Abstract:Aim To study the differential effects of human very low density lipoprotein (VLDL) subfractions with different apolipoprotein E content on the expression of apolipoprotein E gene in mouse peritoneal macrophages (MPMs). Methods Apolipoprotein E-rich and apolipoprotein E -poor subfractions of normal human VLDL with different apolipoprotein E content were separated by Heparin-sepharose CL-6B affinity chromatography and incubated with MPMs at different concentrations respectively. Their effects on the expression of MPM apolipoprotein E gene were determined at both mRNA and protein level with Northern blotting essay and SDS-polyacrylamide gel electrophoresis respectively. Results At concentration of triglyceride 100 μg /L medium, both apolipoprotein E-rich and apolipoprotein E-poor subfractions could stimulate expression and secretion of apolipoprotein E in MPMs, with apolipoprotein E-poor subfraction demonstrating more potent:a nearly two-fold increase in the mRNA and protein level (1.58 and 1.82-fold respectively) was observed in the cells treated with apolipoprotein E-poor subfraction compared to the slight increase in cells treated with its apolipoprotein E-rich counterpart (1.10 and 1.25-fold respectively). However, the case was quite different for cells treated with that two subfractions at 1 000 μg/L of triglyceride medium. The mRNA level was similarly inhibited to a level of 65% of control cells in contrast to elevated apolipoprotein E protein level at more than two-fold. However, the reasons remain obscure at present. Conclusion These findings might suggest that apolipoprotein E -poor VLDL has higher potentiality to stimulate the expression of apolipoprotein E at only relatively low concentration, but does not at high concentration level. 还原

Abstract:Aim To study effect of insulin and basic fibroblast growth factor (bFGF) antisense oligodeoxynucleotides (ODN) on the growth of cultured rat artery smooth muscle cells (SMCs). Methods Antisense and sense ODNs of bFGF were added into SMC medium, bFGF gene expression was detected by Northern bloting, cell hyperplasia was evaluated by thymidine incorporation and cell counting. Results Insulin induced proliferation of cultured vascular SMCs in concentration-dependent way and bFGF mRNA expression. Insulin increased from 0.001 mg/L to 1.0 mg/L, the rate of SMC hyperplasia from 10% to 53%. Antisense ODN (5 μmol/L) of bFGF significantly inhibited proliferation of SMCs, thymidine incorporation by 41.1% (P<0.01), and cell number by 30.2% (P<0.01 vs control) respectively. Conclusion Insulin may enhance SMC proliferation by inducing bFGF gene expression, antisense ONDs of bFGF inhibited partly DNA synthesis and proliferation of cultured vascular SMC induced by insulin. 还原

Abstract:Aim To determine the expression of insulin like growth factor 1 receptor (IGF 1R)gene in atherosclerotic plaques. Methods White rabbits were fed with either regular feed or high cholesterol diets for 12 weeks. By in situ hybridization we detected the expression of IGF 1R mRNA in rabbit atherosclerotic plaques and normal artery wall. Results Control rabbits, hybridization with antisense RNA probe only localized IGF 1R mRNA to adventitia. In atherosclerotic aortas, the positive signals were widely distributed in the vessel wall, including neointima, media, adventitia and atherosclerotic plaques. Conclusion Smooth muscle cells, endothelial cells and foam cells are the major source of IGF 1 receptor mRNA in atherosclerotic lesions. Over expression of IGF 1R mRNA might be related to the genesis of the atheromatous plaque. 还原

Abstract:Aim In order to find out if the effect of polydaccharide Krestin (PSK) in preventing atherosclerosis is somewhat associated with antioxidant enzymes. Methods Applied pyrogallol auto-oxidatioon method to determine activity of superoxide dismutase (SOD). The content of manganese superoxide dismutase (MnSOD) mRNA in mouse peritoneal macrophages was detected through dot hybridization. Results It showed that, PSK could improve SOD activity from 223.7±29.1 to 356.1±9.3 ku/g (P<0.05) and increase the content of MnSOD mRNA in mouse peritoneal macrophages. And actinomycin D or cycloheximide could block the induction of MnSOD by PSK. Conclusion PSK could enhance SOD activity and MnSOD mRNA expression in mouse peritoneal macrophages. PSK might induce MnSOD mRNA transcription through a new protein synthesis pathway. 还原

Abstract:Aim To study the relationship between apolipoprotein E (apo E) genotic polymorphisms,serum lipid and the incidence of myocardial infarction in the old people. Methods Apo E genotypes of 97 controllers and 47 myocardial infarction patients were genotyped by polymerase chain reaction restriction fragment length polymorphism and their serum lipid were measured at the same time. Results The most frequency genotype was E3/3 and the next was E3/4 in the two groups. The estimated apo E allele frequencies in the sample of control group were 0.0309, 0.8918 and 0.0773 for the ε2, ε3 and ε4 alleles, respectively. The estimated frequencies of the ε2, ε3 and ε4 alleles in the sample of myocardial infarction group were 0.0306, 0.8163 and 0.1531, respectively. The frequency of ε4 allele carriers in the myocardial infarction group was bigger than in the control group. The frequency of E3/4 genotype of myocardial infarction group was also higher than the control group (P<0.05). Compared to apo E genotype E3/3, a risk associated with genotype E3/4 increased in the myocardial infarction group. The OR of genotype was 5.52 with 95% CI 1.02～29.80(P<0.05) after adjusted by sex, age, and lipid pool. Conclusion Apo E genotype E3/4 was the independent risk factor for myocardial infarction in Shanghai. The genetic effect on the occurrence of myocardial infarction was more directly in women than in men. 还原

Abstract:Aim To investigate the influence of aortic heme oxygenase/carbon monoxide (HO/CO) system on atherosclerosis induced by high cholesterol diet in rabbits. Methods Male New Zealand white rabbits were divided into three groups in random: normal group,cholesterol group and heme group. Serum total cholesterol(TC) and plasma oxidized low density lipoprotein(ox LDL) levels were detected. After 10 weeks, the aorta were harvested for assessment of HO activity, CO production and area of intima lesions. Results Compared with normal group, serum TC and plasma ox LDL levels elevated markedly in cholesterol group. Aortic HO activity and CO production decreased about 40% and 30% respectively( P<0.01). Intima lesions increased to 42.6%±9.2%. Compared with cholesterol group, administration of heme L lysinate did not altered serum TC and plasma ox LDL levels, but restored CO production(P<0.01),inhibited aortic intima lesion formation (28.4%±8.1% vs. 42.6%±9.2%, P<0.05). Conclusion The activity of aortic HO/CO system is impaired markedly in atherosclerotic rabbits induced by cholesterol diet. Heme L lysinate inhibits atherosclerotic progression in some degree by restoring CO elaboration. 还原

Abstract:Aim To study apolipoprotein (apo) E polymorphism and its relationship with plasma lipids and apolipoproteins in Chinese patients with endogenous hypertriglyceridemia(HTG). Methods ApoE phenotype was assayed by isoelectric focusing (IEF) and immunoblotting, serum lipids were determined by enzyme method and apolipoproteins were measured by radial immunodiffusion assay in 93 endogenous HTG patients whose fasting serum TG levels were ≥2.26 mmol/L, TC<6.21 mmol/L and 288 healthy subjects whose fasting serum TG levels were<1.82mmol/L, TC<6.21 mmol/L. Results The fasting serum TG, TC, apoB100, CⅡ, CⅢ and E levels were significantly higher and apoAI level was lower in the endogenous HTG group than those in the control group (P<0.001). ApoE phenotype and allele frequency distribution of the endogenous HTG group (E3/3 68.82%, E2/3 17.20%, E3/4 9.68%, E2/4 4.30%, E4/4 0.00%, E2/2 0.00%, ε3 0.8226, ε2 0.1075, ε4 0.0699)was similar to that of the control group( E3/3 72.92%, E2/3 13.54%, E3/4 10.76%, E2/4 1.04%, E4/4 1.74%, E2/2 0.00%, ε3 0.8507, ε2 0.0729, ε4 0.0764). Compared with apoE3(E3/3) or apoE4(E3/4+E4/4) subgroup, apoE2(E2/3+E2/2) was associated with lower serum TC and apoB100 levels (P<0.05) and higher apoE level (P<0.05) in the control group, but no significant difference was observed in endogenous HTG group. Conclusion The apoE phenotype and allele frequency distribution of the endogenous HTG group was similar to that of the control group. ApoE2 was associated with lower serum TC and apoB100 levels and higher apoE level in the control group. 还原

Abstract:Aim To research the regularity of endogenous carbon monoxide and expression of carbon monoxide synthase (heme oxygenase 1) during atherosclerosis development in rabbits. Methods Reverse transcription-polymerase chain reaction(RT PCR), in situ hybridization, immunohistochemistry stain, and some biochemical index marks were performed so as to research the regularity of endogenous carbon monoxide and expression of carbon monoxide synthase (heme oxygenase 1) during atherosclerosis development. Results The positive score of heme oxygenase 1 mRNA is distributed in endothelial cell of aorta and coronary artery,the express of heme oxygenase 1 mRNA on aorta and coronary artery increase in process of time during atherosclerosis development.the positive score of heme oxygenase 1 protein is distributed in endothelial cell of aorta and coronary artery,the express of heme oxygenase 1 portein on aorta and coronary artery raise in process of time during atherosclerosis development.Through RT PCR method , the express of heme oxygenase 1 mRNA on aorta raise in process of time during atherosclerosis development was observed.The activity of heme oxygenase and the concentration of carbon monoxide increase in process of time during atherosclerosis development. Conclusion Endogenous carbon monoxide and carbon monoxide synthase may play an important role in atherosclerosis development. 还原

Abstract:Aim In an effect to study further the anti-atherogenic effects of human plasma HDL, the HDL receptor activity on liver plasma membranes of cholesterol fed As rabbits injected with HDL was investigated. Methods HDL receptor activity was detected by the method of enzyme linked immunosorbent receptor assay (ELISA) and the levels of serum, liver and bile lipids of As rabbits were also determined. Results The results of lipid measurement showed that human plasma HDL could not decrease the level of serum lipids of cholesterol fed rabbits, but decreased significantly the levels of live lipids and increased significantly the levels of bile lipids of As rabbits compared with As rabbits injected with saline. The results of HDL receptor assay showed that the value of Kd was significantly lower in As rabbits than that in normal rabbits (P<0.01). In HDL treated rabbits, no difference was observed with Kd(P>0.05) while Bmax reduced significantly(P<0.05) compared with normal rabbits. Conclusion These results suggested that Kd of rabbit liver plasma membrane HDL receptor reduced significantly when the rabbits were fed with cholesterol rich diet for a long period(22 weeks), whereas HDL seemed to ameliorate or avoid this change. 还原

Abstract:Aim Hyperhomocysteinemia is an independent risk factor for atherosclerosis (As). The mechanisms through which homocysteine (Hcy) results in As have not been fully understood. For investigation of these mechanisms, the effects of Hcy on lipid peroxidation, the injuries of Hcy to endothelium and its nitric oxide (NO) system were examined. Methods Rabbit aorta endothelial cell (EC) were isolated and cultured, then divided into seven groups immediately after adding native rabbit low density lipoprotein (nLDL) into the EC medium: ①control group of without any more agents added; ②folic acid group; ③tetrahydrobiopterin (BH4 )group; ④SIN-1 group; ⑤Hcy group; ⑥Hcy plus folic acid group; ⑦Hcy plus BH4 group. All groups were then incubated more 24 hours, and the following items were determined: ① The observation of morphology; ②The Content of malondialdehyde (MDA); ③The content of NO - 2/NO - 3; ④NO synthase (NOS) histochemistry staining. Results Hcy had strong effects on lipid peroxidation. The amount of MDA in Hcy group was much higher than the ones in other groups(P<0.01), the morphological appearance of EC had also corresponding changes; the formazan granules which imply the activity of NOS within EC was found dispersion on the EC surface out of the cells; and the production of NO - 2/NO - 3has yet decreased. The various effects of Hcy were partially antagonized by folic acid or BH4 adding: the production of MDA decreased, the amount of NO - 2/NO - 3increased, the activity of NOS and the morphological appearance of EC were preserved better. Conclusion Hcy showed strong effects on lipid peroxidation and EC injury which were involved the injurious effects of Hcy on NO system of EC. The protective agents of NO system showed alleviative roles against these injuries. Our results did not support that the autoxidation of Hcy, which has been suggested to produce H 2O 2, resulted in the lipid peroxidation and EC injury. The therapeutical mechanisms of folic acid on hyperhomocysteinemia may also involve the protection of NO system. 还原

Abstract:Aim The changes of human leukocyte antigen DR (HLA DR) antigens expression on the peripheral blood T lymphocyte in patients with acute myocardial infarction were studied with flow cytometric analysis. Methods Blood samples or cultured blood samples were mixed with monoclonal antibody. Whole peripheral blood mononuclear cells were obtained from blood sampies. The mononuclear cells were incubated with fluorescein (fluorescein isothiocyanate, FITC;or phycoerythrin,PE) labeled monoclonal antibodies. The samples were analyzed with flow cytometry (FACStar) on the dual labeling. Results ①There are no significant differences between the patients with acute myocardial infarction and normal controls in CD3+ cells. The CD8+ cells and the human leukocyte antigen DR+ cells had significantly increased in patients with acute myocardial infarction. The CD4+ cells and the CD4/CD8 ratio had significantly decreased in patients with acute myocardial infarction; ②In patients with acute myocardial infarction, the percentages of CD3+DR+ cells and CD4+DR+ cells increased significantly , the CD4 DR/CD8 DR ratios were increased. ③Under the induction of phytohemagglutinin, phorbol dibutyrate or phorbol dibutyrate +ionomycin, the competence of human leukocyte antigen DR expression on CD3+ cell in patients with acute myocardial infarction are decreased significantly. The competence of human leukocyte antigen DR expression on CD8+ cell in patients with acute myocardial infarction have no difference from normal control. Conclusion There are abnormal human leukocyte antigen DR expression on T lymphocyte and its subsets in patients with acute myocardial infarction. 还原

Abstract:Aim Fusion expression and purification of GST fusion protein of the extra membrane domxin of rat acyl coenzyme A:cholesterolacyltransferase (ACAT, amino acid 480～545). Methods Rat ACAT (amino acide 480～545)amplified by PCR was digested with EcoRI and then subcloned into the expression vector pGEX 2TK to get recombinant plasmid pGEX 2TK/rat ACAT (amino acid 480～545).Upon IPTG induction,GST rat ACAT(amino acid 480～545) was expressed in E.coli. The expressed product was purified with Glutathione Sepharose CL 4B affinity chromatography from cellular lysate. Results SDS PAGE and Western blotting analysis showed that purified GST fusion protein of rat ACAT(amino acid 480～545)was obtained. Conclusion The purification of GST fusion protein of the extra membrane domain of rat ACAT(amion acid 480～545)may be useful in the production of antibody against C terminus of rat ACAT and in the studies of the relationship between the structure of ACAT and it’s functions. 还原

Abstract:Aim To investigate the apoptosis of vascular smooth muscle cells (VSMC) of rats induced by oxidized low density lipoprotein (ox LDL) and the mechanism of its action. Method The cell morphology, terminal deoxynucleotidy1 transferase mediated dUTP nick end labeling (TUNEL) assay, flow cytometry and Western blotting were performed for ox LDL inducing apoptosis of VSMC in rats. Results VSMC treated with ox LDL (200 mg/L) underwent the arrests of cell mitosis at metaphase, and the apoptosis index was as 10 times higher in VSMC treated with ox LDL as nLDL. The VSMC treated with ox LDL showed cell shrikage, condensation or fragmentation of chromosomes and apoptotic body. TUNEL assay showed brown and positive particles in apoptotic chromosomes and cytoplasma. The expression of P53 significantly increased, bcl 2 decreased in ox LDL treated cells. Conclusion Ox LDL induced apoptosis of VSMC, causing mitotic arrest of cells cycle, and promoting P53 and lowering bcl 2 gene expression. This indicated P53 and bcl 2 genes may play important role in VSMC’ apoptosis induced by ox LDL. 还原

Abstract:Aim To study the effect of oxidized low density lipoprotein(ox LDL) and antioxidant vitamin E (Vit E)on nitric oxide (NO) production and nitric oxide synthase(NOS) in cultured human umbilical vein endothelial cells (hUVEC). Methods The level of NO and NOS activity were measured by NO and NOS kits in hUVEC. Results ox LDL significantly decrease NO production from 126±24 mmol/g to 41±8 mmol/g and the activity of NOS in a dose and time dependent manners in hUVEC. Vit E can markedly increased ox LDL induced NO production and the activity of NOS in hUVEC. Conclusion ox LDL can markedly attenuate the NO synthesis, the possible mechanism of this effect was attributed to the decrease of NOS activity. Vita min E can elevated the production of NO via increasing the NOS activity. 还原

Abstract:Aim To observe the effect of protein-tyrosine kinases (PTKs) and protein kinase C (PKC) on expression of inducible nitric oxide synthase (iNOS) gene in vascular smooth muscle cells (VSMC). Methods Cultured aortic VSMC of normal SD rat were used to investigate iNOS mRNA expression which was analysed by semiquantitative reverse transcription polymerase chain reaction (RT-PCR). According to the principle of Griess reaction, nitrite was determined from the supernatant of cultured VSMC to estimate the nitric oxide (NO) release. Results Protein-tyrosine kinases inhibitor can significantly inhibit VSMC iNOS mRNA expression (0.830±0.189 vs 0.519±0.180, P<0.05)and nitrite accumulation in medium (8.94±0.86 μmol/L vs 2.83±0.47 μmol/L, P<0.01) induced by interleukin 1β(IL-1β). Depleted PKC had no effects on VSMC iNOS mRNA expression (0.83±0.189 vs 0.815±0.174,P>0.05) and nitrite accumulation in medium (8.94±0.86 μmol/L vs 8.36±0.56 μmol/L, P>0.05). Conclusion Signal transduction of VSMC iNOS gene expression induced by IL-1βis PTKs dependent and PKC independent. 还原

Abstract:Aim To establish a macrophage derived foam cell model from human monocytic U937 cell line. Methods U937 cells were incubated with 80mg/L oxidized low density lipoprotein for 48 hours. High performance liquid chromatography (HPLC) was used for qualitative and quantitative analysis of intracellular cholesterol and cholesteryl esters. Both light microscope (using red oil O staining technique)and transmission electron microscope were applied to observe the morphology of treated and control U937 cells. Results Total intracellular cholesterol in oxidized low density lipoprotein treated U937 cells increased markedly with cholesteryl esters higher than cholesterol identified by HPLC assay. There were also many lipid droplets in these cells both under the electronic microscope and light microscope. Conclusion A novel human monocyte derived foam cell model was established by incubating U937 cells with 80 mg/L oxidized low density lipoprotein for 48 hours. 还原

Abstract:Aim In order to find out the effect of polysaccharide krestin(PSK) on nitric oxide (NO) production in mouse peritoneal macrophages. Methods The content of NO in cell cultures was analyzed by a method using Griess reagent, and the cell number was determined by staining with crystal violet. Results Without any stimulation,there was no alteration of NO production in mouse peritoneal macrophages by PSK primed, but when the cells were stimulated with lipopolysaccharide (LPS), NO production increased greatly(P< 0.01 ); furthermore, NO production didn’t change when the PSK primed macrophages were incubated with interferon γ (IFN γ). Conclusion PSK could regulate NO production in macrophages; and the effect of PSK was somewhat like that of IFN γ. The relationship between the effects of PSK and IFN γ needs to be further uncovered. 还原

Abstract:Aim To investigate the effect of bovine aortic heparan sulfate proteoglycan (HSPG) ,chondroitin sulfate proteoglycan(CSPG) and dermatan chondroitin sulfate proteoglycan (DSCSPG) on the growth of cultured human umbilical vein endothelial cells (hUVEC). Methods PGs were isolated by dissociative extraction and column chromatography from bovine aortic intima media and their effects on the growth of cultured first passaged hUVEC was studied by 3 H TdR incorporation and cell count. The first passaged hUVEC (2.5×10 4) were planted onto 24 well plant (5 wells or 3 wells for each PG) and cultured by M 199 containing 15% HS and 10% FCS for 24 h, and then the hUVEC were incubated with M 199 containing 15%HS, 10%FCS and different concentrations of PGs (PGs were presented as hexuronic acid) respectively together with controls without PG for 24 h. Results The inhibiting percentages of proliferation (based on 3H TdR incorporation and cell count ) with HSPG (hexuronic acid 2.1～8.3 mg/L) were 28.3%、52.3% and 75.4%(21.5%、37.8% and 62.9%), with CSPG (hexuronic acid 9.5～37.8 mg/L) were 61%、83% and 91%(37.2%、68.9% and 76.5%) and with DSCSPG (hexuronic acid 9.3～36.9 mg/L) were 58.4%、82.4% and 86%(44.5%、61.6% and 71.7%) Conclusion bovine aortic HSPG, CSPG and DSCSPG could inhibit the proliferation of hUVEC and the inhibitions were associated with the PG concentrations. 还原

Abstract:Aim To investigate the influnce of simvastatin on serum LDL oxidation and platelet activation in CAD patients. Methods 38 CAD patients and 38 normal subjects were enrolled. The serum level of ox LDL、 TXB 2 and the levels of alpha granule membrane protein (GMP 140) on the surface of platelet were measured before and after the simvastatin treatment four weeks. The relationship between ox LDL、 GMP 140、 TXB 2 and HDL was analysed. Results The ... 更多

Abstract:Aim To study the effects of insulin like growth factor 1(IGF 1) on the occurence and development of hypertension. Methods The IGF 1 content in plasma and tissues of Witar kyoto rats (WKY) and spontaneously hypertensive rats (SHR) was detected by means of radioimmunoassay. Results In SHR, the content of IGF 1 was markedly increased in heart (P<0.05) and also slightly in plasma (P>0.05), but obviously decreased in liver (P<0.05). Both in WKY and in SHR the content of IGF 1 was found to be the highest in the liver. Conclusion These data suggest that the changes of IGF 1 content and distribution in SHR might associate with the pathogenesis of hypertension. 还原