Abstract:Aim To study the apoptosis and the expressions of protooncogenes bcl-2, P53 and c-myc of the smooth muscle cells (SMC ) derived from rabbit atherosclerotic (As) plaques. Methods Cell apoptosis was in sute detected with the terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) and DNA agarose gel electrophoresis, respectively. Expressions of protooncogenes bcl-2, c-myc and P53 were observed by Northern blotting.Results It was found that a lot of SMC were positively stained with TUNEL in As plaque tissues. The DNA electropherosis of the plaque tissues showed a ladder pattern. Northern hybridization showed that expressions of protooncogenes bcl-2 and P53 in the plaque tissues were markedly increased, compared with the normal tissues, but the expression of c-myc was decreased. Conclusion Apoptosis of SMC existed in the As plaques. The expressions of protooncogenes bcl-2 - cmyc and P53 play a important role in regulating the apoptosis of SMC in the As plaques.

Abstract:Aim In order to elucidate the role of oxidized low density lipoproteins (OLDL), we have examined the metabolism of OLDL in macrophages and the effect of OLDL on the total cholesterol content in macrophages.Methods The metabolism of low density lipoprotein (LDL) and OLDL in macrophages was assessed with the radio-receptor assays.Results Human macrophage metabolized LDL by a receptor pathway with 390. 6 μg/g cell protein of the Bun and 17. 5 mg/L of kd. The receptor Pathway exhibited satuative kinelies. The metabolism of OLDL was different from LDL. The binding internalization and degradation of OLDL by macrophages increased significantly as compared with the LDL. The total cholesterol content in macrophages incubated with OLDL was also higher than that of cells incubated with native LDL.Conclusion OLDL was metabolized by macrophages with different pathway from LDL and may lead to the formation of foams.

Abstract:Aim To investigate the effect and mechanism of heparan sulfate proteoglycan and heparin on accumulation of lipid in macrophages.Methods Lipid accumulation in macrophages was observed with Sudan Ⅳ stain. Cholesterol content in macrophages was detected with enzyme fluorescence assay. The activity of scavenger receptor of macrophage was assessed by examing the degradation rate of 125 1 labeled oxidized low density lipoprotein (OLDL).Results More red particles stained with Sudan Ⅳ were seen in macrophages incubated with OLDL (100 mg/L). The red particles in macrophages incubated with OLDL (100 mg/L) plus heparan sulfate proteoglycan (HSPG, 15. 5 mg/L) or with OLDL (100 mg/L) plus heparin (HP, 200 mg/L) were far less than that with OLDL only. The total cholesterol and cholesteryl ester in macrophages incubated with OLDL plus HSPG or with OLDL plus HP were significantly lower than that with OLDL (P〈0.05: P〈0.01).The degradation rate of 125 I labeled OLDL of macrophages incubated with HSPG or with HP were lower than control.Conclusion HSPG and HP may inhibit lipid accumulation in macrophage and formation of foam cell possibly through down-regulating the activity of scavenger receptor of macrophage and decreasing the intake of OLDL.

Abstract:Aim To observe whether lipid peroxidation injury to cultured human umbilical vein endothelial cells and bovine aortic endothelial cells has effects on the expression of monocyte chemoattractant protein-l (MCP-1) in them.Methods The lipid peroxidation injury to endothelial cells (EC) was induced by exposure to diamide either at a same concentration but for different incubation time or at different concentrations but for a same incubation time. The EC in different groups were collected and their total RNA was extracted by the single-step method. The MCP-1 mRNA expression in EC was examined by dot blotting using a Y32 P-end la beled 35 mer-oligonucleotide probe, Meanwhile, MCP-1 protein content in the media conditioned by the cultured EC exposed to diamide was determined by sandwich ELISA.Results Cultured EC could express MCP-1 mRNA and protein. Dot blotting showed that exposure of EC to diamide at a concentration of 5 μmol/L for 2, 4 and 8 h, respectively, resulted in a 2. 26 fold, a 2. 41 fold and a 2. 72 fold increase in the levels of MCP-1 mRNA expression in EC respectively. as compared to the control group. On the other hand .exposure of EC to diamide at different concentrations (1 μmol/L, 5 μmol/L and 10 μmol/L) for 3 h, resuited in a 2. 22 fold, 2. 97 fold and 3. 32 fold increase in the levels of MCP-l mRNA expression in EC respectiveiy. The results of ELISA were similar to that of dot blotting.Exposure of EC to diamide at a same concentration but for different incubation time resuited in a 2. 18 fold, a 3. 87 fold and a 5. 87 fold increase in MCP-1 protein content in the conditioned media. By contrast, exposure of EC to diamide at different concentration but for a same incubation time resuiteh in a 2.34 fold, a 3. 44 fold and a 4. 6 fold increase in MCP-1 protein content in the conditioned media. Together these results showed that the MCP-1 expression in EC induced by exposure to diamide was dependent on both dose and the incubation time.Conclusion Lipid peroxidation to EC can induce increased production of MCP-1 in the cells and may play an important role in the recruitment of monocytes into the intima in atherogenesis.

Abstract:Aim To study the association between insertion/deletion polymorphism of angiotensin converting enzyme gene and coronary artery disease.Methods The insertion/deletion of a 287 base pairs segment in 16 intron of angiotensin converting enzyme gene in 79 coronary artery disease patients and 68 healthy subjects was detected by polymerase chain reaction. The subjects were divided into three groups:homozygotes for the deletion, homozygotes for the insertion and heterozygotes, according to its presence orabsence. The serum angiotensin converting enzyme activity was also detected.Results The deletion genotype frequency in coronary artery disease group was marked higher than in controls (p<0.05). The frequency of deletion allele was also greater than that in controls (p<0. 05).The serum angiotensin converting enzyrne activity between coronary artery disease patients and controls was similar. But it was significantly different among different genotypes in two groups. The deletion genotype had the highest, insertion genotype had the lowest.Conclusion The insertion/deletion polymorphism of angiotensin converting enzyme gene is associated with coronary artery disease, and the deletion genotype perhaps is a risk factor for coronary artery diseaes. The elevated serum angiotensin converting enzyme level may contribute to the generation of coronary artery disease in patients who have deletion genotype.

Abstract:Aim To study the gene structures of apolipoproteins of Beijing duck insusceptible to atherosclerosis,the cDNA libraries of duck liver cells were quickly constructed at the first time.Methods With single-step method. total RNA was isolated frorn duck liver tissue. following purification of mRNA which Passed through oligo dT cellulose chromatography. Using mRNA as the template . the first and sceond strands of the cDNAs were synthesized in Time Saver cDNA synthesis Kit. After been ligated with EcoRl /Notl adaptors, the cDNAs were cloned into the arms of λgt10 or λgt11 vectors. Then the recombinants of ADNA were packed to infect E. coli Y 1090 strain. Results Duck liver cDNA libraries with high titer were obtained and rate of positive recombinants was over 98. 5 %. Autoradiography showed that sizes of cDNA products were in 400～ 5000 bp. Each clear plaque DNA which was identified by endorestriction enzyme digestion contained the cDNA insert. Full length cDNA sequence of duck apoAI was cloned and sequenced from the cDNA library constructed.Conclusion Compared with conventional method of constructing cDNA library the present one is simple,time-saving and eady to be used. Some problems of using the method to construct cDNA library were discussed in detail.

Abstract:Aim To study the existence of high density lipoprotein (HDL) receptor in the cell membrane, and this may by provided the evidence for study of HDL receptor function.Methods HDL binding protein was prepared from fresh swine adrenal cortical cell membranes, and purified with affinity chromatography. The soluable membrane protein was analyzed by ligand blotting.Results The dot blotting showed that the membrane protein possessed high affinity with HDL, and western blotting suggested that the molecular weight (MW) of the binding protein by affinity chromatography is 77 kDa. The characters of the HDL binding protein was measured by biotin-avidin enzyme linked immunosorbent assay (BA-ELISA ): Kd= 21. 6 mg/L, Bmax= 678. 2 μg/g protein. Conclusion It is showed that the condition of affinity chromatography was moderate, and the purified swine adrenal cortical cell membrane possessed high affinity with HDL.

Abstract:Aim To observe the characteristic of intracranial large arterial lesions that were induced by experimental hypertension.Methods A morphometric study was performed on middle cerebral artery and basal artery in stroke-prone renovascular hypertensive rats and the morphometric values afert antihypertensive therapy with captopril were compared with those in non-antihypertensive rats.Results It was found that enhanced media thickness was presented in both middle cerebral artery and basal artery. It was mainly induced by hypertrophy and hyperplasia of smooth muscle cells in middle cerebral artery and by mural remodeling in basal artery. The hypertrophy and hyperplasia of smooth muscle cells were depressed, but the remodeling did not reverse to be sufficient to bring about full normalization after antihypertensive therapy.Conclusion These results indicate that the main lesion of intracranial arteries is enhanced media thickness, but different vessels have different reaction to hypertension and antihypertensive therapy.

Abstract:Aim To develop a new method for microdetermination of taurine the composition of antiatherosclerosis, in animal tissues With reverse-phase high performance liquid chromatography (RP-HPLC) Methods Animal tissues were soaked in water to extract taurine. The extraction was concentrated by vacuum freezing and drying. Then using the mixed liquid of Phenylisothiocyanate (PITC), triethylamine,water and ethanol (1: 1:1: 7) as derivative to precolumn derived taurine with PITC. Under the following conditions: 1 mL/min flow rate, 254 nm wavelength and column temperature at 45℃, 991 HPLC was used to separate, determine taurine. Qualitative and quantitative analysls of the content of taurine in sample animal (slug ) tissues was done by using the approaches of peak increase and standard curve respectivelv.Results Under the condition of selected chromatogram, the retention time of taurine was 5.06 min with distinct qualitative phenomenon of peak increasing. The result was reliable. The lowest detectable amount was 4. 48 ng with the concentration of taurine between 5. 58 X 1O3～ 1. 79 X 105μg/L. The linear regression equation between peak area of the chromatogram and the content of taurine was: S = 3. 34 X 10-4W - 1. 09 X 10-3, with r =0. 9998. The relative standard deviation of the sample liquid was 1. 5 % (n =7 ), the added recovery of the sample was P = 99.57 %.Conclusion This approach of soaking the taurine of animal tissues with water is simple and convonient.It decreases the contamination. The derivative reaction is complete; the derivative of taurine is well separated within the column. This approach is a new analytical method for detecting the trace taurine in animal tissues.

Abstract:Aim Detection of the relationship of Chlamydia pneumoniae infection and coronary atherosclerosis.Methods Using three primers that recognize conserved sequence on Chlamydia major outer membrane protein genes, Chlamydia pneumoniae was detected in 32 left anterior descending branch of adult atherosclerotic coronary arteries and 7 normal coronary arteries.Results Fourteen of 32 left anterior descending branch of atherosclerotic coronary arteries were PCR positive, positive rate was 43. 75 %; 7 normal coronary arteries were PCR negative, positive rate was 0%.Conclusion There is a significant difference of chlamydia pneumoniae infection between atherosclerotic coronary arteries and normal coronary arteries.

Abstract:Aim To investigate the relationship between heavier proteinuria or hypoalbuminemis and serum lipid abnormalities in primary nephrotic syndrome.Methods Serum albumin, total cholesterol (TC),low density lipoprotein cholesterol (LDLC) and 24-hour urinary protein content were assayed in nephrotic syndrome patients (n=20) and healthy controls (n=20).Results Hyperlipidemia is a consistent feature of the nephrotic syndrome patients. 24-hour urinary protein content was positive correlated with the level of serum TC (r=0. 43, p<0.05), but no statistically significance was observed between proteinuria and serum LDLC (r =0. 37, P >0.05). The level of serum albumin was inversely significantly correlated with both the level of serum TC and LDLC (r=-0.69, p<0. 01, r=-0. 51, p<0. 01, respectively).Conclusion There were correlated relationship between proteinuria or hypoalbuminemia and serum lipid abnormalities in nephrotic syndrome.

Abstract:Aim To investigate the change of plasma L-arginine, serum NO2 , NO3 and tissue plasminogen activator (t-PA) in patients with coronary heart disease and haperlipemia.Methods The method of Pico-Taq TM amino acid analysis was introduced to measure the level of L-arginine. Serum NO2 and NO3 was observed by catalytic photometric method. Serum t-PA was measured by enzyme linked immunosorbent assay.Results The levels of plasma L-arginine (patients vs control group: 89±8μmol/L vs 113±5 μmol/L),serum superoxide dismutase (patients vs control group: 67 ± 33 ku/L vs 98± 34 ku/L) and tPA (patients vs control group: 0. 94 ±0. 24 ku/L vs 1. 43±0.21 ku/L) were reduced (P〈0. 05 ), but the levels of serum malondialdehyde (patients vs control group: 7. 1 ±1. 5 μmol/L vs 3. 5±0.6μmol/L) and NO2, NO3 (patients vs control group: 38±10μmol/L vs 30± 9 μmol/L) were increased (P〈0. 05).Concluslon Absence of plasma L-arginine in patients heart disease and hyperlipemia may be related with the increase of requirement of nitric oxide and increase of serum NO2 , NO3 may be related with increase of degradation of nitric oxide. The decrease of tPA activity may be related with endothelium disfunction.