The Research On Anti-atherosclerosclerosis With Peroxisome Proliferator-activated Receptor Gamma Activator

Peroxisome proliferator-activated receptor gamma activator has been found to inhibit the atherosclerotic plaque formation and progression in Apo E knock out mice, but it is still not clear about the mechanism of antiatherosclerosis. The development and progression of atherosclerosis involves many biological enzymes such as: nitric oxide syntheses; matrix metallproteinases; however, p-catenin has been found to be relative with the expression of matrix metallproteinases and the migration and proliferation of vascular smooth muscle cells. So we are aimed to investigate PPARt agonist on the expression of nitric oxide syntheses; matrix metallproteinases and P-catenin in hypercholesterolemic rabbit atherosclerotic plaque. Methods: constructing animal models, the control group contains 6 rabbits with normal chows, hypercholesterolemia chows were given to the rabbits for 2 weeks and then under anesthetized with pentobarbital, femoral artery was opened, and the catheter balloon was inserted into abdominal aortic artery for 15 centimeters, then inflated and pulled forward and back for three times. Hypercholesterolemia chows continued to give the rabbits after the procedure. At the same time animals were divided into hypercholesterolemiamodel group (group B), group C with Atovtatin 5 mg/Kg/day; group D with Rosiglitazone 3 mg/Kg/day; combination group with both Atovtatin 5 mg/Kg/day and Rosiglitazone 3 mg/Kg/day. Before balloon injury and after the experiment serum was measured for total cholesterol and triglyceride from each rabbit. Also the thickness of intimal-medium of abdominal aortic artery was determined with SigmaScan Pro 5.0 software under light microscope.lOOmg abdominal aortic artery tissue was grinded with homogenate medium, the supernant was used for eNOS and iNOS activity with a NOS dividing kit.Total RNA was isolated from each group abdominal aortic artery with Trizol regent, RNA concentration was determined spectrophotometrically using absorbance at 260nm and 280nm. The cDNA was obtained with a reverse transcription kit, the PCR were done using special pair primers for VCAM-K ICAM-K eNOS> iNC^ MMP2> MMP^ TIMPK TIMP2, PPAR i > p-catenin > GAPDH. PCR products were detected by semi-quantitative methods.Arterial segment were cut into rings 1 mm long and incubated for 24 hours in 1 mL serum-free DMEM (BioMedia) at 37°Cin humidified 5%CO2/95%air.the medium were measured for MMP2 and MMP9 gelatinolytic activities with SDS-PAGE Zymography; reverse zymographywas performed to measure TIMPl and TIMP2 inhibitory activities in media. Proteins from tissue homogenate were separated on SDS-PAGE gels, transferred to nitrocellose membrance, and Western blotted with momoclone antibodies against PPAR? and P-catenin.Arterial segments were fixed in 4% paraformalsehyde , four-micrometer cross sections were from each block and immunostained with mouse monoclone antibodies directed against MMP2 and P-catenin. Results: 1.there were no difference between hypercholesterolemic chows rabbits on serum total cholesterol and total triglyceride include group B> C> D and E, but they were significantly higher than that in group A. the thickness of intimal-medial in group D and E were significantly decreased than that in group B and C, also there were much less foam cells in atherosclerotic plaque group D and E compared with group B and C. However; the thickness of intimal-medial and foam cells in group C were reduced than that in group B. 2. the eNOS activity were significantly decreased in group B than that in other groups; but the iNOS activity were highest in group B, and there were no significant difference between group C> D and E. 3. the RT-PCR mRNA of ICAM-1 ^ VCAM-1 ^ eNOS> MMP2> MMP9 were significantly decreased in group C^ D and E compared with that in group B. eNOS in group D didn't highly expressed than that in group A. Moreover, TIMPl and TIMP2 were significantly raisedin hypercholesterolemic groups than that in control group. The mRNA and protein of PPARy expressed significantly higher ^ and that of p-catenin expressed significantly lower in group D and E than other groups. Zymography showed underdetected MMP9 activity in group D and E, also MMP2 activity in group C> D and E were significantly lower than that in group B; MMP9 activity could be detected in group C and significantly decreased than that in group B. Immunohistochemistry showed the expression of MMP2 and P-catenin were much reduced in group D and E than that in group B and C.Conclusion: 1. PPARy agonist can reduce the neointimal thinckness and foam cells in the rabbit abdominal aortic artery induced by hypercholesterolemia and injury. 2. one of the important mechanisms of PPARY agonist could inhibit the formation of atherosclerotic plaque is that it can inhibit the expression on mRNA and protein of MMP2 and MMP9. 3. the activation of PPARy inhibit the expression on mRNA and protein of P-catenin should be another signal pathway to anti-atherosclerosclerosis.