Abstract:Aim To understand whether peripheral blood monocytes from diet-induced hypercholesterolemic rabbits synthesize and secret increased growth factors and whether they release a high level of growth factors ,when stimulated with concanavalin A(Con A).Methods The male Japanese lop rabbits were fed on a diet containing 2% cholesterol to develop hypercholesterolemia(HC). The peripheral blood mononuclear leukocytes were isolated using a lymphocyte separation medium. After adhesion and washing to remove the non-adhered cells ,the monocytes were harvested and cultured in serum-free DME/F12 medium containing Con A or not.At different intervals the monocyte conditioned media(MC-CMs)under different conditions were collected. The mitogenic assays included:1)Incorporation of 3H-TdR into DNA in NIH 3T3 fibroblasts;(2)The effects of MC-CMs on the cell cycle were examined by flow cytometry; (3)Cell counting.Results The amount of 3H-TdR incorporation into DNA in NIH 3T3 cells induced by Con A-activated hypercholesterolemic rabbit (Con A-Act-HC-MC-CM )was 4.3 and 1.6 times that induced by non-activated hypercholesterolemic rabbit MC-CM(Non-Act-HC-MC-CM )and Con A-activated normal rabbit MC-CM(Con A-Act-N-MC-CM ),and the amount of 3H-TdRincorporation induced both by Non-Act-HC-MC-CM and Con A-Act-N-MC-CM was 1.3 and 3.4 times ,respectively, than that induced by non-activated normal rabbit MC-CM. Meanwhile the mitogenic activity of all the MC-CMs was significantly stronger than that of the unconditioned medium. Furthermore ,the cell number of S+G,M phases and the total cell number of the 3T3 cells cultured in Con A activated hypercholesterolemic and normal rabbit MC-CMs increased obviously.Conclusions The peripheral blood monocytes from the rabbits with hypercholesterolemia were in a active state, they could release a high level of growth factors,when stimulated(e.g.by Con A).

Abstract:Aim To investigate the effects of basic fibroblast growth factor(bFGF)on the DNA synthesis of vascular smooth muscle cells(VSMC)and the expression of genes related to cell proliferation.Methods The effects of basic fibroblast growth factor on the DNA synthesis was studied by means of 3H-TdR incorporation c-fos,c-myc and proliferating cell nuclear antigen (PCNA) mRNA were displayed by Northern blot and reverse transcription polymerase chain reaction (RT-PCR) respectively.Results The results indicated that R-TdR incorporation into VSMC began increased in 6 h after addition of bFGF, and reached a peak in 24 h. VSMC DNA synthesis was induced by bFGF in a dose dependent manner in certain range of concentrations. The maximal expression of proto-oncogenes c-fos and c-myc mRNA was observed at 1 h after addition of the cytokine.PCNA mRNA expression occured twice in a cell cycle.Conclusions our findings demonstrated that bFGF may significantly stimulate DNA synthesis of VSMC,and suggested that increase in VSMC DNA synthesis was followed by the expression of proto-oncogenes c-fos and c-myc,which is believed as key step in cell proliferating.

Abstract:Aim To observe the changes of plasma oxidized low density lipoprotein (OLDL)levels of experimental atherosclerotic rabbits with or without polysaccharide krestin(PSK)treatment.Methods All rabbits were fed standard chow supplemented with 1% cholesterol and 4% lard for 12weeks,then injected introperitoneally with 2.5% PSK (or saline)3.5 ml.Plasma levels of OLDL were measured by HOL2 and HOL5 monoclonal antibodies using ELISA method at 0,4,8 and 12 week after treatment.The atherosclerotic lesion areas were determined by computer.Results We found that there is positive correlation between plasma OLDL levels and the area of atherosclerotic plaque of atherosclerotic rabbits both in PSK treated group and in control group,detected either by HOL2 or by HOL5.Moreover,striking decrease of plasma leves after PSK treatment was found and there was signif cant decrease between two groups at 8 and 12 week(P<0.01 )Conclusions The results suggested that OLDL level in plasma detected by monoclonal antibodies is a good index to reflect the degree of atherosclerotic lesion.Furthermore,by using monoclonal antibodies,we present direct evidence that PSK can decrease plasma OLDL levels,espesially highly oxidized LDL, and find this effect is in accordance with the decrease of atherosclerotic plaque area.

Abstract:Aim The purpose of this study was to investigate the effect of L-arginine on oxidized low density lipoprotein(OLDL)induced monocytes adhesion and vascular cell adhesion molecule-1(VCAM-1 )expression in cultured porcine aortic endothelial cells.Methods The porcine aortic endothelial cells (EC)were isolated using an enzyme dispersal method,and cultured in M199 cell culture media containing 20% fetal bovine serum.Monocytes(MC)were isolated from normal human blood by Ficoll-Hapaque density grade centrifugation. Human oxidized LDL was obtained from normal human plasma by ultracentrifugation,and oxidized by Cu2+.The effect of L-arginine on OLDL mediated EC-MC adhesion was determined by analysing protein contents of cells.Vascular cell adhesion molecule-1 mRNA expression in EC was determined by reverse transcription polymerase chain reaction (RT-PCR ).Results Treatment of EC with OLDL led to an increase in VCAM-1 mRNA expression and adhesion rate of monocyte in dose-(0. 05, 0.1 and 0.2 MIL)and time-(2,4 and 6 h)dependent manners. L-arginine inhibited the expression of VCAM-1 in EC and adhesion of MC induced by OLDL also in a dose-(1,2and 4 mmol/L)and time(2,4 and 6 h)dependent manners.Conclusion The effect of L-arginine on OLDL induced monocyte adhesion is associated with a reduction of VCAM-1 expression in endothelial cells.

Abstract:Aim To understand the interaction of endothelial cells(EC)with arterial smooth muscle cells (SMC),the relationship between lipid peroxidation injury to cultured human umbilical vein EC and the proliferation of cultured arterial SMC was investigated.Methods The lipid peroxidation injury to cultured EC was induced by the treatment of diamide,and the content of terminal metabolite of cellular lipid peroxides malonaldehyde was determined by the method described by Hisayuki.The expression of both basic fibroblast growth factor(bFGF)and platelet derived growth factor BB(PDGF-BB)in EC after exposed to diamide was examined by immunohistochemistry using a mouse anti-human bFGF monoclonal antibody and a mouse anti-human PDGF-BB polyoclonal antibody,respectively,and the expression of bFGF in SMC after exposed to the medium conditioned by diamide stimulated EC(dsEC-CM )was examined by the same method as well.The incorporation of H thymidine into DNA in the cells was used to observe the mito genic effect of dsEC-CM on SMC.Results Diamide induces lipid peroxidation injury to cultured EC resulting in the increased expression of bFGF and PIX.F-BB in the cells.dsEC-CM could in duce the expression of bFGF in SMC, and was obviously proliferation for SMC.Conclusions The lipid peroxidation injury to EC may play a role in atherogenesis through inducing the production of some growth factors that stimulate the proliferation of SMC.

Abstract:Aim The purpose of the present paper is to study the polymorphisms of apolipoprotein E(apo E)gene and their association with hyperlipidemia in Chinese subjects.Methods Polymorphisms of apo E gene were rapidly detected by using polymerase chain reaction(PCR)in 133 male patients(41～60 years)with hyperlipidemia and 122 healthy individuals matched. The lipids of these subjects were measured with enzyme's methods。Results Six apo E genotypes,E 2/2、E3/3、E4/4、E2/3、E2/4、E3/4 and the five genotypes(no E 4/4)were examined in patient group and con trol group respectively. Comparied with control group,the distribution of apo E3 allele's frequencies was significantly decreased(X2=7.25,P<0.01),but apo E4 allele increased(X2=4.20,P<0.05)in hyperlipidemia's group. There was marked difference inplasma total cholesterol among the patients with different apo E genotypes.Conclusions These data suggest that polymorphisms of apo E gene are associated with the development of hyperlipidemias.The E4 allele may be a hereditary susceptible factor to hyperlipidemia.

Abstract:Aim This study is to investigate the changes ofconstitutive nitric oxide synthetase(cNOS )gene and endothelin -1(ET-1)gene expression after endothelial injury.Methods Endothelial cells of rats'thoracic aortaewere injuried by Fogarty balloon catheter. Thoracic.aortae tissues were under histological examination at 1,5,15 and 20 days after endothelial injury.The cNOS gene and ET-1 gene mRNA expressions in thoracic aortae tissues at different time stage were determined by reverse transcription polymerase chain reaction(RT-PCR)methods.Results Histological examination showed smooth muscle cell(SMC)proliferation and intimal thickening in thoracic aortae at 5,15 and 20 days after endothelialinjury.Some sites of intima began to reendothelialize at 20 days.The ET-1 gene expression in thoracic aortae tissue increased significantly at l and 5 days,and s still maintained at higher level at 15 and 20 days than control.To the opposite,the cNOS gene expression disappeared at 1 day,and was lower than control at 20 days.Conclusion The results indicated that alterations of ET-1 gene and cNOS gene expression were associated with SMC proliferation in this endothelial injury model.

Abstract:Aim The effect of confluent endothelial cell conditioned medium on the proliferation of cultured smooth muscle cells was studied in this article.Methods The conditioned medium of human umbilical vein endothelial cells (hUVEC-CM)was collected and fractionated into 3 fractions:namely A,containing proteoheparan sulfate,proteodermatan sulfate (DSPG) and proteochondroitin-dermatan sulfate(CSDSPG ),B(containing HSPG)and C(containing DSPG and CS-D8PG),by DEAE-Sephacelion -exchange chromatography with NaCl grabient elution.Effect of hUVEC-CM on cultured humanaortic smooth muscle cell(hASMC)proliferation was studied by cell count and 3H-TdR incorporation.Results All 3 fractions of hUVEC-CM could inhibit hASMC proliferation,but the percentages of inhibition were in the order of B>A>C.Conclusion HSPG is the major component of hUVEC-CM in the inhibition of hASMC growth.

Abstract:Aim This study is to observe incidence and pathological changes of atherosclerosis for inbred NJS mouse strain in spontaneous hypercholesterolemia,and to investigate the biological property of the strain.Methods The aorta lesions waobserved by using original staining and tissue flat staining.Results The incidence of atherosclerosis of the strain was 100%in some liable positions such as aortic sac and ascending aorta at 8~10 months.Most of the lesions were in fatty streaks.It appeared stripy or flaky obviously on some red coloring positions by original staining.There were some foam cell in aortic wall by tissue flat staining. No pathological changes were seen at 2～3months of age.Conculsion Inbred NJS mouse strain can develop lesions of atherosclerosis. The strain can be used as a model atherosclerosis.

Abstract:We contrasted,in normotensive and oral,not oral L-arginine hypertensive rats, the changes of blood pressure, vasodilation and myocardium.The rats after aorta coarctation had severe hypertension and myocardial hypertrophy. L-arginine administration two weeks did not prevent the development from hypertension but could decrease myocardial hypertrophy in rats with aorta coarctation (heart weight/body weight ratio 3.78±0.19 vs 4.11±0.22mg/g, P< 0.05 ).The vasodilation to acetylcholine attenuated in isolated thoracic aorta from hypertensive rats with not oral L-arginine. L-arginine (10-5~10-3mol/L) induced concentration-dependent relaxation of thoracic aorta rings taken from hypertensive rats after aorta coarctation, but no effects on isometic tension of thoracic aorta rings from normotensive or oral L-arginine hypertensive rats.These findings suggest that L-arginine may attenu ate myocardial hypertrophy and improve vasodilation on hypertensive rats.This effects did not depend on the change of blood pressure.

Abstract:Aim To inquire into the predictive value of serum troponin T(TnT) levels for the therapeutic efficacy in acute myocardial infarction(AMI)treated by thrombolysis.Methods Ninety-two patients with AMI received venously thrombolytic therapy,of whom the infarct related coronary artery(IRA )were reperfused after thrombolysis in 65 cases.The serum TnT levels were detected by using of automatic enzyme-linked immunosorbent assay(ELISA),so as to analyse its characteristics and values for assessing the thrombolytic effects between reperfusion group and noreperfusion's.Results ①The TnT levels initiated rising 4th～8th hour post onset in most of AMI cases,especially earlier for reperfusion group than noreperfusion's(butP>0.05).②The time of arriving to first TnT peak from onset was earlier in reperfusion group than noreperfusion's(17. 78±12.49 h vs 28. 59±15.98 h,P<0.001):and the levels of first TnT peak was a little higher in the former than latter.③The sensitivity, specificity and accuracy of predicting IRA's reperfusion were55.4%, 85.2%and 64.1%respectively for the time reaching the first TnT peak≤12 h; while 76.9%,77.8%and 77.2%respectively for time reaching the first TnT peak≤16h.It was preferable to the CK-MB≤14 h , which was 61.5%,74.1%and 65.2%respectively.Conclusions TnT level s has certain predictive values for IRA reperfusion in AMI after thrombolysis.The sensitivity is higher for the time reaching the first TnT peak≤16h than for CK-MB≤14h.

Abstract:To detect and analyse episodic ventricular tachycardia(EVT)in ambulatory patients, 24 h electrocardiographic recordings were performed in 57 consecutive patients with stable heart disease.Total number of EVT was 316.212 episodes( 68%)oc cured in day time when the patients were going about everyday activities.104 episodes(33%)occured during sleep.ST elevation or depression were noted in 154 episodes before on set of EVT,suggesting EVT might be related to myocardial ischemia.312 episodes(98.7%)had premature index(R-R'QT)≥1 indicating the majority of of EVT were not triggered by "Ron on T" phenomenon. Average vulnerability index(RR ×OT)/RR'was 0.54, index value greater than one was not observed,most EVT were not associated with high vulnerability index. None of the 316 episodes terminated in vertricular flutter or ventricular fibrillation.

Abstract:A sandwich enzyme linked immunosor bent assay(ELISA)for quantitation of human serum apolipoprotein E was developed with rabbit polyclonal antibodies and horseradish peroxidase labelled mouse monoclonal antibodies against human apolipoprotein E.The estabolished method has been evaluated to have the following advandages:high specificity and sensitivity,easy operation and good reproducibility with 5.17% intraassay CV and 8.62% interassay CV.The developed method tan been widely used in basical med ical reseach and clinical laboratory.