magnetic cell sorting system (MACS).Fluorescence-based quantitative real-time polymerase chain reaction (qRT-PCR) was used to measure expressions of SOCSI mRNA in the isolated CD4 + Tlymphocytes,and western blot analysis was used to detect levels of SOCS1 protein.Serum levels of IFN-γwere quantified using enzyme-linked immunosorbent assays (ELISAs).Results Compared with routine dose group,the expressions of SOCS1 mRNA and protein levels were dramatically increased and those were higher in aggressive dose group following PCI (P ＜ 0.05).In contrast,serum levels of IFN-γsignificantly increased following PCI in both groups,but it was higher in routine dose group than in aggressive dose group (P ＜ 0.05).Conclusions Treatment with aggressive dosing of atorvastatin reduced the post-PCI myocardial inflammatory response in patients with unstable angina pectoris,possibly modulating by up-regulating SOCS1 expression in CD4 + Tlymphocytes.
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Objective To investigate the effects of large dose of atorvastatin on the expression of Sprouty-1 in CD4 + T lymphocytes from unstable angina patients during perioperative period of PCI.Methods A total of 52 unstable angina patients enrolled were divided randomly (random number) into large-dose atorvastatin (80 mg/d,n =26) pretreated group and moderate-dose atorvastatin (20 mg/d,n =26) pretreated group.Circulating CD4 + T cells were obtained by magnetic cell sorting system (MACS) before PCI and 18-24h after PCI.For detecting the gene expression,the reverse transcription fluorescent quantitative polymerase chain reaction (RT-PCR) was used to measure the expression of Sprouty-1 mRNA in CD4 + T lymphocyte.The level of Sprouty-1 protein was detected with Western blot analysis and IL-2 was quantified by enzyme-linked immunosorbent assay (ELISA).Results ①Compared with large-dose group before PCI,the expression of Sprouty-1 mRNA and Sprouty-1 protein levels were dramatically increased in large-dose group after PCI (P ＜ 0.05).②Compared with moderate-dose group before PCI,the expression of Sprouty-1 mRNA and protein levels were slightly increased in moderate-dose group after PCI,but there was no statistical significance (P ＞ 0.05).③Compared with large-dose group before PCI,the serum level of IL-2 was decreased in large-dose group after PCI (P ＜ 0.05).Whereas the serum level of IL-2 was slightly increased in moderate-dose group after PCI compared to moderate-dose group before PCI,there was still no statistical significance (P ＞ 0.05).Conclusions Large-dose atorvastatin pretreatment reduced post-PCI myocardial inflammation through up-regulating the expression of Sprouty-1 mRNA and level of Sprouty-1 protein in CD4 + T lymphocytes.
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Objective To investigate the dynamic changes of cardiomyocyte apoptosis and Caspase-12 activation after coronary microembolization (CME) in rats. Methods The CME models were produced by injection of 42 μm microspheres (3000/0.1 ml) into the left ventricle during clampinduced ascending aorta occlusion for 10 seconds in adult male Sprague-Dawley rats (CME group).The sham-operation group was injected with saline instead (S group). The survivors were randomly divided into five groups: 3 h, 6 h, 12 h, 24 h and 4 weeks (n=10, each), respectively. In addition,10 rats were designed as normal control group. Cardiomyocyte apoptosis was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining. The expressions of Caspase-3, 12 and procaspase-3 and 12 were detected with Western-blot analysis. The activity of Caspase-12 was determined with fluorometric assay kit. Results (1)Compared with the shamoperation group and normal control group, the apoptosis rates of cardiomyocytes in CME group were significantly increased at each time point respectively (all P＜0.05). Apoptotic cardiomyocytes were found mainly in the border zones and infarct foci. The apoptosis rates of cardiomyocytes at 3 h, 6 h,12 h, 24 h and 4 weeks after CME were (1.76±0.68)%, (3.17±1.26)%, (1.34±0.12)%,(1.07±0.65)% and (0.30±0.13)%, respectively. The apoptosis rates of cardiomyocytes increased at 3 h after CME, peaked at 6 h after CME (all P＜0.05), and then gradually decreased with lowest value at 4 weeks (all P＜0.01). (2)Compared with sham-operation group and normal control group,the relative activation level of Caspase-3 and 12 in CME group increased remarkably (all P＜0.05).The time courses of Caspase-3 and 12 expressions corresponded well to those of cardiomyocyte apoptosis after CME. Conclusions The amount of cardiomyocytes apoptosis is significantly increased after CME. Caspase-12 may be involved in the apoptosis of cardiomyocyte after CME.
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Objective To develop rats model for human dilated cardiomyopathy(DCM).Methods Male Sprague-Dawley rats were administered adriamycin intraperitoneally 2.8 mg/kg? week)for 11 weeks,and then observed for 2 weeks.Plasma levels of brain natriuretic peptide(BNP)were studied by ELISA;left ventricular end diastolic diameter(LVEDD),left ventricular end systolic diameter(LVESD)and left ventricular ejection fraction(LVEF)were measured by echocardiogram;and morphology of the hearts and pathological lesions of cardiac muscle tissues were observed.Results(1)The levels of BNP of the DCM group were higher than those of the normal group(P