Heritability of myopia and its relation with GDJ2 and RASGRF1 genes in Lithuania

Refractive error is one of the priority targets of World Health Organization Vision 2020 [1]. More than 150 million people in the world are estimated to be visually impaired because of uncorrected refractive error, 8 million of them are functionally blind [2]. Myopia affects approximately one-third of adults older than 20 years in the United States, and in areas with high prevalence, specifically in urban East Asia, more than 80% of students graduating from school are myopic [3]. It is estimated that by the year 2020, 2.5 billion people - one third of the world’s population will have been affected by myopia alone [4]. Worldwide, myopia affects approximately one in four individuals in western population and is the most common visual disorder [4, 5].

In Lithuania, the data on refractive errors are systematically collected, and the official statistics are published by the Lithuanian Department of Statistics. In the population of Lithuania from 2007 to 2014, the prevalence of myopia increased from 44.3 to 63.9 per 1000 population (www.​sic.​hi.​lt).

Both genetic and environmental factors can affect development of myopia, but the exact causes are not fully understood [6]. The genetic contribution to a trait often is assessed through heritability. Heritability is the proportion of phenotypic variability in a population trait that is due to genetic differences [7]. Up to now, the heritability of myopia has not been studied in Lithuania.

Twin studies allow us to estimate the overall gene influence, and the results can show the heritability of myopia. The twin method assumes that monozygotic (MZ) twins are influenced by largely similar environmental differences as dizygotic (DZ) twins, but MZ twins share the same genes whereas DZ twins on average share only half their genes [8]. Twin method is excellent for the estimation of heritability of myopia, but it does not show the specific genes that may possibly be involved in the heritability of myopia. Thus, we chose genetic markers, in order to establish whether any of them are significantly associated with myopia.

Genome-wide association studies (GWAS) for refractive error showed that single nucleotide polymorphisms (SNPs) in 15q25 and 15q14 were associated with refractive error and myopia [9]. RASGFR1 is a gene made of 28 exons. This gene has a significant influence on development of myopia. RASGFR1 gene encodes Ras protein-specific guanine nucleotide-releasing factor-1, which is highly expressed in the retina and neurons. Then RASGFR1 gene proceeds to activate Ras [10, 11]. Also, RASGRF1 is a nuclear exchange factor that promotes GDP/GTP exchange on the Ras family GTPases and is related to synaptic transmission of the photoreceptor responses [12]. Muscarinic receptors and retinoic acid can regulate RASGRF1 expression as well [11]. Some animal and human studies showed that muscarinic inhibitors prevented the development of myopia [13]. In animal models of myopia there was detected reduced synthesis of choroidal retinoic acid [14]. To date, studies of SNP rs8027411 of RASGRF1 gene associations with high myopia in different populations have provided controversial results [6, 15‐17].

The GJD2 gene at 15q14 encodes a neuron-specific protein connexin 36 (CX36), a 36 kDa protein, which is a neuron-specific protein of a family of integral membrane proteins [18]. CX36 forms gap junction channels between adjacent membranes of neuronal cells. It is present in photoreceptors, bipolar and amacrine cells, and, by enabling intercellular transport of small molecules and ions, plays an essential role in the transmission process of the retinal electric circuitry [18, 19].

The aim of our research was to find associations between the GJD2, RASGRF1 genes and myopia development and to assess the heritability of myopia in Lithuania.

230 pairs of twins (135 MZ and 95 DZ) aged between 18 and 40 participated in the study, their mean age being 25.08 years (SE 0.7 years) (Table 1). The mean spherical equivalent was − 1.324 ± 0.150, with a range from 0.49 D to − 7.375 D. There were no significant differences between MZ and DZ twins in age and spherical equivalent of the left and right eyes.

Refractive errors for twin 1 versus twin 2 for mean of spherical equivalent are shown in figs. 1 and 2. Intrapair correlations for spherical equivalent in twin pairs were significantly higher in MZ twin pairs r = 0.539 (p < 0.001, 95% CI 0.353–0.684) than in DZ twin pairs r = 0.203 (p < 0.01, 95% CI 0.0633–0.442) in myopia group. Correlations for spherical equivalent in emmetropia group were not significant: r = 0.091 (p > 0.05,995% CI -0.215-0.381) in MZ twin pairs and r = − 0.220 (p > 0.05, 95% CI -0.587-0.222) in DZ twin pairs. The correlations of MZ were clearly higher compared to DZ pairs, indicating genetic effects on myopia.

We genotyped two SNPs (rs634990 and rs8027411) in 189 myopia (83 MZ and 106 DZ) and 96 emmetropia (25 MZ and 71 DZ) subjects. The results are shown in Table 2. The distribution of the two SNP genotypes matched the Hardy-Weinberg equilibrium (p ≥ 0.05). High-degree myopia was present in 11 cases, 33 twins had medium-degree 10 and 145 twins mild-degree myopia. But we didn’t find significant correlations between myopia degree and genotypes.

5 models were used to calculate the odds ratios to have myopia separately with each gene (GJD2 or RASGRF1), results were not significant (Table 3, Table 4).

But we found significant association between the combinations of GJD2 CC and RASGRF1 GT and myopia (Table 5). The odds ratio of myopia compared to emmetropia (95% confidence intervals [CIs]) was 2.7 (1.018–7.460) for GJD2 CC and RASGRF1 GT genotypes.

The number with combinations of genotypes rs634990 CC and rs8027411 GT and myopia degrees are shown in Table 6.

We estimated heritability of myopia according to correlations for MZ and DZ twin pairs and our study showed 67.2% heritability of myopia. Three published twin studies of refractive error have found high heritability from 84 to 86% [23], 89 to 94% [24] and 75 to 88% [25]. It is indicate that heritability in Lithuania is lower than in other Europian populations. Dirani et al. have reported that different populations have shown a wide range of heritability estimates ranging from 50 to 90% [25]. Results shown that the samples of the population and different methods may affect the estimates of heritability [25]. In our study 77% twins had mild-degree myopia, 17% - medium-degree and 6% - high-degree myopia. Meanwhile, the medium-degree myopia accounted for the largest portion in the mentioned studies of heritability in Europe.

Study showed that the gene GJD2, located nearest to the locus 15q14, and RASGRF1 15q25 are important for the transmission and processing of visual signals [23, 26]. The studies of genetic associations in some European and Japanese populations showed that common genetic variations located in GJD2 and RASGRF1 were associated with common myopia and refractive error [5, 11, 15].

A study of genome-wide associations (GWASs) showed associations of SNP with refractive error in 5328 individuals of the Dutch population which were not related. They found that carriers of the C allele of rs634990 have a higher risk of myopia [5]. Qiang et al. found that RASGRF1 gene was significantly associated with high-degree myopia (risk allele T) but GJD2 gene was not [15]. Also, results of meta-analysis, which included 2529 individuals with high-degree myopia and 3127 controls, showed that RASGRF1 was significantly associated with high-degree myopia in Chinese and Japanese populations. However, carriers of the RASGRF1 G allele had a lower risk of high-degree myopia compared to carriers of the T allele (G versus T) [27]. Also, Hysi et al. found that individuals carrying TT alleles on the RASGRF1 were significantly more likely to have myopia than those homozygous for the non-susceptibility GG alleles. We found a significant association between combinations of GJD2 and RASGRF1 genotypes and myopia. Our study showed that individuals with combinations of GJD2 CC and RASGRF1 GT genotypes were 2.7 times more likely to have myopia (p = 0.046). This indicates that some of our results are consistent with the previous reports. Individuals carrying CC alleles on the GJD2 were significantly more likely to have myopia than carriers of TT alleles. But carriers of GT allele on the RASGRF1 gene had more risk to have myopia than carriers of wild type alleles.

Our studies have shown that the heritability of myopia makes 66.4% in Lithuania. We detected significant associations between the combinations of GJD2 CC and RASGRF1 GT and odds ratio of developing myopia.