Effect Of Periostin On Angiotensin Ⅱ-Mediated ApoE^-/- Mouse EL/HL Expression And The Discussion Of Its Mechanism

Objective:ApoE^-/-atherosclerosis model was established by high-fat diet.Adopt methods of gene expression silencing and the overexpression of periostin to investigate the effect of periostin on endothelial lipase and hepatic lipase.In order to elucidate the effection of periostin on angiotensin II inhibition of reverse cholesterol transport.Furthermore,whether periostin regulates cholesterol homeostasis through p38MAPK signal transduction pathway is further discussed,in order to provide a potential aim for clinical prevention and treatment of atherosclerosis.Methods:1.Molding:Fifty male C57BL/6J,6-8 weeks old ApoE^-/-mice were selected and randomly divided into normal diet group（ND group）,hyperfat diet group（HD group）,normal expression of periostin group（HD+Ang Ⅱ group）,periostin group about silence（HD+POSTN-mus-769+Ang Ⅱ group）,and overexpression of periostin group（HD+POSTN mus+Ang Ⅱ group）.Each group had 10 mice;2.Processing method:0.9%sodium chloride was administered to groups I,II and III by tail vein injection,LV3-POSTN-mus-769 was injected into group IV to silence the lentivirus suppressor gene,and LV5-Postn-mus overexpressed lentivirus gene was injected into group V by tail vein injection to explore the effect of periostin on Ang Ⅱ mediated atherosclerosis.Based on this model,the grouped mice（group III,IV,V）were injected with the same concentration of Ang Ⅱ subcutaneously.3.Detect:1)A TC,TG,HDL-C,LDL-C was detected using a biochemical analyzer to assess lipid levels2).Aortic histopathology（oil red O stain）was performed to assess the formation and characteristics of intra aortic plaques.3).Perform liver pathology（oil red O stain,hematoxylin-eosin stain（HE stain））to assess the extent of fatty lesions.4).Use Western-blot to test the expressions of EL/HL and periostin,and clarify the mediating effect of periostin on Ang Ⅱ inhibition of reverse cholesterol transport.p-p38MAPK was tested to further explore whether periostin regulates cholesterol homeostasis through p38 MAPK signal transduction pathway.5).Use RT-PCR to test the expressions of EL/HL,p-p38MAPK,and periostin mRNA in liver tissue,and further explore the induction mechanism of Ang Ⅱ to periostin.Results:1.High-fat diet can lead to the formation of arterial plaque in ApoE^-/-mice.2.Periostin can affect abnormal lipid metabolism:Compared with HD+Ang Ⅱ group,silent periostin gene decreased the contents of TC,TG,and LDL-C,but increased the contents of HDL-C（P<0.05）,It was shown that silencing of the periostin gene could positively affect in the expression of Ang Ⅱ on lipid metabolism and suppress its expression.In contrast,overexpression of periostin gene significantly increased TC,TG,and LDL-C levels and significantly decreased HDL-C levels（P<0.05）,indicating that it promotes expression of Ang Ⅱ,which in turn affects lipid metabolism.3.Ang Ⅱ can promote the formation of AS plaques:compared with HD+Ang Ⅱ group,there were amount of lipid deposition was observed in the aorta,indicating that Ang Ⅱ is related to AS plaque formation4.Periostin can affect AS plaque formation:Compared with mice in the HD+Ang Ⅱ group,lipid deposition can still be seen in the silent group,but the AS plaque lipid deposition in the aorta is reduced,while in the overexpression group,the AS plaque area can be increased and the aortic intima thickens.5.Periostin can make liver fat lesions more significant:compared with HD+Ang Ⅱ group,HE staining showed that the basic structure of liver tissue was observed in silent group;Oil red O staining showed that there are a little lipid deposition in hepatocytes.In overexpression group,the basic structure of liver tissue disappeared completely in HE staining,and the cell arrangement was faded away.Oil red O staining revealed a large number of unevenly distributed lipid droplets deposited on liver cells.6.Ang Ⅱ could promote the expression of periostin:Western blot of liver tissue showed an increased expression of periostin protein in HD+Ang Ⅱ group compared to HD group（P<0.05）.7.Ang Ⅱ could promote the increase of EL/HL protein expression:compared with ND and HD groups,EL/HL protein expression level was increased in HD+Ang Ⅱ group（P<0.05）,and Ang Ⅱ upregulates EL/HL expression.8.Periostin participated in Ang Ⅱ mediated EL/HL expression in ApoE^-/-mice,and inhibited RCT:compared with HD+Ang Ⅱ group,the content of EL/HL protein in liver tissue of HD+POSTN mus-769+Ang Ⅱ group was reduced（P<0.05）.The content of EL/HL protein in over-expressed periostin group was reduced（P<0.05）.9.Ang Ⅱ can further adjust EL/HL by adjusting the expression of periostin through p38MAPK signaling pathway:Compared with HD group,the expression contents of p-p38MAPK,periostin and EL/HL in HD+Ang Ⅱ group were reduced（P<0.05）,suggesting that Ang Ⅱ can be expressed on the p38MAPK pathway.Compared with the HD+Ang Ⅱ group,the trends of p-p38MAPK,EL/HL and periostin expression in the silent periostin group and overexpressed periostin were consistent,and there was a positive correlation between p-p38MAPK,periostin,and EL/H.10.RT-PCR results showed that the mRNA content of EL/HL,periostin and p-p38MAPK increased with the expression of Ang Ⅱ,and silent periostin could inhibit the expressions of EL/HL and p-p38MAPK.Overexpressed periostin can promote the expression of EL/HL and p-p38MAPK,indicating that silent periostin can positively regulate reverse cholesterol transport and inhibit Ang Ⅱ expression in p38MAPK pathway,and overexpressed periostin can negatively regulate reverse cholesterol transport,promote the expression of Ang Ⅱ in p38 MAPK pathway,and further promote AS.Conclusion:1.High-fat diet can establish ApoE^-/-atherosclerosis model.2.Ang Ⅱ can increase periostin level,and silent periostin can promote RCT,and overexpression can inhibit RCT.3.periostin participates in Ang Ⅱ-mediated EL/HL expression in ApoE^-/-mice.4.Ang Ⅱ can promote the regulation of EL/HL in the p38MAPK signal transduction pathway by modulating periostin,and aggravate AS.