Three Single Nucleotide Polymorphisms of LOXL1’ in a Turkish Population with Pseudoexfoliation Syndrome and Pseudoexfoliation Glaucoma

Yetkin Yaz; Nilgün Yıldırım; Yasemin Aydın Yaz; Oğuz Çilingir; Zafer Yüksel; Fezan Mutlu

doi:10.4274/tjo.83797

ABSTRACT

Objectives:

To investigate the three single nucleotide polymorphisms (SNPs) (rs3825942, rs1048661, and rs2165241) of the LOXL1 gene in pseudoexfoliation syndrome (XFS) and pseudoexfoliation glucoma (XFG) in the Turkish population.

Materials and Methods:

DNA was obtained from blood samples of 48 XFS, 58 XFG, and 171 control subjects. Three LOXL1 SNPs (rs3825942, rs1048661, rs2165241) were investigated with real time PCR, a probe-based genotyping method, and melting curve analysis.

Results:

All three SNPs of LOXL1 were significantly associated with XFS (rs3825942 p=3.54x10^-6, odds ratio [OR]=∞; rs1048661 p=0.008, OR=2.18; rs2165241 p=8.69x10^-9, OR=4.30) and XFG (rs3825942 p=3.41x10^-7, OR=∞; rs1048661 p=1.75x10^-5, OR=3.78; rs2165241 p=3.85x10^-11 OR=4.90). No significant differences were observed between the XFS and XFG groups for any of the SNPs. The GG genotype of rs3825942 was more valuable for distinguishing pseudoexfoliative cases from healthy individuals. The homozygous TT genotype of rs2165241 was associated with 6-fold increased XFS risk (p=8.15x10^-8, OR=6.32) and 7-fold increased XFG risk (p=1.45x10^-10 OR=7.95). The GGT haplotype consisting of all three risk alleles was associated with a 7.45-fold higher risk of XFS/XFG (p=8.65x10^-14, OR=7.45). Presence of T allele of rs2165241 conferred 3 times higher risk for men than women (p=6.78x10^-5, OR=3.202).

Conclusion:

LOXL1 SNPs are associated with increased risk for pseudoexfoliation in the Turkish population. T allele of rs2165241 was found to be the most important characterized risk factor for our cohort. All SNP distributions were similar to other European and American populations.

Keywords:

Pseudoexfoliation syndrome, pseudoexfoliation glaucoma, LOXL1 gene

Introduction

Pseudoexfoliation glaucoma (XFG) is the most common identifiable cause of secondary open-angle glaucoma worldwide.¹ Pseudoexfoliation is an age-related systemic disorder characterized by progressive accumulation of abnormal fibrillar extracellular material in ocular and extraocular tissues.^1,2 Pseudoexfoliation syndrome (XFS) is diagnosed based on the appearance of pseudoexfoliation material (PXM) on anterior segment structures like the pupillary border of the iris or the anterior lens capsule. XFS has been associated with increased risk of cataract and glaucoma^3,4 and predisposition to a broad range of intraocular complications during surgery.^1,2

The structure and origin of PXM has not been determined. A recent genome-wide association (GWA) study identified the LOXL1 gene, which encodes an enzyme necessary for elastogenesis and collagen cross-linking, as a major genetic risk factor for developing PXF.^5,6 Three single nucleotide polymorphisms (SNPs) have been associated with PXF: one intronic SNP, rs2165241 located in intron 1, and two nonsynonymous coding SNPs, rs3825942 (G153D) and rs1048661 (R141L) located in exon 1.⁵ However, the frequency of risk alleles for both variants varies among different populations. Based on these studies, LOXL1 is necessary but not sufficient for PXM development.⁵ Due to the relationship between PXM, the extracellular matrix, and basement membrane, the role of different genes were investigated in PXM formation. These genes were related to elastic fiber and extracellular matrix metabolism, such as MMP1, MMP3, FBN1, LTBP2, MFAP2, TGM2, TGF-b1, and CLU.^7,8,9 In addition, LOXL1 promoter haplotypes, which influence gene expression and lead to reduced enzyme activity, are associated with XFS/XFG.⁵

The purpose of this study was to investigate variant SNPs (rs2165241, rs3825942 and rs1048661) of the LOXL1 gene to determine the association of LOXL1 polymorphism with XFG and XFS in the Turkish population.

Materials and Methods

Results

A total of 277 individuals (58 XFG, 48 XFS, 171 controls) over 40 years old were recruited for this study in Eskişehir Osmangazi University Department of Glaucoma. The mean age was 68 (66.5-70) years for the XFS group, 69 (68-73.25) years for the XFG group, and 67 (64-70) years for control subjects.

The genotype distribution of all SNPs conformed to Hardy-Weinberg equilibrium. In both XFS and XFG, a strong association with the risk allele of each individual SNP (rs2165241T, rs1048661G, and rs3825942G) was observed (Table 2). The G allele of rs3825942 was present in all XFS and XFG patients, thus OR could not be calculated (OR=∞ p=3.54x10^-6, OR=∞, p=3.41x10^-7). In the control group, G allele for rs3825942 was more common than the A allele. For rs3825942, all patients in the XFS and XFG groups had the GG genotype, and again ORs could not be calculated (OR=∞ p=1.57x10^-6, OR=∞, p=1.45x10^-6). The G allele and GG genotype of rs1048661 were detected more frequently in all groups. The T allele of rs1048661 was underrepresented in patients with XFS (OR=2.18 95% CI=1.21-3.91, p=0.008) and XFG (OR=3.78 95% CI=1.99-7.18, p=1.75x10^-5) when compared to control subjects. No TT genotype of rs1048661 was detected in XFS (OR=1.93 95% CI=0.98-3.77, p=0.076) or XFG (OR=3.71 95% CI=1.83-7.47, p=2.75x10^-14) patients. In control subjects, the TT genotype of rs1048661 was detected more frequently when compared with XFS and XFG patients. However, the GG and GT genotype of rs1048661 were overrepresented in control subjects when compared to the TT genotype of rs1048661. For rs2165241, the T allele was more frequent than C allele in XFS (OR=4.30 95% CI=2.55-7.25 p=8.69x10^-9) and XFG (OR=4.90 95% CI=2.98-8.06 p=3.85x10^-11) patients, while in control subjects the C allele was more common. The genotype distribution of rs2165241 was different in XFS and XFG patients and control subjects. The TT genotype of rs2165241 was overrepresented and the CC genotype of rs2165241 was underrepresented in XFS (OR=6.32 95% CI=3.16-12.64, p=8.15x10^-8) and XFG (OR=7.95 95% CI=4.10-15.42, p=1.45x10^-10) patients when compared to control subjects. The CT genotype was the most frequent genotype in control subjects. For three SNPs the TT genotype of rs2165241 was detected more frequently in XFS (58%) and XFG (63%) when compared to control subjects (19%). Likewise, the homozygous TT genotype of rs2165241 was associated with 6.32-fold higher risk of XFS (95% CI=3.16-12.64) and 7.95-fold higher risk of XFG (95% CI=4.10-15.42).

For three SNPs, rs3825942, rs1048661, rs2165241, haplotype analysis of risk alleles was calculated to determine the combined effects on pseudoexfoliation patients (XFS+XFG) and control subjects (Table 3). The haplotypes consisting of risk alleles were overrepresented in pseudoexfoliation patients as compared to control subjects for each SNP (p=8.65x10^-14) and were associated with 7.45-fold increased risk of pseudoexfoliation (95% CI=4.22-12.99).

According to previous studies, females are affected by XFS more often than males, whereas XFG is more severe in males than females.² In our allele analysis of all pseudoexfoliation patients (XFS+XFG) based on gender, the T allele of rs2165341 was detected in 75% of males and 49% of females (Table 4). Existence of T allele was associated with 3.2 times higher risk for men than women (p=6.78x10-5, c²=15.871, OR=3.202 95% CI=1.719-5.989).

Discussion

After Thorleifsson et al.⁵ from Iceland identified three associated polymorphisms of LOXL1 in XFS/XFG, many studies were performed on Caucasian, Asian, and African populations.^{10,11,12,13,14,15,16,17,18,19,20,21,22,23,24,25,27,28,29,30,31,32} In our study we investigated the association between three SNPs of LOXL1 (rs1048661, rs3825942, rs2165241) and XFS/XFG in the Turkish population. Our findings show a significant association with XFS, as in other recent studies. However, geographically Turkey is located in both Asia and Europe, and this association in our population was similar to that observed in Caucasians, as in European populations.

In our study, the relationship between G allele and GG genotype of rs3825942, G allele and GG genotype of rs1048661, and T allele and TT genotype of rs2165241 was found in XFS/XFG, as in the Caucasian population.^{10,11,12,13,14,15,16,17,18,19,20,21,22} In the present study the G allele and GG genotype of rs3825942 was present in all XFS and XFG cases. According to the results of an African study and another Turkish study¹¹ for rs3825942, the A allele carries an increased risk for XFS.^11,32 However, in our study and another study from Turkey, the A allele was not detected in any XFS and XFG cases and was suggested to be protective.¹⁰ Similarly, in another Turkish study, the G allele GG genotype of rs1048661 were detected in exon 1 of LOXL1, as seen in the patients in our study group.¹²

Studies in European and American populations reported that G allele and GG genotype of rs1048661 and T allele and TT genotype of rs2165241 were associated with XFS, whereas studies in Asian populations identified the opposite relationship for risk alleles and genotypes for these SNPs.^{23,24,25,27,28,29,30,31} Our results appear to be similar to Caucasians but different from Asians in terms of allelic and genotypic distributions of rs1048661 and rs2165241.^{10,11,12,13,14,15,16,17,18,19,20,21,22} Another study from Turkey indicated that T allele and TT genotype of rs2165241 were associated with XFS/XFG, like our study, but revealed no significant relationship with G allele and GG genotype of rs1048661.¹³ For this reason, the pathogenesis of XFS cannot be explained by genetic factors alone.

In several studies researching whether LOXL1 gene polymorphism has any role in predicting XFG development, no significant association was found in the differentiation of XFS and XFG.¹ Likewise, in our study, no differences in LOXL1 polymorphism were found between the XFS and XFG groups. On the other hand, recent studies have shown that females were affected more frequently by XFS than males, whereas XFG was more severe in males compared to females.² We observed a strong relationship between the T risk allele in the rs216341 SNP and gender in XFS and XFG, with men carrying the T allele showing at 3 times higher risk of disease. However, a study from Japan did not show significant gender differences in any of the three SNPs.²⁸

In our study, the haplotype (GGT) consisting of all three risk alleles of LOXL1 SNPs (rs3825942 G, rs1048661 G and rs2165341 T) was associated with a 7.45-fold higher risk of XFS/XFG. A study in an American population reported a 3-fold higher risk⁶ and a study in a Polish population determined a 4-fold higher risk with the GGT haplotype.²²

Conclusion

Our findings are similar to previous Turkish studies that investigated two non-synonymous coding SNPs rs3825942 and rs1048661, but we observed intronic SNP rs2165341 as well. Our results support the existence of a significant association between three SNPs of LOXL1 with both XFS and XFG, though no significant differences were found between the XFS and XFG patients. Also different from the genotypes of exonic SNPs (rs3825942, rs1048661), the TT genotype of rs2165341 was detected more frequently in pseudoexfoliation and was associated with 6-fold and 7-fold increases in risk of XFS and XFG, respectively. Likewise, we observed a relationship between the T allele of rs2165341 and gender, as presence of the T allele was associated with 3 times higher risk for men compared to women. To the best of our knowledge, rs2165341 is an important and risk-modifying factor in our cohort. In conclusion, LOXL1 gene polymorphism has a significant influence on XFS and XFG pathogenesis, but is inadequate to explain the exact mechanism. Therefore, further genetic and epigenetic studies are needed.

References

Ritch R, Schlötzer-Schrehardt U. Exfoliation syndrome. Surv Ophthalmol. 2001;45:265–315. [PubMed] [Google Scholar]

Schlötzer-Schrehardt U, Naumann GO. Ocular and systemic pseudoexfoliation syndrome. Am J Ophthalmol. 2006;141:921–937. [PubMed] [Google Scholar]

Schlötzer-Schrehardt UM, Koca MR, Naumann GO, Volkholz H. Pseudoexfoliation syndrome. Ocular manifestation of a systemic disorder? Arch Ophthalmol. 1992;110:1752–1756. [PubMed] [Google Scholar]

Ritch R. Exfoliation syndrome: The most common identifiable cause of openangle glaucoma. J Glaucoma. 1994;3:176–178. [PubMed] [Google Scholar]

Thorleifsson G, Magnusson KP, Sulem P, Walters GB, Gudbjartsson DF, Stefansson H, Jonsson T, Jonasdottir A, Jonasdottir A, Stefansdottir G, Masson G, Hardarson GA, Petursson H, Arnarsson A, Motallebipour M, Wallerman O, Wadelius C, Gulcher JR, Thorsteinsdottir U, Kong A, Jonasson F, Stefansson K. Common sequence variants in the LOXL1 gene confer susceptibility to exfoliation glaucoma. Science. 2007;317:1397–1400. [PubMed] [Google Scholar]

Fan BJ, Pasquale L, Grosskreutz CL, Rhee D, Chen T, DeAngelis MM, Kim I, del Bono E, Miller JW, Li T, Haines JL, Wiggs JL. DNA sequence variants in the LOXL1 gene are associated with pseudoexfoliation glaucoma in a U.S. clinicbased population with broad ethnic diversity. BMC Med Genet. 2008;9:5. [PMC free article] [PubMed] [Google Scholar]

Morrison JC, Green WR. Light microscopy of the exfoliation syndrome. Acta Ophthalmol Suppl. 1988;66:5–27. [PubMed] [Google Scholar]

Schlötzer-Schrehardt U. Genetics and genomics of pseudoexfoliation syndrome/glaucoma. Middle East Afr J Ophthalmol. 2011;18:30–36. [PMC free article] [PubMed] [Google Scholar]

Fingert JH, Alward WL, Kwon YH, Wang K, Streb LM, Sheffield VC, Stone EM. LOXL1 mutations are associated with exfoliation syndrome in patients from the Midwestern United States. Am J Ophthalmol. 2007;144:974–975. [PubMed] [Google Scholar]

Kasım B, İrkeç M, Alikaşifoğlu M, Orhan M, Mocan MC, Aktaş D. Association of LOXL1 gene polymorphisms with exfoliation syndrome/glaucoma and primary open angle glaucoma in a Turkish population. Mol Vis. 2013;19:114–120. [PMC free article] [PubMed] [Google Scholar]

Asfuroglu M, Cavdarli B, Koz OG, A Yarangumeli A, Ozdemir EY. Association of Lysyl Oxidase-Like 1 Gene Polymorphism in Turkish Patients With Pseudoexfoliation Syndrome and Pseudoexfoliation Glaucoma. J Glaucoma. 2017;26:54–57. [PubMed] [Google Scholar]

Yilmaz SG, Palamar M, Onay H, Ilim O, Aykut A, Ozkinay FF, Yagci A. LOXL1 gene analysis in Turkish patients with exfoliation glaucoma. Int Ophthalmol. 2016;36:629–635. [PubMed] [Google Scholar]

Yaylacioğlu Tuncay F, Aktaş Z, Ergün MA, Ergün SG, Hasanreisoğlu M, Hasanreisoğlu B. Association of polymorphisms in APOE and LOXL1 with pseudoexfoliation syndrome and pseudoexfoliation glaucoma in a Turkish population. Ophthalmic Genet. 2017;38:95–97. [PubMed] [Google Scholar]

Aragon-Martin JA, Ritch R, Liebmann J, O’Brien C, Blaaow K, Mercieca F, Spiteri A, Cobb CJ, Damji KF, Tarkkanen A, Rezaie T, Child AH, Sarfarazi M. Evaluation of LOXL1 gene polymorphisms in exfoliation syndrome and exfoliation glaucoma. Mol Vis. 2008;14:533–541. [PMC free article] [PubMed] [Google Scholar]

Challa P, Schmidt S, Liu Y, Qin X, Vann RR, Gonzalez P, Allingham RR, Hauser MA. Analysis of LOXL1 polymorphisms in a United States population with pseudoexfoliation glaucoma. Mol Vis. 2008;14:146–149. [PMC free article] [PubMed] [Google Scholar]

Yang X, Zabriskie NA, Hau VS, Chen H, Tong Z, Gibbs D, Farhi P, Katz BJ, Luo L, Pearson E, Goldsmith J, Ma X, Kaminoh Y, Chen Y, Yu B, Zeng J, Zhang K, Yang Z. Genetic association of LOXL1 gene variants and exfoliation glaucoma in a Utah cohort. Cell Cycle. 2008;7:521–524. [PubMed] [Google Scholar]

Hewitt AW, Sharma S, Burdon KP, Wang JJ, Baird PN, Dimasi DP, Mackey DA, Mitchell P, Craig JE. Ancestral LOXL1 variants are associated with pseudoexfoliation in Caucasian Australians but with markedly lower penetrance than in Nordic people. Hum Mol Genet. 2008;17:710–716. [PubMed] [Google Scholar]

Mossböck G, Renner W, Faschinger C, Schmut O, Wedrich A, Weger M. Lysyl oxidase-like protein 1 (LOXL1) gene polymorphisms and exfoliation glaucoma in a Central European population. Mol Vis. 2008;14:857–861. [PMC free article] [PubMed] [Google Scholar]

Pasutto F, Krumbiegel M, Mardin CY, Paoli D, Lämmer R, Weber BH, Kruse FE, Schlötzer-Schrehardt U, Reis A. Association of LOXL1 common sequence variants in German and Italian patients with pseudoexfoliation syndrome and pseudoexfoliation glaucoma. Invest Ophthalmol Vis Sci. 2008;49:1459–1463. [PubMed] [Google Scholar]

Lemmelä S, Forsman E, Onkamo P, Nurmi H, Laivuori H, Kivelä T, Puska P, Heger M, Eriksson A, Forsius H, Järvelä I. Association of LOXL1 gene with Finnish exfoliation syndrome patients. J Hum Genet. 2009;54:289–297. [PubMed] [Google Scholar]

Wolf C, Gramer E, Muller-Myhsok B, Pasutto F, Gramer G, Wissinger B, Weisschuh N. Lysyl Oxidase-like 1 gene polymorphisms in German patients with normal tension glaucoma, pigmentary glaucoma and exfoliation glaucoma. J Glaucoma. 2009;19:136–141. [PubMed] [Google Scholar]

Malukiewicz G, Lesiewska-Junk H, Linkowska K, Mielnik M, Grzybowski T, Sulima N. Analysis of LOXL1 single nucleotide polymorphisms in Polish population with pseudoexfoliation syndrome. Acta Ophthalmol. 2011;89:64–66. [PubMed] [Google Scholar]

Fuse N, Miyazawa A, Nakazawa T, Mengkegale M, Otomo T, Nishida K. Evaluation of LOXL1 polymorphisms in eyes with exfoliation glaucoma in Japanese. Mol Vis. 2008;14:1338–1343. [PMC free article] [PubMed] [Google Scholar]

Hayashi H, Gotoh N, Ueda Y, Nakanishi H, Yoshimura N. Lysyl oxidase-like 1 polymorphisms and exfoliation syndrome in the Japanese population. Am J Ophthalmol. 2008;145:582–585. [PubMed] [Google Scholar]

Mabuchi F, Sakurada Y, Kashiwagi K, Yamagata Z, Iijima H, Tsukahara S. Lysyl oxidase-like 1 gene polymorphisms in Japanese patients with primary open angle glaucoma and exfoliation syndrome. Mol Vis. 2008;14:1303–1308. [PMC free article] [PubMed] [Google Scholar]

Mori K, Imai K, Matsuda A, Ikeda Y, Naruse S, Hitora-Takeshita H, Nakano M, Taniguchi T, Omi N, Tashiro K, Kinoshita S. LOXL1 genetic polymorphisms are associated with exfoliation glaucoma in the Japanese population. Mol Vis. 2008;14:1037–1040. [PMC free article] [PubMed] [Google Scholar]

Tanito M, Minami M, Akahori M, Kaidzu S, Takai Y, Ohira A, Iwata T. LOXL1 variants in elderly Japanese with exfoliation syndrome/glaucoma, primary open-angle glaucoma, normal tension glaucoma, and cataract. Mol Vis. 2008;14:1898–905. [PMC free article] [PubMed] [Google Scholar]

Ozaki M, Lee KY, Vithana EN, Yong VH, Thalamuthu A, Mizoguchi T, Venkatraman A, Aung T. Association of LOXL1 gene polymorphisms with pseudoexfoliation in the Japanese. Invest Ophthalmol Vis Sci. 2008;49:3976–3980. [PubMed] [Google Scholar]

Chen L, Jia L, Wang N, Tang G, Zhang C, Fan S, Liu W, Meng H, Zeng W, Liu N, Wang H, Jia H. Evaluation of LOXL1 polymorphisms in exfoliation syndrome in a Chinese population. Mol Vis. 2009;15:2349–2357. [PMC free article] [PubMed] [Google Scholar]

Lee KY, Ho SL, Thalamuthu A, Venkatraman A, Venkataraman D, Pek DC, Aung T, Vithana EN. Association of LOXL1 polymorphisms with pseudoexfoliation in the Chinese. Mol Vis. 2009;15:1120–1126. [PMC free article] [PubMed] [Google Scholar]

Sagong M, Gu BY, Cha SC. Association of lysyl oxidase-like 1 gene polymorphisms with exfoliation syndrome in Koreans. Mol Vis. 2011;17:2808–2817. [PMC free article] [PubMed] [Google Scholar]

Williams SE, Whigham BT, Liu Y, Carmichael TR, Qin X, Schmidt S, Ramsay M, Hauser MA, Allingham RR. Major LOXL1 risk allele is reversed in exfoliation glaucoma in a black South African population. Mol Vis. 2010;16:705–712. [PMC free article] [PubMed] [Google Scholar]

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