The Study Of The Effect Of LKB1 On Atherosclerosis And Its Mechanism

BACKGROUNDAtherosclerosis is a disease with the highest morbidity and mortality in the world,and is closely related to cardio-cereovascular diseases.Unfortunately,its incidence rate is increasing year by year,and has a younger trend,which brings a heavy burden to human society.With the increase in the number of patients and the change in the structure of patients,the application of traditional treatments gradually restricted.Therefore,it is of great social significance to find more targeted treatment strategies.Thanks to the application of genetic techniques such as pedigree tracing,more and more studies have shown that vascular smooth muscle cells（VSMCs）are involved in the whole process of atherosclerosis,and the phenotypic transformation under the stimulation of various factors is an important pathogenesis of atherosclerosis.As a well-known tumor suppressor gene,LKB1 has also been found to have an important impact on cardiovascular disease in recent years,but it is not clear whether LKB1 can participate in the occurrence and development of atherosclerosis by affecting VSMCs.OBJECTIVETo explore the effect and mechanism of LKB1 on vascular smooth muscle cells in atherosclerosis.METHODSThirty 8-week-old male Apo E^-/-mice were divided into control group and overexpressed group randomly.The model of VSMCs-specific LKB1 overexpression was established by injecting empty adeno-associated virus（AAV）and overexpressed LKB1 AAV.The mice were fed with high-fat diet for 8 weeks,and the body weight of the mice was weighed and recorded every Tuesday and Friday.Serum was taken at the beginning and end of the experiment to detect the level of blood lipids.Aortic specimens were taken to make pathological sections and carry out follow-up experiments:Oil red O staining and HE staining were used to observe the formation of atherosclerotic plaques;the expression of LKB1in aorta was detected by fluorescence quantitative PCR and Western Blot;the expressions of AMPK,ABCA1,α-SMA,OPN,MMP-2,and CD68 in plaques were detected by Western Blot,the co-expression ofα-SMA and CD68 in plaques were detected by immunofluorescence double staining.RESULTS（1）The expression levels of m RNA and protein of LKB1 in vascular specimens of overexpression group were significantly higher than those of control group.（2）There was no significant difference in body weight and blood lipid level between the two groups before and after the experiment（P>0.05）.（3）Oil red O staining showed that the pathological changes of aorta in the overexpression group were slighter than the control group,and the lipid accumulation was not obvious.A large area of pathological changes and obvious lipid accumulation were found in the aortic sections of mice in the control group,and there was significant difference between the two groups（P<0.05）.HE staining showed that the degree of atherosclerosis in the overexpression group was slighter than that in the overexpression group.Compared with the overexpression group,the vascular lumen in the control group was narrowed,the intima was continuously destroyed,foam cells could be seen under the intima,the media thickened obviously,and the smooth muscle cells proliferated and arranged disorderly.（4）Western Blot showed that compared with the control group,the expression ofα-SMA of overexpression group increased,while the expression of OPN,MMP-2 and CD68 decreased significantly（P<0.05）.The results of immunofluorescence double staining showed that compared with the overexpression group,there was obvious expression ofα-SMA in the whole layer of vascular specimens in the control group,and obvious CD68 positive cells could be seen in the expression site ofα-SMA.The results of Image Pro Plus software analysis showed that there was a clear co-localization relationship betweenα-SMA and CD68 in the control group,but not in the overexpression group.（5）Western Blot showed that there was no significant difference in AMPK protein expression level between the two groups（P>0.05）.CONCLUSIONVSMCs participate in the pathophysiological process of atherosclerosis through phenotypic transformation,and LKB1 can delay the occurrence and development of atherosclerosis by inhibiting phenotypic transformation of VSMCs.In addition,our study also found that LKB1 may inhibit the formation of atherosclerosis independently of AMPK.