Study On The Mechanism Of Vav1/Rac1 Pathway In N~?-Carboxymethyl-Lysine Inhibiting Foam Cells Migration

Objective:To investigate the effect of Vav1/Rac1 pathway on the inhibition of foam cell migration by N^?-carboxymethyl lysine and its mechanism.Methods:?1?Clinical study:recruited patients from January 2015 to December 2017 in the department of orthopaedics of the affiliated hospital of jiangsu university?zhenjiang,China?,hospitalized patients with type 2 diabetes with below knee amputation?n=16?and patients with below knee amputation due to car accident?n=20?.Baseline data of age,gender,serum low-density lipoprotein,high-density lipoprotein,triglyceride,total cholesterol,fasting blood glucose and serum CML were calculated.HE staining and Masson staining were used to detect the size of vascular plaque and fibrous cap,and the phosphorylation degree of Vav1,Vav2 and Vav3 was detected by Western blot.Rac1,Cdc42 and RhoA were detected by G-lisa.?2?In vivo experiments:6-week-old ApoE^-/-mice were used to construct a diabetes model through STZ,and were divided into control group,model group,CML group,and anti-Vav1 group.The lipid deposition in blood vessels and paravascular lymph nodes was detected by oil red staining,and the expression of macrophages in blood vessels and lymph nodes was detected by CD68 immunofluorescence staining.?3?In vitro experiments:primary macrophages in the abdominal cavity were extracted by inducing aseptic inflammation in the abdominal cavity.Oil red staining was used to verify the effect of CML on foam cell formation.Vav1 siRNA was used to interfere with the expression of Vav1,Western blot and qRT-pcr were used to verify the interference efficiency.Cell migration was detected by scratch assay and Transwell assay.Peritoneal macrophage reflux assay was used to detect the migration ability of macrophages.Rac1,RhoA and Cdc42 were detected by G-lisa.Results:?1?Compared with the accident amputees,the levels of serum low-density lipoprotein,triglyceride,total cholesterol,fasting blood glucose and serum CML were significantly higher in diabetic amputees.?2?diabetes leads to the increase of atherosclerotic plaque,the thinning of fibrous cap,the increase of CML in serum,the increase of phosphorylation level of Vav1 protein,and the increase of Rac1 activity.?3?compared with the model group,the accumulation of foam cells in the vascular plaques of diabetic atherosclerosis mice was significantly increased,and the accumulation of lipid in the paravascular lymph nodes was decreased.Vav1 antibody inhibited intraplate accumulation and promoted foam cell flow to paravascular lymph nodes.?4?CML promoted the formation of foam cells derived from primary abdominal macrophages and inhibited the migration of foam cells.CML promoted the phosphorylation of Vav1 in foam cells,inhibited the expression of Vav1 and restored the migration ability of foam cells.?5?Rac1 activity was increased in CML stimulated foam cell migration,and inhibition of Rac1 activity promoted the recovery of foam cell migration ability.?6?ghost peptide staining showed that CML promoted the reduction of plate-like pseudopodia in foam cells,and Vav1 and Rac1 interference could respectively restore plate-like pseudopodia.Conclusion:?1?Vav1/Rac1 pathway is activated in diabetic atherosclerotic plaques;?2?Effect of Vav1 on the progression of diabetic atherosclerosis by affecting foam cell migration;?3?Activating the Vav1/Rac1 pathway,CML inhibits the formation of plate-like pseudopodia,thus inhibiting the migration of foam cells.