Association Between Body Levels of Trace Metals and Glaucoma Prevalence

Importance  Abnormal body levels of essential elements and exposure to toxic trace metals have been postulated to contribute to the pathogenesis of diseases affecting many organ systems, including the eye.

Objective  To investigate associations between body levels of trace metals and the prevalence of glaucoma in a cross-sectional population-based study.

Design, Setting, and Participants  Blood or urine metallic element levels and information pertaining to ocular disease were available for 2680 individuals 19 years and older participating in the fourth Korea National Health and Nutrition Examination Survey between January 1, 2008, and December 31, 2009, the second and the third years of the survey (2007-2009). Glaucoma diagnosis was based on criteria established by the International Society of Geographic and Epidemiologic Ophthalmology. Demographic, comorbidity, and health-related behavior information was obtained via interview. Multivariable logistic regression analyses were performed to determine associations between blood and urine trace element levels and the odds of glaucoma diagnosis. All analyses were performed between September 2014 and December 2014.

Main Outcome and Measure  The presence or absence of glaucoma.

Results  After adjustment for potential confounders, blood manganese level was negatively associated with the odds of glaucoma diagnosis (odds ratio [OR], 0.44; 95% CI, 0.21-0.92). Blood mercury level was positively associated with glaucoma prevalence (OR, 1.01; 95% CI, 1.00-1.03). No definitive association was identified between blood cadmium or lead levels or urine arsenic level and a diagnosis of glaucoma.

Conclusions and Relevance  These findings in a cross-sectional study of the South Korean population suggest that a lower blood manganese level and a higher blood mercury level are associated with greater odds of glaucoma. For more confidence that trace metals may have a role in the pathogenesis of glaucoma, prospective studies would need to confirm that the presence of such trace metals increases the chance of developing glaucoma.

Glaucoma is an optic neuropathy associated with retinal ganglion cell loss and structural alteration of the optic nerve head. It is the leading cause of irreversible blindness worldwide¹ and a growing public health concern because of an aging global population.² The etiology of glaucoma is multifactorial, with validated risk factors that include greater intraocular pressure (IOP), older age, and a positive family history.³ Other possible concomitant factors include high glutamate levels,⁴ alterations in nitric oxide metabolism,⁵ and vascular changes.⁶ Findings from a prior study⁷ suggest that oxidative stress and free radical accumulation may harm the trabecular meshwork, resulting in an increase in IOP. Such oxidative stress may also be harmful to retinal ganglion cells, with resultant cell death that leads to optic nerve damage characteristic of glaucoma.⁸

Trace metals demonstrated to be essential for normal cellular function of the human body include iron, copper, magnesium, and manganese. These elements serve as activators or cofactors for many transporters, transcription factors, and enzymes. Disruption of essential element metabolism can result in severe dysfunction and diseases in many organ systems. Abnormal essential element levels or exposure to toxic trace metals (eg, lead, mercury, and arsenic) has been postulated to have a role in neurodegenerative diseases,⁹ coronary heart diseases,¹⁰ type 2 diabetes mellitus,¹¹ and diseases involving the eye such as pseudoexfoliation syndrome,^12,13 age-related macular degeneration,¹⁴ and glaucoma.¹⁵ However, the effect of body metal levels on the pathogenesis of glaucoma has remained poorly understood. Yuki et al¹⁵ reported that hair lead levels were significantly higher in women with primary open-angle glaucoma compared with healthy control subjects. An animal study¹⁶ demonstrated abnormally low manganese levels in the primary visual pathway of glaucomatous mice. Ceylan et al¹³ also reported increased levels of blood manganese and mercury in patients with pseudoexfoliation syndrome but not in those with pseudoexfoliative glaucoma. To help facilitate further understanding of the potential association between essential metals and glaucoma, we investigated the relationship between body levels of 5 trace metals (manganese, mercury, lead, cadmium, and arsenic) and the prevalence of glaucoma using a large population-based survey conducted in South Korea.

Box Section Ref ID

All analyses were based on data from the second and third years of the fourth Korea National Health and Nutrition Examination Survey (KNHANES IV) obtained between January 1, 2008, and December 31, 2009. The KNHANES is a cross-sectional survey that examines the health and nutritional status of the noninstitutionalized civilian population of South Korea. The survey has been conducted annually since 2007 under the auspices of the Korea Centers for Disease Control and Prevention, with approval by its institutional review board. The KNHANES consists of the health interview, health behavior and nutrition surveys, and a health examination. The survey adheres to the principles outlined in the Declaration of Helsinki for research that involves humans, and all participants provide written informed consent. This nationwide representative study uses a stratified, multistage probability sampling design, with a rolling survey sampling model.

In the KNHANES, the 1-year survey data and integrated information from the 2008 through 2009 surveys representing the entire adult population of Korea were included. Response rates of 77.8% in 2008 and 82.8% in 2009 were obtained.¹⁷ Among 11 163 individuals 19 years and older who participated during the 2-year study period and received an ophthalmologic examination, 3079 underwent measurement of body levels of 5 trace elements, including blood manganese, mercury, lead, and cadmium and urine arsenic. An additional 399 participants who had a history of retinal disease or stroke were excluded to minimize the chance that nonglaucomatous visual field (VF) defects might result in an erroneous diagnosis of glaucoma.

Data on demographic characteristics and health-related variables were collected through personal interview and a self-administered questionnaire. Physical examination and blood and urine sampling collections were performed at a mobile examination center. Ophthalmologic interview questions and examinations were added in the second half of 2008 and thus available for the KNHANES IV. The methods describing ophthalmologic examinations conducted in the KNHANES have been described in prior publications.^18,19 After an ophthalmology-focused interview, participants underwent visual acuity measurement, automated refraction, slitlamp examination, IOP measurement, fundus photography, and (when deemed appropriate) VF examination. A digital nonmydriatic fundus camera (TRC-NW6S; Topcon) and a standard digital camera (D-80; Nikon) were used to obtain the digital fundus images, which were obtained from all participants 19 years and older under physiological mydriasis. For each participant, one 45° nonmydriatic digital retinal image centered on the fovea was obtained per eye (2 images per person). Vertical cup-disc ratios (VCDRs) were measured from digital fundus images.

The primary outcome variable was the presence or absence of glaucoma diagnosis as defined by the International Society of Geographic and Epidemiologic Ophthalmology (ISGEO) criteria, as described in prior work.²⁰ Category 1 criteria are defined by a VF defect consistent with glaucoma and either a VCDR of at least 0.7 (97.5th percentile) or VCDR asymmetry between the right and left eyes of at least 0.2 (97.5th percentile). Category 2 criteria, used when VF results are not definitive or are unavailable, require a VCDR of at least 0.9 (99.5th percentile) or VCDR asymmetry between the right and left eyes of at least 0.3 (99.5th percentile). Category 3 criteria, used when no information is available on VF testing or optic disc examination, require a visual acuity of less than 3/60 and an IOP exceeding the 99.5th percentile for this population (21 mm Hg). Testing of visual function was performed with frequency-doubling technology in participants with suspected glaucoma as defined by criteria described in a previous study,¹⁸ and a defect was deemed present if 2 different test locations were abnormal. The test was repeated if the proportion of fixation errors or false-positive responses exceeded 0.33. If an individual failed the fixation cutoff on 2 VF testing attempts, the test result was considered invalid. Individuals who were unable to perform valid VF testing were then considered for glaucoma diagnosis using the ISGEO category 2 or category 3 criteria.

Primary predictor variables included blood levels of 4 trace elements (manganese, mercury, lead, and cadmium) and urine arsenic level. To assess the levels of trace elements in whole blood, 3-mL blood samples were drawn into standard commercial vacuum tubes containing sodium heparin (Vacutainer; Becton, Dickinson & Co). Graphite furnace atomic absorption spectrometry with Zeeman background correction (AAnalyst 600; Perkin Elmer) was used for measurement of blood manganese, lead, and cadmium and urine arsenic. A gold amalgam collection method with a direct mercury analyzer (DMA-80; Milestone) was used for measurement of blood mercury. More than 20 mL of clean midstream urine was collected as the sample for total arsenic at the time of health checkup.¹¹ The limits of detection were 0.16 μg/L for manganese, 0.05 μg/L for mercury, 0.02 μg/dL for lead, 0.09 μg/L for cadmium, and 1.68 μg/L for arsenic. To convert manganese level to nanomoles per liter, multiply by 18.202; mercury level to nanomoles per liter, multiply by 4.985; lead level to micromoles per liter, multiply by 0.0483; cadmium level to nanomoles per liter, multiply by 8.896; and arsenic level to micromoles per liter, multiply by 0.0133. The concentrations in all samples were higher than the limits of detection. The interassay coefficients of variation were 2.2% to 4.8% for manganese, 1.1% to 4.1% for mercury, 2.2% to 6.0% for lead, 3.0% to 11.9% for cadmium, and 2.5% to 3.2% for arsenic. Creatinine-adjusted urine arsenic values were calculated to correct for the effect of differences in creatinine clearance among study participants.²¹ All blood metal analyses were performed at the NeoDin Medical Institute, Seoul, a laboratory certified by the Korean Ministry of Health and Welfare. An internal quality assurance and control program at this institute has been described in previous studies.^14,22,23

Blood samples were collected by venipuncture after 10 to 12 hours of fasting, and all blood analyses were performed by the Seoul Medical Science Institute, a laboratory certified by the Korean Ministry of Health and Welfare. Demographic and socioeconomic information was obtained from a health interview. Body mass index was calculated as weight in kilograms divided by height in meters squared. Smoking status was determined by self-report, and individuals were classified as smokers or nonsmokers. Alcohol consumption was assessed by participants’ drinking behavior during the month before the interview. Blood pressure was measured 3 times at 5-minute intervals using a sphygmomanometer with the patient in a sitting position, and the average of the second and third measurements was used for the analysis. Refractive status was categorized as emmetropia (−0.99 to +0.99 diopter [D]), mild myopia (−1.00 to −2.99 D), moderate myopia (−3.00 to −5.99 D), high myopia (less than −5.99 D), or hyperopia (+1.00 D or higher).

Complex sample analysis was used for the KNHANES data to weight all values according to statistical guidance from the Korea Centers for Disease Control and Prevention. A regression model was constructed after the identification of potential confounding variables. All risk factors that were identified as being associated with glaucoma diagnosis by univariate analysis with P < .10 as the cutoff point were then included in the multivariable analysis to assess the possible independent association between body levels of trace metals and glaucoma. After ascertainment of such a possible association, 95% CIs of odds ratios (ORs) were identified for each possible association. All statistical tests were 2-sided with 95% CIs and were performed using a software program (SPSS, version 21.0; SPSS Inc).

Of 2680 right eyes included in our analysis, 36 met the ISGEO criteria for glaucoma diagnosis, representing a mean (SE) of 1.5% (0.3%) of the population sample with required information. Table 1 summarizes unadjusted demographic characteristics, comorbidities, and health-related behavior in those with vs without glaucoma diagnosed using the ISGEO criteria. Participants with glaucoma were older and exercised less than those without the disease. The presence of glaucoma was associated with a slightly higher prevalence of moderate to high myopia. Individuals with glaucoma also had higher serum ferritin and aspartate aminotransferase levels than those without the disease.

Among 5 trace metals, blood manganese and mercury were associated with glaucoma prevalence among the study population.

After adjustment for age, sex, exercise, and ferritin and aspartate aminotransferase levels, blood manganese levels were significantly lower (OR, 0.44; 95% CI, 0.21-0.92) in participants with (mean, 12.36 µg/L) vs without (mean, 13.49 µg/L) glaucoma, while positive associations were observed for blood mercury levels in those with (mean, 6.18 µg/L) vs without (mean, 5.31 µg/L) glaucoma (OR, 1.01; 95% CI, 1.00-1.03) (Table 2). We found no such associations between glaucoma diagnosis and blood lead, blood cadmium, or urine arsenic levels.

Blood manganese levels were further categorized into tertiles, and the distribution of tertiles also differed based on glaucoma diagnosis. Multivariable logistic regression models were constructed to assess a possible independent association between the odds of glaucoma diagnosis among those with various blood manganese levels. The third tertile of blood manganese level (≥14.46 µg/L) was associated with lower odds of glaucoma diagnosis compared with the first tertile (≤11.41 µg/L) even after adjustment for potential confounding variables (P = .01 for trend) (Table 3). P value for trend analysis was performed using multivariable logistic regression with tertile level of manganese as a continuous variable. In contrast, further stratifying blood mercury levels into tertiles did not definitively identify an association with glaucoma diagnosis when the first tertile (≤3.37 µg/L) was used as the reference for comparison with the second tertile (3.38-5.40 µg/L; OR, 1.46; 95% CI, 0.52-4.07) and the third tertile (≥5.41 µg/L; OR, 1.36; 95% CI, 0.55-3.37) (P = .85 for trend).

The present study investigating associations between body levels of trace metals and the risk of glaucoma found that lower blood manganese levels and higher blood mercury levels are associated with greater glaucoma prevalence. We found no such associations between glaucoma diagnosis and blood lead, blood cadmium, or urine arsenic levels in our population.

Manganese is an essential trace metal present in all tissues and required for the maintenance of proper cell function. It is a cofactor of many enzymes, including superoxide dismutase, which inhibits neuronal death presumably by interfering with a superoxide signal for apoptosis.²⁴ The etiology and symptoms of manganese toxicity are well defined.²⁵ Manganese-induced parkinsonism is characterized by progressive neurological deterioration, with symptoms that include bradykinesia, tremor, impaired postural reflexes, and dystonia.^26,27 While high manganese-induced toxicity has been investigated extensively, changes in physiological function occurring from manganese deficiency are less well understood.

We found a negative association between blood manganese level and the prevalence of glaucoma in a multivariable analysis. A second analysis that divided the study population into tertiles confirmed that the group having the highest manganese level showed decreased odds of glaucoma diagnosis compared with the group having the lowest level. Ceylan et al¹³ previously reported that increased levels of blood manganese and mercury were found in patients with pseudoexfoliation syndrome. However, manganese levels were not higher in the pseudoexfoliative glaucoma group relative to control subjects without glaucomatous disease, while mercury levels were slightly higher in the pseudoexfoliative glaucoma group compared with the controls. Namuslu et al²⁸ observed decreased levels of manganese in resected pterygial tissue and suggested that manganese depletion may result in a reduced antioxidant effect and resultant accumulation of harmful reactive oxygen species in the conjunctiva, thereby contributing to pterygium formation. An animal study¹⁶ revealed that retinal manganese levels were significantly deficient in 10-month-old glaucomatous DBA/2J mice compared with aged-matched C57BL/6J mice and 5-month-old DBA/2J mice. Further work has demonstrated neuroprotective effects and superoxide scavenging activity of manganese corroles in vitro and in vivo, suggesting that they could be candidates for therapeutic agents against diseases associated with superoxide-related axonal injury.²⁹ Neuroprotective effects of manganese porphyrins as superoxide dismutase mimetics have also been demonstrated in animal models of ischemia.^30,31 Further prospective studies will be necessary to assess the antioxidant role and potential neuroprotective effect of manganese in glaucoma.

Our findings also suggest that high blood mercury levels may have a role in glaucoma, but this association was not as strong as that seen with low blood manganese levels. Furthermore, the dose-response relationship was not confirmed in a tertile analysis. Mercury, lead, cadmium, and arsenic are xenobiotic metals with no physiological functions. Such metals often enter organisms by molecular mimicry using inherent transporters for essential metals.³² Exposures to xenobiotic metals are frequently related to the development of toxicity and pathologic conditions.³³ Humans are generally exposed to organic species of mercury (methylmercury) from digesting seafood. Although we found no associations between body levels of lead, cadmium, or arsenic and glaucoma prevalence, toxicity from exposure to these metals has been identified. Rhee et al²¹ found that blood lead level is significantly associated with a metabolic syndrome in Korean adults. Yuki et al¹⁵ reported significantly higher lead levels in a group of women with primary open-angle glaucoma (P = .03) and in a glaucoma group with low IOP (P = .02) compared with a control group. Evidence from epidemiological studies^34,35 revealed that chronic exposure to inorganic arsenic in drinking water (100 μg/L) was associated with diabetes mellitus. Cadmium accumulates with age in the retinal pigment epithelium,³⁶ and findings of studies^14,37,38 have led to the hypothesis that it is a possible contributor to age-related macular degeneration. While the association between body cadmium level and glaucoma has not been well investigated, one study³⁹ showed that it was detected in a small number of aqueous humor samples from individuals with glaucoma.

Our study has some limitations. First, only one frequency-doubling technology test was obtained for each participant in the KNHANES. The more rigorous 2-2-1 rule for VF interpretation suggested by previous investigators was not used.⁴⁰ In an effort to minimize false-positive diagnoses, we excluded participants with retinal disease or a history of stroke, thereby reducing error related to inclusion of individuals who may have had VF defects due to nonglaucomatous conditions. However, such exclusion may have introduced error into our findings because 3.0% (9 of 399 individuals) of the excluded group were diagnosed as having glaucoma as defined by the ISGEO criteria. Nevertheless, we have no reason to believe that this error would be related to blood or urine trace metals. Second, VF examination was to be administered to all participants with suspected glaucoma in the KNHANES, but some individuals for unknown reasons did not receive this test. Among 2680 included participants, 564 (representing 21.5% of this population) had suspected glaucoma, of whom 345 did not undergo VF testing and thus were presumably categorized as such based solely on structural characteristics of the optic nerve. Furthermore, cross-sectional population studies cannot determine causation, and this work simply demonstrates an association between body levels of trace metals and glaucoma diagnosis. A prospective study with risk factors noted before the development of glaucoma would be necessary to establish a causal nature of such associations. Third, another potential shortcoming is that body metal levels were measured at only a single time point, which may not represent long-term status with regard to these variables. However, there are no definitive biomarkers for assessing chronic manganese exposure, and the most commonly used measure in animal studies is the manganese concentration in tissue.^41,42 Unfortunately, human tissue is generally accessible only in a biopsy specimen. Serum manganese level is the most easily measured surrogate for body manganese status in humans.⁴³ In our study, heavy metal exposure status was evaluated only by blood or urine samples without confirmation of tissue levels. Fourth, only one chemical form of mercury was measured, which may not be an accurate surrogate for overall exposure.

After adjusting for potential confounders, our findings suggest that a lower blood manganese level and a higher blood mercury level are associated with greater odds of glaucoma diagnosis in a representative sample of the South Korean population 19 years and older. Future prospective investigations will be necessary to confirm these associations and to explore the role of trace elements in the pathogenesis of glaucoma, as well as possible neuroprotective effects, which could lead to novel therapeutic targets in glaucoma management.

Submitted for Publication: February 23, 2015; final revision received June 4, 2015; accepted June 11, 2015.

Corresponding Author: Shan C. Lin, MD, Department of Ophthalmology, University of California, San Francisco, 10 Koret St, Room K301, San Francisco, CA 94143-0730 (LinS@vision.ucsf.edu).

Published Online: August 6, 2015. doi:10.1001/jamaophthalmol.2015.2438.

Author Contributions: Dr S. C. Lin had full access to all the data in the study and takes responsibility for the integrity of the data and the accuracy of the data analysis.

Study concept and design: All authors.

Acquisition, analysis, or interpretation of data: S.-C. Lin, S. C. Lin.

Drafting of the manuscript: S.-C. Lin.

Critical revision of the manuscript for important intellectual content: S. C. Lin, Singh.

Statistical analysis: S.-C. Lin.

Administrative, technical, or material support: S. C. Lin, Singh.

Study supervision: S. C. Lin.

Conflict of Interest Disclosures: All authors have completed and submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest, and none were reported.

Funding/Support: This study was supported by core grant EY002162 from the National Eye Institute, by That Man May See, and by Research to Prevent Blindness (all to Dr S. C. Lin).

Role of the Funder/Sponsor: None of the sponsors or funding organizations had a role in the design or conduct of the study; collection, management, analysis, or interpretation of the data; preparation, review, or approval of the manuscript; and decision to submit the manuscript for publication.