Study On The Stability Of Protein Tyrosine Phosphatase SHP-2 On Atherosclerotic Plaque In ApoE Gene Knockout Mice

Objective: Atherosclerosis(AS)is a common disease with a high rate of disability and mortality.The researches on its pathogenesis、prevention and treatment have been a hot issue in society.The main harm of AS comes from the instability of plaques,which can fall off and form emboli,leading to serious clinical complications and affecting people’s lives.In recent years,autophagy has been found in all stages and major cells of AS.Autophagy and apoptosis have a complex relationship of reinforcing and inhibiting each other.Excessive cell apoptosis is a powerful promoting factor for plaque instability,so it is extremely important to explore the regulation of autophagy and apoptosis about atherosclerotic plaque.Protein tyrosine phosphatase SHP-2(Src homology 2 domain-containing protein tyrosine phosphatase,SHP-2),a non-receptor tyrosine phosphatase,has been proved that it can regulate the progression of disease by participating in major cellular events such as migration 、 differentiation and metabolism through complex signaling pathways.The role of SHP-2 in the stability of atherosclerotic plaques has not been reported in detail.Therefore,from the perspective of autophagy and apoptosis,the study which studied the effect of SHP-2 inhibitor PHPS1(Phenyl hydrazone pyrazolone sulfonate 1)on the stability of atherosclerotic plaque in Apo E gene knockout(Apo E-/-)mice can provide a new theoretical basis and prevention strategy for stabilizing atherosclerotic plaques and reducing the complications.Methods: Twenty male Apo E-/-mice aged 6-8 weeks were fed adaptively for one week and randomly divided into control group and PHPS1 group.Mice in PHPS1 group were intraperitoneally injected with PHPS1(0.3mg/kg/d),while mice in control group were intraperitoneally injected with the same dose of normal saline.The mice were fed with 1.25% cholesterol high-fat diet for 8 weeks.Routine paraffin sections were made from mice aortic roots.Atherosclerotic plaque area was determined by hematoxylin-eosin(HE)staining.The percentage changes of collagen in atherosclerotic plaques were analyzed by Sirius red staining.The changes of macrophage expression level in the plaques were detected by CD68 immunofluorescence.Cell apoptosis in the plaque was observed by TUNEL staining.The expression levels of autophagy-related proteins(LC3-II,ATG-5),apoptosis-related proteins(CHOP,Caspase-3)and PI3K/Akt/m TOR pathway proteins in atherosclerotic plaques were detected by Western Bolt.Results: The mice of PHPS1 group were compared with those of control group:1.Changes in atherosclerotic plaques area: HE staining of paraffin section showed an increase in plaque area.The difference between the groups was statistically significant(P<0.01).2.Changes in the composition of atherosclerotic plaques: Sirius red staining showed a decrease in the percentage of collagen in the plaque(P<0.01);CD68 immunofluorescence showed an increase in the macrophage content in the plaque(P<0.01).The differences between the groups were statistically significant.3.TUNEL staining showed increased apoptosis in the plaques,and the difference was statistically significant(P<0.01).4.Western Bolt results showed that the protein expression levels of PI3K/Akt/m TOR pathway were inhibited.The relative protein expression levels of p-PI3K/t-PI3K、 p-Akt/t-Akt and p-m TOR/t-m TOR were decreased,and the differences were statistically significant(P<0.01).5.Western Bolt results indicated that the relative protein expression levels of autophagy-related proteins(LC3-II and ATG-5)and apoptosis-related proteins(CHOP and Caspase-3)in atherosclerotic plaques were increased.The differences were statistically significant(P<0.01).Conclusions:1.SHP-2 can inhibit plaque formation,reduce macrophage content and increase collagen fiber content.2.SHP-2 can inhibit autophagy through PI3K/Akt/m TOR pathway and inhibit apoptotic,so as to stabilize plaques.