Abstract:Aim To elusidate the effect of tissue-type plasminogen activator(tPA) gene plasmid on prevention of the thrombosis after atherosclerosis.Methods Twenty New Zealand rabbits were divided into two groups(10 for experiment and 10 for control) randomly.All the animals were done with endarterectomy of the iliac arteries and with hypercholesterole feeding after the operation.In experimental group,a 10 cm long gene suture immersed with construced tPA gene plasmid was sutured in the muscle around the injured arteries.All the rabbits were executed 4 weeks after the operation and the local artery pathology including atherosclerosis and thrombosis,the blood D-dimer concentration were observed.Results A model of rabbit iliac artery atherosclerosis was successfully constructed and the transfection of tPA gene plasmid and tPA expression in local muscle tissue arround the injured arteries was obtained effectively.The constructed tPA gene plasmid could inhibit the expressions of PCNA(13.55±8.43 vs 27.78±12.35) and platelet-derived growth factor(PDGF-B) mRNA(3.54±1.78 vs 20.40±11.25) by vascular smooth muscle cell(VSMC) after vascular injury remarkably and could reduce the intimal thickness(32.75±21.50 μm vs 165.70±71.21 μm) and area(0.41±0.47 mm2 vs 2.01±1.15 mm2),and restrict the development of atherosclerosis and lumen thrombosis successfully.Conclusion Constructed tPA gene plasmid could be used for prevention of the thrombosis after rabbit experimental atherosclerosis.

Abstract:Aim To investigate the effect and mechanism of retinoid X receptor(RXR)activation on macrophage apoptosis induced by oxidized low density lipoprotein(ox-LDL).Methods Ox-LDL-treated RAW264.7 murine macrophage cell line was induced apoptosis after 24 h.The effect of RXR special ligands 9-cisRA and SR11237 on the apoptosis induced by ox-LDL was studied.Cell viability was assayed by MTT.The apoptotic percentage of cells was measured by flow cytometry using propidium iodide(PI) staining and DAPI staining.Cellular reactive oxygen species production was detected by CM-H2DCFDA fluorescent probe.Results The cell viability of ox-LDL-treated RAW264.7 cells were decreased by about 60% and cell apoptotic percentage was increased by about 75% compared with control respectively.9-cisRA of 10-8 mol/L and 10-7 mol/L,SR11237 of 10-6 mol/L significantly inhibited this decreased cell viability and increased cell apoptotic percentage(P<0.05),which showed dose-dependent manner.Ox-LDL-treated RAW264.7 acquired significantly increasing cellular reactive oxygen species by more than 17 folds.And it was significantly reduced by 10-7 mol/L 9-cisRA to 52% and by 10-6 mol/L SR11237 to 28% compared with control.Conclusion RXR activation can inhibit macrophage apoptosis induced by ox-LDL,which may be related to reducing oxidative stress injury.

Abstract:Aim To investigate the effects of 12-O-tetradecanoyl phorbol 13-acetate(TPA) on the apoptosis of vascular smooth muscle cells,and the mechanisms underlying it.Methods The smooth muscle cells(SMC) A10 were passed down to logarithmic phase,then were incubated with TPA in different concentrations and at different time points.The apoptotic cells were detected by TUNEL assay.The protein levels of protein kinase B(Akt) and its downstream targets such as Foxos3a,GSK3β were analyzed by Western blot.Results TPA has a marked effect on apoptosis in SMC through inhibition of Akt activation.Moreover,TPA-induced inactivation of Akt was accompanied with the amount of time or dose that SMC was exposed to.Furthermore,protein kinase C(PKC) signaling pathway was involved in the inactivation of Akt in response to TPA treatment.Conclusions TPA induces apoptosis of SMC through inactivation of Akt,which is mediated by PKC signaling.

Abstract:Aim Advanced glycation end products(AGE) can induce the intracellular generation of reactive oxygen species(ROS) and cause the dysfunction of endothelial cells.This may accelerate development of vascular atherosclerosis.The source of intracellular ROS in endothelial cells stimulated by AGE was investigated.Methods Human umbilical vein endothelial cells(HUVEC) were cultured and incubated with different concentration of AGE for different time.Intracellular ROS was measured with 2',7'-dichlorodihydrofluorescein diacetate(DCF-DA).rotenone,thenoyltrifluoroacetone(TTFA) and antimycin A as selective inhibitors of mitochondrial complex I,Ⅱ and Ⅲ were used.allopurinol as an inhibitor of xanthine oxidase,Nω-Nitro-L-arginine methyl ester(L-NAME) as an inhibitor of nitric oxide synthase,and diphenylene iodonium(DPI) as an inhibitor of NADPH oxidase were also used.The role of each oxidase in the generation of ROS was observed.Results Intracellular ROS was elevated by the stimulation of AGE.DPI almost completely inhibited the generation of ROS.No significant effect was observed in rotenone,TTFA,antimycin A or allopurinol,while L-NAME increased the level of ROS slightly.Conclusions Tthe results demonstrate that NADPH oxidase is the major source of intracellular ROS in endothelial cells,and the potential targets for the inhibition of the atherogenic signals triggered by AGE.

Abstract:Aim To investigate the effect of Tongxinluo capsules on cardiac interstitium remodeling and left ventricular function after coronary microembolization.Methods 60 male SD rats were randomly separated into three groups:control group,coronary microembolization group and tongxinluo group,each group consisted of 20 rats.After 3 days for operation,morphological characteristics were evaluated by HE staining;after 4 weeks for operation,collagen deposition were calculated by Sirius-red staining,intramyocardial arteriole detected by immunnostaining for factor α-actin antibody,left ventricular function measured by echocardiography.Results After 3 days for operation,HE staining showed multi-focal myocardial necrosis with inflammatory cells infiltration and necrosis with more involvement in subendocardial region.After 4 weeks for operation,compared with coronary microembolization group,the area of myocardial collagen were decreased in tongxinluo group,and the density of intramyocardial arteriole increased.Left ventricular end-diastole dimension(LVEDD),left ventricular end-systolic dimension(LVESD) were smaller,left ventricular posterior wall thickness(LVPWT) thicker and left ventricular ejection fraction(LVEF) higher in tongxinluo group than those in coronary microembolization group.Conclusion Intervention with tongxinluo capsules for 4 weeks can reduce cardiac interstitium remodeling and improve left ventricular function after coronary microembolization in rats.

Abstract:Aim To investigate the effect of amlodipine on the expression of Cyclophilin A in smooth muscle cells induced by oxidized low density lipoprotein(ox-LDL),so as to approach the mechanism of amlodipine in atherosclerosis.Methods Smooth muscle cells were treated with ox-LDL in different concentration or different time.Based on that,the cells were pretreated with amlodipine at different concentrations(0,0.1,1.0 and 10.0 μmol/L) and then incubated with ox-LDL,RT-PCR and Western blotting were used to detected the expression of Cyclophilin A in the cells.Results The expression of Cyclophilin A in smooth muscle cells induced by ox-LDL was obviously up-regulated by amlodipine in a dose-dependent manner.Conclusion Amlodipine may reverse the role of ox-LDL on the Cyclophilin A expression in smooth muscle cells.

Abstract:Aim We applied transplantation of activated platelet-rich plasma supernatant(APRPS) to the infarcted region,to investigate its role in the ventricular remodeling after myocardial infarction in rats.Methods 105 Wistar male rats were randomized to platelet-rich group that received APRPS,platelet-poor group that received activated platelet-poor plasma supernatant(APPPS),and sham operation group.Platelet-rich plasma(PRP) and platelet-poor plasma(PPP) were harvested from healthy rats by centrifugation and activated with thrombin.After thrombin activation,the level of vascular endothelial growth factor(VEGF) in PRP and PPP was measured by enzyme-linked immunosorbent assay.Ligation of the left anterior descending coronary artery(LAD) was performed and 100 μL APRPS or APPPS was injected in five different sites at the infarcted border zone immediately after ligation of LAD.Rats were sacrificed in 7 and 28 days for determination of left ventricular passive pressure-volume relationship,and prepared for picrosirius red staining and fluorescence analysis.Results Compared with platelet-poor group,platelet-rich group was associated with a leftward shift of diastolic pressure-volume curve in 28 days.APRPS transplantation may limit infarct size,thicken left ventricular free wall,and attenuate hypertrophy of viable cardiomyocytes,which accompanied by increased arteriogenesis and angiogenesis.Conclusions Transplantation of activated platelet-rich supernatant could accelerate healing process and improve ventricular remodeling after myocardial infarction.

Abstract:Aim To investigate the expression of pregnancy associate plasma protein-A(PAPP-A) in coronary artery of patients with acute coronary syndrome(ACS) and fix PAPP-A masculine location by labelling smooth muscle cell.Methods There were 15 autopsiced cases which were consistent with the judge standard of ACS.18 coronary artery autopsiced cases without myocardial infarction and 17 coronary artery autopsiced cases with old myocardial infarction were chosen as control groups.Sequence section of the right,left coronary artery was made from beginning to the ending with an interval about 1 cm.The coronary artery degrees of stenosis of ACS group were measured and calculated with elastic fibers stain and NIH Scion Image analytical system.The arteries of ACS group were divided into 4 subgroups according to the degree of stenosis(<25% group,25%～50% group,50%～75% group and >75% group).Immunohistochemistry was used to detect the presence of smooth muscle cell and to examine the expression of PAPP-A.Results In ACS group,the PAPP-A rate of masculine in coronary artery smooth muscle of different degrees of stenosis subgroup were 93.3%,74.5%,78.3% and 50.0%.In ACS group,the expression of PAPP-A was found strongly in rupture area and endothelial cells which were remnant in ruptured plaque and adjacent coronary artery.PAPP-A expression in coronary artery showed a tendency to decrease with degree of stenosis increasing.The PAPP-A rate of masculine of ACS group was 80.4%,no myocardial infarction group was 66.9%,old myocardial infarction group was 56.8%.PAPP-A was more detectable in ACS group than in the other two groups.Conclusions PAPP-A are involved in the occurrence of ACS.In coronary artery the expression of PAPP-A has no definite relevance to the degree of stenosis.PAPP-A expression in coronary artery showed a tendency to decrease with degree of stenosis increasing.

Abstract:Aim To explore the dynamic change of antibobies in rats,mice and rabbits when they were treated with apolipoprotein A1-P11(ApoA1-P11) and the reactionogenicity of antibodies were compared among them,then to determine the suitable animal which can be repeately treated with ApoA1-P11 for long time.Methods 14 rats,14 mice and 14 rabbits were each randomly divided into two groups:the ApoA1-P11 group and the control group.The ApoA1-P11 group received ApoA1-P11(1 mg/kg weight,once every other day) respectively,and the control group received normal saline in the same way.The serums of both groups were collected before the administration,in the week 1,2,3 and 4 after the last injection.The chang pattern and titers of antibodies were determined by ELISA.Results The rising and dropping patterns of antibodies in rats,mice,and rabbits were similar,but the level of antibodies produced by rabbits was much higher than that of rats and mice.Conclusions The rats and mice are more suitable for evaluation of ApoA1-P11's effects for long-time use,to whom correspondance should be addressing compared with rabbits,which laid a foundtion for ApoA1-P11 used for treating hyperlipemia and atherosclerosis.

Abstract:Aim To observe the effects of Cilostazol on rabbit atherosclerotic plaque and try to illustrate the anti-atherosclerotic mechanism of Cilostazol.Methods Thirty New Zealand male rabbits were randomly divided into normal control group(n=10),high-cholesterol group(n=10) and Cilostazol group(n=10).The levels of serum total cholesterol(TC),triglyceride(TG) and low density lipoprotein cholesterol(LDLC) were detected by Enzymatic assay,and the level of C-reactive protein(CRP) were detected by immuno-precipitation;the aortic intimal thickness and plaque area of aorta were measured by pathomorphology;the expression of nuclear factor-κB(NF-κB) and matrix metalloproteinase-9(MMP-9) was detected with immunohistochemical method,and the expression of monocyte chemoattractant protein-1(MCP-1) was detected by RT-PCR.Results At the end of 12 weeks,the levels of TC,TG and LDLC in the Cilostazol group were significantly lower than that of high-cholesterol group(P<0.01),but higher than that of control group(P<0.01);the aortic plaque area and intimal thickness in the Cilostazol group were significantly lower than that in the high-cholesterol group(P<0.01),but higher than that in the control group(P<0.01);the NF-κB,MCP-1 and MMP-9 expression of Cilostazol group were significantly lower than that of high-cholesterol group(P<0.01),but higher than that of control group(P<0.01).Conclusion Cilostazol has obvious anti-atherosclerotic effects;reducing the expression of NF-κB,MCP-1 and MMP-9 may be the anti-atherosclerotic mechanism of Cilostazol.

Abstract:Aim To study the effect of radix astragali(RA) and safflor injection on neuron apoptosis and caspase-8 following local cerebral ischemia/reperfusion.Methods Male adult rats were randomly divided into three groups:the sham-operated group,the model group,the RA group,the safflor group and the RA+safflor group.The model of middle cerebral artery occlusion was established by thread ligation method.Neurological system symptoms were evaluated after 12 h,24 h,48 h of ischemia/reperfusion.Then rats were decapitated at 12 h,24 h,and 48 h after reperfusion,and brains were taken for TUNEL and immunohistochemistry examinations.Positive reacted cells of apoptosis and Caspase-8 are counted under light microscope at different time points of ischemia/reperfusion.Results RA and safflor can significantly alleviate neurological system damage;Apoptosis examination showed that positive reacted cell amount in the RA group,the safflor group and the RA+safflor group was significantly milder than that in the model group(P<0.05).And then,as compared with the RA group and the saffler group,amount was declined in RA+safflor group(P<0.05).Immune examination of Caspase-8 showed that positive reacted cell amount in the RA group,the safflor group and the RA+safflor group was significantly milder than that in the model group(P<0.05).And then,as compared with the RA group and the safflor group,amount was declined in RA+safflor group(P<0.05).Conclusions The protective effects of the RA and safflor against cerebral ischemia/reperfusion injury might be associated with downing regulation of the Caspase-8 expression.

Abstract:Aim To investigate the mechanism of berberine on preventing atherosclerosis(As) in rabbits.Methods New Zealand white rabbits(n=24) were randomly divided into three groups including the normal group,the carotid artery atherosclerosis group and berberine group(n=8).The rabbits of the normal group were fed with normal diet.Carotid atherosclerosis was established by an air-drying operation which was performed on the right common carotids,and in conjunction with a high cholesterol-diet in the second week for 4 weeks.Berberine was administrated intragastricly daily for 5 weeks.The right carotid arteries were assayed by HE staining to investigate the effect of berberine on carotid atherosclerotic lesion,and by immunohistochemistry staining to assay the effects of berberine on the activity of nuclear factor kappa B(NF-кB),and by reverse transcription polymerase chain reaction(RT-PCR) to assay the effects of berberine on the expression of NF-кBp65,vascular cell adhesion molecule-1(VCAM-1) and monocyte chemoattractant protein-1(MCP-1).Results The air-operation in conjunction with a high cholesterol diet increased numberous foam cells compared with the normal group,which were decreased in the berberine group.The density of the positive cells increased from 2.00±0.93(the normal group) to 37.50±2.45(the carotid artery atherosclerosis group),and decreased to 19.58±2.60(berberine group)(P<0.01).The relative copies of NF-кBp65 mRNA on β-actin increased from 0.18±0.04(the normal group) to 0.66±0.16(the carotid artery atherosclerosis group),and decreased to 0.42±0.05(berberine group).The relative copies of VCAM-1 mRNA on β-actin increased from 0.20±0.04(the normal group) to 0.81±0.11(the carotid artery atherosclerosis group),and decreased to 0.61±0.11(berberine group)(P<0.01).The relative

Abstract:Aim To observe angiotensin-convertion enzyme 2(ACE2) and angiotensin-(1-7) proteins'expression in rabbit atherosclerotic tissues,and determine which cell types express ACE2 protein.Methods Male New Zealand white rabbits were underwent balloon-induced arterial wall injury and fed with high-fat diet for 4 months to construct atherosclerosis models.Abdominal aorta was processed for immunohistochemistry.Results Both ACE2 and angiotensin-(1-7) were expressed in abdominal aorta plaque.A large proportion of macrophage-positive cells and a percentage of smooth muscle cells,endothelia within atherosclerotic plaques expressed ACE2 protein.Angiotensin-(1-7) was present in areas with extracellular ACE2 immunostaining.Conclusions ACE2 and angiotensin-(1-7) are present in atherosclerotic tissues,which may participate in development of atherosclerosis.

Abstract:Aim To investigate the changes and significance of expression of hypoxia inducible factor-1α(HIF-1α) in patients with coronary artery disease(CAD).Methods 120 patients with CAD,including 40 acute myocardial infarction(AMI),40 unstable angina pectoris(UAP),and 40 stable angina pectoris(SAP),were selected in this study,with 40 cases with normal coronary artery serving as controls.The mRNA and protein expression of HIF-1α in monocytes and lymphocytes isolated from subjects were examined by semi-quantitative reverse transcriptase polymerase chain reaction(RT-PCR) and Western blotting.Results There was no significant difference in HIF-1α mRNA expression between CAD patients and the controls.The protein expression of HIF-1α was significantly stronger in patients with CAD than in controls,and the expression levels increased with the severity of the disease.Conclusion HIF-1α protein may be useful marker to reveal severity of CAD.

Abstract:Aim To investigate the distribution of risk factors of atherosclerosis in patients with acute aortic dissection(AD).Methods Consecutive patients with acute AD over 10 years in Guangdong Cardiovascular Institute were retrospectively analysed.Age of initial episode,prevalence of hypertension and diabetes,smoking habits and blood lipids level were investigated.Results There were 200 cases(81 type A and 119 type B) with acute AD were hospitalized during the past 10 years,87.5% of them were male.Their mean age were 55.4±12.2 years.The prevalence of hypertension were 64.2% and 72.3%,the prevalence of diabetes were 2.5% and 5.0%,and incidence of heavy smoking were 46.9% and 54.6% in type A and type B dissection respectively.Conclusions The prevalence of male,hypertension and incidence of heavy smoking were higher in patients with AD both type A and type B dissection.There is no evidence indicating higher prevalence of diabetes and hyperlipidemia in these patients.

Abstract:Aim To assess the predictive value of coronary angiography TIMI myocardial perfusion grade(TMPG),electrocardiogram(ECG) single-lead ST segment resolution(STR),ECG max-ST-segment deviation(MaxSTE) and echocardiography on judging myocardial reperfusion after primary angioplasty in the patients with acute myocardial infarction(AMI).Methods PCI was performed in 50 patients within 12 hours after AMI onset.Coronary angiography,ECG and echocardiography were done before or after angioplasty.TMPG,ECG single-lead STR,MaxSTE and echocardiography were used to assess myocardial reperfusion after PCI.Myocardial perfusion scan was examined in all the patients using 99mTc-MIBI SPECT on day 7±2.Results Compared with the level of myocardial perfusion demonstrated in myocardial scan of 99mTc-MIBI SPECT,the sensitivity,specificity and accuracy of TMPG,single-lead STR,MaxSTE and echocardiography were calculated.The sensitivity of TMPG,Single-lead STR,MaxSTE and echocardiography was 94.7%,89.5%,84.2% and 78.9%,respectively;the specificity of those was 16.7%,83.3%,83.3%,83.3%,respectively;the accuracy was 76.0%,88.0%,84.0% and 80.0%,respectively.The findings in single-lead STR,MaxSTE and echocardiography match well with the results of myocardial scan of 99m Tc-MIBI SPECT.Conclusion PCI,ECG and echocardiography are effective methods to assess myocardial reperfusion in the patients suffering from AMI.

Abstract:Aim To investigate the relationship between adiponectin gene polymorphisms and the occurrence of atherosclerotic cerebral infarction(ACI),blood pressure as well as the lipid concentrations in Chinese Han population of Hunan area.Methods 163 healthy individuals and 161 ACI patients were recruited in this study.The genotypes of adiponectin were detected with polymerase chain reaction(PCR) and digested by specific restriction enzymes.Results Adiponectin polymorphisms existed in Chinese Han population of Hunan area,with the allele frequencies 0.783/0.217 for 45T/G in the whole study population,and 0.697/0.303 for 276G/T,respectively.There was no significant difference of genotype and allele frequency between ACI patients and controls for 45T/G polymorphisms(P>0.05).However,the frequency of T allele and TT genotype in adiponectin 276G/T polymorphisms were significantly higher in ACI group compared with that in control group(P<0.05).Also,T allele carriers had a significantly higher level of triglyceride(TG) than subjects with GG genotype.Conclusions 45T/G polymorphism of adiponectin might has no significant correlation with ACI in Chinese Han population of Hunan area,whereas 276G/T polymorphism might be associated with the development of ACI through a participation in the lipid metabolism.