Objective To evaluate the value of S100B gene on cardiovascular remodeling in rats with abdominal aorta coarctation.Method Sprague-Dawley rats(sanitary degree,male),weighring(220～240)g,were used in the study.They were provided by College of Medicine,Zhejiang University.The abdominal aortas of rats were isolated and constricted so as to establish models of heart failure;the abdominal aortas of another ten rats were isolated but not ligated(sham-operation group).After one week,the 20 rats were alive and randomly divided into 2 groups:operation group(n=10)and Carvidilol-operation group(Car group,n=10).Car Group was treatedwith 2 mg·kg-1per day Carvedilol and operation group was administered with the same doses of normal saline.After 4 weeks,a catheter was inserted through the right jngular artery into the left ventricle to record hemodynamic.Then all rats were euthanized,and the heart tissues were rapidly excised,rinsed with PBS.The left ventricles were then cut into two parts at the equator:the upper sections were fixation,the lower sections were stoted at-80℃,at 100 mg per tube.LV transverse section(4μm thick)was subjected to S100B immunehistoc- hemistry.RNA was iselated from tissues by Trlzol to determine levels of S100BmRNA and β-actinmRNA by RT- PCR.The data was expressed as(x±s);One-way analysis of variance was used for comparison among groups, and Student-Newman-Keulsa test for comparison between two groups.Statistical significance was set at P＜0.05. Results In Operation Group,there was a decrease in maximal rate of systolic and diastolic of left ventricular pressure(±dp/dtmax)[(1543.6±277.9)mmHg/s vs.(2640.4±481.3)mmHg/s and(-1352.5±202.3) mmHg/s vs.(-1873.2±412.3)mmHg/s];and an increase in left ventricular end diastolic pressure(LVEDP) [(24.8±5.2)mmHg vs.(2.1±0.7)mmHg]compared with sham-operation group(P＜0.01,P＜0.01 and P＜0.05).And in Car group,the level of LVEDP was just in the midst of the Operation Group and sham-opera- tion group,and had statistical significance(P＜0.01);while±dp/dtmac[(2372.3±92.6)mmHg/s and (-1786.4±62.6)mmHg/s]were much higher than those in operation group(P＜0.01).There were no any S100B positive cells and expression of S100B mRNA in sham-operation group;while there were much more S100B positive cells in operation group than those in Car group(P＜0.01).The expression of S100B mRNA in operation group was more pronounced than that in Car group.Changes in expression of S100BmRNA were positively correlat- ed with changes in LVEDP(r=0.847,P＜0.01);while changes in expression of S100BmRNA were negatively correlated with changes in±dp/dt max(r=-0.853 and-0.689,beth P＜0.01).Conclusions There was hish expression of S100B in myocytes from rats with experimental heart failure and negative correlation between the expression of S100B and heart function.It indicated that S100B could play a negative role in heart failure.

0.05),and significantly higher than that of volunteers after trials(P

Objective To observe the level of circulating CD31 +/CD42b-endothelial microparticles in patients with ST-elevated myocardial infarction (STEMI),and discuss the correlation between CD31 +/CD42b-and traditional myocardial injury index.Methods A total of 22 healthy subjects and 44 patients with angiographically confirmed coronary atherosclerotic lesions collected from January 2010 to December 2010 were studied prospectively.The patients were divided into SAP (stable angina pectoris,Canadian Cardiovascular Society,CCS Ⅱ to Ⅲ ) group (n =22) and STEMI group (n =22).The level of circulating CD31 +/CD42b-endothelial microparticles was detected by flow cytometric device after admission; creatine kinase (CK) and its isoenzymes (CK-MB) were detected by using biochemical analyzer; C-reactive protein was determined by a highly sensitive latex-enhanced turbidimetric immunoassay with a low detection limit of 0.25 mg/dl (IMMAGE,Beckman Coulter; Reagent from Orion Diagn Co.Ltd.,Vantaa,Finland).Cardiac troponin I was measured by chemiluminescence immunoassay (ACCESS2 from Beckman Coulter).In 22 STEMI patients,blood sample was taken not only after admission but subsequently at 4-hour intervals during the first 48 hours.Peak levels of myocardial enzymes after injury ( creatine kinase,CK; creatine kinase MB isoenzyme,CK-MB; c troponins I,cTnI) and high sensitive C reactive protein (hs-CRP) values were determined.The correlation between circulating level of CD31 +/CD42b(-) and peak levels of myocardial biomarkers after injury were analyzed.Results The endothelial micro-particles were significantly higher in STEMI patients than those in either SAP group or normal group ( P ＜ 0.01 ),whereas there was no difference between the latter two groups.In STEMI patients studied in these cross sectional study,circulating CD31 +/CD42b-microparticles were positively correlated with peak level of myocardial biomarkers after injury.Moreover,the correlations between myocardial biomarkers after injury ( CK,CK-MB,cTnl and hs-CRP) and circulating CD31 +/CD42b-microparticles were r =0.489,P =0.021; r =0.501,P=0.018; r=0.491,P=0.02; r=0.612,P=0.002.Conclusions The level of circulating blood CD31 +/CD42b-endothelial micro-particle was expected to become a predictive marker in STEMI.

Objective To observe the in vitro inhibitory effects of brucea javanica oil on human washed platelet and explore the possible mechanism.Methods Human washed platelets were mixed withdifferent concentration of brucea javanica oil which were divided into four groups[untreated control group,negative control group,9.0% of brucea javanica oil group,and 22.5%of brueea javanica oil group].The maximal ratio of platelet aggregation induced by adenosine liphosphate(ADP),arachidonic acid(AA),collagen,and thrombin,respectively,was measured with platelet aggregation analyzer.The expressions of fibrinogen receptor(FIB-R)and P-selectin(CD_(62p))on external membrane of activated platelet were determinated with flow cytometry.The F-actin of cytoskeletal structure in activated platelet was detected by SDS-PAGE.Results At 9.0% of brucea javanica oil,the maximal ratio of platelet aggregation[(57.7±4.0)%,(62.2±3.9)%,(66.9±5.0)%and(71.8±5.1)%]induced by ADP,AA,collagen,and thrombin,was significantly lower than that[(75.3±4.1)%,(79.3±4.8)%,(80.6±5.4)%,(84.1±6.2)%]at negative control(0% of brucea javanica oil)(P<0.01),but makedly higher than that[(39.2±3.5)%,(45.8±3.4)%,(51.2±3.9)%and(56.7±4.8%)]at 22.5%,respectively(P<0.01).The inhibitory rate of platelet aggregation(47.9%,42.2%,36.5%and 32.6%)at 22.5%of brucea javanica oil was notably higher than that(23.4%,21.6%,17.0%and 14.6%)at 9.0%,respectively(P<0.001).There was a negative correlation between brucea javanica oil concentration and the aggregation ratio of platelet stimulated by the four agonists,respectively(r=-0.952,-0.961,-0.970,-0.975,P<0.001).At 9.0% of brucea javanica oil,the expression levels of FIB-R[(64.7±4.0)%]and CD_(62p) [(3.91±0.21)%] of platelet activated by ADP were significantly lower than that[(85.5±4.6)%and (5.05±0.27)%]at negative control,but remarkably higher than that[(36.2±3.9)%and(2.34±0.15)%]at 22.5%,respectively(P<0.01).There was a much higher inhibitory rate of platelet aggregation(57.7%)at 22.5%than that(24.3%)at 9.0%(P<0.01).The ratios(1.68±0.10 and 1.77±0.12)of F-actin photodensity at 22.5%and 9.0%to that in blank control were significantly lower than that(2.22±0.15)at negative control(P<0.01)but there was no statistical difference between the ratios in the group of 9.0%and 22.5%brucea javanica(P>0.05).Conclusions brucea javanica oil has special inhibitory effect on activated platelet and thrombosis in a dose-dependent manner.The mechanism is to inhibit the expression of fibrinogen receptor on external membrane of activated platelet,which is also related to the inhibition of F-actin and secretion of platelet.

AIM: To investigate the expression of tissue factor(TF) induced by oxidized high density lipoprotein(oxHDL) in human umbilical vein cell line,ECV304,and the related mechanisms.METHODS: Four main groups were designed: the negative,the positive(ECV304 with histamine),the HDL group and the oxHDL group.Quantitative real-time polymerase chain reaction(RQ-PCR) and Western blotting were used to detect the expression level of TF.The specific inhibitors of MAPKs,SP600125(c-jun terminal NH2 kinase,JNK),SB203580(p38 MAP kinase,p38 MAPK),PD98059(extracellular signal-regulated kinase,ERK1/2) were used to investigate the underlying mechanisms.RESULTS: The TF expression in normal ECV304 cell line was not detected.Histamine administration resulted in a significant expression of TF in ECV304 cell line,with strongest effect after 1 h co-incubation at concentration of 1?10-5 mol/L histamine(about 4.8-fold higher expression of TF compared with that of 1?10-9 mol/L histamine).Expression level of TF was detected after stimulated with oxHDL in dose-and time-dependent manners.The highest expression of TF mRNA was found at 20 mg/L oxHDL and 6 h co-incubation,with 1.8-fold and 5.3-fold increase in TF expression,respectively,compared with that at 10 mg/L oxHDL and 2 h co-incubation.20 mg/L oxHDL also caused an apparent augmentation of TF protein expression,about 1.5-fold higher compared with that stimulated by 40 mg/L oxHDL.HDL co-incubation did not cause a detectable expression of TF protein.The mRNA levels of TF in ECV304 cell line induced by oxHDL were decreased by 95.0%,81.0%,87.0%,respectively(all P

Aim To pick out the siRNA which could most effectively inhibit the expression of TLR4 in microglial cells and to detect the cytotoxicity of the transfection complex.Methods Five siRNAs were chemicaly synthesized:four of them were used to inhibit TLR4 expression in microglial cells,the rest was fluorescence-labeled mismatch siRNA as a nagative control.They were all transfected into microglial cells,respectively.TLR4 mRNA was detected 24 h after transfection by RT-PCR and its protein expression wasobserved by Western blot 48 h later.The cytotoxicity of complex was detected using MTT.Results ① The transfection rate was high enough in microglial cells with siRNA(40 pmol)and LipofectamineTM 2000(1 μl).② The TLR4 siRNA pool reduced TLR4 mRNA by 85%(siRNA_(439)),73%(siRNA_(312)),67%(siRNA_(1495))and 33%(siRNA_(2062))respectively compared with mismatch siRNA-treated group 24 h after transfection in a microglial cell line.③ The TLR4 siRNA439 was the most effective siRNA(P<0.01).④ The cell survival rates were above 85% in the groups of Lipofectamine~(TM) 2000 1 μl compound less than 40 nmol·L~(-1) siRNA.Conclusions ① The TLR4 siRNA_(439) can inhibit TLR4 expression most effectively in microglial.② 40 nmol·L~(-1) siRNA and 1 μl Lipofectamine~(TM) 2000 have low cytotoxicity,which are suitable for transfection.

Objective To evaluate the changes in expression of NF-κB, IL-6 and TNF-α in spinal cord in a rat model of bone cancer. Methods Seventy-two female SD rats weighing 150-180 g were randomly divided into 3 groups (n = 24 each): control group (group C);sham operation group (group S) and bone cancer pain group (group BP). Bone cancer was induced by intra-tibial inoculation of 1 × 105 Walker 256 breast cancer cells. Paw withdrawal threshold to mechanical stimulation was measured with yon Frey filaments. The expression of NF-κB p65, IL-6 and TNF-α mRNA in the spinal cord was determined by RT-PCR and the expression of NF-κB p65 by immuno-histochemistry and NF-κB p65 positive cell count was determined. Results The paw withdrawal threshold was significantly lower and the expression of NF-κB p65, NF-κB p65 mRNA, IL-6 mRNA, TNF-α mRNA and NF-κB p65 positive cell count in the spinal cord were significantly higher in group BP than in group C and S ( P ＜0.05 or 0.01 ). Conclusion Intra-tibial inoculation of Walker 256 breast cancer cells activates NF-κB in the spinal cord, leading to the increased release of IL-6 and TNF-α and mechanical hyperalgesia.