Study Of Relationship Between Wnt Signal Pathway And Artery Restenonsis After Balloon Injury

Percutaneous coronary intervention therapy(PCI) has been further developed to become one of the important procedures in treatment of coronary heart disease, however, restenosis,still the biggest problem,is the limiting factor to long-term follow-up result of PCI。Although drug-eluting stent is a milestone of reducing the accident rate of restenonsis,we haven't found a fundamental solution to restenonsis yet now.Intensive study the genomic mechanism of vascular restenosis after injury may be the key to resolve the difficult problem ultimately.Base on the fact that mass smooth muscle cells can be seen proliferating in artery intima after PTCA,most scholars believe vascular restenosis after PTCA formed by thickening neonatal intima consisting with proliferating SMC from tunica media and its secreting extracellular matrix.Experiment studies conformed certain drugs which can inhibit proliferation of SMC may reduce vascular restenosis after PTCA.Regulatory effect of wnt siginal pathway on vascular restenosis is a new research field. Although researches suggest wnt siginal pathway possibly involved in the proliferation of SMC after vascular injury, evidence is not sufficient and in vivo research information is lacking.Inflammation is a necessary step in the process of intima restoration after vascular injury.Some Chinese herbs, such as emodin, has anti-inflammatory effect. It has not been known whether these herbs have effect on prevention of vascular restenosis and their possible mechanisms.The purpose of this study:(1) Find new therapeutic target of anti-vascular restenosis.(2) Provide new idea and information for depth research and development of Chinese herbs in this field.Part1Role of wnt signal pathway in proliferation and collagen secretion of smooth muscle cells from rats aorta induced by AngⅡCultured cells were divided into seven groups:control group;AngⅡ (10-6mol/L) group;AngⅡ+atrovastatin(0.1umol/L)group; AngII+atrovastatin(1umol/L)group;AngⅡ+atrovastatin (10umol/L) group;AngII (10-6mol/L)+SiRNA-dvl-l group; AngⅡ (10-6mol/L)+SiRNA-dvl-1+atrovastatin (10umol/L) group. Another three groups were added in experiment of MTT.The results show:(1) After Dvl-1Gene was silenced by SiRNA after transcription, proliferation of VSMC induced by AngⅡ was inhibited significantly; Proliferation of VSMC induced by AngⅡ were inhibited significantly by lumol/L concentrations of atorvastatin in comparison with the control group(P<0.05), the inhibited effect showed time and concentration dependent manners;(2) High expression of Wnt4, Dvl-1and β-catenin mRNA and proteins were induced by AngⅡ, which could be inhibited by atrovastatin in concentration dependent manner, and the latter two could be reversed significantly by siRNA-Dvl-1.(3) High expression of collegan Ⅰ,collegan Ⅲ mRNA and proteins in VSMC induced by AngⅡ could be inhibited siganificantly by siRNA-Dvl-1and atrovastatin (1,10umol/L)。The above results indicate:1) Induced by AngⅡ,VSMC proliferate accompanying Wnt signal moleculars highly expressed.2) Expression of intracellular wnt signal moleculars reduced significantly when AngⅡ-induced proliferation of VSMC was inhibited by siRNA-Dvl-1and atrovastatin;3) Activation of Wnt pathway involved with AngⅡ-induced high expression of collegan genes in VSMC.Conclusion:Wnt signal pathway involved with AngⅡ-induced VSMC proliferation.Part2Expression of Wnt signal moleculars in rat carotid artery injuried by balloonRat model of balloon-injuried carotid artery was eatablished. SD Rats were divided into3groups:sham operation group; model group, statin group.The result show:(1)6weeks after injured by balloon, intimal of the injured artery became thickening. Thickness ratio of intima/tunica media and collegan volume in intima increased significantly compared with that of sham's (p<0.05); Compared with model group, thickness ratio of intima/tunica media and collegan volume in statin group reduced significantly (p<0.05);(2) Immunohistochemistry showed expression of wnt signals as Wnt4,dvl-1and?-catenin increased significantly in injured intima and tunica of carotid artery of model group and reduced in statin group;(3) Expression of wnt4. dvl-1and?-catenin in injured artery increased significantly in comparison with sham operation group while reduced in statin group;(4) Proteins expression of collegan Ⅰ and collegan Ⅲ in model group was higher than statin's. The above results indicate:1) Pathological changes of injured rat's carotid artery associated with expression level of wnt signal moleculars.Conclusion:Wnt signal pathway involved with pathological changes of balloon-injuried artery in vivo.Part3Study on mechanisms of emodin's preventive effect on artery restenosis after injured.Cell experiment in vitroExperement groups:control, AngⅡ groups, emodin group, AngⅡ+SiRNA group, AngⅡ+SiRNA+emodin group.The result show:(1) VSMC proliferation induced by Angll could be inhibited by Emodin in a concentrated-dependent manner;(2) High expression of wnt4. dvl-1,β-catenin and cyclinDl's mRNA and proteins induced by AngⅡ could be down-regulated significantly by emodin with certain concentration that inhibit SMC proliferation;(3) High expression of collegan Ⅰ,collegan Ⅲ's mRNA and proteins could be down-regulated significantly by emodin.Animal experiment in vitroRat model of balloon-injuried carotid artery was established. SD Rats were divided into3groups:sham operation group; model group, emodin (ig) group.The result show:Compared with model group,(1) hyperplasia of artery intima area of neointimal,thickness of intima and area ratio of intima/tunica media in emodin group were all reduced (p<0.05);(2) immunohistochemistry showed expression of wnt signals as Wnt4. dvl-1and β-catenin in injured artery of emodin group were reduced (p<0.05);(3) expression of wnt4,dvl-1and β-catenin in injured artery of emodin group reduced;(4):Collagen specific staining with Picric Sirius red showed collagen volume fraction reduced in emodin group.The above results indicate:1) Emodin could inhibit VSMC proliferation through down-regulation of wnt signal pathway;2) Emodin could inhibit collagen synthesis and secretion of VSMC though down-regulation of wnt signal pathway.Conclusion:Pathological changes of balloon-injured carotid artery could be abated by emodin.