Association of LIPC and advanced age-related macular degeneration

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Abstract

Purpose

To determine whether there is an association between hepatic lipase (LIPC) and age-related macular degeneration (AMD) in two independent Caucasian cohorts.

Methods

A discovery cohort of 1626 patients with advanced AMD and 859 normal controls and a replication cohort of 2159 cases and 1150 controls were genotyped for two single-nucleotide polymorphisms (SNPs) in the promoter region of LIPC. The associations between the SNPs and AMD were examined by χ2 tests.

Results

In the discovery cohort, rs493258 and rs10468017 were both associated with advanced AMD (P=9.63E−3 and P=0.048, respectively). The association was corroborated in the replication cohort (P=4.48E−03 for rs493258 and P=0.015 for rs10468017). Combined analysis resulted in even more significant associations (P=1.21E−04 for rs493258 and P=1.67E−03 for rs10468017).

Conclusion

The LIPC promoter variants rs493258 and rs10468017 were associated with advanced AMD in two independent Caucasian populations, confirming that LIPC polymorphisms may be a genetic risk factor for AMD in the Caucasian population.

Introduction

Age-related macular degeneration (AMD) is the leading cause of irreversible blindness for elderly individuals in the developed world. It is estimated that >9 million people in the United States suffer from intermediate or advanced AMD.1, 2 Depending on the severity, AMD can be classified into two separate stages. Early AMD is characterized by soft drusen and pigmentary changes in the retinal pigment epithelium (RPE). Advanced AMD leads to vision loss and can be further subdivided into geographic atrophy (GA, dry AMD) and choroidal neovascularization (wet AMD). Despite the high prevalence and significant public health burden of AMD, its etiology and pathophysiology remain poorly understood.

AMD is a multi-factorial progressive disease that involves complex interactions between genetic and environmental factors.3, 4 Candidate gene association studies have identified multiple genes related to AMD, including CFH,5, 6 ARMS2/HTRA1,7, 8, 9 C2I,10 and CFB.11 Identification of these genes has led to extensive studies on the alternative complement pathway and mitochondria-related oxidation pathways in relation to AMD.12, 13, 14 Despite significant progress in identifying AMD-associated genes, genetic susceptibility loci discovered thus far only account for approximately half the heritability of AMD.15 Recent studies demonstrated that antioxidant micronutrients and lipids may also have an important role in AMD;16, 17, 18, 19 a genome-wide association study found a significant association between advanced AMD and hepatic lipase (LIPC), the gene encoding hepatic triglyceride lipase.20 In this study, we confirmed the association of LIPC with advanced AMD in two independent Caucasian cohorts.

Materials and methods

Subjects and clinical diagnosis

This study was approved by the Institutional Review Board of the University of California, San Diego, CA, USA. The research adhered to the tenets of the Declaration of Helsinki. Informed consent was signed by all subjects before participation in the study. We certify that all applicable institutional and governmental regulations concerning the ethical use of human volunteers were followed during this research.

In total, 1626 nonfamilial advanced AMD patients and 859 normal age-matched controls (age 50 years or older without drusen or RPE changes) were recruited at the University of California, San Diego, CA, USA. Demographics, medical history, and a blood sample were collected at the baseline visit. All participants underwent standard ophthalmic examinations, including slit lamp exams and indirect ophthalmoscopy. A pair of stereoscopic color fundus photographs (50°) was taken, centered on the fovea using a Topcon fundus camera (Topcon TRV-50VT, Topcon Optical Company, Tokyo, Japan). Grading was carried out using a standard grid classification suggested by the International Age-related Maculopathy Epidemiological Study Group.21 In total, 2159 cases and 1150 controls from the Michigan, Mayo, AREDS, and Pennsylvania data set were included in this study as a replication cohort.

Genotyping

Genomic DNA samples were extracted from peripheral blood leukocytes with the Qiagen kit (Qiagen Inc., Chatsworth, CA, USA), according to the manufacturer’s instructions. The discovery cohort of 1626 advanced AMD patients were genotyped for the LIPC variants rs10468017 and rs493258. Variance and position of the two single-nucleotide polymorphisms (SNPs) are shown in Supplementary Table 1. Allele frequencies were compared with 859 age and ethnicity-matched normal controls by laboratory personnel blinded to case/control status. The findings were tested for replication in an independent cohort of 2159 advanced AMD patients and 1150 controls.

Genotyping of both SNPs was achieved by primer extension of multiplex PCR products followed by SNaPshot on an ABI 3100 genetic analyzer (Applied Biosystems, Foster City, CA, USA). All genotyping results were of high quality, with an average call rate of 98.5%.

Statistical analysis

Deviation from Hardy–Weinberg equilibrium was assessed with a statistical significance level of 0.01. χ2 tests under the additive model were performed to assess evidence for association between the LIPC genotype and advanced AMD between cases and controls. The statistical significance level was adjusted by Bonferroni correction. Odds ratios and 95% confidence intervals were also calculated to estimate risk size of the risk alleles by using SPSS 11.5 software (Chicago, IL, USA). Linkage disequilibrium (LD) patterns were defined by Haploview 4.1 software (Cambridge, MA, USA).

Results

The demographics of the discovery cohort are presented in Table 1. The promoter variant rs493258 was found to be significantly associated with advanced AMD in the discovery cohort (P=9.63E−03), replication cohort (P=4.48E−03), and combined cohort (1.21E−04; Table 2a). For rs10468017, a significant association with advanced AMD was also found in the discovery cohort (P=0.048), replication cohort (P=0.015), as well as the combined cohort (P=1.67E−03; Table 2b).

Table 1 Discovery cohort demographics
Table 2 (a) Association between LIPC-rs493258 and advanced AMD; (b) association between LIPC-rs10468017 and advanced AMD

Overall, the frequency of the minor (A) allele of rs493258 was 43.4% in cases vs 47.1% in controls, whereas the frequency of the minor (T) allele of rs10468017 was 26.7% in cases vs 29.5% in controls. Haplotype analysis did not show strong LD between the two SNPs (r2=0.369). Genotype counts are shown in Supplementary Tables 2a and b.

Discussion

Here, we confirmed the genetic association between AMD and LIPC and expand upon previous reports by analyzing two independent Caucasian populations. Our findings were consistent with previous studies in which both SNPs were reported to be significantly associated with AMD.20, 22

The LIPC gene encodes hepatic triglyceride lipase, which is expressed in the liver. LIPC catalyzes the hydrolysis of phospholipids, mono-, di-, and triglycerides, and acyl-CoA thioesters, and it has an important role in the metabolism of lipoproteins, including high-density lipoprotein (HDL)23, 24, 25 and low-density lipoprotein.26 Previous studies have demonstrated that LIPC is associated with metabolic syndrome,27, 28 atherosclerosis,29, 30 and other cardiovascular disorders.31, 32

The mechanism(s) underlying the relationship between LIPC and AMD is unknown. Despite the association of LIPC and serum HDL levels, the correlation between AMD and serum HDL levels is inconsistent.21, 33 These findings suggest that LIPC may modulate AMD risk through a mechanism independent from its effect on HDL levels. Central to AMD progression is the buildup of cellular debris, proteins, and lipids within Bruch’s membrane, which results in the formation of drusen.34 These changes in Bruch’s membrane result in tissue hypoxia and the production of angiogenic signaling molecules (eg, VEGF) that promote aberrant blood vessel growth.35 In addition, oxidative stress in the eye can cause the oxidation of phospholipids. These oxidation products are pro-inflammatory and can also lead to RPE apoptosis, VEGF production, and angiogenesis.36, 37 As LIPC is an important enzyme in lipid metabolism and has been shown to be related to both the accumulation of drusen and progression from large drusen to advanced AMD,38 it may modulate AMD risk by affecting lipid homeostasis and the accumulation of damaging biomolecules in the eye. Further studies evaluating the role of lipoproteins in the pathogenesis and progression of AMD will elucidate the role of LIPC in AMD and may lead to novel strategies for AMD prevention and treatment.

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Acknowledgements

We thank Chao Zhao, Kevin Wang, Daniel Kasuga, and Jean Guan for their assistance with this study. We also thank all the participating AMD patients and their families. This study is supported by 973 Program (2011CB510200, 2013CB967504); Genentech, NEI/NIH (Bethesda), KACST -UCSD Center of Excellence in Nanomedicine, Research to Prevent Blindness (New York), and VA Merit Award (San Diego).

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Correspondence to K Zhang.

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Supplementary Information accompanies this paper on Eye website

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Keywords

  • LIPC
  • hepatic lipase
  • advanced age-related macular degeneration
  • genetics

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