Apolipoprotein E genotype and macrophage inflammatory status: Impact of antioxidant/anti-inflammatory dietary components

心血管疾病(CVD)包括冠心病和中风，在欧盟每年约有190万人死亡，占总死亡率的42%。个人患病的风险由环境(运动、吸烟、药物、饮食等)和遗传因素共同决定。我们DNA中包含的基因变异(多态)携带了制造体内所有蛋白质所需的信息，可以影响合成蛋白质的结构或产生的水平。这可能会随后影响身体新陈代谢和心血管疾病等疾病的风险。一种这样的多态是apoE多态，通常观察到该基因有3个版本，即E2、E3或E4。载脂蛋白E4基因携带者(E3/E4或E4/E4，占英国人口的25%)与常见的E3/E3(占英国人口60%)基因相比，发生心血管疾病的可能性高约50%。然而，这种风险增加的生理原因却鲜为人知。我们和其他人主要使用实验室中培养的细胞(细胞培养技术)产生的初步数据表明，apoE蛋白可能在体内起到天然抗氧化剂的作用，E4版本的抗氧化能力明显低于E3或E2版本。低抗氧化状态被认为是导致氧化应激的原因，氧化应激促进了心血管疾病的发展。此外，我们到目前为止进行的研究表明，E4蛋白的这种低抗氧化能力可能会影响巨噬细胞的行为方式。巨噬细胞是一种白细胞，它会在我们的动脉壁上积聚胆固醇形成斑块，这是动脉粥样硬化(动脉结垢)的主要特征。除了积累胆固醇外，巨噬细胞还会产生一系列称为炎性化合物的物质，加速动脉粥样硬化的进程。由于巨噬细胞在体内产生高达20%的载脂蛋白E，并且已知对氧化应激敏感，因此推测E4个体心血管疾病风险增加的部分原因是基因对巨噬细胞功能的影响。这项研究的主要目的是调查这一假说。此外，还将调查饮食中存在的一系列物质对E4基因对巨噬细胞的拟议负面影响的抵消能力。简而言之，将使用已插入人类E3或E4基因的小鼠巨噬细胞进行一系列细胞培养实验。这些细胞将在模拟体内条件的条件下生长，并将研究基因对巨噬细胞代谢的许多方面的影响。人类的饮食中含有许多天然的抗氧化剂或发炎剂，如维生素C、鱼油脂肪和水果和蔬菜中的类黄酮类。将进行第二系列实验，以确定这些化合物是否可以抵消E4基因对巨噬细胞功能的负面影响。最后，人类志愿者将被分型，10名E3/E3、E3/E4和E4/E4个体将被要求提供血液样本，从中分离单核细胞(前巨噬细胞集落刺激因子)。将进行最后一系列测试，以确保我们使用方便的小鼠细胞进行的实验中的主要发现在使用人类细胞时也是明显的。从科学的角度来看，本研究的成果将扩大我们目前对载脂蛋白E4基因导致心血管疾病风险的机制的认识。从公共卫生的角度来看，这项研究将调查天然饮食成分在一定程度上抵消载脂蛋白E4个体增加的疾病风险的能力。如果要将更广泛的人群基因图谱作为减少或推迟慢性病发病的政策，那么抵消“高危”基因所带来的风险增加的策略(饮食或其他生活方式)是必不可少的。

Cardiovascular disease (CVD), which includes coronary heart disease and stroke, accounts for about 1.9 million deaths per year in the EU, which represents 42% of total mortality. An individual's risk of disease is determined by both environmental (exercise, smoking, medication, diet etc) and genetic factors. Variations in genes (polymorphisms), contained within our DNA, which carry the information necessary for the manufacture of all proteins in the body, can affect both the structure of the resultant protein or the levels produced. This may subsequently impact on body metabolism and risk of diseases such as CVD. One such polymorphism is the apoE polymorphism with 3 versions of the gene i.e. E2, E3 or E4, commonly observed. Carriers of the apoE4 version (E3/E4 or E4/E4, 25% of the UK population) are about 50% more likely to develop CVD compared to the common E3/E3 (60% UK population) genotype. However, the physiological reasons for this increased risk are poorly understood. Preliminary data generated by ourselves and others, mainly using cells grown in the laboratory (cell culture techniques), suggest that the apoE protein may act as a natural antioxidant in the body, with the antioxidant capabilities of the E4 version significantly lower than either the E3 or the E2 version. Low antioxidant status is known to contribute to oxidative stress which promotes the development of CVD. Furthermore our studies conducted thus far are suggestive that this low antioxidant capacity of the E4 protein may affect the way macrophages behave. Macrophages are a type of white blood cell, which accumulate cholesterol in our artery walls to form a plaque, which is a main feature of atherosclerosis ('furring' of the arteries). In addition to accumulating cholesterol macrophages produce a range of substances, called inflammatory compounds, which speed up the atherosclerotic process. As macrophages produce up to 20% of apoE in the body, and are known to be sensitive to oxidative stress it is hypothesised that the increased CVD risk in E4 individuals is in part due to an impact of genotype on macrophage function. The main aim of the study is to investigate this hypothesis. Furthermore the ability of a range of substances present in the diet to counter-act the proposed negative effect of the E4 genotype on macrophages will be investigated. In brief, a range of cell culture experiments will be conducted using mouse macrophages, into which the human E3 or E4 gene has been inserted. The cells will be grown under conditions which simulate conditions in the body, and the effect of genotype on many aspects of macrophage metabolism will be investigated. The human diet contains many components which act as natural antioxidant or inflammatory agents, such as vitamin C, fish oil fats, and flavonoids found in fruit and vegetables. A second series of experiments will be conducted to ascertain if these compounds can counteract the negative effect of the E4 genotype on macrophage function. Finally human volunteers will be genotyped, and ten E3/E3, E3/E4 and E4/E4 individuals will be asked to provide a blood samples from which monocytes (pre-macrohpages) will be isolated. A final series of tests will be conducted to ensure that the main findings in our experiments using the convenient mouse cells, are also evident using human cells. From a scientific point of view the output from the current study will expand our current knowledge regarding the mechanisms by which an apoE4 genotype contributes to CVD risk. From a public health viewpoint the study will investigate the ability of natural dietary components to in part counteract the increased disease risk observed in apoE4 individuals. If more widespread population genetic profiling is going to be adopted as a policy to reduce or delay the onset of chronic diseases, then strategies (dietary or other lifestyle) to counteract the increased risk posed by an 'at-risk' genotype are essential.