Expression Of E-FABP And FAS In Atherosclerotic Plaques Of Human Femoral Arteries

Background:Due to the improvement of living standards and an aging population, critical lower limb ischemia associated with arteriosclerosis obliterans(ASO) has frequently been encountered. The main clinical manifestations of ASO are intermittent claudication, rest pain, ischemic ulcers or even gangrene. In addition,since severe complications often develop, the current teatmeat for ASO included medical therapy, surgery, gene and stem cell transplantation, is not optimistic. The pathophysiological mecanism of ASO has remained unclear,the main theory including damage response theory, lipid infiltration theory, mononuclear macrophage function theory and so on. Recent studies suggest that arteriosclerosis (AS) is the chronic diseases caused by lipids disorder and inflammatory response.Actively study the pathophysiological mecanism of ASO, is of great significance to the early detection, early diagnosis and early treatment which is important to decrease the amputation rate and improve the life quality of patients.Epidermal fatty acid binding protein (E-FABP),as one subtype of the fatty acid binding protein (FABPs),consists of small cytoplasmic proteins that bind long-chain fatty acid, facilitate their solubilization and transport to the cell,which plays an important role in the metabolism of long chain fatty acids. Some studies indicate that the adipocyte fatty acid binding protein (A-FABP) is present in large amounts in human carotid atherosclerotic plaques, especially in macrophages. E-FABP is highly homologous to A-FABP.Fatty acid synthase (FAS) is the key enzyme to catalyze the synthesis of long chain saturated fatty acids, studies show that the increased expression of E-FABP may be related to a higher mobilization of long chain fatty acids to the intracellular metabolic sites in the breast cancer.In the Femoral artery atherosclerotic plaque, whether or not the change of the expression E-FABP is associated with the expression of FAS, has not yet been illustrated.Objective:To explore the expressions and significance of Epidermal Fatty Acid Binding Protein (E-FABP) and Fatty Acid Synthase(FAS) in atheroselerotic plaques of human femoral arteries.Methods:1.We examined the femoral arterial atheroselerotic plaques of21men(n=12) and women(n=9) who had underwent the femoral intima-ablative in the Second Affiliated Hospital of Zhengzhou University between Novermber2010and May2011.The subject’s average age56.73±6.23years.All cases confirmed by color Doppler ultrasound diagnosis and lower artery angiopraphy.Paired samples of human splenic arterial tissues were obtained from13patients(8men,5women, age52.12±3.41years) who undergoing splenectomy for traumatic rupture of spleen. All the volunteers had been invited to attend a clinical examination and to provide a fasting blood sample.There was no clinical or analytical evidence of tumor, systemic inflammatory disease, autoimmune disease.All subjects who were informed the relevant consent,signed the "femoral artery atherosclerosis research informed consent form". All specimens got rid of remained bloodstains simply when obtained in surgery. And the specimen have be divided into two parts.one parts were fixed by10%formalin,while the other part were immediately frenzen in liquid nitrogen,and stored at-80℃refrigerator.2.The human femoral arterial atheroselerotic plaques were divided into stable(n=9) and unstable(n=12) plaques according to HE staining,and13cases normal splenic arterial specimens as control group. 3.The expressions of E-FABP and FAS protein were detected by immunohistochemical SP staining method.The change and correlations of expression of E-FABP and FAS protein were analyzed respectively in plaques.4. The expressions of E-FABP and FAS mRNA were detected by Reverse Transcription-Polymerase Chain Reaction(RT-PCR).The change and correlations of expression of E-FABP and FAS mRNA were analyzed respectively in plaques.5.Statistical analysis:The statistical analyses and graphics were performed using the program SPSS17.0.Data are expressed as mean士standard deviation(x±s). The one way analysis of variance(ANOVA) were used for comparisons between groups, and the sattistical significance of differences between the two groups was assessed with the least—significant difference(LSD) test.Pearson’s correlation coefficients were calculated to estimate the linear correlation. A value of a less than0.05was considered sattistically significant.Results:1.The pathmorphological observationHE staining show that three layer of splenic arterial tissues (intima,media,adventitia) were intact, with the visible internal and external elastic,cell arranged in linear array in control group.The cells of unstable plaques group arranged in disorder and endothelial cells were not complete.The lipid core more than40%of plaques area which were eccentric type were observed.There are also amount of foam cells,inflammatory cells, cholesterol crystal and neovessels in plaques,with the thin fibrous cap.The lipid core of stable plaques less than40%of plaques or no lipid core,which had a thickening fibrous cap, foam and inflammatory cells were infrequent and calcification can be found.2.Immunohistochemistry results2.1The amount of E-FABP in unstable and stable plaques were higher than those in control group.The immunopositive staining of E-FABP mainly located in the area infiltrated by inflammatory cells and foam cells.The differences for expression of E-FABP in three group were statistical significance (F=35.554, P<0.001).Compared with control group (1.950±0.069),the protein of E-FABP expressions were higher in atherosclerotic plaques (P<0.001). The expressions of E-FABP protein in unstable plaques group (2.155±0.060) were higher than those in stable plaques group (2.081±0.051)(P=0.011)2.2The amount of FAS in unstable and stable plaques were higher than those in control group.The immunopositive staining of FAS mainly located in the area infiltrated by inflammatory cells and foam cells.The differences for expression of FAS in three group were statistical significance (F=125.843, P<0.001).Compared with control group (1.971·0.038),the protein of FAS expressions were higher in atherosclerotic plaques(P<0.001).The expressions of FAS protein in unstable plaques group (2.170±0.028) were higher than those in stable plaques group (2.083±0.024)(P<0.001).2.3There was a positive correlation between the expressions of E-FABP and FAS protein in atherosclerotic plaques (r unstable group=0.907,P<0.001; r stable group=0.675,P=0.046), and there was no correlation in control group (r=0.138,P=0.654)3.RT-PCR results3.1The differences for expression of E-FABP mRNA in three group were statistical significance (F=11.142, P<0.001). Compared with control group (1.156±0.144).the protein of E-FABP mRNA expressions were higher in atherosclerotic plaques (P<0.001).The expressions of E-FABP mRNA in unstable plaques group (1.593±0.312) were higher than those in stable plaques group (1.365±0.206)(P=0.033)3.2The differences for expression of FAS mRNA in three group were statistical significance (F=11.176, P<0.001). Compared with control group (1.103±0.149).the protein of FAS mRNA expressions were higher in atherosclerotic plaques (P<0.001).The expressions of FAS mRNA in unstable plaques group (1.423±0.201) were higher than those in stable plaques group (1.260±0.147)(P=0.037)3.3There was a positive correlation between the expressions of E-FABP and FAS mRNA in atherosclerotic plaques (r unstable group=0.650,P=0.022; r stable group=0.778,P=0.014), and there was no correlation in control group (r=0.120,P=0.696).Conclusion:1.The expressions of E-FABP and FAS in femoral arteries atherosclerotic plaques were up-regulation.2. The enhanced expressions of E-FABP and FAS in atherosclerotic plaques of human femoral arteries may be associated with the instability of plaques.