Experimental and Molecular Pathology 87 (2009) 234–238

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Experimental and Molecular Pathology

j o u r n a l h o m e p a g e : w w w. e l s e v i e r. c o m / l o c a t e / y e x m p

Association between vascular endothelial growth factor gene polymorphisms and

age-related macular degeneration in a Polish population
Katarzyna Janik-Papis a, Malgorzata Zaras b, Anna Krzyzanowska a, Katarzyna Wozniak a, Janusz Blasiak a,⁎,
Jerzy Szaflik b, Jacek P. Szaflik b
a
Department of Molecular Genetics, University of Lodz, Banacha 12/16, 90-237 Lodz, Poland
b
Department of Ophthalmology, Medical University of Warsaw and Samodzielny Publiczny Szpital Okulistyczny, Sierakowskiego 13, 03-710 Warsaw, Poland

a r t i c l e i n f o a b s t r a c t

Article history: The pathogenesis of age-related macular degeneration (AMD) is thought to be determined by an array of
Received 12 May 2009 environmental and genetic factors. The association of increased expression of vascular endothelial growth
and in revised form 7 September 2009 factor (VEGF) with AMD, especially the wet form of AMD, was reported in several studies. The VEGF gene is
Available online 15 September 2009
highly polymorphic and some of its polymorphisms may affect its expression. In our work, we searched for
an association between the −460CN (rs833061) and −634GNC (rs2010963) polymorphisms of the VEGF
Keywords:
Age-related macular degeneration
gene and the occurrence of AMD and its dry and wet forms. We have chosen these polymorphisms because
AMD they were shown to be significant in other studies and we previously showed their association with diabetic
Vascular endothelial growth factor retinopathy. A total of 401 individuals were enrolled in this study: 136 controls, and 88 patients with dry and
VEGF 177 with wet AMD. The polymorphisms were determined with DNA from peripheral blood lymphocytes by
Gene polymorphism allele-specific and restriction fragment length polymorphism polymerase chain reaction. The significance of
the polymorphisms was assessed by multiple logistic regression, producing odds ratios (ORs) and 95%
confidence intervals (CIs). We observed a weak association (OR 2.90) between AMD occurrence and the C/T
genotype of the −460CNT polymorphism. An association (OR 3.77) between the C/T genotype of the
−460CNT polymorphism and the occurrence of dry AMD was observed. The T/T genotype considerably
lowered the risk of dry AMD (OR 0.19). Dry AMD was associated with the C/C genotype of the −634GNC
polymorphism (OR 3.68). Another weak association (OR 2.63) was found between the C/T genotype of the
−460CNT polymorphism and the occurrence of wet AMD. The occurrence of AMD was correlated with the
presence of the combined C/T–G/G genotype of both polymorphisms (OR 2.41), whereas the T/T–G/G and
T/T–G/C genotypes exerted a protective effect against the disease (OR 0.22 and 0.48, respectively). The
presence of the C/T–G/G and C/T–C/C combined genotypes increased the risk of dry AMD (OR 2.08 and 3.77,
respectively), whereas the presence of the T/T–G/G and T/T–G/C genotypes decreased the risk (OR 0.15 and
0.28, respectively). In the wet form of AMD, the combined genotype C/T–G/G slightly favored the disease
(OR 2.61) and the T/T–G/G genotype had a protective effect (OR 0.25). Analysis of haplotypes of both
polymorphisms yielded similar results for AMD in general as well as for the dry and wet forms of the disease:
the CG haplotype favored both forms of AMD, whereas the TG haplotype protected against both forms of
AMD. The results obtained indicate that the −460CNT and −634GNC polymorphisms of the VEGF gene may
be associated with the dry and wet forms of AMD in a Polish population.
© 2009 Elsevier Inc. All rights reserved.

Introduction
Although age-related macular degeneration (AMD) might take a
long time to develop, the onset of symptoms, including loss of vision,
often occurs suddenly . Nowadays, AMD is the leading cause of
blindness in individuals older than 55 years in developed countries
(Montezuma et al., 2007). In 2004, 1.75 million people in the United
States were affected by AMD and 7 million people were at risk of
⁎ Corresponding author. Fax: +4842 635 44 84.
E-mail address: januszb@biol.uni.lodz.pl (J. Blasiak).
developing AMD (Friedman et al., 2004). Generally, two types of AMD
can be considered: dry (non-exudative), which is the most common
form, and wet (exudative). In the dry form, there is progressive
atrophy of the retinal pigment epithelium (RPE), with subsequent loss
of the choriocapillaris and photoreceptors within the macula. Patients
with dry AMD experience a slow, progressive and inexorable decline
in central vision, whereas patients with the wet form suffer from
sudden loss of vision from either bleeding or fluid leakage from
abnormal vessels originating from the choriocapillaris beneath the
RPE and macula (Lopez et al., 1996). No remedy is known for dry
AMD; its wet form is treated to prevent the loss of vision or,
eventually, its restoration, but with modest success.

0014-4800/$ – see front matter © 2009 Elsevier Inc. All rights reserved.
doi:10.1016/j.yexmp.2009.09.005
 K. Janik-Papis et al. / Experimental and Molecular Pathology 87 (2009) 234–238
Table 1
Characteristics of patients with age-related macular degeneration (AMD) and
individuals without visual disturbances (controls; mean ± SD).
Individuals Number Age (years) Gender (females + males)
All 401 72.8 ± 9.5 266F + 135M
AMD 265 72.8 ± 8.4 175F + 90M
Dry AMD 88 74.3 ± 8.7 59F + 29M
Wet AMD 177 72.5 ± 8.0 116F + 61M
Controls 136 71.7 ± 10.2 91F + 45M
There are many theories to explain the pathogenesis of AMD. An
array of growth factors has been shown to be involved in the
development of the disease (see Ding et al., 2009, for review). Because
the wet AMD involves neovascularization, vascular endothelial
growth factor (VEGF) is a candidate for both diagnosing and treating
this most devastating form of AMD. VEGF, known also as VEGF-A,
plays a critical role in angiogenesis, vasculogenesis, and lymphangio-
genesis in normal and pathological cells. The gene encoding VEGF, the
VEGF-A gene, belongs to the VEGF gene family, which also contains the
VEGF-B, -C, and -D genes, as well as the PIGF (placental growth factor)
gene. The VEGF-A gene has eight exons separated by seven introns
(Houck et al., 1991). All genes of the family undergo alternative
splicing, producing many isoforms. Of these the VEGF165 form is the
most abundant and corresponds to a 23 kDa polypeptide, constituting
a monomer of homodimeric human VEGF-A, referred hereafter in this
paper as VEGF (Ferrara, 2004).
Polymorphisms of the VEGF gene have been investigated in AMD,
but data obtained in this research seem controversial (Penn et al.,
2008). The data obtained in these reports seem controversial.
Recently, no association between AMD and the 3 common functional
polymorphisms of the VEGF gene was found in a large prospective
population-based cohort study, The Rotterdam Study (Boekhoorn
et al., 2008). No association between AMD and 7 single nucleotide
polymorphisms (SNPs) of the VEGF gene, located in its promoter and
coding regions, was reported for an Anglo-Celtic subpopulation
(Richardson et al., 2007). In a study of a relatively small English
population, only 1 of 14 SNPs of the promoter and coding regions of
the VEGF gene was associated with the wet form of AMD (Churchill
et al., 2006). However, when haplotypes were analyzed, several
proved to be associated with the disease. In a large study testing the
association of 8 candidate genes with AMD, VEGF proved to have the
strongest association with the disease in both a family-based study
and a case–control study (Haines et al., 2006).
The data on the importance of VEGF in the pathogenesis of AMD
and contradictory report on the role of genetic variability of VEGF in
the risk of AMD prompted us to search for an association between the
occurrence of AMD and genetic variants of the VEGF gene in a Polish
subpopulation. We investigated 2 common polymorphisms of this
Fig. 1. Genotypes of the −460CNT polymorphism in the promoter region of the VEGF gene determined by allele-specific PCR detection and analyzed by 3% agarose gel
electrophoresis, stained with ethidium bromide and viewed under UV light. Lane M displays ΦX 174/BsuRI molecular weight marker; lanes marked by C and T show the results of
amplification with primers specific to the C and T alleles, respectively. Genotypes of patients numbered 1–4 are indicated in the lower part of the picture.
235
gene: a C → T transition at −1498 (the −460CNT polymorphism,
GenBank SNP ID rs833061) and a G → C transversion at 405 (the
−634GNC polymorphism, rs2010963). The somewhat confusing
notation of the polymorphisms is related to the different methods
of counting their location relative to the start codon in either
complementary DNA (cDNA) or DNA. The first polymorphism is
located in the promoter region of the gene, whereas the second is
placed in the 5′ untranslated region (5′UTR) of exon 1. Both
polymorphisms are located in the regulatory regions of the VEGF
gene; therefore, if they affect the structure of the gene, they may
primarily affect the level of transcription. Moreover, patients with
AMD have been reported to have polymorphisms within the
untranslated portions of the VEGF gene (introns and the 5′UTR),
where growth factors and transcription factors may bind to initiate
the transcription of VEGF (Churchill et al., 2006). There are other
known functional polymorphisms of the VEGF gene; however, we
chose the described polymorphisms because we previously reported
an association between them and diabetic retinopathy (Szaflik et al.,
2008) and thought it would be interesting to check whether they may
be significant in another type of retinal disturbance.
Methods
Clinical subjects
Peripheral blood samples were obtained from 401 patients with
AMD (n = 265) who were examined in the Department of Ophthal-
mology, Medical University of Warsaw, Poland, and from sex- and age-
matched individuals with exclusion of AMD (n = 136, controls). Of the
patients, 88 had dry AMD and the remaining 177 had the wet form
(Table 1). Medical history was obtained from all subjects, and no one
reported current or previous cancer or any genetic disease. The
patients underwent ophthalmic examination, including best-cor-
rected visual acuity, intraocular pressure, slit lamp examination, and
fundus examination, performed with a slit lamp equipped with either
non-contact or contact fundus lenses. Diagnosis of AMD was
confirmed by optical coherence tomography (OCT) and, in some
cases, by fluorescein angiography (FA) and indocyanin green angiog-
raphy (ICG). OCT evaluated retinal thickness, the presence of RPE
atrophy, drusen, or subretinal fluid and intraretinal edema; angiogra-
phy assessed the anatomical status of the retinal vessels, the presence
of choroidal neovascularization and leakage. The OCT examinations
were performed with Stratus OCT model 3000, software version 4.0
(Oberkochen, Germany). The FA and ICG examinations were complet-
ed with a Topcon TRC-50I IX fundus camera equipped with the digital
Image Net image system, version 2.14 (Topcon, Tokyo, Japan).
The study was approved by the Bioethics Committee of the
Medical University of Warsaw, Poland, and each patient gave written
informed consent.
236 K. Janik-Papis et al. / Experimental and Molecular Pathology 87 (2009) 234–238

Fig. 2. Genotypes of the −634GNC polymorphism in the 5′ untranslated region of the VEGF gene determined by PCR-based BsmFI restriction fragment length polymorphism and
analyzed by 12% polyacrylamide gel electrophoresis, stained with ethidium bromide and viewed under UV light. Lane M displays ΦX 174/BsuRI molecular weight marker; all
remaining lanes present genotypes of the indicated polymorphisms.

DNA preparation
Peripheral blood lymphocytes (PBLs) were isolated by centrifuga-
tion in a density gradient of Histopaque-1077 (15 min, 280 × g). The
pellet containing the PBLs was resuspended in Tris–EDTA buffer, pH 8,
to yield about 1–3 × 105 cells/ml. Genomic DNA was extracted from
the PBLs by phenol/chloroform extraction and proteinase K digestion.
The final samples were kept in Tris–EDTA buffer, pH 8, at − 20 °C until
use.
Determination of the VEGF genotype
Allele-specific polymerase chain reaction (PCR) was applied to
determine the genotypes of the −460CNT polymorphism; restriction
fragment length polymorphism PCR was performed to reveal the
genotypes of the −634GNC polymorphism. All PCR reactions were
carried out in an MJ Research, INC thermal cycler, model PTC-100
(Waltham, MA, USA).
To determine the genotype of the −460CNT polymorphism, we
used 25 μl of a PCR mixture that contained 10 ng genomic DNA; 1 U
MasterAmp™ Taq polymerase (Epicentre, Madison, WI, USA) in 1×
PCR buffer (100 mM Tris–HCl, pH 8.3, 500 mM KCl, 0.1% gelatin); 4 μl of
25 mM MgCl2, 3 μl of 2.5 mM dNTPs, and 1 μl (20 μM) of each primer.
Thermal cycling conditions were as follows: initial denaturation step
at 95 °C for 5 min, 30 cycles at 95 °C for 30 sec, 45 sec at 62 °C annealing
temperature and 45 sec at 72 °C. The following primers were used:
allele-specific sense oligonucleotides 5′-TGCGTGTGGGGTTGAGGGC-
3′ for C variant and 5′-TGCGTGTGGGGTTGAGGGT-3′ for T variant and
reverse primer 5′-CCCGCCGCAATGAAGGGGA-3′. The PCR products
were separated onto a 3% agarose gel. Fig. 1 presents a representative
gel obtained after genotyping of this polymorphism.
To determine the −634GNC polymorphism, we used 35 μl of a PCR
mixture that contained 10 ng genomic DNA; 1.5 U MasterAmp™ Taq
polymerase (Epicentre, Madison, WI, USA) in 1× PCR buffer (100 mM
Tris–HCl, pH 8.3, 500 mM KCl, 15 mM MgCl2, 0.1% gelatin); 7 μl of
25 mM MgCl2; 4.2 μl of 2.5 mM dNTPs; and 1.4 μl (20 μM) of each
primer. Thermal cycling conditions were as follows: initial denatur-
ation step at 95 °C for 5 min, 30 cycles at 95 °C for 45 sec, 1 min at 50 °C
annealing temperature, and 1 min at 72 °C . The final extension step
was performed at 72 °C for 5 min. We used the following primers:
sense 5′-GGCGCTCGGTGCTGGAATTT -3′, antisense 5′-AGCTAGCA-
CTTCTCGCGGCT-3 (POLGEN, Lodz, Poland). The 634 bp PCR product
was digested 3 hours with 2 U of the restriction enzyme BsmFI
(Fermentas, Vilnius, Lithuania). The G allele was digested into 449,
344, 291, 187, and 158 bp fragments, whereas the C variant was
digested into fragments of 635, 449, 158, and 187 bp. Digested
products were separated onto a 12% polyacrylamide gel. A photograph
of a representative gel for this polymorphism is presented in Fig. 2.
Statistical analysis
The allelic frequencies were estimated by gene counting and the
genotypes were scored. The χ2 analysis was used to compare the
observed number of genotypes with that expected for a population in
a Hardy–Weinberg equilibrium. The χ2 analysis was also used to test
the significance of the differences of observed alleles and genotypes
between groups. A logistic regression model was used to calculate the
odds ratios (ORs) and 95% confidence intervals (CIs). Analyses were
performed with STATISTICA 6.0 software (Statsoft, Tulsa, OK, USA).
Results
The patients were divided into three groups according to
genotypes of the −460CNT polymorphism of the VEGF gene promoter
(C/C, C/T, and T/T) and the 3 genotypes of the other polymorphism of
this gene, −634GNC (G/G, G/C, and C/C). The distributions of these

Table 2
Distribution of genotypes, frequency of alleles of the −460CNT and −634GNC
polymorphisms of the VEGF gene and odds ratio (OR) with 95% confidence interval
(95% CI) in patients with age-related macular degeneration (AMD) and individuals
without visual disturbances (controls).
Table 3
Distribution of genotypes, frequency of alleles of the −460CNT and −634GNC
polymorphisms of the VEGF gene and odds ratio (OR) with 95% confidence interval
(95% CI) in patients with dry age-related macular degeneration (AMD) and individuals
without visual disturbances (controls).

Genotype Controls (n = 134) AMD (n = 265) OR (95% CI) Genotype Controls (n = 134) Dry AMD (n = 88) OR (95% CI)
or allele or allele
Number Frequency Number Frequency Number Frequency Number Frequency

−460CNT −460CNT
C/C 11 0.08 26 0.10 1.22 (0.58–2.54) C/C 11 0.08 8 0.09 1.13 (0.44–2.94)
C/T 63 0.47 191 0.72 2.90 (1.89–4.48) C/T 63 0.47 67 0.76 3.77 (2.06–6.9)
T/T 60 0.45 48 0.18 0.86 (0.46–1.58) T/T 60 0.45 13 0.15 0.19 (0.10–0.40)
C 85 0.32 243 0.46 1.82 (1.34–2.48) C 85 0.32 83 0.47 1.92 (1.30–2.84)
T 183 0.68 287 0.54 0.55 (0.40–0.75) T 183 0.68 93 0.53 0.52 (0.35–0.78)
−634GNC −634GNC
G/G 85 0.63 164 0.62 0.93 (0.61–1.44) G/G 85 0.63 47 0.53 0.66 (0.38–1.14)
G/C 44 0.33 84 0.32 0.94 (0.61–1.48) G/C 44 0.33 30 0.34 1.05 (0.60–1.87)
C/C 5 0.04 17 0.06 1.76 (0.64–4.90) C/C 5 0.04 11 0.13 3.68 (1.23–11.0)
G 214 0.80 412 0.22 0.88 (0.61–1.27) G 214 0.80 124 0.7 0.60 (0.39–0.93)
C 54 0.20 118 0.78 1.14 (0.79–1.63) C 54 0.20 52 0.3 1.66 (1.07–2.58)

Significant ORs are bolded. Significant ORs are bolded.

 K. Janik-Papis et al. / Experimental and Molecular Pathology 87 (2009) 234–238 237

Table 4
Distribution of genotypes, frequency of alleles of the −460CNT polymorphism of the
VEGF gene and odds ratio (OR) with 95% confidence interval (95% CI) in patients
with wet age-related macular degeneration (AMD) and individuals without visual
disturbances (controls).
disease: the CG haplotype favored both forms of AMD, whereas the TG
haplotype protected against both forms (Table 6).
Discussion

Genotype Controls (n = 134) Wet AMD (n = 177) OR (95% CI)

The pathogenesis of AMD is not completely known, and several
or allele
Number Frequency Number Frequency genetic factors in combination with environmental, lifestyle, and
C/C 11 0.08 18 0.10 1.26 (0.58–2.78) physiological factors may be involved. Growth factors are natural
C/T 63 0.47 124 0.70 2.63 (1.65–4.21) candidates for contributors to the wet form of AMD because it
T/T 60 0.45 35 0.20 0.30 (0.18–0.50) involves neovascularization. However, the relationship between dry
C 85 0.32 160 0.45 1.78 (1.27–2.47)
T 183 0.69 194 0.55 0.56 (0.40–0.78)
and wet forms of AMD is not unequivocal. If wet AMD is a later stage of
dry AMD, which can be justified in some cases, the changes involving
Significant ORs are bolded.
expression of the VEGF gene and the function of its product could not
be detected by clinical methods. However, we cannot exclude the
possibility that these changes may be revealed by methods of
genotypes did not differ significantly from those expected by the molecular biology. That is why we decided to study genetic variability
Hardy–Weinberg equilibrium. in the VEGF gene in both forms of AMD. We studied genotype
We observed a weak association (OR 2.90, 95% CI 1.89–4.48) distribution of these two polymorphisms as well as combined
between AMD and the C/T genotype of the −460CNT polymorphism genotypes and haplotypes because susceptibility to complex diseases
of the VEGF gene (Table 2). In general, the C allele of this may be attributed to multiple alleles whose concerted action may
polymorphism seemed to be positively correlated with the occurrence contribute to the disease. The prevalence of women in all groups
of AMD, whereas the T allele had a rather protective effect against (Table 1) follows from the fact that female gender is a risk factor for
AMD. We did not observe any association between the occurrence of AMD (Jager et al., 2008).
overall (dry and wet) AMD and genotypes of the other polymorphism, The role of genetic variability in AMD has been extensively studied.
−634GNC (Table 2). However, a direct link between AMD occurrence and mutations in
An association (OR 3.77) between the C/T genotype of the genes responsible for macular dystrophies has been established only
−460CNT polymorphism and occurrence of the dry form of AMD in some cases of familial AMD (Allikmets et al., 1997). However, most
was observed (Table 3). Furthermore, the T/T genotype considerably AMD cases have no known family connotation.
lowered the risk of dry AMD (OR 0.19). Generally, the C allele favored As we mentioned previously, the −634GNC polymorphism was
the dry form of AMD and the T allele protected against it. The not associated with AMD in the largest study performed so far, The
occurrence of dry AMD was correlated with the C/C genotype of the Rotterdam Study (Boekhoorn et al., 2008). This is in apparent contrast
−634GNC polymorphism (OR 3.68; Table 3). The G allele protected with the results we obtained. We think that the main reason for this
against this form of the disease and the C allele favored it. discrepancy might be ethnic differences in the studied populations.
Another weak association (OR 2.63) was found between the C/T Inhabitants of the Ommoord region (Rotterdam suburb) were
genotype of the −460CNT polymorphism and the occurrence of wet enrolled in the Rotterdam Study. We invited people living in Warsaw
AMD (Table 4). The C allele of this polymorphism was positively and its close vicinity. However, we might expect some ethnic
associated with the occurrence of wet AMD, whereas the T allele differences in the studied populations because the Dutch people are
protected against it. There was a lack of any association between wet in general more ethnically diverse than the Polish people. In the
AMD and the other polymorphism of the VEGF gene (results not Rotterdam Study, the population was 99% white, whereas our
shown). population was 100% white. Next, as we mentioned previously,
The occurrence of AMD was correlated with the presence of AMD may be regarded as a result of a complex interaction of genetic
the combined C/T-G/G genotype of both polymorphisms (OR 2.41, and environmental factors. We might suspect that the local
Table 5), whereas the T/T-G/G genotype exerted a protective effect environment of Warsaw, with a population of more than 2 million
against the disease (OR 0.22). The presence of the C/T-G/G genotype and heavy industry plants, may be quite different from that of
increased the risk of dry AMD (OR 2.08), whereas the presence of the Ommoord, with about 25 thousand inhabitants. However, our
T/T-G/G genotype decreased the risk (OR 0.15). In the case of wet research does not have the high statistical power of the Rotterdam
AMD, the combined genotype C/T-G/G slightly favored the disease Study. In light of the likely implication of VEGF in the process of
(OR 2.61) and the T/T-G/G genotype had a protective effect (OR 0.25). pathogenesis of at least the wet form of AMD, the association of some
The analysis of haplotypes of both polymorphisms yielded similar polymorphisms of the VEGF gene with the level of expression of AMD
results for AMD in general as well as for the dry and wet forms of the and the contradictory results obtained so far, the research on the role

Table 5
Distribution of haplotypes of the −460CNT and −634GNC polymorphisms of the VEGF gene and odds ratio (OR) with 95% confidence interval (95% CI) in patients with dry or wet age-
related macular degeneration (AMD) and individuals without visual disturbances (controls).

Haplotype Controls (n = 134) AMD (n = 265) OR (95% CI) Dry AMD (n = 88) OR (95% CI) Wet AMD (n = 177) OR (95% CI)

Number (frequency) Number (frequency) Number (frequency) Number (frequency)

C/C–G/G 7 (0.05) 17 (0.07) 1.24 (0.50–3.08) 4 (0.05) 0.86 (0.25–3.04) 13 (0.07) 1.44 (0.59–3.71)
C/C–G/C 4 (0.03) 9 (0.04) 1.14 (0.35–3.78) 4 (0.05) 1.55 (0.38–6.36) 5 (0.03) 0.94 (0.25–3.59)
C/C–C/C 0 0 – 0 – 0 –
C/T–G/G 35 (0.26) 122 (0.48) 2.41 (1.53–3.80) 37 (0.42) 2.05 (1.16–3.64) 85 (0.48) 2.61 (1.61–4.25)
C/T–G/C 25 (0.19) 60 (0.23) 1.28 (0.76–2.15) 23 (0.26) 1.54 (0.81–2.94) 37 (0.21) 1.15 (0.65–2.03)
C/T–C/C 3 (0.02) 9 (0.04) 1.54 (0.41–5.77) 79 (0.08) 3.77 (0.95–15.0) 2 (0.01) 0.50 (0.08–3.03)
T/T–G/G 43 (0.32) 25 (0.10) 0.22 (0.13–0.38) 6 (0.07) 0.15 (0.06–0.38) 19 (0.11) 0.25 (0.14–0.46)
T/T–G/C 15 (0.11) 15 (0.06) 0.48 (0.23–1.01) 3 (0.03) 0.28 (0.08–1.00) 12 (0.07) 0.58 (0.26–1.28)
T/T–C/C 2 8 (0.03) 2.05 (0.43–9.81) 4 (0.05) 3.14 (0.56–17.5) 4 (0.02) 1.53 (0.28–8.46)

“ – ” not estimated.
238 K. Janik-Papis et al. / Experimental and Molecular Pathology 87 (2009) 234–238

Table 6
Distribution of haplotypes of the −460CNT and −634GNC polymorphisms of the VEGF gene and odds ratio (OR) with 95% confidence interval (95% CI) in patients with wet or dry age-
related macular degeneration (AMD) and individuals without visual disturbances (controls).

Haplotype Controls (n = 134) AMD (n = 265) OR (95% CI) Dry AMD (n = 88) OR (95% CI) Wet AMD (n = 177) OR (95% CI)

Number (frequency) Number (frequency) Number (frequency) Number (frequency)

CG 131 (0.24) 390 (36.8) 1.80 (1.43–2.27) 121 (0.34) 1.62 (1.21–2.18) 269 (0.38) 1.89 (1.48–2.43)
CC 33 (0.06) 78 (7.4) 1.21 (0.79–1.84) 31 (0.09) 1.47 (0.88–2.45) 47 (0.07) 1.08 (0.68–1.72)
TG 297 (0.56) 434 (40.9) 0.56 (0.45–0.69) 127 (0.36) 0.45 (0.34–0.60) 307 (0.43) 0.62 (0.49–0.77)
TC 75 (0.14) 158 (14.9) 1.08 (0.80–1.45) 73 (0.21) 1.61 (1.13–2.29) 85 (0.12) 0.84 (0.60–1.17)

Significant ORs are bolded.

of genetic variability of the VEGF gene in the pathogenesis of AMD
should be continued.
Conclusion
The C/T genotype of the −460CNT polymorphism of the VEGF gene
may be associated with enhanced risk of dry and wet AMD in the
Polish population, whereas the T/T genotype may reduce the risk of
the dry form of this disease. Dry AMD may be associated with the C/C
genotype of another polymorphism of the VEGF gene, −634GNC. Both
polymorphisms may interact, producing haplotypes that modulate
the risk of AMD.
Conflict of interest statement
The authors declare that there are no conflicts of interest.
Acknowledgment
This work was supported by the Ministry of Science and Higher
Education, grant number 402 071 32/2210.
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