purpose. To study to what extent genetic factors determine the retinal nerve fiber layer thickness (RNFLT) in healthy subjects.

methods. In vivo peripapillary optical coherence tomography (OCT), clinical examination, lens fluorescence, and fundus photography were performed on both eyes of 25 monozygotic and 25 dizygotic same-sex pairs of twins. The cross-sectional study included twins aged from 20 to 45 years recruited from a population-based register. Only healthy eyes were included. Main outcome variables: peripapillary OCT RNFLT, reproducibility, and heritability (the proportion of the total observed variance statistically attributable to genetic factors).

results. The within-pair difference in RNFLT was 4.6% (0.7%–15.2%; median [range]) in monozygotic versus 7.3% (0.2%–20%) in dizygotic twins (*P* = 0.032, Mann-Whitney test). The RNFLT heritability was 66%. The RNFLT measurement was found to decrease 3.8 μm per decade (*P* = 0.003). The RNFLT heritability increased to 82%, when corrected for the effect of age and excluding within-pair refractive differences of 2 D or more. The signal-to-noise ratio correlated with lens transmittance (*r* = 0.25, *P* = 0.012), age (*r* = −0.29, *P* = 0.004), and RNFLT (*r* = 0.43, *P* < 0.001). Intravisit RNFLT reproducibility was 4.2%.

conclusions. Peripapillary RNFLT in healthy adults, as measured by OCT, was determined predominantly by genetic factors in this study population. Theoretically, these factors may involve variations in the number of ganglion cells and nerve fiber formations early in life and/or in the rate at which these structures are subsequently lost.

^{ 1 }Specific gene loci involved include the loci 1p36 and 2q21 in primary congenital glaucoma (buphthalmia), 1q23-q25/TIGR in primary juvenile glaucoma and adult onset POAG, and the optineurin gene on 10p14 in adult-onset POAG.

^{ 2 }

^{ 3 }

^{ 4 }

^{ 5 }Rare families with adult-onset POAG demonstrate patterns of Mendelian inheritance, that suggest single-gene disease, but multiple gene involvement, is probably involved in the typical patient. Thus, defects in currently identified genes account for only a small fraction of glaucoma cases. There are few data available to asses the overall effect of hereditary factors in the development of glaucoma, but several studies show that a family history of adult-onset POAG is a major risk factor.

^{ 6 }

^{ 7 }

*n*= 100) of 25 monozygotic (MZ) and 25 dizygotic (DZ) pairs of twins were examined in a cross-sectional study including twins aged 20 to 45 years. All twin pairs were same-sex and recruited from a population-based register comprising twins born in Denmark between 1870 and 1996 (The Danish Twin Registry, University of Southern Denmark, Odense, Denmark).

^{ 8 }Exclusion criteria included cataract and other opacities near the optical axis of the eye, as well as other manifest eye disease, with such findings leading to the exclusion of both twins in a pair.

^{ 9 }

^{ 10 }

^{ 11 }

^{ 12 }

^{ 13 }

^{ 14 }Using an internal fixation light, the scan circle was centered on the optic nerve head (ONH). No scaling was made for variation in subject refraction and fundus magnification. All scans were obtained at the maximum power of 750 mW and stored in a digital archive for later processing.

^{ 15 }

^{ 16 }

^{ 17 }

^{ 18 }The excitation wavelength was 430 to 490 nm, and detection was at 530 to 630 nm. Lens transmittance was calculated as the square root of the ratio between the posterior and anterior peak in lens fluorescence, using the mean of six scans (three per eye).

*t*-test or the Mann-Whitney test (two-sided), linear regression (least square), and correlation analysis (Pearson’s correlation coefficient). The proportion of total variation attributable to genetic factors is expressed as heritability (h

^{2}), which is twice the difference in interclass correlation (

*r*) between MZ and DZ twins

^{ 19 }

*r*is defined as

^{ 20 }Within a twin pair, the A and B status was randomly chosen. To obtain a symmetric distribution around the identity line, both of the two coordinates A,B and B,A for any one pair of twins was used in the correlation analysis. The 95%-confidence interval (ci

_{95}) of the heritability is calculated as

_{t}is the SE of the heritability,

^{ 21 }which is calculated as

*n*is the number of offspring per family (

*n*= 2) and

*t*is 1/2h

^{2}.

*N*is twice the number of families (100) because h

^{2}is calculated using both of the two coordinates A,B and B,A for any one pair of twins.

^{ 22 }Data analysis was made using R computer software version 1.2.3 (http://www.r-project.org).

*n*= 100) was 104.4 ± 9.9 μm. We found no significant difference between the mean RNFLT in MZ twins (105.6 ± 10.1 μm; mean ± SD) and that in DZ twins (103.1 ± 9.7 μm;

*P*= 0.21,

*t*-test). Furthermore, there was no difference in RNFLT between women and men (

*P*= 0.70,

*t*-test). For the superior, nasal, inferior, and temporal quadrants the thicknesses were 126.1 ± 13.3, 79.6 ± 13.5, 129.5 ± 14.0, and 80.1 ± 12.9 μm, respectively. The RNFLT did not differ between the twin A and B groups (

*P*= 0.82,

*t*-test), thus justifying the pooling of all 100 subjects in the analysis. All mention of RNFLT in the text that follows relates exclusively to the mean thickness of the full circle.

*r*= 0.43,

*P*< 0.001) and with increasing age (

*r*= −0.30,

*P*= 0.003; Fig. 1 ). Thus, the RNFLT was found to decrease 3.8 μm per decade, from 110 μm at age 20 to 100 μm at age 45. The RNFLT decreased slightly with decreasing lens transmittance (

*r*= 0.25,

*P*= 0.014) and with increasingly negative refraction (

*r*= 0.20,

*P*= 0.047). For the total study population, the OCT S/N ratio was (mean ± SD, [range]) 55.4 ± 2.2 dB (50.8–59.5) and lens transmittance was 0.91 ± 0.04 (0.76–0.99).

*r*= 0.25,

*P*= 0.012), and both parameters correlated negatively with increasing age (

*r*= −0.29,

*P*= 0.004;

*r*= −0.36,

*P*< 0.001). Age explained 8% and 13% of the total variation in OCT S/N ratio and lens transmittance, respectively.

*P*= 0.056).

*P*= 0.83, Mann-Whitney test). Reproducibility was calculated as the numeric difference in RNFLT (full-circle mean) between the two scans chosen for analysis relative to the mean of the two scans. Reproducibility decreased significantly with age (

*r*= 0.24,

*P*= 0.017), with decreasing lens transmittance as determined by lens fluorometry (

*r*= −0.31,

*P*= 0.002), and with decreasing OCT S/N ratio (

*r*= −0.48,

*P*< 0.001). These factors explain 6%, 9%, and 23%, respectively, of the total variation in RNFLT reproducibility. The RNFLT reproducibility decreased slightly with decreasing thickness of the retinal nerve fiber layer (

*r*= −0.23,

*P*= 0.020).

^{ 23 }By World Health Organization definitions, this asymptomatic subject was diabetic, but was classified in the study as having borderline diabetes.

*P*= 0.032, Mann-Whitney test; Table 2 ). Interclass correlation (

*r*) on RNFLT (crude data) was 0.812 and 0.481 in MZ versus DZ twin pairs (Fig. 2) . Thus, the heritability (h

^{2}) was 66% (ci

_{95}48%–84%) when based on raw data. After correction for the effect of age, using linear regression residuals from RNFLT analyzed as a function of age, the interclass correlation (

*r*) was 0.802 and 0.411 in MZ versus DZ, yielding a heritability of 78% (61%–95%).

*P*= 0.078; Table 2 ), potentially confounding the analysis on RNFLT heritability because RNFLT was found to correlate with refraction (as mentioned earlier). Consequently, we performed an additional analysis on age-corrected residuals after exclusion of all twin pairs with an intrapair difference in refraction (spherical equivalent) equal to or greater than 2 D. The remaining 24 MZ and 17 DZ twin pairs demonstrated an interclass correlation (

*r*) of 0.800 in MZ versus 0.388 in DZ twin pairs, yielding a heritability of 82% (64%–100%).

*P*= 0.003).

^{ 24 }

^{ 25 }A cohort effect cannot be ruled out in this cross-sectional study, but it seems unlikely. A follow-up of the present study should clarify this matter.

*P*= 0.056).

^{ 22 }The design of our study does not permit any conclusion regarding the potential relation between such factors and age-related loss of nerve fibers.

^{ 11 }

^{ 26 }

^{ 27 }

^{ 28 }It is not known whether this phenomenon can be reproduced in viable tissue. The effect may be caused by the abrupt increase in refractive index between the vitreous and the retina, notably the specular surface reflection of the retina.

^{ 28 }

^{ 29 }Consequently, this may lead to problems for the data analysis algorithms defining the thickness of the RNFL by OCT. OCT has been shown systematically to yield lower RNFLT values than histologic tissue sections,

^{ 30 }

^{ 31 }

^{ 32 }but other potential causes could involve optical artifacts in the living human eye as well as artifacts introduced by histologic tissue preparation. In addition, the exact location of histologic sections in relation to the landmarks of the fundus is notoriously difficult. Furthermore, effects of variations in data analysis algorithms have been demonstrated (Hougaard JL, Sander B, ARVO Abstract 271, 2002). Unfortunately, the fundamental characteristics of the proprietary algorithms used in current instruments have not been documented, but empiric testing demonstrates a high level of agreement with visual evaluation of the RNFL on biomicroscopy or fundus photography and with automated perimetric localization of RNFL defects. Our results are amenable, however, to confirmation by independent methods, and we have found a comparable level of heritability of the optic nerve head rim area as assessed by stereoscopic inspection of fundus photographs in the present study sample (Dejgaard N, Hougaard JL, Larsen M, unpublished data, 2002). In addition, a genetic influence on the cup-to-disc area ratio was found in 17 healthy twin pairs in a study by Teikari and Airaksinen.

^{ 33 }

^{ 34 }found a heritability of open-angle glaucoma of 13%, based on concordance rates in MZ and DZ twins.

Monozygotic Twins | Dizygotic Twins | P ^{*} | |
---|---|---|---|

Number of twins | 50 | 50 | |

Female/male (n/n) | 24/26 | 32/18 | — |

Age (y) | 36 (20–45) | 35 (20–45) | NS |

Refraction (spherical equiv.; D) | 0.00 (−5.28–1.50) | −0.56 (−6.38–4.38) | NS |

Fasting blood glucose (mmol/L) | 4.9 (4.2–5.6) | 4.9 (4.2–6.2) | NS |

Diastolic blood pressure (mm Hg) | 70 (55–81) | 69 (51–91) | NS |

Body mass index (kg/m^{2}) | 23.5 (18.9–28.0) | 23.8 (17.9–35.3) | NS |

Smoking habits (Pack-years) | 0.4 (0.0–29.0) | 0.0 (0.0–24.0) | NS |

**Figure 1.**

**Figure 1.**

Monozygotic Twin Pairs | Dizygotic Twin Pairs | P ^{, ‡} | |
---|---|---|---|

Within-pair absolute numerical difference | |||

Refraction (spherical equivalent; D) | 0.4 (0.0–2.4) | 0.7 (0.0–6.1) | 0.078 |

Lens transmittance (%) | 3.2 (0.1–6.0) | 3.8 (0.3–18.1) | 0.35 |

S/N (dB)^{*} | 1.00 (0.00–5.50) | 1.00 (0.25–4.25) | 0.68 |

Within-pair relative numerical difference | |||

RNFLT (%)^{, †} | 4.6 (0.7–15.2) | 7.3 (0.2–20.0) | 0.032 |

**Figure 2.**

**Figure 2.**