An Experimental Study On The Treatment Of Vascular Smooth Muscle Phenotype Transformation And Improvement Of Atherosclerosis From The Perspective Of The Spleen

Purpose:Atherosclerosis(AS)Mice model was established by high fat feedin g Apo E-/-mice,the indexers of vascular smooth muscle cells(VSMCS)such a s proliferator-activated receptor γ(PPARγ),smooth muscle 22α(SM22α),matri x metalloproteinase-2(MMP-2),matrix metalloproteinase-9(MMP-9)were detec ted in arterial tissue to explore the mechanism of JPTMF in preventing and re versing AS in early stage.Material and method:50 8-week-old Apo E-/-male mice were high-fat fed for one week a nd then randomly divided into 5 groups(10 for each),which are model group,JPTMF high-dosage group,JPTMF medium-dosage group,JPTMF lowdosage group and positive control group.Ten 8-week-old SPF male C57 B L/6J from same strain as normal group.The model group and the normal group were gavaged with equal amount of normal saline.JPTMF high-do sage group,JPTMF medium-dosage group,JPTMF low-dosage group and po sitive control group were treated with the same amount of high,medium and low-dosage solutions of JPTMF and atorvastatin calcium tablet sol ution.The high,medium and low dosage of JPTMF was set as 3:1:1/3.Aft er one week of adaptive feeding,after 12 weeks treatment and tissue we re collected at the end of 13 weeks.HE staining and oil red O staining were used to observe the morphological changes and lipiddeposition of aorta in mice.Contents of PPARγ,MMP-2 and MMP-9 in aorta were det ected by enzyme-linked immunosorbent assay(ELISA)and real-time qua ntitative PCR(RT-PCR).Contents of SM22α in aorta were detected by re al-time quantitative PCR(RT-PCR).Results:1.HE staining results:Normal group:Aorta tissue structure is normal.No degeneration,necrosis or hyperplasia was observed in the epithelial cells on tunica intima.The subendothelial inner elastic membrane is normal,and no plaque was found in subendothelial and middle membrane.Model group:Aortic structure is disordered and intima is not smooth.Foam cells were seen in the tunica middle membrane,and obviouly atheromatous plaques are formed.JPTMF high-dosage group:Aorta is normal structured,the wall of the tube is not thickened,the intima is smooth,and no plaque is formed.JPTMF medium-dosage group:The aortic structure is relatively normal and intima is smooth,several lipid cells in the tunica middle membrane,and plaque formation occasionally occurs in the intima.JPTMF low-dosage group:Aorta is structurally disordered,the intima i s not smooth.A plurality of foam cells can be seen and atherosclerotic plaquse are formed on the surface of the intima.Positive control group:Aorta has normal structure,no thickening of tube wall,the intima is smooth,normal arrangement of elastic fibers and no plaque formation.2.Oil red O results :Compared with the normal group,the model group had a large amount of lipid deposition in the aorta.Compared with the model group,the lipid level i n aorta of each drug group has been decreased especially in JPTMF high-dosa ge group.3.ELISA results :(1)Compared with the normal group,the content of PPARγ in aorta of m odel group decreased significantly(P<0.01).Compared with the model group,th e content of PPARγ in aorta of all the four medication groups was significantl y increased(P<0.01),high to low ranking as:JPTMF high-dosage group,Positive control group,JPTMF medium-dosage group,and JPTMF low-dosage group.(2)Compared with the normal group,the content of MMP-2 in aorta of the model group increased significantly(P<0.01).Compared with the model group,the content of MMP-2 in aorta of all the four medication groups decreased significantly(P<0.05),high to low ranking as:JPTMF high-dosage group,Positive control group,JPTMF medium-dosage group,and JPTMF low-dosage group.(3)Compared with the normal group,the content of MMP-9 in aorta of the model group increased significantly(P<0.01).Compared with the model group,the content of MMP-9 in aorta of all the four medication groups decreased significantly(P<0.01),high to low ranking as:JPTMF high-dosage group,Positive control group,JPTMF medium-dosage group,and JPTMF low-dosage group.4.RT-PCR results show that:(1)Compared with the normal group,RNA expression of PPARγ in aorta of model group decreased significantly(P<0.01).Compared with the model group,RNA expression of PPARγ in aorta of all the four medication groups were significantly increased(P<0.01),high to low ranking as:JPTMF high-dosage group,Positive control group,JPTMF medium-dose group,JPTMF low-dosage group.(2)Compared with the normal group,RNA expression of SM22α in aorta of model group decreased significantly(P<0.01).Compared with the model group,RNA expression of SM22α in aorta of all the four medication groups were significantly increased(P<0.05),high to low ranking as:JPTMF high-dosage group,Positive control group,JPTMF medium-dosage group,and JPTMF low-dosage group.(3)Compared with the normal group,RNA expression of MMP-2 in aorta of the model group increased significantly(P<0.01).Compared with the model group,RNA expression of MMP-2 in aorta of all the four medication groups de creased significantly(P<0.05),high to low ranking as:JPTMF high-dosage group,P ositive control group,JPTMF medium-dose group,and JPTMF low-dose group.(4)Compared with the normal group,RNA expression of MMP-9 in aorta of the model group increased significantly(P<0.01).Compared with the modelgr oup,RNA expression of MMP-9 in aorta of all the four medication groups decr eased significantly(P<0.05),high to low ranking as:JPTMF high-dosage group,Po sitive control group,JPTMF medium-dose group,and JPTMF low-dose group.Conclusion:1.The occurrence and development of AS pathogenesis is related to the theory of "spleen-ying-vessel correlation",which demonstrated as ‘locating in vessel,rooting from spleen,starting in qi and changing in ying ’.2.One of the mechanisms of AS may be related to phenotype transformation o f VSMCS.3.JPTMF can inhibit the phenotypic transformation of VSMCS by reducing the production of VSMCS contraction marker SM22α and regulate regulate the rele ase of MMP-9 and MMP-2,so as to prevent AS.4.The high,medium and low doses of JPTMF can prevent and reverse the for mation of AS in early stage,with clear dosage related effect.JPTMF high-dosage group is the best,and its mechanism may be related to inhibiting the phenotyp ic transformation of VSMCS in Apo E-/-mice.