Discussion
In this cohort, detection of atrophy of the peripapillary nerve fibre layer was more successful and was more closely associated with carriage of LHON-associated pathogenic mitochondrial variants than perfusion measurement, and also more closely correlated with optic neuropathy. The evaluation of peripapillary retinal nerve fibre layer perfusion by OCTA measurement was not possible in the majority of LHON-affected patients due to fixation difficulty, usually associated with poor vision. In contrast, of the patients who could fixate for scanning by OCTA (33%), we found that the LHON-affected individuals had reduced peripapillary VD compared with controls. A greater proportion of the asymptomatic relatives were able to fixate for OCTA measurement (69%), and overall the peripapillary VD was not significantly different to that of controls, of whom all could fixate for perfusion measurement. Of the asymptomatic relatives, 17% were found to have reduced VD, and increased VD was observed in 10%. Abnormal VD had a variable association with visual function and was present with a VA level varying from normal VA through to blindness, and also occurred in a subject with normal vision but subclinical signs of optic neuropathy including prolongation of VEP. Normal nerve fibre layer thickness, with a lack of atrophy, showed greater specificity for signs of normal optic nerve function (92%) versus measurement of normal perfusion (66%) for normal nerve function.
The problem with using OCTA measurement in LHON is obtaining measurements when VA is poor and fixation is difficult, as a scan requires steady fixation for several seconds. Notwithstanding this difficulty, OCTA findings in the peripapillary circulation have been described in LHON, demonstrating variable associations including negative findings of the asymptomatic stage of LHON,23 and reduced vessel density in established LHON.17 Furthermore, cohorts of LHON families have been studied, demonstrating reduced vessel density in asymptomatic individuals,23 or increased vessel density affecting the temporal ONH, after correcting for retinal nerve fibre layer thickness, in asymptomatic male carrier relatives.24
Peripapillary RNFL vessel density was lower in LHON than in normal eyes in our study. We suspect that the reduced perfusion is secondary to atrophic changes of the ONH and secondary loss of the RNFL vascular plexus, resulting in relative ischaemia. This ischaemia may be a recovery-limiting factor when considering therapies that promote mitochondrial respiratory chain function (ie, idebenone) and reduced OCTA VD may suggest limited benefit from such treatments. Indeed, each eye that showed reduced RNFL perfusion in the LHON and asymptomatic relative groups was also found to have RNFL thinning relative to controls. In this study, 66% of the LHON group could not fixate adequately for scanning and were therefore excluded from analysis. We suspect these unmeasured individuals may have had low peripapillary VD also based on the clinical characteristics of ONH pallor and peripapillary RNFL thinning.12 Our control group was older than our LHON group (median age 48 vs 21 years), which may lead to the LHON group having undetected cases of even lower VD, as with advancing age there is a reduction in peripapillary VD.25 There were sex differences in the comparison groups, with only males included in the measured LHON group. In healthy subjects, VD parameters measured by OCTA do not vary by sex,25 but LHON shows some sex-dependent penetrance with more vision loss seen in males.10
Asymmetric changes in VD were observed in some asymptomatic LHON relatives. One case had discordant VD changes across both ONHs. This was associated with optic nerve dysfunction and poor vision of LogMAR 2.4 in one eye, which had low VD, and normal vision with an abnormally short VEP of 94 ms in the contralateral eye, which had raised VD. The implications of this are not clear, as our study was not longitudinal. Onset of LHON is usually sequential, with one eye affected within 3 months of the fellow eye, though cases of more than 10 years between eyes losing vision have been reported.26 A shortened VEP implicit time implies increased conductivity of retinal ganglion cells. This may represent compensatory increased mitochondrial biomass as mitochondrial dysfunction develops, prior to visual loss.
The measurement of peripapillary RNFL perfusion did not equate to that of RNFL thickness and each provided different data, and correlated differently with optic nerve function. Unlike RNFL perfusion, where we found no difference between controls and asymptomatic relatives, RNFL thickness was significantly reduced versus controls in both asymptomatic relatives and LHON-affected individuals. RNFL thinning was present in the chronic cases of LHON who could fixate (two eyes only in each of C9 and C10) and was present in 6 eyes of 19 who had measurement in the asymptomatic relatives sample. The presence of normal RNFL showed greater specificity (92%) for normal optic nerve function, including VA and VEP latency, in the asymptomatic relatives, compared with the specificity of normal vessel density for normal optic nerve function (66%). In this cohort, lower perfusion did not give any additional insights into the aetiology of thinned RNFL. RNFL thinning may occur at an earlier stage of progressing neuropathy than vessel density changes, which may follow later when atrophy is established. Individuals in our asymptomatic relative cohort may represent subclinical cases of LHON.
None of the asymptomatic relatives had raised RNFL thickness above the derived normal range, whereas one of the samples had raised vessel density, that was found in association with shortened P100 latency. These asymptomatic relatives may have early preclinical optic neuropathy due to impaired mitochondrial function. Distinctly, increased perfusion may represent an earlier again stage prior to manifestation of vision loss.
Future studies could investigate correlation of perfusion and thickness with onset of ONH telangiectasia, classically seen at the onset of symptoms of LHON. ONH analysis by quadrant would be useful, and longitudinal studies of changing perfusion over time may give more insight into the significance of our findings. Analysis of these measurements could be done with other tests of visual function, including perimetry.
Limitations of this study include its small size and cross-sectional nature, and that age-matching was limited to the control group comparisons; the LHON-affected group was significantly younger than the asymptomatic relative group. LHON is already a rare condition and fixation difficulties meant that accurate peripapillary OCTA could be captured in only a small subset of affected patients. We evaluated the peripapillary RNFL, as the primary site of pathology, but more information may be derived from also evaluating perfusion changes at the macula and within the papillomacular bundle. Strengths of the study include the evaluation of multiple individuals who have LHON and also a group of asymptomatic maternal relatives when compared with a normative database assessed on the same imaging platform.
Potential utility of OCTA and OCT in LHON could be in monitoring asymptomatic relatives for optic nerve RNFL perfusion and thickness changes over time. Those eligible for monitoring may be as many as 1 in 10 000 based on population studies of LHON prevalence,27 or as high as 1 in 300 based on prevalence of LHON-associated mitochondrial variants.28 While historically there was no treatment available for LHON, monitoring and screening may become of more relevance as more treatment options are being investigated and developed, including idebenone and intravitreal gene replacement therapy.29,30 Longitudinal studies are required to evaluate whether microcirculatory abnormalities may be static or progress over time and how reduction of peripapillary RNFL VD may impact outcomes of these novel treatments. Our finding of raised peripapillary RNFL VD associated with shortened P100 latency is interesting but requires longitudinal studies to determine its significance.