Research
Original Investigation | OPHTHALMIC MOLECULAR GENETICS
Challenges in Elucidating the Genetics of Diabetic Retinopathy Jane Z. Kuo, MD, PhD; Tien Y. Wong, MD, PhD; Jerome I. Rotter, MD
IMPORTANCE In the past decade, significant progress in genomic medicine and technologic
developments has revolutionized our approach to common complex disorders in many areas of medicine, including ophthalmology. A disorder that still needs major genetic progress is diabetic retinopathy (DR), one of the leading causes of blindness in adults. OBJECTIVE To perform a literature review, present the current findings, and highlight some key challenges in DR genetics. DESIGN, SETTING, AND PARTICIPANTS We performed a thorough literature review of the genetic factors for DR, including heritability scores, twin studies, family studies, candidate gene studies, linkage studies, and genome-wide association studies (GWASs). MAIN OUTCOME MEASURES Environmental and genetic factors for DR. RESULTS Although there is clear demonstration of a genetic contribution in the development and
progression of DR, the identification of susceptibility loci through candidate gene approaches, linkage studies, and GWASs is still in its infancy. The greatest obstacles remain a lack of power because of small sample size of available studies and a lack of phenotype standardization. CONCLUSIONS AND RELEVANCE The field of DR genetics is still in its infancy and is a challenge
because of the complexity of the disease. This review outlines some strategies and lessons for future investigation to improve our understanding of this complex genetic disorder. JAMA Ophthalmol. 2014;132(1):96-107. doi:10.1001/jamaophthalmol.2013.5024 Published online November 7, 2013.
D
iabetic retinopathy (DR), an important microvascular complication of diabetes mellitus (DM), is a leading cause of visual impairment in adults 20 to 74 years of age.1 More than 93 million people worldwide have DR, 17 million of whom have proliferative diabetic retinopathy (PDR) and 28 million of whom have vision-threatening DR.2 This number will continue to escalate with an aging population, increasing obesity, and a rapidly progressing DM epidemic. More individuals, especially Hispanics, people of African descent, and Asians, will be vulnerable to blinding DR in coming years.3,4 There is clearly a need to develop strategies to identify at-risk individuals for early interventions. Compared with other major causes of visual loss, such as age-related macular degeneration,5 myopia6,7 and glaucoma,8,9 the search for genetic clues to DR has not progressed as rapidly. To date, the few studies that have reported on possible susceptibility genes for DR have yielded inconsistent results. There is clearly a familial relationship in DR because twin and family studies10-17 indicate a genetic basis. Several candidate gene studies have reported promising genes,18-21 but few of them have been replicated, and the few positive findings reveal only weak genetic associations.18-20,22 In genome-wide approaches, 3 linkage studies performed in Pima Indians and 96
Author Affiliations: Author affiliations are listed at the end of this article. Corresponding Author: Jerome I. Rotter, MD, Institute for Translational Genomics and Population Sciences, Los Angeles Bio Medical Research Institute, Harbor–UCLA Medical Center, 1124 W Carson St, Torrance, CA 90502 ([email protected]). Section Editor: Janey L. Wiggs, MD, PhD.
Mexican Americans have identified regions on chromosomes 1, 3, and 12 to be suggestive of DR.13,23,24 In contrast to age-related macular degeneration, myopia, and glaucoma, few genome-wide association studies (GWASs) have been conducted thus far on DR. The few GWASs are of modest sample sizes in Hispanics, Chinese, and white populations and have reported borderline associations with DR in either type 1 or type 2 DM.25-28 In this review, we highlight these key genetic studies of DR with an emphasis on the most recent developments. We also discuss issues and challenges with elucidating the genetics of DR and indicate approaches that will provide the opportunity to advance our knowledge of this complex genetic disorder.
Definition and Classification of DR The diagnosis of DR is clinically defined by the presence of retinal microvascular lesions in patients with DM; however, these retinal lesions are not specific and may also be present in individuals without DM.29,30 The classification of DR is graded by severity and divided into nonproliferative diabetic retinopathy (NPDR) and PDR. Key retinal changes in NPDR include micro-
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Diabetic Retinopathy Genetics
Original Investigation Research
aneurysms, hard exudates, cotton wool spots, intraretinal microvascular abnormalities, and venous beading; these further subdivide NPDR into mild, moderate, and severe forms. Key retinal changes in PDR include neovascularization of the optic disc or elsewhere, preretinal hemorrhage, or vitreous hemorrhage. On the other hand, clinically significant macular edema, which is graded as its own entity, can develop at any stage of the DR spectrum. Thus, the various classifications in DR grading, resulting in heterogeneity of DR phenotype, pose a significant challenge in genetic studies of DR. The assessment of DR via a standardized stereoscopic photograph has been proposed to overcome this issue, and more researchers have used this approach by adopting and grading DR using the Early Treatment Diabetic Retinopathy Study severity scale or a similar modification. Recently, the assessment of DR and diabetic macular edema via optical coherence tomography has been proposed as an imaging modality to better visualize the intraretinal morphologic changes in patients with DM31; however, the classification of DR and diabetic macular edema via optical coherence tomography has not been clearly defined or adopted for use.
Genetics of DM Diabetic retinopathy occurs on the background of DM. That genetic factors play a major role in the etiology of DM has long been appreciated because of ethnic differences in frequency, increased familial aggregation, and a markedly higher concordance in monozygotic vs dizygotic twins. This is true for each of the major subforms of DM (type 1 and type 2). To date, approximately 60 loci have been successfully identified for type 2 DM, of which only 3 were discovered before the GWAS era.32,33 Although most of these studies were performed in individuals of European descent, more recent studies of Asians,34 Hispanics,35 and African Americans35-37 have also demonstrated some level of associations for these signals, supporting the hypothesis that these signals (or the causal variants that are in high linkage disequilibrium with these signals) are likely common alleles that are widely distributed in the human population and each contributing a small effect on disease risk.32 These important discoveries through large collaborative efforts by GWAS approaches have led to substantial progress in the understanding of genetics in type 2 DM, leading to the identification of novel pathways, demonstrating mechanistic associations, and supporting prior epidemiologic studies.38 These findings have illustrated some important key lessons that are useful in genetic studies of DR. First, joining forces by international collaborative efforts is necessary to increase statistical power by increasing sample size.38 Second, both analysis by treating the phenotype as a dichotomous trait and analysis of a related, quantitative trait are useful.38 Third, connection of genetic findings with more defined physiologic parameters increases understanding.38 Elucidating the genetic basis of type 2 DM offers an ideal model to approach the genetic study of DR. However, it should be apparent that the phenotype of type 2 DM has several advantages, such as the ease of classification and readily available large samples of patients even without detailed assessment. jamaophthalmology.com
Environmental Factors for DR The origin of DR remains complex and poorly understood. Large epidemiologic studies2,39,40 have consistently demonstrated that the duration of DM and adequacy of glycemic control are 2 of the major contributors to the development and progression of DR. This was robustly documented in the Wisconsin Epidemiological Study of Diabetic Retinopathy (WESDR), which found that duration of DM was the strongest predictor for progression of DR, with prevalence of DR ranging from 17% in type 1 DM to 29% in type 2 DM for patients with DM for less than 5 years and increasing markedly to almost 100% for type 1 DM and 78% for type 2 DM in patients with DM for more than 15 years.41,42 In other landmark studies,2,39,40,43 such as the Diabetes Control and Complications Trial, UK Prospective Diabetes Study, the Action to Control Cardiovascular Risk in Diabetes (ACCORD) Eye Study, and Meta-Analysis for Eye Disease Study, intensive glycemic control was effective in reducing the rate of DR progression in both type 1 and type 2 DM. Other studies2,44,45 have demonstrated that blood pressure control is another risk modifier, although some studies43 did not support this finding. Recently, increasing evidence has supported an association between dyslipidemia and DR46; the Fenofibrate Intervention and Event Lowering in Diabetes Study47 and the ACCORD Eye Study43 demonstrated that reduction in lipids could also limit DR progression. Despite strong evidence of DR susceptibility, these environmental risk modifiers by themselves do not account for the complete risk susceptibility. First, this is exemplified by clinical observations that some individuals develop DR despite good glycemic control and short duration of disease, whereas others do not develop DR, even with poor glycemic control and longer duration of DM.48 Second, the strongest environmental factors (duration of DM and glycosylated hemoglobin) only explained approximately 11% of the variation in retinopathy risk in the Diabetes Control and Complications Trial.49,50 Similarly, a combination of glycosylated hemoglobin, blood pressure, and total cholesterol only explained approximately 10% of the variation in retinopathy risk in the WESDR,51 suggesting that the remaining approximately 90% of the variation in retinopathy risk is presumably explained by other risk factors. Finally, population studies indicate that retinopathy signs, such as microaneurysms, were detectable in 7% to 13% of patients without DM and patients in whom the glycosylated hemoglobin level was well below 5%.29 In addition, singlenucleotide polymorphisms (SNPs) associated with DM or hypertension were not associated with retinopathy in individuals without DM,52 suggesting that other risk factors, independent of hyperglycemia and DM, contribute to the development and progression of retinopathy signs similar to DR.
Genetic Factors for DR Attempts to identify gene(s) in the development of DR have been conducted during the past few decades. To date, these studies have been limited to twin studies,10 family studies,11-17 JAMA Ophthalmology January 2014 Volume 132, Number 1
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Research Original Investigation
Diabetic Retinopathy Genetics
Table 1. Summary of Familial Aggregation Studies in Diabetic Retinopathy Ethnicity (Study)
Type of Diabetes
Heritability
Evidence of Familial Aggregation
11
Strong evidence of familial clustering in PDR
12
1
White (FinnDiane study)
1
PDR, 0.52
Pima Indian
2
Any DR, 0.18
Modest evidence of familial clustering
13
South Indian
2
Strong evidence of familial clustering (OR = 3.37 in sibling of diabetes with and without DR)
14
Chinese
2
Strong evidence of familial clustering in any DR; DR diagnosed in 7.1% of siblings without DR vs 29.7% of siblings with DR
15
Mexican American
2
Strong evidence of familial clustering in severe DR (OR = 1.72) but not any DR
16
Multiethnic (FIND-Eye Study)a
2
Strong evidence of familial clustering
17
Any DR, 0.27 PDR, 0.25 Mexican American, 0.24
Figure. Heritability Estimates for Diabetic Retinopathy 1.00
Heritability of DR
0.75
0.52 0.50
0.27 0.25
0.25
0.24
0.18
0.00 DCCT, PDR
Pima Indian, Any DR
FIND-Eye Study, Any DR
FIND-Eye FIND-Eye Study, Study, PDR Mexican American
Study
DCCT indicates Diabetes Control and Complications Trial; DR, diabetic retinopathy; FIND-Eye Study, Family Investigation of Nephropathy and Diabetes–Eye Study; PDR, proliferative diabetic retinopathy.
candidate gene studies,18-22 linkage studies,13,23,24 and smallscale GWASs with modest sample sizes.25-28 In support of a genetic hypothesis of DR, several studies3,4 have found a discrepant rate of the prevalence of DR among US populations, with a significantly higher prevalence observed in Hispanics, African Americans, and Chinese Americans. Compared with whites, in African Americans other risk modifiers, such as duration of DM, glycemic control, and blood pressure, appear to account for the higher prevalence of DR, but these factors do not explain the higher prevalence seen in Hispanics,53-56 suggesting that other factors, including genetic factors, may influence susceptibility to DR.
Twin and Family Studies In twin studies, Leslie and Pyke10 found the same degree of severity in 35 of 37 concordant twins with type 2 DM (95%) compared with only 21 of 31 concordant twins with type 1 DM (68%). This early observation was extended by familial aggregation 98
Reference
Low evidence of increased risk of any DR; strong evidence of familial clustering for severe DR (OR = 3.1 in relatives of diabetic patients with and without DR)
Whites (DCCT)
Abbreviations: DCCT, Diabetes Control and Complications Trial; DR, diabetic retinopathy; FIND-Eye Study, Family Investigation of Nephropathy and Diabetes–Eye Study; FinnDiane, Finnish Diabetic Nephropathy; OR, odds ratio; PDR, proliferative diabetic retinopathy. a Mexican Americans, European Americans, African Americans, and American Indians.
studies, with siblings and relatives of diabetic patients with DR having as high as a 3-fold increased risk of DR compared with siblings and relatives of diabetic patients without DR11-17 (Table 1). This trend was seen in either type 1 or type 2 DM and across different ethnicities. Furthermore, evidence of a familial aggregation is more consistently seen in the presence of more severe retinopathy and less in the presence of any retinopathy, with heritability scores ranging from 18% to 27% for any DR13,17 and 25% to 52% for PDR12,17 in either type 1 or type 2 DM (Figure). Thus, as previously mentioned, it is important to not only provide a standardized assessment of DR but also compare those without DR with those with more severe stages of DR. The Family Investigation of Nephropathy and Diabetes– Eye study, in which nearly half of the 2368 diabetic patients are Mexican Americans, demonstrated that the heritability of any DR in this population of type 2 DM is as high as 24%.17
Candidate Gene Studies Most genetic research in DR has used the candidate gene approach. Several pathways and processes have been proposed to play an important role in the pathogenesis of DR, which led to the testing of a number of hypothesized candidate genes. Although a number of candidate genes and genetic variants have been proposed in the literature, few of them have been replicated, and the few positive findings only revealed weak genetic associations with DR.18,19 In this approach, the analysis compares the frequency of a particular genetic variant in individuals with (cases) or without (controls) DR. Several pathways and processes have been proposed, including the renin-angiotensin system, glucoseinduced pathways, vascular endothelial dysfunction, tissue matrix remodeling, and angiogenesis.18,19 Potential candidate genes involved in these pathways and processes include ACE, AGTR1, AGT, VEGF, AKR1B1, RAGE, GLUT1, APOE, MTHFR, PAI-1, ITGA2, PPAR-γ, and NOS3. Their associations or lack thereof with DR have been extensively documented in prior reviews.18-21 We summarize the most important findings with a focus on AKR1B1 and VEGF because of their biological implications.
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Research
Original Investigation | OPHTHALMIC MOLECULAR GENETICS
Challenges in Elucidating the Genetics of Diabetic Retinopathy Jane Z. Kuo, MD, PhD; Tien Y. Wong, MD, PhD; Jerome I. Rotter, MD
IMPORTANCE In the past decade, significant progress in genomic medicine and technologic
developments has revolutionized our approach to common complex disorders in many areas of medicine, including ophthalmology. A disorder that still needs major genetic progress is diabetic retinopathy (DR), one of the leading causes of blindness in adults. OBJECTIVE To perform a literature review, present the current findings, and highlight some key challenges in DR genetics. DESIGN, SETTING, AND PARTICIPANTS We performed a thorough literature review of the genetic factors for DR, including heritability scores, twin studies, family studies, candidate gene studies, linkage studies, and genome-wide association studies (GWASs). MAIN OUTCOME MEASURES Environmental and genetic factors for DR. RESULTS Although there is clear demonstration of a genetic contribution in the development and
progression of DR, the identification of susceptibility loci through candidate gene approaches, linkage studies, and GWASs is still in its infancy. The greatest obstacles remain a lack of power because of small sample size of available studies and a lack of phenotype standardization. CONCLUSIONS AND RELEVANCE The field of DR genetics is still in its infancy and is a challenge
because of the complexity of the disease. This review outlines some strategies and lessons for future investigation to improve our understanding of this complex genetic disorder. JAMA Ophthalmol. 2014;132(1):96-107. doi:10.1001/jamaophthalmol.2013.5024 Published online November 7, 2013.
D
iabetic retinopathy (DR), an important microvascular complication of diabetes mellitus (DM), is a leading cause of visual impairment in adults 20 to 74 years of age.1 More than 93 million people worldwide have DR, 17 million of whom have proliferative diabetic retinopathy (PDR) and 28 million of whom have vision-threatening DR.2 This number will continue to escalate with an aging population, increasing obesity, and a rapidly progressing DM epidemic. More individuals, especially Hispanics, people of African descent, and Asians, will be vulnerable to blinding DR in coming years.3,4 There is clearly a need to develop strategies to identify at-risk individuals for early interventions. Compared with other major causes of visual loss, such as age-related macular degeneration,5 myopia6,7 and glaucoma,8,9 the search for genetic clues to DR has not progressed as rapidly. To date, the few studies that have reported on possible susceptibility genes for DR have yielded inconsistent results. There is clearly a familial relationship in DR because twin and family studies10-17 indicate a genetic basis. Several candidate gene studies have reported promising genes,18-21 but few of them have been replicated, and the few positive findings reveal only weak genetic associations.18-20,22 In genome-wide approaches, 3 linkage studies performed in Pima Indians and 96
Author Affiliations: Author affiliations are listed at the end of this article. Corresponding Author: Jerome I. Rotter, MD, Institute for Translational Genomics and Population Sciences, Los Angeles Bio Medical Research Institute, Harbor–UCLA Medical Center, 1124 W Carson St, Torrance, CA 90502 ([email protected]). Section Editor: Janey L. Wiggs, MD, PhD.
Mexican Americans have identified regions on chromosomes 1, 3, and 12 to be suggestive of DR.13,23,24 In contrast to age-related macular degeneration, myopia, and glaucoma, few genome-wide association studies (GWASs) have been conducted thus far on DR. The few GWASs are of modest sample sizes in Hispanics, Chinese, and white populations and have reported borderline associations with DR in either type 1 or type 2 DM.25-28 In this review, we highlight these key genetic studies of DR with an emphasis on the most recent developments. We also discuss issues and challenges with elucidating the genetics of DR and indicate approaches that will provide the opportunity to advance our knowledge of this complex genetic disorder.
Definition and Classification of DR The diagnosis of DR is clinically defined by the presence of retinal microvascular lesions in patients with DM; however, these retinal lesions are not specific and may also be present in individuals without DM.29,30 The classification of DR is graded by severity and divided into nonproliferative diabetic retinopathy (NPDR) and PDR. Key retinal changes in NPDR include micro-
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Research Original Investigation
Diabetic Retinopathy Genetics
Table 2. Summary of Individual Candidate Gene Studies of Diabetic Retinopathya Gene
Polymorphism
Comments
AKR1B1/ALR2
(CA)n dinucleotide repeats
z − 2 microsatellite confers risk in all DR, NPDR, and PDR of type 1 and type 2 DMb
Reference(s) 20
z + 2 and z microsatellite protective against all DR in type 2 DM; no ethnic differenceb
VEGF
rs759853
T allele protective against DR in type 1 DMb
rs35839483
Not associated with DR
rs2010963 (−634 C/G)
Insignificant finding for DR or PDR, but C allele confers risk for NPDR in meta-analysisb
rs25648
T allele increases risk of DR but finding insignificant
20, 21, 63-71 20, 63, 68
rs1570360 (−116A>G)
inconsistent and insignificant finding
20, 63, 72
rs3095039
T allele increases risk, but finding inconsistent and insignificant
20, 63, 70
rs35569394
−2549DEL increases risk but finding insignificant
rs699947 (−2578 A/C)
A allele increases risk but finding insignificant
20, 66, 74-77
rs13207351 (−152A)
Associated with PDR but insignificant in other studies
63, 72, 75
rs735286 (+4618)
Haplotype-tagged SNP associated with severity of DR
72
rs2146323 (+5092)
Haplotype-tagged SNP associated with severity of DR, associated with early progression of DR
72, 77
20, 73
rs833061 (−1498 C/T)
Inconsistent and Insignificant finding
rs3025021
Inconclusive
rs10434
G allele associated with blinding DR
76
rs833068
G allele confers risk in DR
76
rs833070
Associated with early progression of DR but weak association
77
ITGA2
rs2910964
A allele increases risk a but one study in Chinese was not significant
20, 78, 79
AGTR1
rs5186
C allele confers protection, but finding insignificant
ACE
63, 68, 69, 75 75, 76
20
INS/DEL at intron 16
Inconsistent and insignificant finding
20
rs4343
Associated with DR in Chinese
80
ADRB3
rs4994
Inconsistent and insignificant finding
20
AGT
rs4762
C allele confers protectionb
20
APOE
E2/E3/E4
Inconsistent and insignificant finding
FGF2
rs41456044
A allele increases risk, but finding insignificant
20
rs308395
G allele increases risk, but finding insignificant
20
NOS3
20, 81, 82
rs1799983
G allele increases risk, but finding insignificant
20, 79
rs41322052
Inconsistent and insignificant finding
20, 83
rs3138808
Inconsistent and insignificant finding
20
rs841853
Insignificant finding
20
HLA
DR1-8
Insignificant finding
20
SDH
rs2055858 and rs3759890
Weak association
84
ICAM1
rs13306430
G allele confers protectionb
20 85
SLC2A1
rs5498
Not associated with DR
MTHFR
rs1801133
T allele increases risk but finding insignificant
20
NPY
rs16139
C allele increases risk but finding insignificant
20
PAI-1
rs1799768
4G/5G allele increases risk but finding insignificant
PON1
rs662
Inconsistent and insignificant finding
20
PPARG
rs1801282
G allele confers protection but finding insignificant
20
AGER/RAGE
VDR
20, 86
rs1800624 (−374T/A)
Inconsistent and insignificant finding
20, 85, 87, 88
rs1800625 (−429T/C)
Inconsistent and insignificant finding
20, 85, 89
rs2070600
Associated with DR of Indian ethnicity
85
rs10735810
T allele increases risk, but finding insignificant
20 (continued)
100
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Diabetic Retinopathy Genetics
Original Investigation Research
Table 2. Summary of Individual Candidate Gene Studies of Diabetic Retinopathya (continued) Gene
Polymorphism
Comments
EDN1
rs5370 (Lys198Asn)
Reduced risk in Chinese
Reference(s) 79
ROCK2
Thr431Asn and Arg83Lys
No association
90
CPVL/CHN2
rs39059
Increases risk of DR, significant in meta-analysis
91
FRMD3
rs10868025
Significant in stage 1, but insignificant in stage 2
91
CARS
rs451041
Not significant
91 91
IRS2
rs1411766
Not significant
SOD2
rs4880
C allele reduced risk
92
MnSOD
Ala16Val
Significant with DR
65
CA
rs2403104, rs17741410, rs1496533, rs17814594, rs12544332, rs1496529, rs725605, rs2645050, rs2645049, rs13278559
Not associated with DR
93
PEDF
rs12150053, rs12948385, rs8697961, rs1136287
Not associated with DR
85
EPO
rs1617640
Not associated with DR
85
HTRA1/ARMS2
rs11200638, rs10490924
Not associated with DR
85
CFH
rs1061170, rs3753394
Not associated with DR
85
PSMD9
rs74421874, rs3825172, rs14259
Associated with DR
94
Abbreviations: DM, diabetes mellitus; DR, diabetic retinopathy; NPDR, nonproliferative diabetic retinopathy; PDR, proliferative diabetic retinopathy; SNP, single-nucleotide polymorphism. a
Genome-wide approaches not included.
b
Significant and consistent direction in more than 1 study.
Table 3. Summary of Linkage Studies of DR Ethnicity Pima Indian
Type of Diabetes
Sample Size
Definition of DR
2
103 sib-pairs
Any DR
Region
Logarithm of Odds Score
3q26
1.36
D3S3053, D3S2427
9q21
1.46
D9S1120, D9S910
3.01 and 2.58
Pima Indian
2
211 sibships
Retinopathy score in worst eye
1p36
Mexican American
2
282 sib-pairs
Any DR
1p36 3q12
Severe NPDR or PDR
Nearest Markers
a
Reference 24
D1S3669
13
1.24
GGAT2A07
23
2.41
GATA68D03
7p15
1.02
GATA41G07
12p13
2.47
GATA49D12
15q25
1.07
ATA28G05
15q26
1.16
GATA22F01
2q37
1.11
AFM112yd4
3p26
1.29
GATA22G12
3q12
1.40
GATA68D03
12q23
1.03
GATA85A04
Abbreviations: DR, diabetic retinopathy; NPDR, nonproliferative diabetic retinopathy. a
For single-point and multiple-point analysis, respectively.
the Pima Indians, indicating suggestive evidence of linkage for DR in this population. This approach has several limitations. First, linkage studies only offer rough estimates of the genomic region because the mapping resolution is generally low, literally a test of millions of basepairs. Much more extensive efforts are required to pinpoint specific causal variants responsible for DR. Second, linkage studies often benefit from large families. Linkage studies on DR have thus far been conducted in Pima Indians and Mexican Americans, where large families are available for study. It has not been reported in other ethnicities. Third, the effect size (penetrance) of individual variants may be of sufficiently small magnitude that most study would be underpowered to detect genomic locations via cosegregation expected for complex multifactorial disorders such as DR. jamaophthalmology.com
Genome-Wide Association Studies More recent technologic advances have revolutionized the field toward the second hypothesis-free generating approach, GWASs, in which up to millions of SNPs can be tested against traits such as DR. These developments include microarraybased technology with tag SNPs, using the concept of linkage disequilibrium where adjacent or correlated SNPs cosegregate together in populations. Data from publicly available databases, such as the HapMap and the 1000 Genome Project, have been instrumental in developing such arrays. Since the first reported success of a well-designed GWAS, more than 2000 loci have demonstrated significant and often replicated associations with one or more common complex disorders.32 Although this field has received a number of critiJAMA Ophthalmology January 2014 Volume 132, Number 1
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Table 4. Summary of Genome-Wide Association Studies of Diabetic Retinopathy Ethnicity Mexican American
Taiwanese (Chinese)
White (GoKinD and EDIC)
Taiwanese (Chinese)
102
Type of Sample Size, Diabetes Cases/Controls 2
2
1
2
103 (Moderate to severe NPDR and PDR)/183 (normal to early NPDR)
174 (NPDR and PDR)/575 diabetic patients (no DR) and 100 nondiabetic patients
437 (PDR)/570 diabetic patients ≥8 years (no DR)
Odds Ratio
P Value
Gene(s)
Reference
rs6909083
6p12/NA
NA
2E-05
TINAG
25
rs17083119
6q22/NA
NA
3E-05
C6orf170
rs2300782
5q22/A
2.64
6E-05
CAMK4
rs10519765
15q13/G
3.33
3E-04
FMN1
rs899036
11p12/A
3.13
3E-04
API5
rs10501943
11q22/C
3.04
3E-04
CNTN5
rs17376456
5q15/A
3.63a
3E-15b
C5orf36
rs2038823
13q32/C
2.33a
5E-11b
HS6ST3
rs4838605
10q11/C
1.58a
2E-09b
ARHGAP22
rs12219125
10p12/T
1.62a
9E-09b
PLXDC2-NEBL
rs4462262
10q21/C
1.54a
Original Investigation | OPHTHALMIC MOLECULAR GENETICS
Challenges in Elucidating the Genetics of Diabetic Retinopathy Jane Z. Kuo, MD, PhD; Tien Y. Wong, MD, PhD; Jerome I. Rotter, MD
IMPORTANCE In the past decade, significant progress in genomic medicine and technologic
developments has revolutionized our approach to common complex disorders in many areas of medicine, including ophthalmology. A disorder that still needs major genetic progress is diabetic retinopathy (DR), one of the leading causes of blindness in adults. OBJECTIVE To perform a literature review, present the current findings, and highlight some key challenges in DR genetics. DESIGN, SETTING, AND PARTICIPANTS We performed a thorough literature review of the genetic factors for DR, including heritability scores, twin studies, family studies, candidate gene studies, linkage studies, and genome-wide association studies (GWASs). MAIN OUTCOME MEASURES Environmental and genetic factors for DR. RESULTS Although there is clear demonstration of a genetic contribution in the development and
progression of DR, the identification of susceptibility loci through candidate gene approaches, linkage studies, and GWASs is still in its infancy. The greatest obstacles remain a lack of power because of small sample size of available studies and a lack of phenotype standardization. CONCLUSIONS AND RELEVANCE The field of DR genetics is still in its infancy and is a challenge
because of the complexity of the disease. This review outlines some strategies and lessons for future investigation to improve our understanding of this complex genetic disorder. JAMA Ophthalmol. 2014;132(1):96-107. doi:10.1001/jamaophthalmol.2013.5024 Published online November 7, 2013.
D
iabetic retinopathy (DR), an important microvascular complication of diabetes mellitus (DM), is a leading cause of visual impairment in adults 20 to 74 years of age.1 More than 93 million people worldwide have DR, 17 million of whom have proliferative diabetic retinopathy (PDR) and 28 million of whom have vision-threatening DR.2 This number will continue to escalate with an aging population, increasing obesity, and a rapidly progressing DM epidemic. More individuals, especially Hispanics, people of African descent, and Asians, will be vulnerable to blinding DR in coming years.3,4 There is clearly a need to develop strategies to identify at-risk individuals for early interventions. Compared with other major causes of visual loss, such as age-related macular degeneration,5 myopia6,7 and glaucoma,8,9 the search for genetic clues to DR has not progressed as rapidly. To date, the few studies that have reported on possible susceptibility genes for DR have yielded inconsistent results. There is clearly a familial relationship in DR because twin and family studies10-17 indicate a genetic basis. Several candidate gene studies have reported promising genes,18-21 but few of them have been replicated, and the few positive findings reveal only weak genetic associations.18-20,22 In genome-wide approaches, 3 linkage studies performed in Pima Indians and 96
Author Affiliations: Author affiliations are listed at the end of this article. Corresponding Author: Jerome I. Rotter, MD, Institute for Translational Genomics and Population Sciences, Los Angeles Bio Medical Research Institute, Harbor–UCLA Medical Center, 1124 W Carson St, Torrance, CA 90502 ([email protected]). Section Editor: Janey L. Wiggs, MD, PhD.
Mexican Americans have identified regions on chromosomes 1, 3, and 12 to be suggestive of DR.13,23,24 In contrast to age-related macular degeneration, myopia, and glaucoma, few genome-wide association studies (GWASs) have been conducted thus far on DR. The few GWASs are of modest sample sizes in Hispanics, Chinese, and white populations and have reported borderline associations with DR in either type 1 or type 2 DM.25-28 In this review, we highlight these key genetic studies of DR with an emphasis on the most recent developments. We also discuss issues and challenges with elucidating the genetics of DR and indicate approaches that will provide the opportunity to advance our knowledge of this complex genetic disorder.
Definition and Classification of DR The diagnosis of DR is clinically defined by the presence of retinal microvascular lesions in patients with DM; however, these retinal lesions are not specific and may also be present in individuals without DM.29,30 The classification of DR is graded by severity and divided into nonproliferative diabetic retinopathy (NPDR) and PDR. Key retinal changes in NPDR include micro-
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Diabetic Retinopathy Genetics
Original Investigation Research
aneurysms, hard exudates, cotton wool spots, intraretinal microvascular abnormalities, and venous beading; these further subdivide NPDR into mild, moderate, and severe forms. Key retinal changes in PDR include neovascularization of the optic disc or elsewhere, preretinal hemorrhage, or vitreous hemorrhage. On the other hand, clinically significant macular edema, which is graded as its own entity, can develop at any stage of the DR spectrum. Thus, the various classifications in DR grading, resulting in heterogeneity of DR phenotype, pose a significant challenge in genetic studies of DR. The assessment of DR via a standardized stereoscopic photograph has been proposed to overcome this issue, and more researchers have used this approach by adopting and grading DR using the Early Treatment Diabetic Retinopathy Study severity scale or a similar modification. Recently, the assessment of DR and diabetic macular edema via optical coherence tomography has been proposed as an imaging modality to better visualize the intraretinal morphologic changes in patients with DM31; however, the classification of DR and diabetic macular edema via optical coherence tomography has not been clearly defined or adopted for use.
Genetics of DM Diabetic retinopathy occurs on the background of DM. That genetic factors play a major role in the etiology of DM has long been appreciated because of ethnic differences in frequency, increased familial aggregation, and a markedly higher concordance in monozygotic vs dizygotic twins. This is true for each of the major subforms of DM (type 1 and type 2). To date, approximately 60 loci have been successfully identified for type 2 DM, of which only 3 were discovered before the GWAS era.32,33 Although most of these studies were performed in individuals of European descent, more recent studies of Asians,34 Hispanics,35 and African Americans35-37 have also demonstrated some level of associations for these signals, supporting the hypothesis that these signals (or the causal variants that are in high linkage disequilibrium with these signals) are likely common alleles that are widely distributed in the human population and each contributing a small effect on disease risk.32 These important discoveries through large collaborative efforts by GWAS approaches have led to substantial progress in the understanding of genetics in type 2 DM, leading to the identification of novel pathways, demonstrating mechanistic associations, and supporting prior epidemiologic studies.38 These findings have illustrated some important key lessons that are useful in genetic studies of DR. First, joining forces by international collaborative efforts is necessary to increase statistical power by increasing sample size.38 Second, both analysis by treating the phenotype as a dichotomous trait and analysis of a related, quantitative trait are useful.38 Third, connection of genetic findings with more defined physiologic parameters increases understanding.38 Elucidating the genetic basis of type 2 DM offers an ideal model to approach the genetic study of DR. However, it should be apparent that the phenotype of type 2 DM has several advantages, such as the ease of classification and readily available large samples of patients even without detailed assessment. jamaophthalmology.com
Environmental Factors for DR The origin of DR remains complex and poorly understood. Large epidemiologic studies2,39,40 have consistently demonstrated that the duration of DM and adequacy of glycemic control are 2 of the major contributors to the development and progression of DR. This was robustly documented in the Wisconsin Epidemiological Study of Diabetic Retinopathy (WESDR), which found that duration of DM was the strongest predictor for progression of DR, with prevalence of DR ranging from 17% in type 1 DM to 29% in type 2 DM for patients with DM for less than 5 years and increasing markedly to almost 100% for type 1 DM and 78% for type 2 DM in patients with DM for more than 15 years.41,42 In other landmark studies,2,39,40,43 such as the Diabetes Control and Complications Trial, UK Prospective Diabetes Study, the Action to Control Cardiovascular Risk in Diabetes (ACCORD) Eye Study, and Meta-Analysis for Eye Disease Study, intensive glycemic control was effective in reducing the rate of DR progression in both type 1 and type 2 DM. Other studies2,44,45 have demonstrated that blood pressure control is another risk modifier, although some studies43 did not support this finding. Recently, increasing evidence has supported an association between dyslipidemia and DR46; the Fenofibrate Intervention and Event Lowering in Diabetes Study47 and the ACCORD Eye Study43 demonstrated that reduction in lipids could also limit DR progression. Despite strong evidence of DR susceptibility, these environmental risk modifiers by themselves do not account for the complete risk susceptibility. First, this is exemplified by clinical observations that some individuals develop DR despite good glycemic control and short duration of disease, whereas others do not develop DR, even with poor glycemic control and longer duration of DM.48 Second, the strongest environmental factors (duration of DM and glycosylated hemoglobin) only explained approximately 11% of the variation in retinopathy risk in the Diabetes Control and Complications Trial.49,50 Similarly, a combination of glycosylated hemoglobin, blood pressure, and total cholesterol only explained approximately 10% of the variation in retinopathy risk in the WESDR,51 suggesting that the remaining approximately 90% of the variation in retinopathy risk is presumably explained by other risk factors. Finally, population studies indicate that retinopathy signs, such as microaneurysms, were detectable in 7% to 13% of patients without DM and patients in whom the glycosylated hemoglobin level was well below 5%.29 In addition, singlenucleotide polymorphisms (SNPs) associated with DM or hypertension were not associated with retinopathy in individuals without DM,52 suggesting that other risk factors, independent of hyperglycemia and DM, contribute to the development and progression of retinopathy signs similar to DR.
Genetic Factors for DR Attempts to identify gene(s) in the development of DR have been conducted during the past few decades. To date, these studies have been limited to twin studies,10 family studies,11-17 JAMA Ophthalmology January 2014 Volume 132, Number 1
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Table 1. Summary of Familial Aggregation Studies in Diabetic Retinopathy Ethnicity (Study)
Type of Diabetes
Heritability
Evidence of Familial Aggregation
11
Strong evidence of familial clustering in PDR
12
1
White (FinnDiane study)
1
PDR, 0.52
Pima Indian
2
Any DR, 0.18
Modest evidence of familial clustering
13
South Indian
2
Strong evidence of familial clustering (OR = 3.37 in sibling of diabetes with and without DR)
14
Chinese
2
Strong evidence of familial clustering in any DR; DR diagnosed in 7.1% of siblings without DR vs 29.7% of siblings with DR
15
Mexican American
2
Strong evidence of familial clustering in severe DR (OR = 1.72) but not any DR
16
Multiethnic (FIND-Eye Study)a
2
Strong evidence of familial clustering
17
Any DR, 0.27 PDR, 0.25 Mexican American, 0.24
Figure. Heritability Estimates for Diabetic Retinopathy 1.00
Heritability of DR
0.75
0.52 0.50
0.27 0.25
0.25
0.24
0.18
0.00 DCCT, PDR
Pima Indian, Any DR
FIND-Eye Study, Any DR
FIND-Eye FIND-Eye Study, Study, PDR Mexican American
Study
DCCT indicates Diabetes Control and Complications Trial; DR, diabetic retinopathy; FIND-Eye Study, Family Investigation of Nephropathy and Diabetes–Eye Study; PDR, proliferative diabetic retinopathy.
candidate gene studies,18-22 linkage studies,13,23,24 and smallscale GWASs with modest sample sizes.25-28 In support of a genetic hypothesis of DR, several studies3,4 have found a discrepant rate of the prevalence of DR among US populations, with a significantly higher prevalence observed in Hispanics, African Americans, and Chinese Americans. Compared with whites, in African Americans other risk modifiers, such as duration of DM, glycemic control, and blood pressure, appear to account for the higher prevalence of DR, but these factors do not explain the higher prevalence seen in Hispanics,53-56 suggesting that other factors, including genetic factors, may influence susceptibility to DR.
Twin and Family Studies In twin studies, Leslie and Pyke10 found the same degree of severity in 35 of 37 concordant twins with type 2 DM (95%) compared with only 21 of 31 concordant twins with type 1 DM (68%). This early observation was extended by familial aggregation 98
Reference
Low evidence of increased risk of any DR; strong evidence of familial clustering for severe DR (OR = 3.1 in relatives of diabetic patients with and without DR)
Whites (DCCT)
Abbreviations: DCCT, Diabetes Control and Complications Trial; DR, diabetic retinopathy; FIND-Eye Study, Family Investigation of Nephropathy and Diabetes–Eye Study; FinnDiane, Finnish Diabetic Nephropathy; OR, odds ratio; PDR, proliferative diabetic retinopathy. a Mexican Americans, European Americans, African Americans, and American Indians.
studies, with siblings and relatives of diabetic patients with DR having as high as a 3-fold increased risk of DR compared with siblings and relatives of diabetic patients without DR11-17 (Table 1). This trend was seen in either type 1 or type 2 DM and across different ethnicities. Furthermore, evidence of a familial aggregation is more consistently seen in the presence of more severe retinopathy and less in the presence of any retinopathy, with heritability scores ranging from 18% to 27% for any DR13,17 and 25% to 52% for PDR12,17 in either type 1 or type 2 DM (Figure). Thus, as previously mentioned, it is important to not only provide a standardized assessment of DR but also compare those without DR with those with more severe stages of DR. The Family Investigation of Nephropathy and Diabetes– Eye study, in which nearly half of the 2368 diabetic patients are Mexican Americans, demonstrated that the heritability of any DR in this population of type 2 DM is as high as 24%.17
Candidate Gene Studies Most genetic research in DR has used the candidate gene approach. Several pathways and processes have been proposed to play an important role in the pathogenesis of DR, which led to the testing of a number of hypothesized candidate genes. Although a number of candidate genes and genetic variants have been proposed in the literature, few of them have been replicated, and the few positive findings only revealed weak genetic associations with DR.18,19 In this approach, the analysis compares the frequency of a particular genetic variant in individuals with (cases) or without (controls) DR. Several pathways and processes have been proposed, including the renin-angiotensin system, glucoseinduced pathways, vascular endothelial dysfunction, tissue matrix remodeling, and angiogenesis.18,19 Potential candidate genes involved in these pathways and processes include ACE, AGTR1, AGT, VEGF, AKR1B1, RAGE, GLUT1, APOE, MTHFR, PAI-1, ITGA2, PPAR-γ, and NOS3. Their associations or lack thereof with DR have been extensively documented in prior reviews.18-21 We summarize the most important findings with a focus on AKR1B1 and VEGF because of their biological implications.
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Research
Original Investigation | OPHTHALMIC MOLECULAR GENETICS
Challenges in Elucidating the Genetics of Diabetic Retinopathy Jane Z. Kuo, MD, PhD; Tien Y. Wong, MD, PhD; Jerome I. Rotter, MD
IMPORTANCE In the past decade, significant progress in genomic medicine and technologic
developments has revolutionized our approach to common complex disorders in many areas of medicine, including ophthalmology. A disorder that still needs major genetic progress is diabetic retinopathy (DR), one of the leading causes of blindness in adults. OBJECTIVE To perform a literature review, present the current findings, and highlight some key challenges in DR genetics. DESIGN, SETTING, AND PARTICIPANTS We performed a thorough literature review of the genetic factors for DR, including heritability scores, twin studies, family studies, candidate gene studies, linkage studies, and genome-wide association studies (GWASs). MAIN OUTCOME MEASURES Environmental and genetic factors for DR. RESULTS Although there is clear demonstration of a genetic contribution in the development and
progression of DR, the identification of susceptibility loci through candidate gene approaches, linkage studies, and GWASs is still in its infancy. The greatest obstacles remain a lack of power because of small sample size of available studies and a lack of phenotype standardization. CONCLUSIONS AND RELEVANCE The field of DR genetics is still in its infancy and is a challenge
because of the complexity of the disease. This review outlines some strategies and lessons for future investigation to improve our understanding of this complex genetic disorder. JAMA Ophthalmol. 2014;132(1):96-107. doi:10.1001/jamaophthalmol.2013.5024 Published online November 7, 2013.
D
iabetic retinopathy (DR), an important microvascular complication of diabetes mellitus (DM), is a leading cause of visual impairment in adults 20 to 74 years of age.1 More than 93 million people worldwide have DR, 17 million of whom have proliferative diabetic retinopathy (PDR) and 28 million of whom have vision-threatening DR.2 This number will continue to escalate with an aging population, increasing obesity, and a rapidly progressing DM epidemic. More individuals, especially Hispanics, people of African descent, and Asians, will be vulnerable to blinding DR in coming years.3,4 There is clearly a need to develop strategies to identify at-risk individuals for early interventions. Compared with other major causes of visual loss, such as age-related macular degeneration,5 myopia6,7 and glaucoma,8,9 the search for genetic clues to DR has not progressed as rapidly. To date, the few studies that have reported on possible susceptibility genes for DR have yielded inconsistent results. There is clearly a familial relationship in DR because twin and family studies10-17 indicate a genetic basis. Several candidate gene studies have reported promising genes,18-21 but few of them have been replicated, and the few positive findings reveal only weak genetic associations.18-20,22 In genome-wide approaches, 3 linkage studies performed in Pima Indians and 96
Author Affiliations: Author affiliations are listed at the end of this article. Corresponding Author: Jerome I. Rotter, MD, Institute for Translational Genomics and Population Sciences, Los Angeles Bio Medical Research Institute, Harbor–UCLA Medical Center, 1124 W Carson St, Torrance, CA 90502 ([email protected]). Section Editor: Janey L. Wiggs, MD, PhD.
Mexican Americans have identified regions on chromosomes 1, 3, and 12 to be suggestive of DR.13,23,24 In contrast to age-related macular degeneration, myopia, and glaucoma, few genome-wide association studies (GWASs) have been conducted thus far on DR. The few GWASs are of modest sample sizes in Hispanics, Chinese, and white populations and have reported borderline associations with DR in either type 1 or type 2 DM.25-28 In this review, we highlight these key genetic studies of DR with an emphasis on the most recent developments. We also discuss issues and challenges with elucidating the genetics of DR and indicate approaches that will provide the opportunity to advance our knowledge of this complex genetic disorder.
Definition and Classification of DR The diagnosis of DR is clinically defined by the presence of retinal microvascular lesions in patients with DM; however, these retinal lesions are not specific and may also be present in individuals without DM.29,30 The classification of DR is graded by severity and divided into nonproliferative diabetic retinopathy (NPDR) and PDR. Key retinal changes in NPDR include micro-
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Table 2. Summary of Individual Candidate Gene Studies of Diabetic Retinopathya Gene
Polymorphism
Comments
AKR1B1/ALR2
(CA)n dinucleotide repeats
z − 2 microsatellite confers risk in all DR, NPDR, and PDR of type 1 and type 2 DMb
Reference(s) 20
z + 2 and z microsatellite protective against all DR in type 2 DM; no ethnic differenceb
VEGF
rs759853
T allele protective against DR in type 1 DMb
rs35839483
Not associated with DR
rs2010963 (−634 C/G)
Insignificant finding for DR or PDR, but C allele confers risk for NPDR in meta-analysisb
rs25648
T allele increases risk of DR but finding insignificant
20, 21, 63-71 20, 63, 68
rs1570360 (−116A>G)
inconsistent and insignificant finding
20, 63, 72
rs3095039
T allele increases risk, but finding inconsistent and insignificant
20, 63, 70
rs35569394
−2549DEL increases risk but finding insignificant
rs699947 (−2578 A/C)
A allele increases risk but finding insignificant
20, 66, 74-77
rs13207351 (−152A)
Associated with PDR but insignificant in other studies
63, 72, 75
rs735286 (+4618)
Haplotype-tagged SNP associated with severity of DR
72
rs2146323 (+5092)
Haplotype-tagged SNP associated with severity of DR, associated with early progression of DR
72, 77
20, 73
rs833061 (−1498 C/T)
Inconsistent and Insignificant finding
rs3025021
Inconclusive
rs10434
G allele associated with blinding DR
76
rs833068
G allele confers risk in DR
76
rs833070
Associated with early progression of DR but weak association
77
ITGA2
rs2910964
A allele increases risk a but one study in Chinese was not significant
20, 78, 79
AGTR1
rs5186
C allele confers protection, but finding insignificant
ACE
63, 68, 69, 75 75, 76
20
INS/DEL at intron 16
Inconsistent and insignificant finding
20
rs4343
Associated with DR in Chinese
80
ADRB3
rs4994
Inconsistent and insignificant finding
20
AGT
rs4762
C allele confers protectionb
20
APOE
E2/E3/E4
Inconsistent and insignificant finding
FGF2
rs41456044
A allele increases risk, but finding insignificant
20
rs308395
G allele increases risk, but finding insignificant
20
NOS3
20, 81, 82
rs1799983
G allele increases risk, but finding insignificant
20, 79
rs41322052
Inconsistent and insignificant finding
20, 83
rs3138808
Inconsistent and insignificant finding
20
rs841853
Insignificant finding
20
HLA
DR1-8
Insignificant finding
20
SDH
rs2055858 and rs3759890
Weak association
84
ICAM1
rs13306430
G allele confers protectionb
20 85
SLC2A1
rs5498
Not associated with DR
MTHFR
rs1801133
T allele increases risk but finding insignificant
20
NPY
rs16139
C allele increases risk but finding insignificant
20
PAI-1
rs1799768
4G/5G allele increases risk but finding insignificant
PON1
rs662
Inconsistent and insignificant finding
20
PPARG
rs1801282
G allele confers protection but finding insignificant
20
AGER/RAGE
VDR
20, 86
rs1800624 (−374T/A)
Inconsistent and insignificant finding
20, 85, 87, 88
rs1800625 (−429T/C)
Inconsistent and insignificant finding
20, 85, 89
rs2070600
Associated with DR of Indian ethnicity
85
rs10735810
T allele increases risk, but finding insignificant
20 (continued)
100
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Table 2. Summary of Individual Candidate Gene Studies of Diabetic Retinopathya (continued) Gene
Polymorphism
Comments
EDN1
rs5370 (Lys198Asn)
Reduced risk in Chinese
Reference(s) 79
ROCK2
Thr431Asn and Arg83Lys
No association
90
CPVL/CHN2
rs39059
Increases risk of DR, significant in meta-analysis
91
FRMD3
rs10868025
Significant in stage 1, but insignificant in stage 2
91
CARS
rs451041
Not significant
91 91
IRS2
rs1411766
Not significant
SOD2
rs4880
C allele reduced risk
92
MnSOD
Ala16Val
Significant with DR
65
CA
rs2403104, rs17741410, rs1496533, rs17814594, rs12544332, rs1496529, rs725605, rs2645050, rs2645049, rs13278559
Not associated with DR
93
PEDF
rs12150053, rs12948385, rs8697961, rs1136287
Not associated with DR
85
EPO
rs1617640
Not associated with DR
85
HTRA1/ARMS2
rs11200638, rs10490924
Not associated with DR
85
CFH
rs1061170, rs3753394
Not associated with DR
85
PSMD9
rs74421874, rs3825172, rs14259
Associated with DR
94
Abbreviations: DM, diabetes mellitus; DR, diabetic retinopathy; NPDR, nonproliferative diabetic retinopathy; PDR, proliferative diabetic retinopathy; SNP, single-nucleotide polymorphism. a
Genome-wide approaches not included.
b
Significant and consistent direction in more than 1 study.
Table 3. Summary of Linkage Studies of DR Ethnicity Pima Indian
Type of Diabetes
Sample Size
Definition of DR
2
103 sib-pairs
Any DR
Region
Logarithm of Odds Score
3q26
1.36
D3S3053, D3S2427
9q21
1.46
D9S1120, D9S910
3.01 and 2.58
Pima Indian
2
211 sibships
Retinopathy score in worst eye
1p36
Mexican American
2
282 sib-pairs
Any DR
1p36 3q12
Severe NPDR or PDR
Nearest Markers
a
Reference 24
D1S3669
13
1.24
GGAT2A07
23
2.41
GATA68D03
7p15
1.02
GATA41G07
12p13
2.47
GATA49D12
15q25
1.07
ATA28G05
15q26
1.16
GATA22F01
2q37
1.11
AFM112yd4
3p26
1.29
GATA22G12
3q12
1.40
GATA68D03
12q23
1.03
GATA85A04
Abbreviations: DR, diabetic retinopathy; NPDR, nonproliferative diabetic retinopathy. a
For single-point and multiple-point analysis, respectively.
the Pima Indians, indicating suggestive evidence of linkage for DR in this population. This approach has several limitations. First, linkage studies only offer rough estimates of the genomic region because the mapping resolution is generally low, literally a test of millions of basepairs. Much more extensive efforts are required to pinpoint specific causal variants responsible for DR. Second, linkage studies often benefit from large families. Linkage studies on DR have thus far been conducted in Pima Indians and Mexican Americans, where large families are available for study. It has not been reported in other ethnicities. Third, the effect size (penetrance) of individual variants may be of sufficiently small magnitude that most study would be underpowered to detect genomic locations via cosegregation expected for complex multifactorial disorders such as DR. jamaophthalmology.com
Genome-Wide Association Studies More recent technologic advances have revolutionized the field toward the second hypothesis-free generating approach, GWASs, in which up to millions of SNPs can be tested against traits such as DR. These developments include microarraybased technology with tag SNPs, using the concept of linkage disequilibrium where adjacent or correlated SNPs cosegregate together in populations. Data from publicly available databases, such as the HapMap and the 1000 Genome Project, have been instrumental in developing such arrays. Since the first reported success of a well-designed GWAS, more than 2000 loci have demonstrated significant and often replicated associations with one or more common complex disorders.32 Although this field has received a number of critiJAMA Ophthalmology January 2014 Volume 132, Number 1
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Table 4. Summary of Genome-Wide Association Studies of Diabetic Retinopathy Ethnicity Mexican American
Taiwanese (Chinese)
White (GoKinD and EDIC)
Taiwanese (Chinese)
102
Type of Sample Size, Diabetes Cases/Controls 2
2
1
2
103 (Moderate to severe NPDR and PDR)/183 (normal to early NPDR)
174 (NPDR and PDR)/575 diabetic patients (no DR) and 100 nondiabetic patients
437 (PDR)/570 diabetic patients ≥8 years (no DR)
Odds Ratio
P Value
Gene(s)
Reference
rs6909083
6p12/NA
NA
2E-05
TINAG
25
rs17083119
6q22/NA
NA
3E-05
C6orf170
rs2300782
5q22/A
2.64
6E-05
CAMK4
rs10519765
15q13/G
3.33
3E-04
FMN1
rs899036
11p12/A
3.13
3E-04
API5
rs10501943
11q22/C
3.04
3E-04
CNTN5
rs17376456
5q15/A
3.63a
3E-15b
C5orf36
rs2038823
13q32/C
2.33a
5E-11b
HS6ST3
rs4838605
10q11/C
1.58a
2E-09b
ARHGAP22
rs12219125
10p12/T
1.62a
9E-09b
PLXDC2-NEBL
rs4462262
10q21/C
1.54a
