Assessment of Proteins Associated With Complement Activation and Inflammation in Maculae of Human Donors Homozygous Risk at Chromosome 1 CFH-to-F13B
Assessment of Proteins Associated With Complement Activation and Inflammation in Maculae of Human Donors Homozygous Risk at Chromosome 1 CFH-to-F13B
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DOI：
10.1167/iovs.15-17009
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发表时间：
2015-07-01
影响因子：
4.4
通讯作者：
Hageman, Gregory S.
中科院分区：
文献类型：
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作者：
Keenan, Tiarnan D. L.;Toso, Marc;Hageman, Gregory S.
目的.研究1号染色体基因型和吸烟对人黄斑区补体激活和炎症的影响。供体黄斑组织根据AMD相关CFH-至-F13 B基因座的双倍型分为三组：纯合“风险”（n = 9，56-78岁）、纯合中性（n = 2，64-79岁）和纯合“保护”（n = 6，61-78岁）双倍型。重要的是，所有供体在ARMS 2/HTRA 1基因座上均为纯合子无风险，因此检查了纯1号染色体定向途径。通过使用14种抗体进行免疫组织化学，主要是针对补体和炎症的标志物，然后进行共聚焦显微镜和免疫荧光定量（均对供体状态进行掩蔽）。与纯合子受保护供体相比，CFH至F13 B纯合子风险供体在黄斑布鲁赫膜（BM; P = 0.03）、脉络膜毛细血管（CC; P = 0.04）和脉络膜毛细血管间隔（CC IS; P = 0.03）中显示出显著更高水平的末端补体复合物（TCC）。吸烟与BM（P = 0.05）、CC IS（P = 0.03）和脉络膜基质（CS; P = 0.01）中TCC增加相关，与RPE（P = 0.04）、BM（P = 0.01）、CC（P = 0.05）和CS（P = 0.05）中C反应蛋白（CRP）水平显著升高相关。吸烟与黄斑部RPE（P = 0.04）和CS（P = 0.01）中较高水平的氧化应激相关。CFH-至-F13 B基因座的遗传风险与人黄斑RPE-脉络膜界面的较高水平的补体激活相关，吸烟也是如此。有吸烟史的高危献血者CRP水平显著升高。对来自具有“纯”二倍体型供体的人黄斑组织的检查允许评估仅由CFH至F13 B驱动的AMD相关途径。这些发现对于确定1号染色体导向的通路和治疗靶点具有重要意义。
PURPOSE. To determine the effects of chromosome 1 genotype and cigarette smoking on levels of complement activation and inflammation in the human macula.METHODS. Donor macular tissue was stratified into three groups by diplotype at the AMD-associated CFH-to-F13B locus: homozygous "risk" (n = 9, 56-78 years), homozygous neutral (n = 2, 64-79 years), and homozygous "protective" (n = 6, 61-78 years) diplotype. Importantly, all donors were homozygous nonrisk at the ARMS2/HTRA1 locus, so that purely chromosome 1-directed pathways were examined. Immunohistochemistry was performed by using 14 antibodies, mostly against markers of complement and inflammation, followed by confocal microscopy and immunofluorescence quantification (all masked to donor status).RESULTS. Donors homozygous risk at CFH-to-F13B exhibited significantly higher levels of terminal complement complex (TCC) in macular Bruch's membrane (BM; P = 0.03), choriocapillaris (CC; P = 0.04), and choriocapillaris intercapillary septa (CC IS; P = 0.03), compared to homozygous protected donors. Smoking was associated with increased TCC in BM (P = 0.05), CC IS (P = 0.03), and choroidal stroma (CS; P = 0.01), and with substantially elevated C-reactive protein (CRP) levels in RPE (P = 0.04), BM (P = 0.01), CC (P = 0.05), and CS (P = 0.05). Smoking was associated with higher levels of oxidative stress in macular RPE (P = 0.04) and CS (P = 0.01).CONCLUSIONS. Genetic risk at the CFH-to-F13B locus was associated with higher levels of complement activation at the human macular RPE-choroid interface, as was cigarette smoking. Levels of CRP were substantially elevated in risk donors with smoking history. Examination of human macular tissue from donors with "pure" diplotypes allows assessment of AMD-associated pathways driven solely by CFH-to-F13B. These findings have important implications for identifying chromosome 1-directed pathways and therapeutic targets.