Adaptive optics scanning laser ophthalmoscopy images in a family with the mitochondrial DNA T8993C mutation

Abstract

Purpose: This study was designed to assess the effect of mitochondrial DNA (mtDNA) mutation T8993C on cone structure in a family expressing neurogenic muscle weakness, ataxia, and retinitis pigmentosa (NARP) syndrome.

Methods: Five family members were studied, using clinical examination, nerve conduction studies, perimetry, optical coherence tomography (OCT) measures of central retinal thickness, and electroretinography. High-resolution images of cone structure using adaptive optics scanning laser ophthalmoscopy (AOSLO) were obtained in four subjects with stable fixation. Cone spacing was compared to 18 age-similar normal subjects and converted to z-scores at each location where unambiguous cones were identified. Tissue levels of T8993C mutant heteroplasmy in blood and hair follicles were quantified using real-time allele-refractory mutations system (ARMS) quantitative polymerase chain reaction (qPCR).

Results: Subjects expressing the T8993C mutation showed varying levels of disease severity. The subject with the lowest mutant load (42%-54%) showed no neurologic or retinal abnormalities. The remaining four subjects with over 77% mutant load all expressed severe neurologic and/or retinal abnormalities. AOSLO images revealed three patterns of cone spacing: pattern 1, normal; pattern 2, increased cone spacing within a contiguous cone mosaic; and pattern 3, patchy cone loss with increased cone spacing. Visual function was most severely affected in pattern 3.

Conclusions: High levels of T8993C mutant load were associated with severe neurologic or visual dysfunction, while lower levels caused no detectable abnormalities. Visual function was better in patients with a contiguous and regular cone mosaic. Patients expressing high levels of the mtDNA T8993C mutation show abnormal cone structure, suggesting normal mitochondrial DNA is necessary for normal waveguiding by cones.

Figures

Figure 1

Family Pedigree. Squares indicate males, circles indicate females. Filled symbols on the left half indicate neuropathy while filled symbols on the right half indicate retinopathy. Diagonal lines indicate deceased individuals. Examined individuals are indicated numerically (I-1, II-1, II-2, II-3, II-4) as in the text.

Figure 2

The cone receptor mosaic was imaged using AOSLO in the region outlined on the fundus photograph belonging to subject I-1, showing RPE mottling surrounding a small central island of preserved RPE (A); GVF testing showed a dense pericentral scotoma to the V4e and I4e targets with a preserved central island corresponding to the preserved RPE seen on examination (B); AOSLO images showed increased cone spacing with patchy cone loss, with corresponding absolute scotomas (indicated as 0) shown with superimposed microperimetry results (C). Scale bar, 1°.

Figure 3

OCT images of four of the subjects. The OCT scans are 20° horizontal scans across the retina including the foveal center. IR fundus images with the scan line indicated are shown on the left. Arrows indicate the extent of the AOSLO images shown in Figure 4 for subjects II-3, II-4, and I-1. (A) In subject II-3, a central region of preserved photoreceptor outer nuclear layer (ONL), inner segment layer (ISL), and outer segment layer (OSL) extends to approximately 2° eccentricity in all directions, while a ring of approximately 1° width between 2° to 3° shows loss of the ONL, ISL, and OSL. More normal outer retinal structure is restored at 3° in all directions. Subject II-4 (B) shows preserved outer retinal structure at the fovea with progressive attenuation of the ONL, ISL, and OSL beginning gradually around 5° eccentric to fixation and extending throughout the region imaged. Subject I-1 (C) shows more abrupt loss of the ONL, ISL, and OSL beginning within 1° from a preserved region at the fovea. (D) An external fixation target was used to align the scan with the foveal center in subject II-1. The OCT shows complete loss of the ONL, ISL, and OSL with disruption of the RPE layer throughout the region imaged, including the foveal center.

Figure 4

Comparison of cone structure near the fovea in patients with T8993C mutations. The small black dot in each image indicates the preferred fixation point. All images are to the same scale. (A) Subject II-2 has normal cone spacing at all eccentricities (cones within 0.5° of the foveal center are often difficult to resolve in this and other normal subjects). (B) Subject II-3 shows coarse cones with increased cone spacing at the foveal center, surrounded by a ring where cones are only dimly visible. A contiguous array of cones peripheral to this ring is visualized, although cone spacing is increased. Subjects II-4 (C) and I-1 (D) show similar patterns of sparse cones with increased cone spacing near fixation. Peripheral to regions of preserved cones are regions where unambiguous cones cannot be seen. (D) represents the central 2.5° of the image shown in Figure 2C. Scale bar, 1°.

Figure 5

(A) Cone spacing of NARP patients and normal subjects versus eccentricity. Data from the normal subjects are plotted as small dots. Solid line: best fit to normal data. Dashed lines: 95% confidence limits of the best fit. Subject II-2 shows normal cone spacing, but all others show increased spacing. (B) Plots of fundus-guided microperimetry sensitivity loss in decibels versus cone spacing z-scores reveal the functional consequences of patchy cone loss (see Fig. 3) on visual performance. Subject II-3 performs better than subjects I-1 and II-4, likely due to the presence of a contiguous cone mosaic.