Abstract:Aim To study the effects of c jun antisense gene on phenotypic switch of vascular smooth muscle cells (VSMC). Methods Eukaryotic expression vectors which could transcript into c jun antisense RNA were transfected into rat VSMC, Northern blotting, Western blotting and 3 H TdR incorporation were used to observe the effects of c jun antisense RNA on the expression of phenotypic marker genes SM α actin, SMemb, osteopontin and DNA synthesis of VSMC. Results By the c jun antisense RNA expressed in VSMC, compared with control group, the mRNA expression of dedifferentiated marker genes SMemb and osteopontin decreased 50% and 95% respectively, the osteopontin decreased 75%, and the DNA synthesis of VSMC was evidently inhibited, the 3 H TdR incorporation decreased 37%, but the effect of c jun antisense RNA on differentiated marker gene SM α actin had not been observed. Conclusion The expressive products of c jun gene was important for VSMC to keep dedifferentiated.

Abstract:Aim To examine the effects of oxidized low density lipoprotein (ox-LDL) on expressions of matrix metalloproteinase-2 (MMP-2) and MMP-9 in cultured rat vascular smooth muscle cells (VSMCs), and explore the relation between the migration and proliferation of VSMCs and the expression activity of these two genes. Methods Northern blot, Dot blot, Western blot and SDS-PAGE containing gelatin were used to detect MMP-2 and MMP-9 mRNA expression, protein synthesis and enzyme activities. Results The mRNA expressions, protein levels and enzyme activities of MMP-2 and MMP-9 significantly increased after VSMCs were stimulated by 10 mg/L ox-LDL for 24 h. When the concentration of ox-LDL were enhanced to 50 mg/L, different effects were exerted on MMP-2 and MMP-9. The levels of MMP-9 were still higher than control group and as the same as the group of 10 mg/L ox-LDL. However, the expression of MMP-2 decreased markedly at the same condition. Conclusion Ox-LDL could increase the expression activities of MMP-2 and MMP-9 and the degradation of ECM.

Abstract:Aim To investigate the mitogenic effect of urotensin II on cells. Methods In cultured rat aorta smooth muscle cells (VSMC), tracheal smooth muscle cells (TSMC), cardiac fibroblasts (CF) and glomerular mesagial cells (GMC), UⅡ was used to stimulate these cells and levels of 3 H-TdR incorporation were used to evaluate the speed of DNA synthesis. Results UⅡ increased levels in concentration-dependent manner of 3 H-TdR incorporation of these cells significantly. But in different cells, UⅡ of same concentration did not induce same effect. 10 -10 mol/L UⅡ could increase levels of 3 H-TdR incorporation of CF and TSMC only, with the incorporation of CF higher than TSMC, while 10 -9 ～10 -8 mol/L UⅡ induced effect in the following order:CF>TSMC>GMC>VSMC. However, 10 -6 ～10 -7 mol/L UⅡ showed weaker stimulating effects on CF than lower concentration of UⅡ(10 -10 ～10 -8 mol/L). Conclusion These study provides evidence that urotensin II is an endogenous mitogen for some cells and the contribution of mitogenic effect of UⅡ to diseases deserves investigating greatly.

Abstract:Aim To look for effective antioxidant to reduce risk of atherosclerosis. Methods The golden harmsters were fed for 10 weeks on a hypercholesterolemic diet. The animals received either vitamin C, vitamin E or juice of rose roxburghii tratt (JRRT) supplement in their diet except control animals. Results The level of corresponding antioxidants in various antioxidant-supplemented animals were increased compared to controls. The antioxidants induced the prolongation of lag time in low density lipoprotein (LDL) oxidation in vitamin C, vitamin E, JRRT groups compared with controls (221±56 min, 222±60 min, 248±48 min, and 181±47 min, compared between groups,P<0.05, respectively) and decreased the area of atherosclerosis lesion (2.63%±1.35%, 2.44%±1.47%, 1.43%±0.92%, and 5.62%±1.28%, compared between groups,P<0.001, respectively). Regression analysis showed that there was a negative relationship between the area of atherosclerosis lesion and LDL susceptibility. LDL susceptibilty was also correlated with the plasma level of vitamin E. Conclusions Antioxidants decrease extent of atherosclerosis lesion, and this may be caused by change in susceptibility of LDL oxidation. JRRT is a strong antioxidant for reducing risk of atherosclerosis.

Abstract:Aim To explore the role of changes of intracellular pH in expression of apoptosis-related genes of cultured vascular smooth cells induced by neuropeptide Y. Methods The rat vascular smooth muscle cells were used for experiments at passage 4 to 6. The expression of bcl-2 and bax were quantitatively detected in cultured vascular smooth muscle cells and the intracellular pH were also quantitatively detected with the immunofluorescent quantitative skill through Laser scanning confocal microscope (ACAS570). Results It was found that at the action of NPY (10 -6 mmol/L), bcl-2 and bax were markedly expressed in cultured smooth muscle cells compared with control group. The fluorescent value of bcl-2 and bax were 1 349.67±108.28 and 1397.50±104.43, much higher than 921.72±80.31 and 1 060.44±107.54 in the control group respectively (P<0.01). Intracellular fluid pH in VSMC was decreased by NPY. Conclusion The expression of apoptosis-related genes induced by neuropeptide Y is mediated, in part, by intracellular acidosis in cultured vascular smooth muscle cells.

Abstract:Aim To investigate the characters of Ca 2+ -activated K + channels (KCa) of artery smooth muscle cell(SMC) from normal human mesentery. Methods Human normal mesenteric artery branch was digested by enzyme. Patch clamp technique was used to pull cell-attached and inside-out patches on the normal human mesenteric artery SMC respectively. The signal channel open and close times, open probability (P 0), open channel number per patch were recorded. Results The channels showed obviously voltage dependent, the amplitude voltagerelation curve can be linearly fitted during the voltage range from 10～60 mV(in symmetrical [K +] 0 140 mmol/L and [Ca 2+ ] free < 10 -9 mol/L). The KCa channels have slope conductance of 192.3±29.2 Ps and 202.5±58.3 Ps, in cell-attached and inside-out patches respectively. The channels were Ca 2+ sensitive . The single channel P 0 and open channel number per patch were increased apparently according to the increase of Ca 2+ (10 -8 ～10 6 mol/L) in cytoplasm . TEA in 2×10 -4 mmol applied in the out side of the channel can reduce the signal channel current amplitude. Conclusion The characters of KCa from normal human mesenteric artery SMC were mainly big conductance KCa, which have voltage and Ca 2+ concentration dependent and similar to other artery smooth muscle cell of human.

Abstract:Aim To study the protective effects of Onychin on vasular endothelium-dependent relaxation relaxation damage induced by lysophophatidylcholine(LPC). Methods The vasorelaxation responseto acetylcholine(ACh)were investigation in the rabbit thoracic aorta; The LDH level in conditioned media of cultured endothelial cells was measured by DGKC assay. Results On the rabbit aortic rings, Onychin alone did not have effect on relaxation response to Ach and on contraction response to phenylephrine. LPC 4 mg/L significantly attenuated the endothelium-dependent relaxation of rabbit aortic rings as shown by decreasing the relaxation percentage from 39.1±10.1, 67.1±9.6 and 76.7±10.0 to 2.1±1.0, 10.0±3.9 and 16.1±3.5 response to 0.1, 1.0 and 3.0 μmol/L ACh, respectively; Pretreatment of onychin 3 μmol/L for 10 min markedly increase the relaxation percentage to 14.6±2.6,32.2±2.8,42.1±8.0. The effect of Onychin was blocked by nitric oxide synthase inhibitor N ω-nitro-L-arginine(N-L-A) and prostacyclin synthetase inhibitor indomethacin ( maximal relaxalion percentage: 22.4±7.2, 24.8±2.3 vs 42.1±7.9 to ACh 3 μmol/L). Furthermore,Onychin obviously decreased LPC-induced LDH release of cultured endothelial cells as shown by lowering LDH level from 115.3±19.3 to 30.8±5.4 IU/L. Both N-L-A and indomethacin also inhibited the effect of onychin on LDH release. Conclusion Onychin protects the endothelium-dependent relaxation against elicited-LPC injury with a mechanism related to the activation of nitric oxide and prostacyclin.

Abstract:Aim To elucidate the proatherogenic effect of oxidized low density lipoprotein (ox-LDL)on the apoptosis of cultured endothelium cells and protect effect of nimodipine. Methods ox-LDL was obtained by incubated with Cu²⁺ ,different concentrations of ox-LDL incubated with endothelium cells. MTT test was carried out to evaluate the cell growing state. PI staining test and ELISA test were conducted to analyze apoptosis index. Results The inhibiting effect of ox-LDL on the endothelium growth by ox-LDL was in a dose- dependent manner. ELISA and PI staining showed that ox-LDL induced endothelium cell apoptosis at the same concentration (P<0.05), Nimodipine could inhibited the ox-LDL induced endothelium cell apoptosis. Conclusions The cytotoxicity of ox-LDL to endothelium cell was related to the dosage concentration of ox-LDL: the ox-LDL could inhibit the cultured endothelial cells growth,and even resulted in apoptosis. ox-LDL could start the atherosclerosis in vivo due to the endothelium cells impairment and apoptosis.Nimodipine could protect endothelium from ox-LDL induced apoptosis.

Abstract:Aim To investigate the dynamic changes of nuclear factor-κB(NF-κB)activity in balloon-injured arteries,rabbit aortas(n=18)were subjected to abrasion injury using 4F Forgarty catheter. The arterial tissues were collected and nuclear protein of the tissue was extracted at 0, 12, 24, 48 and 72 h following injury. NF-κB activity in balloon-injured arteries were examined by electrophoretic mobility shift assay. No detectable NF-κB activity was found in normal arteries of rabbits,while NF-κB binding activity in balloon-injured arteries increased apparently,which was detected immediately following injury,peaked at 12 hours,lasted 48 h, returned to baseline at 72 h. The results of density scan for binding blot showed that NF-κB activity was enhanced more than 3.7 fold at 12 h(n=4,P<0.01),2.5 fold at 24 h (n=4,P<0.01),and 1.4 fold at 48 h (n=4,P<0.05), compared with baseline activity (0 h)of NF-κB. The present data showed that balloon injury could trigger NF-κB activation in arteries, which was likely to be involved in the proliferation of arterial smooth muscle cells.

Abstract:Aim To study the effect of calcineurin (CaN)-dependent signaling passway on proliferation of rat vascular smooth muscle cells(VSMC) under stimulation of Ryanodine (RY). Methods Upon the model of cultured rat VSMC, Ca 2+ releasing from Ca 2+ stores stimulated with RY, CaN signaling pathway was blocked with cyclosporine A (CsA) and Ca 2+ channel with verapamil(Ver), detecting the activities of VSMC CaN, mitogen activated protein kinase (MAPK) and protein kinase C(PKC), 3 H-Leucine( 3 H-Leu) and 3 H-Thymidine( 3 H-TdR) incorporation as the target to evaluate VSMC proliferation. Results Synthesis rate of protein and nucleic acid stimulated by RY in VSMC increased significantly in contrast to control (P<0.01); CsA and Ver markedly inhibited syntheses of protein and nucleic acid mediated by RY in VSMC with a significant difference from RY-stimulated group(P<0.01). CsA and Ver also suppressed VSMC CaN activity mediated by RY, and Ver suppressed VSMC PKC activity mediated by RY. Conclusions The study indicates CaN signaling pathway play an important role in VSMC proliferation induced by RY, but it is not the only signaling pathway in regulating VSMC proliferation, MAPK-centered signaling pathway is also involved in VSMC proliferation induced by RY.

Abstract:Aim β-amyloid protein (β AP) may play some important role in the pathogenesis of atherosclerosis. The purpose of this study is to investigate the effect of β AP on U937 cells. Methods The β-amyloid protein was added to the media of U937 cells and the apoptosis of the cells was determined by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) and agarose gel electrophoresis, the cell cycle of U937 cells was analyzed with flow cytometry. Results TUNEL and flow cytometry showed that β-amyloid protein was able to induced apoptosis of U937 cells. Electrophoresis of genomic DNA showed internucleosomal fragments of DNA. Conclusions β-amyloid protein could induce apoptosis in U937 cells.

Abstract:Aim To study apolipoprotein (apo) E polymorphism and its relationship with plasma lipids and apolipoproteins in Chinese patients with type Ⅱa hyperlipoproteinemia. Methods and Subjects Apo E genotype were assayed by polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP), serum lipids were determined by enzyme method and apolipoproteins were measured by radial immunodiffusion assay in 87 type Ⅱa hyperlippoproteinemia patients whose fasting serum lipids levels were TG≤1.82 mmol/L,TC ≥6.21 mmol/L and 230 healthy subjects whose fasting serum lipids levels were TG≤1.82 mmol/L,TC<6.21 mmol/L from a population of Chinese Han nationality in Chengdu area. Results Compared with the control group the values of serum TG, TC, HDLC, LDLC, nHDLC, apo B100, apo CⅡ, apo CⅢ and apo E levels in patients with typeⅡa hyperlipoproteinemia were significantly increased (P<0.01) and the values of apo E/CⅢ ratio were significantly decreased (P<0.05). Apo E3/3 genotype and allele ε3 frequency in type Ⅱa hyperlipoproteinemia group and the control group were both the highest. Allele ε4 frequency in type Ⅱa hyperlipoproteinemia group was tended to increase than that in the control group and allele ε2 frequency was tended to decrease (P>0.05). In type Ⅱa hyperlipoproteinemia group the genotype of apo E₂ was with higher serum apo E levels compared with the genotype of apo E3, and the genotype of apo E4 was with lower serum apo E levels than those in genotypes E3 and E₂ (P<0.001). Conclusion Apo E gene polymorphism was associated with serum apo E levels in type Ⅱa hyperlipoproteinemia.

Abstract:Aim To investigate the lipoprotein lipase gene HindⅢ polymorphism associated with Chinese endogenous hypertriglyceridemics (HTG). Methods Lipoprotein lipase gene Hind Ⅲ polymorphism was studied using polymerase chain reaction (PCR) and restriction fragment length polymorphisms (RFLP) in 200 endogenous hypertriglyceridemics and 202 healthy subjects from a population of Chinese Han nationality in Chengdu area. Results The results showed that both in HTG group and control group, the H2H2 homozygous was the major alleles. H2 allelic frequencies in HTG group were higher than that of control group (0.855 vs 0.745,P<0.01).but H1 allelic frequencies in HTG group was significantly lower than that of control group (0.145 vs 0.255,P<0.01). the plasma triglycerides(TG) level, apo CⅡ and apo CⅢ levels, TG/HDLC ratio in H2H2 genotypes were significantly higher than that of H1H1 genotypes (P<0.01), also higher than that of H1H2 genotypes (P<0.05). Conclusions Therfore, it is suggest that the HindⅢ RFLP at intron 8 of LPL gene is associated with endogenous hypertriglyceridemics to some extent in Chinese population.

Abstract:Aim To assess the extent of atherosclerosis in elderly hypertensive patients. Methods With high resolution ultrasound, not only intima-media thickness (IMT), plaque of carotid arteries, but also endothelium-dependent relaxing function were measured in 82 subjects, consisted of 62 patients with essential hypertension and 20 controls. Results IMTs of carotid arteries were higher in hypertensive patients than those in normal subjects (P<0.05). There was an increasing trend for IMT with the severity of plaque. Flow mediated dialtation in the groups with essential hypertension was much reduced compared with the control group (P<0.05). Conclusions Increases in IMT of carotid arteries and impairment of endothelium-dependent vasolidation are related to atherosclerosis in early stage in elderly hypertensive patients.

Abstract:Aim To investigate the preventive effect of V Flex stent implanted into coronary vessels on coronary arteriosclerosis disease. Methods One hundred and seventy four V Flex stents implantations were attempted in 124 patients (age 62.5±6.2) with 138 arteriosclerosis wounded vessels. Lesion types were A in 8 vessels, B in 86 vessels and C in 44 vessels. Acute angiographic success was 100%. Results Smooth angiographic results were obtained in all cages. Complications were one non stent related death, four non Q wave myocardial infarction. Six month control angiography was performed in 108 (27%) cases of eligible patients and revealed a restenosis rate of 24%. Conclusion Our study show that V Flex is an excellent coronary stent for treatment of complex lesions, providing excellent results with a low complication rate.

Abstract:Aim To study the effect of hypercholesterolemia (HC) on platelet quantitative stereologic parameters and its change after treatment by pravastatin. Methods 20 patients with HC were selected and treated with pravastatin 10～20 mg/d for 4～8 weeks. The changes of platelet ultrastructure were observed by electron microscopic and analysed with quantitative stereologic method. Results After 4～8 weeks treatments, the levels of serum total cholesterol (TC) and platelet aggregation function were lowered and most of platelet quantitative stereologic parameters were improved. Conclusion Altered platelet function and quantitative stereologic parameters with pravastatin therapy in patients with HC can be corrective with its mechanism of enhancing atherosclerosis regression.

Abstract:Aim To investigate the relationships between plasma calcitonin generelated peptide (CGRP) and the severity of coronary lesions and left ventricular function in patients with coronary heart disease. Methods In 42 patients with fixed stenosis coronary artery ≥70% of luminal diameter in the major coronary arteries and 15 normal persons, the concentration of plasma CGRP was measured using radioimmunoassay (RIA). The relationships between plasma CGRP and the Leaman coronary artery score as well as left ventricular ejection fraction (LVEF) were assessed. Results The concentration of plasma CGRP was obviously lower in the coronary disease group than in the control group (276.12±164.75 ng/L vs 411.44±110.32 ng/L,P<0.01). The concentration of plasma CGRP was negatively related to the Leaman score (r=-0.61,P<0.001) and positively to LVEF (r=0.57,P<0.001). The Leaman score had no significant correlation to LVEF in the whole group. However, in the subgroup with the level of plasma CGRP<250 ng/L, the Leaman score had significantly negative correlation to LVEF (r=-0.49,P<0.05). Conclusions The plasma CGRP had correlation to both the severity of coronary lesions and left ventricular function in patients with coronary heart disease. In the patients with decreased concentration of plasma CGRP, the left ventricular function deteriorated with the progress of coronary lesions.

Abstract:Aim To establish a cell cultured container with the condition of carbon monoxide (CO) or hypoxia (H) and to explore the effects of CO and hypoxic on rat aortic endothlial cell (RAEC). Methods By the use of drawing-charging method to obtain desired gas concentration in the container in a reletively short time, and ultrastructural features of RAEC was observed by transmission electron microscopy. Results 1) The concentration of CO and O 2 in container can reach the necessariexia, the typical constituents of mitochondria, vesicles, endoplasmic reticulums, neclei and cytomembrane was seen in RAEC. But there are large vacuoles increased,swollen and brolen mitochondria occurred only some of cells. Many lipid bodies formed in cells. Cell surface blebbing and many vacuoles could be seen in some cells. Chromatin was located to the rim in the neclei. Intercellular conjection was obviously separate. After 12 h of low-density carbon monoxide (1% CO) compund hypoxia, intercellular conjection was lightly solitary,a few vacuoles architectionics was seen in cytoplasm, and few chromation rimmed in the nuclei. [WT5”HZ]Conclusion The cell architectionics and conjection was injured in hypoxia, And cell injured level decreased in low-density CO compound hypoxia.It suggest that low-density CO can partly protect RAEC from being damaged by hypoxia.