Each participant signed the informed consent that was part of the protocol approved by the National Eye Institute (NEI) Institutional Review Board, the AREDS clinical site, or the University of Sydney. All human research adhered to the tenets of the Declaration of Helsinki.
The clinic-based NEI case–control study samples included 103 clinically diagnosed advanced AMD cases and 137 age-matched controls from the greater Washington, DC, area who were evaluated by the AREDS ophthalmologists at the NEI.
19 20 35 In addition, DNA samples of 488 subjects (296 with advanced AMD and 192 control subjects) from the AREDS Genetic Repository were obtained and included as the second clinic-based sample in the study. NEI and AREDS patients and control subjects were self-identified as whites of non-Hispanic descent. Because AREDS participants were recruited under criteria identical with the NEI subjects, we combined all subjects from the two sources, to reach an adequate sample size for the study (NEI+AREDS). NEI+AREDS included 172 subjects with geographic atrophy, 227 subjects with neovascular AMD, and 329 control subjects.
The patients with AMD in the NEI and AREDS case–control studies all were diagnosed with the advanced stage of the disease as defined by the guidelines of the AREDS study, based on fundus photographs.
35 Patients with advanced AMD had geographic atrophy of at least 175 μm in diameter, involving the center of the macula and/or choroidal neovascularization defined as nondrusenoid retinal pigment epithelial (RPE) detachment, serous or hemorrhagic retinal detachment, hemorrhage under the retina or the RPE, or subretinal fibrosis, with drusen in at least one eye.
35 The normal control subjects were clinically evaluated showing an absence of drusen or less than five small drusen (<63 μm), no evidence of significant extramacular drusen, and an absence of all other retinal disease affecting the photoreceptors and/or outer retinal layers such as high myopia, retinal dystrophies, central serous retinopathy, vein occlusion, diabetic retinopathy, uveitis, and other retinal diseases. Retinal photographs were obtained in all patients and the AREDS control subjects. Smoking data were collected for AREDS patients by a detailed administered questionnaire.
3 Smoking data for the NEI patients were collected similarly. However, the questionnaire contained only three questions that asked whether the study participants had ever smoked, if they currently smoked, and, if so, how often. Demographic information on all study groups is summarized in
Table 1 .
Collection and clinical evaluation of subjects in the BMES has been described.
36 37 Briefly, the BMES is a population-based cohort study of common eye diseases and health-related parameters among suburban residents aged 49 years or older in the Blue Mountains region of Australia, near Sydney. The area has a stable and ethnically homogenous population. Retinal photographs were taken of at least one eye in 99%, or of both eyes in 98%, of study participants. During the second survey of the BMES (1997–2000), 2334 of 3654 baseline participants (75% of survivors), together with 1174 (85% of newly eligible) persons who moved to the area and who were within the eligible age group at the time, were examined and photographed. From this second survey population of the BMES (
n = 3508), AMD was diagnosed by grading the retinal photographs, with the graders masked to the participant’s identity. All advanced AMD cases were adjudicated and confirmed by a retinal specialist (PM). Early AMD was defined in either eye from (1) the presence of large (125 μm or larger in diameter), soft indistinct or reticular drusen within the macular area or (2) the presence of both large soft distinct drusen within the macula and retinal pigment abnormalities in the absence of late AMD. This closely followed the definition of early AMD used in the Beaver Dam, Wisconsin, population.
38 Advanced (late) AMD was defined as the presence of neovascular or atrophic AMD. Neovascular AMD was defined as serous or hemorrhagic detachment of the sensory retina or retinal pigment epithelium (RPE), the presence of subretinal or sub-RPE hemorrhages, or subretinal fibrous scar tissue. Geographic atrophy was defined as a discrete area greater than 175 μm in diameter characterized by a sharp border and the presence of visible choroidal vessels.
36 Smoking history was obtained with an interviewer-administered questionnaire. Participants were asked whether they had ever smoked and, if so, at what age they commenced smoking, as well as whether and when they had quit smoking.
39 DNA samples of 851 subjects consisting of 283 cases and 568 control subjects were sent to the NEI for genotyping. After 16 subjects who could not be genotyped were excluded, 835 subjects including 278 cases (54 late and 224 early AMD) and 557 age-, sex-, and smoking status–matched control subjects were included in the analyses.