低浓度镉在kdrl：EGFP转基因斑马鱼血管粥样硬化中的作用及其机制研究

Role and mechanism of low concentration of cadmium in promoting atherosclerosis in kdrl:EGFP transgenic zebrafish

摘要:

目的 探究低浓度镉在kdrl：EGFP转基因斑马鱼血管粥样硬化中的作用及其机制。方法 选取18只健康转基因品系内皮细胞特异性绿色荧光［Tg（kdrl：EGFP）］成年斑马鱼复制动脉粥样硬化斑马鱼模型，按照不同处理方式分为A组（空白对照组）、B组（30 μmol/L镉）、C组（30 μmol/L镉+ 阿托伐他汀），每组6只。手术切取各组斑马鱼血管和斑块组织标本，采用荧光显微镜观察各组荧光斑马鱼血管胆固醇累积情况并测量血管内皮层厚度，苏木精-伊红（HE）染色观察各组斑马鱼血管斑块的病理学变化。分别采用实时荧光定量聚合酶链反应和Western blotting检测各组斑马鱼血管斑块组织HIF-1α、p-Akt、p-P70、VEGF mRNA和蛋白的表达。结果 B、C组血清镉浓度高于A组（P <0.05）。B组斑马鱼血管内皮层厚度较A、C组增厚（P <0.05），且血管中胆固醇积累明显。HE染色结果显示，与A、C组比较，B组炎症细胞浸润的脂肪变性更明显，且细胞核悬挂在中心或挤压到侧面。B组斑马鱼血管斑块组织HIF-1α、p-Akt、p-P70、VEGF mRNA和蛋白相对表达量均高于A、C组（P <0.05）。结论 低浓度镉可促进kdrl：EGFP转基因斑马鱼的动脉粥样硬化进程，相关机制与HIF-1α、p-Akt、p-P70、VEGF有关，阿托伐他汀可逆转镉的促AS进程。

Abstract:

Objective To investigate the role and mechanism of low concentration of cadmium in promoting atherosclerosis in kdrl:EGFP transgenic zebrafish.Methods Eighteen healthy adult zebrafish with endothelial cell-specific expression of enhanced green fluorescent protein [Tg (kdrl:EGFP)] were selected to establish zebrafish models of atherosclerosis. According to different treatment methods, they were divided into group A (blank control group), group B (30 μmol/L of cadmium) and group C (30 μmol/L of cadmium + atorvastatin), with six zebrafish in each group. Surgical removal of blood vessels and plaque tissues from each group of zebrafish was performed. The accumulation of cholesterol in the blood vessels of each group of zebrafish was observed using a fluorescence microscope, and the thickness of the vascular endothelium was measured. Hematoxylin-eosin (HE) staining was used to detect the pathological changes of plaques in the blood vessels of each group of zebrafish. The quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting were applied to detect the mRNA and protein expression levels of HIF-1α, p-Akt, p-P70 and VEGF in the plaques in the blood vessels of each group, respectively.Results Compared with the group A, the serum concentration of cadmium was higher in the group B and the group C (P < 0.05). Compared with the group A and the group C, the thickness of the vascular endothelium was significantly increased and there was a significant accumulation of cholesterol in the blood vessels of zebrafish (P < 0.05). The HE staining revealed that the group B exhibited more significant infiltration of inflammatory cells and steatosis, with the cell nucleus suspended in the center or squeezed to the side compared with the group A and the group C. The relative mRNA and protein expressions of HIF-1α, p-Akt, p-P70 and VEGF in plaques in the blood vessels of the group B were higher than those in the group A and the group C (P < 0.05).Conclusions Low concentrations of cadmium facilitates atherosclerosis in kdrl:EGFP transgenic zebrafish, and the underlying mechanisms are related to HIF-1α, p-Akt, p-P70 and VEGF. Atorvastatin could reverse the atherosclerosis promoted by the cadmium.