Effect Of Different ApoE Genotype And 5XFAD Genetic Background On Atherosclerosis And Expression Of Inflammatory Molecules In Mouse Aorta

[Objective] Apolipoprotein E(Apo E), which is one of the important human plasma lipoproteins, is closely related to lipid metabolism. Aop E gene polymorphism is involved in a variety of diseases, including atherosclerosis(AS) and alzheimer’s disease(AD). This study is aimed at investigating the effect of different Apo E genotype on serum lipid, aortic atherosclerosis and the expression of inflammatory molecules, such as Intercelluar Adhesion Molecule-1(ICAM-1), Vascular Cell Adhesion Molecule-1(VCAM-1) and Monocyte Chemoattractant Protein-1(MCP-1)in 5XFAD or non-5XFAD transgenic mice, and exploring the possible mechanism involved in MAPKs signaling pathways.[Methods] The SPF transgenic male mices were divided into control group(C57BL/6J), the Apo E-TR group(Apo E-KO, Apo E2, Apo E3, Apo E4), the Apo E-TR/5XFAD group(Apo E-KO/5XFAD, Apo E2/5XFAD, Apo E3/5XFAD,Apo E4/5XFAD). Animals were fed with normal diet for 12 months. Serum total cholesterol(TC), triglyceride(TG), low density lipoprotein cholesterol(LDL-C), high density lipoprotein cholesterol(HDL-C) were measured with enzymic method. The aorta(from aortic root to aorta arch) frozen sections were stained with hematoxylin-eosin(HE staining) and Sudan IV to observe and detecte histologic characteristics and pathological changes of atherosclerosis plaque. Aortic intimal area were analyzed by Image Pro Plus; Intraaortic inflammatory molecules(VCAM-1,MCP-1) were detected with immunofluorescence stain. ICAM-1, VCAM-1 protein expression, MAPKs(ERK1/2, JNK, P38) phosphorylation levels were determined by western blot.[Results]1. Blood lipid levels:(1) Serum TC, TG, and LDL-C levels increases significantly in Apo E-/- and mice compared with control group, which was 8.3, 4.9, 26.1 and 19.5, 5.2, 31.0 folds higher than control group, respectively. Non-HDL-C level was 33.4(Apo E-/-), 54.0(Apo E-/-/ 5XFAD) folds higher than the wild type group respectively(P<0.05);(2) In Apo E2 mice and Apo E/5XFAD mice, serum TC, TG, LDL- C, non-HDL-C level was 6.3, 4.4, 16.3, 18.4 and 7.0, 2.8, 10.1, 12.4 folds higher than the control group mice respectively(P<0.05);(3) Among Apo E-TR group and Apo E-TR/5XFAD group, serum TC, TG, LDL-C,HDL-C, non-HDL-C level and TC/HDL-C ratio were not significantly different in E3 or E4 carrier mice compared to control group(P>0.05);(4) TC and non-HDL-C level in Apo E-/-/5XFAD mice were significantly higher than those in Apo E-/-mice, which was 2.3 and 1.6 folds, respectively(P<0.05).2.Atherosclerosis lesion in aorta:(1) There were obvious atherosclerotic plaques in the aortic arch of Apo E-/- andApo E-/-/5XFAD mice. Intimal area in both groups significantly increased compare to the control group, which was 58107.6±7339.0μm2(Apo E-/-), 96208.6±32032.5μm2(Apo E-/-/5XFAD) VS 18819.3±2162.7μm2(control), respectively(P<0.05);(2) Compared with control group, a small plaque formation were observed in Apo E2/5XFAD mouse, but there was no significant difference in intimal area(40078.7±25272.5μm2 VS 18819.3±2162.7μm2, P>0.05);(3) For Apo E3 and Apo E4 mice, there was no obvious atherosclerotic plaque in aorta,and no significant difference in the intima area compared with control group(P>0.05).3.The expression of inflammation molecules in aorta:(1) In Apo E-/-and Apo E-/-/5XFAD mice, ICAM-1 protein expression was higher than control group(P<0.05); However, with AD genes background or not, in Apo E-TR groups(Apo E2, Apo E3, Apo E4) and Apo E-TR/5XFAD groups(Apo E2/5XFAD,Apo E3/5XFAD, Apo E4/5XFAD), ICAM-1 protein was comparable to control group(P>0.05);(2) VCAM-1 protein expression increased significantly in Apo E-/- mice,Apo E-/-/5XFAD mice and Apo E2/5XFAD mice compared with control group(P< 0.05);Among Apo E-TR groups(Apo E2, Apo E3, Apo E4) and Apo E3/5XFAD,Apo E4/5XFAD mice VCAM-1 protein was comparable to control group(P > 0.05);(3) In aorta immunofluorescence staining, the fluorescence intensity of VCAM-1,MCP-1 were stronger among Apo E-/-, Apo E-/-/5XFAD and Apo E2/5XFAD mice.4. MAPKs phosphorylation level:(1) ERK1/2 phosphorylation level increased in Apo E-/- mice, Apo E2 mice and all Apo E-TR/5XFAD mice compared to control group(P<0.05);(2) JNK phosphorylation level only increased in Apo E-/- mice(P<0.05).(3) P38 phosphorylation level has no significant difference among all groups(P>0.05).[Conclusions]1.In Apo E-/- mice with/without AD genes background, there were much higher blood lipid level, more severe aortic atherosclerotic lesion/plaque, and higher expression of ICAM-1 and VCAM-1; Apo E2 genotype can cause the blood lipid disorder, but AD genetic background was needed for proatherosclerosis effect and evaluated inflammatory molecules. In Apo E3, E4 mice, there was no obvious lipid disorder, AS lesions and abnormal expression of ICAM-1 and VCAM-1.2. To some extend, 5XFAD genetic background worsen Lipid disorder in Apo E-/-mice,advanced AS lesion of Apo E-/- and Apo E2 mice, and increased the expression of inflammatory molecules.3. ERK1/2 pathway was activated in Apo E-/-, Apo E2 and Apo E-TR/5XFAD mouse aorta.