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. 2023 Feb 24;18(2):e0282175. doi: 10.1371/journal.pone.0282175

Is retina affected in Huntington’s disease? Is optical coherence tomography a good biomarker?

Pavel Dusek 1,#, Ales Kopal 1,2,#, Michaela Brichova 3, Jan Roth 1, Olga Ulmanova 1, Jiri Klempir 1, Jana Lizrova Preiningerova 1,*
Editor: Alfred S Lewin4
PMCID: PMC9955964  PMID: 36827300

Abstract

Aim of the study

Comparative cross-sectional study of retinal parameters in Huntington’s disease and their evaluation as marker of disease progression.

Clinical rationale for the study

Huntington’s disease (HD) is a neurodegenerative disorder with dominant motor and neuropsychiatric symptoms. Involvement of sensory functions in HD has been investigated, however studies of retinal pathology are incongruent. Effect sizes of previous findings were not published. OCT data of the subjects in previous studies have not been published. Additional examination of structural and functional parameters of retina in larger sample of patients with HD is warranted.

Materials and methods

This is a prospective cross-sectional study that included: peripapillary retinal nerve fiber layer thickness (RNFL) and total macular volume (TMV) measured by spectral domain optical coherence tomography (OCT) of retina, Pelli-Robson Contrast Sensitivity test, Farnsworth 15 Hue Color discrimination test, ophthalmology examination and Unified Huntington’s disease Rating Scale (UHDRS). Ninety-four eyes of 41 HD patients examined in total 47 visits and 82 eyes of 41 healthy controls (HC) examined in total 41 visits were included. Analyses were performed by repeated measures linear mixed effects model with age and gender as covariates. False discovery rate was corrected by Benjamini-Hochberg procedure.

Results

HD group included 21 males and 20 females (age 50.6±12.0 years [mean ± standard deviation], disease duration 7.1±3.6 years, CAG triplet repeats 44.1±2.4). UHDRS Total Motor Score (TMS) was 30.0±12.3 and Total Functional Capacity 8.2±3.2. Control group (HC) included 19 males and 22 females with age 48.2±10.3 years. There was no statistically significant difference between HD and HC in age. The effect of the disease was not significant in temporal segment RNFL thickness. It was significant in the mean RNFL thickness and TMV, however not passing false discovery rate adjustment and with small effect size. In the HD group, the effect of disease duration and TMS was not significant. The Contrast Sensitivity test in HD was within normal limits and the 15-hue-test in HD did not reveal any specific pathology.

Conclusions

The results of our study support possible diffuse retinal changes in global RNFL layer and in macula in Huntington’s disease, however, these changes are small and not suitable as a biomarker for disease progression. We found no other structural or functional changes in retina of Huntington’s disease patients using RNFL layer and macular volume spectral domain OCT and Contrast Sensitivity Test and 15-hue-test.

Clinical implications

Current retinal parameters are not appropriate for monitoring HD disease progression.

1. Introduction

Huntington’s disease (HD) is an autosomal dominant hereditary neurodegenerative disorder caused by CAG repeat expansion in the huntingtin (HTT) gene encoding the HTT protein. HD manifests itself dominantly by motor, neuropsychiatric and cognitive symptoms. In the last decades, a substantial number of studies examining sensory functions in various neurodegenerative diseases including HD were published.

There is evidence of the presence of HTT in retina and the impairment of retinal functions in various animal models of HD. Helmlinger et al [1] found strong deficiencies in vision, retinal dystrophy, and mutant huntingtin in the retina of R6 transgenic mice. Johnson et al [2] showed reduced oscillatory potential amplitudes and disinhibition of the photopic response and accumulation of huntingtin in the horizontal cells of the retina of HD rats.

Several studies have indicated an impairment of visual evoked potentials (VEP) [3], retinal increment thresholds for a foveal blue test light [4], impaired color differentiation (CD) [5], an impairment of contrast sensitivity (CS) for moving gratings [6] and optical coherence tomography (OCT) parameters [7–9] in HD humans. However, other studies reported no significant findings in OCT parameters [10] or OCT angiography [11]. All these studies included small number of participants or were methodologically insufficient, see S1 Table for details. Additionally, patients with HD do not report any kind of visual difficulties. Therefore, we decided to study both structural and functional parameters of retina in larger sample of patients with HD and compare them with normal controls.

2. Materials and methods

We recruited 44 patients with HD from a cohort of patients followed at our university HD clinic, to participate in the study, four of the patients were measured twice during the study period. This study protocol was reviewed and approved by ethical committee of General University Hospital in Prague IORG0002175 and all subjects gave written informed consent prior to the enrolment to the study.

Inclusion criteria were as follows: genetically confirmed HD patients (CAG triplet count 35 or more) with Unified Huntington’s disease Rating Scale Total Motor Score greater than 5 and older than 18 years of age willing to participate in the study between 3rd of October 2012 and 18th of April 2018.

Exclusion criteria were as follows: history of central nervous system disorder other than HD, inflammatory disorder of the eye in the last 3 months, history of optic neuritis, glaucoma, diabetic retinopathy, age-related macular degeneration, retinal and macular oedema, macular holes, vitreomacular traction syndrome, retinoschisis, retinal detachment, retinal neovascularization, and tumors. Patients were assessed by Unified Huntington’s disease Rating Scale (UHDRS). Ophthalmological examination included best corrected visual acuity (100% contrast), intraocular pressure measurement and fundus examination. Pelli-Robson Contrast Sensitivity test Chart 4K, which uses a single large letter size (20/60 optotype) with contrast varying across groups of letters, was used to evaluate CS. A Pelli-Robson score of 1.5 to 2.25 indicates normal CS, a score of 0.9 to 1.35 indicates moderate contrast loss, a score of 0.3 to 0.75 severe contrast loss and a score of 0.15 and less indicates profound contrast loss. Farnsworth D-15 Color test was performed to evaluate CD. The results of the Farnworth D-15 test determine color perception or defects in deutan, protan or tritan discrimination.

Using a spectral-domain OCT machine (Heidelberg Spectralis), without pupil dilation, we obtained a circular scan manually centered on the optic nerve head (diameter 3.4mm, Automated Real-Time—ART 100) and a macular volume scan centered on the fovea (63 lines, ART 20). The thickness of the peripapillary retinal nerve fiber layer (RNFL) in μm and total macular volume (TMV) in mm3 was automatically determined by the instrument (Heyex version 5.8). All retinal images were checked for scan quality. We analyzed the mean peripapillary RNFL thickness (RNFL-G) as the main measure of axonal health in the retina and the thickness of RNFL in the temporal peripapillary segment (RNFL-T) that represents the most vulnerable retinal fibers. We also used total macular volume (TMV) measured in the area of a 6 mm wide circular mask to represent global retinal changes.

As a control group, we selected 41 healthy subjects matched by age and gender from the clinic database of OCT measurements.

Study data were collected and managed using REDCap electronic data capture tool [12]. The data cleaning procedure was done using SciPy (Python for scientific computing) version 1.0.0 [13] and statistical analysis was done in R version 4.0.3. Inter-group difference was assessed as a hierarchical model of a subject consisted of two eyes (and repeated measurements in time in 4 cases) using repeated measures linear mixed-effects model (library lmerTest version 3.1.3 [14]) with group, age, and gender as fixed effects and subject as a random effect. Effect size was calculated as Hedges’ g (library effectsize version 0.6.0.1 [15]). Figures were generated using library ggplot2 version 3.4.0. Within HD group, we assessed the effect of UHDRS TMS and disease duration using repeated measures linear mixed-effects model with subject as a random effect. False discovery rate was corrected by Benjamini-Hochberg procedure, results were considered significant with adjusted P lower than 0.05.

3. Results

Forty-one patients from our cohort of 44 HD patients completed the study. Three patients were excluded from the study due to vitreoretinal pathology (because of possible distortion of OCT measurements). Measurements of HD patients were compared with 41 healthy controls (HC). Demographic data, UHDRS Total Motor Score (TMS) and Total Functional Capacity (TFC) listed in Table 1. The age and gender of HC group did not differ from the HD group (Mann-Whitney U-test p = 0.369 and Fisher’s exact test p = 0.8253 respectively). Summary of the OCT measurements and effect sizes of the intergroup difference are listed in Table 2.

Table 1. Demographic data in the Huntington’s disease group.

Demographic parameter Value
Number of HD patients (male: female) 41 (21:20)
Mean age in years (±SD) 50.6 (±12.0)
Mean disease duration in years (±SD) 7.1 (±3.6)
Mean CAG triplet repeats (±SD) 44.1 (±2.4)
Mean UHDRS TMS (±SD) 30.0±12.3
Mean UHDRS TFC (±SD) 8.2±3.2
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SD–standard deviation, TFC–Total Functional Capacity, TMS–Total Motor Score, UHDRS–Unified Huntington’s Disease Rating Scale

Table 2. RNFL-G thickness, RNFL-T thickness and macular volume parameters in Huntington’s disease patients and in healthy controls.

OCT parameter HD patients (mean ± SD) Control group (mean ± SD) Hedges’ g (0.95-CI)
RNFL thickness G 96.7±7.7 μm 101±8.8 μm 0.475 (0.157–0.792)
RNFL thickness T 69.6±10.4 μm 71.1±11.1 μm 0.138 (-0.175–0.451)
Total Macular volume 8.58±0.387 mm3 8.71±0.423 mm3 0.317 (-0.0145–0.647)
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RNFL–Peripapillary Retinal Nerve Fiber Layer, G–global segment, T–temporal segment, SD–standard deviation, CI–confidence interval.

The linear mixed effects models for global mean RNFL thickness (RNFL-G) and for TMV showed significant difference between HD patients and HC (P = 0.0272, P = 0.03894 respectively), however, this effect did not pass the false discovery rate adjustment (P = 0.0576, P = 0.06814 respectively) and the effect size was small. The model for temporal segment RNFL thickness (RNFL-T) was not significant. Interestingly, there was a very significant effect of gender on the macular volume. All P-values of the three intergroup models are listed in Table 3. Effect sizes of intergroup differences for RNFL-T, RNFL-G and TMV are listed in Table 3. Mean TMV in HD patients was 8.5±0.37 mm3.

Table 3. Linear mixed effects models for RNFL-G thickness, RNFL-T thickness, and macular volume with group, age and gender as fixed effects.

Response OCT variable Fixed effect P-value Adjusted P-value
RNFL-G thickness Group (HD vs. HC) 0.0272 0.0575
RNFL-G thickness Age 0.220 0.281
RNFL-G thickness Gender 0.700 0.735
RNFL-T thickness Group (HD vs. HC) 0.403 0.470
RNFL-T thickness Age 0.013 0.0346
RNFL-T thickness Gender 0.762 0.762
Total Macular Volume Group (HD vs. HC) 0.0389 0.068
Total Macular Volume Age 0.0548 0.0886
Total Macular Volume Gender 0.00154 0.004642
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RNFL–Peripapillary Retinal Nerve Fiber Layer, OCT–optical coherence tomography, G–global segment, T–temporal segment, HD–Huntington’s Disease, HC–healthy controls.

We analyzed the effect of disease duration and UHDRS TMS on RNFL thickness and TMV within the HD group also using linear mixed effects models. The effect of UHDRS TMS on temporal segment RNFL and on TMV was significant. None of these within group effects passed the false discovery rate adjustment. Other effects were not significant. All P-values of the three within HD group models are listed in the Table 4.

Table 4. Linear mixed effects models for RNFL-G thickness, RNFL-T thickness, and macular volume in the Huntington’s disease group with UHDR TMS and disease duration as fixed effects.

Response OCT variable Fixed effect P-value Adjusted P-value
HD RNFL-G thickness UHDRS TMS 0.0776 0.116
HD RNFL-G thickness Disease Duration 0.524 0.580
HD RNFL-T thickness UHDRS TMS 0.0294 0.0576
HD RNFL-T thickness Disease Duration 0.152 0.213
HD Total Macular Volume UHDRS TMS 0.0302 0.0576
HD Total Macular Volume Disease Duration 0.228 0.281
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RNFL–Peripapillary Retinal Nerve Fiber Layer, OCT–optical coherence tomography, G–global segment, T–temporal segment, HD–Huntington’s Disease, UHDRS–Unified Huntington’s Disease Rating Scale, TMS–Total Motor Score.

Pelli-Robson Contrast Sensitivity test Chart 4K was performed in 13 HD patients. Contrast sensitivity for both eyes was 1.64±0.04, which is within normal limits. Farnsworth D-15 Color test was performed in 14 HD patients. 6 patients had unspecified pathology, 6 had normal results, 1 patient had tritanopia and 1 patient had deuteranopia.

4. Discussion

There are a few studies proving the presence of mutant HTT in retina in animal HD models [1, 2]. Unfortunately, there is no post-mortem human study of retina in HD patients.

Even in long term clinical observation, HD patients report no subjective complaints about vision. On the other hand, several studies using various paraclinical tools (VEP, OCT etc.) found evidence for structural changes of retina or a functional impairment of visual pathways in HD patients [3–9]. However, a comparison of results between individual studies reveals conflicting findings in various parameters.

Total macular volume (TMV) was reduced in HD patients in study by Haider et al [8], but there was no difference between HD an HC in studies by Kersten et al [7], Andrade et al [10]. Our study showed significant difference (not passing the false discovery rate adjustment), but with small effect size, which may explain the discrepancy. To further confirm that the significant difference is indeed a false positive and false discovery rate adjustment was appropriate, we performed a post-hoc analysis. A correlation analysis of TMV and age (Pearson’s r = -0.24, P = 0.04) or triplet expansion length (Pearson’s r = -0.15, P = 0.20) in the patients group shows that changes of TMV are driven by aging only and not by changes associated with Huntington’s disease. To evaluate effect of Huntington’s disease on accelerated macular aging, we created a linear mixed-effects model with interaction of age and disease effects, however, this interaction was insignificant (P = 0.84). As can be noted in the Fig 1, the rate of macular loss with aging is even higher (not significantly) in the control group, which contradicts the notion that Huntington’s disease could accelerate the macular aging.

Fig 1. Scatter plot of macular volume vs. age in patient and control group.

Fig 1

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Note that the rate of macular volume loss with aging is steeper in the control group than in the patient group. The difference in slopes is not significant.

Additionally, there was a very significant effect of gender on TMV. As previously described in the literature [16], changes in male maculae are on average greater than in female maculae, and we have reproduced these results.

Mean total RNFL thickness was reduced in HD patients with Total Functional Score Stage III in study by Gatto et al [9] but there was no difference in studies by Kersten et al [7], Haider et al [8], Andrade et al [10], and Di Maio [11]. Our study showed significant difference (again not passing the false discovery rate adjustment), but with small effect size, which may, again, explain the discrepancy.

Temporal RNFL thickness was hypothesized to be the part of RNFL most vulnerable to neurodegeneration [9]. Kersten et al [7] and Gatto et al [9] found it reduced in HD patients but there was no difference in the study by Haider et al [8]. Our study didn’t show a significant intergroup difference. However, there was a significant effect of age in the intergroup model. Additionally, we found a significant effect of UHDRS TMS in the within HD group model, but this effect did not pass the false discovery rate adjustment. Our results support the notion that temporal RNFL atrophy is driven by age (rather than disease), which could explain the incongruity of the studies with different populations.

Peripapillary RNFL thickness was not different in HD patients from healthy controls in studies by Kersten et al [7] and Haider et al [8]. Inferior and nasal RNFL thickness were not altered by HD in the study by Gatto [9], were not examined in other studies and are not reported as significant to neurodegeneration. Therefore, we did not examine them to limit multiple statistical comparisons and increase statistical power of our study. Referenced studies, their sample size and their main findings are summarized and compared in S1 Table.

In our prospective cross-sectional study, we examined 82 eyes–the largest study sample on the topic. In 41 HD, we measured both retinal structural and visual functional parameters, six subjects were examined twice leaving some possibility of longitudinal evolution of the parameters. We focused not only on the detection on the impairment but also on mutual relationship of functional and structural parameters. To our best knowledge, only single study by Kersten et al [7] investigated OCT and color discrimination together as in our study. Our findings in larger sample size did not confirm the results of the study by Kersten et al [7], who reported a negative correlation between macular volume and disease duration and UHDRS motor score. However, in Kersten study, there were no macular volume changes between HD and controls. It could be speculated that the macular volume changes may be effect of aging rather than effect of HD, since they are in both groups of subjects. We also cannot corroborate the correlation between temporal RNFL thickness and disease duration [7].

Furthermore, we found no pathology for contrast sensitivity evaluated by Pelli-Robson Contrast Sensitivity test Chart 4K in HD patients. This test examines the stationary CS, which reflects the functional integrity of parvocellular pathways, sensitive to high spatial frequencies and low temporal frequencies. O’Donnell et al [6] reported a selective deficit in CS for moving gratings in HD patients, dependent on the selective dysfunction in the magnocellular pathway, sensitive to low spatial frequencies, high temporal frequencies, and luminance.

Our results for color discrimination evaluated by Farnsworth D-15 Color test were nonspecific and ambiguous. Cognitive impairment may be an important factor interfering with the color discrimination. Büttner et al [5] found the mean total error scores and the partial scores for the red-green and the blue-yellow axes in Farnsworth-Munsell 100 Hue test which is more sensitive than our Farnsworth D-15 Color test. Kersten et al [7] found incorrect identification of Ishihara plates in HD patients, but they did not distinguish among types of color discrimination impairments. Ishihara plates method is used mainly for testing color discrimination in congenital disorders and may not be applicable in HD.

Haider et al [8] found no difference in ophthalmological findings in HD patients compared with HC, such as in our study.

5. Study limitations

Even though this study is the largest done on the topic, the sample size still may be underpowered to show significant difference. HD is a rare disease, and it is a challenge to reach larger group of patients willing to participate in the study. Additionally, if OCT and functional parameters correlated with disease progression, the findings would be most profound in later stages of the disease, when it is most difficult for the patients to undergo all possible examinations and endure long examination time. Therefore, we omitted some of the functional and structural examinations (VEP, foveal blue test, OCT angiography) that may have been stressful or exhausting for the study participants. Scotopic/photopic testing described in rats [2] was not performed. Only adult patients were included in the cohort and maximum triplet expansion length was 51, so these results are not transferable to juvenile Huntington’s disease or late disease stages. Retinal ganglion cell complex was not analyzed. Analysis of the individual retinal layers of the macula requires a high-quality scan. Examination of the retina using OCT had to be adapted to involuntary movements and the ability to cooperate during examination in patients with HD. In many patients, due to the limitations of the current technology, the quality of the scans does not allow reliable segmentation of the retinal layer in the macula.

6. Conclusion

The results of our study support the notion that there are abnormalities in the temporal RNFL layer of Huntington’s disease patients. However, it seems that, within the limits of the current technology and software, the magnitude of these abnormalities is very small and not clinically useful. There were no other changes in the structural parameters of RNFL layer and total macular volume by means of spectral domain OCT and no changes in selected functional parameters in HD patients. According to our results, retinal spectral domain OCT at its present development for measurement of RNFL layer and total macular volume is not appropriate as a marker of HD progression. D-15 color testing is not a good marker of retinal changes associated with Huntington’s disease. Surprisingly, our findings differ from most of the previous published studies. Prospective longitudinal studies and/or meta-analyses of all published studies are needed to solve this discrepancy. Additionally, further studies may focus more on other retinal layers, such as retinal ganglion cells.

Supporting information

S1 Table. Comparison of published studies and their findings in the main OCT parameters.

HD–Huntington’s disease patients, HC–healthy controls

(DOCX)

Click here for additional data file. (14.2KB, docx)

Data Availability

The data that support the findings of this study are available in the GitHub repository (https://doi.org/10.5061/dryad.ncjsxksxr). Analysis source code may be found in the GitHub repository (https://github.com/PavelDusek/hd-oct).

Funding Statement

Czech Science Foundation project number and number 19–01747S Ministry of Health of the Czech Republic project number AZV–NU20–04–0136 Joint Programme – Neurodegenerative Disease Research (JPND) project number 8F19004 National Institute for Neurological Research (Programme EXCELES, ID Project No. LX22NPO5107) Funded by the European Union – Next Generation EU Charles University: Cooperatio Program in Neuroscience General University Hospital in Prague project MH CZ-DRO-VFN64165. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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PONE-D-22-18702Is Retina Affected in Huntington’s Disease? Is Optical Coherence Tomography a good biomarker?PLOS ONE

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Reviewer #1: The authors evaluate retinal OCT findings as a possible biomarker for HD severity and progression. This is the largest number of patients included in a retinal evaluation of HD, and thus deserves a comprehensive analysis of the different layers, in addition to the RNFL and RGC complex. The temporal RNFL changes they report are small. And there is enough variability that 4 referenced studies were evenly divided in terms of these changes. The expansion repeats (~44) are in agreement with adult onset/complete penetrance, but do not take into consideration more severe/juvenile forms, nor does it consider late disease. However, this is a minor consideration, particularly since the authors are talking about identifying biomarkers for HD. To compare with expansion repeat numbers, age and disease duration, the authors use SD-OCT, contrast sensitivity (for acuity) and D15 color testing. Scotopic/photopic testing was not performed. This is of interest because the authors quote a previous functional study released only as an abstract (Johnson et al, 2012) which suggested inner retinal dysfunction that may be unassociated with RGCs.

Previous HD models have shown retinal degeneration, but it is unclear whether the changes in the early models (in the fly and mouse) are true indicators of the disease, since their expansion repeats were huge, and may have contributed to unusual retinal degeneration patterns similar to retinitis pigmentosa.

Previous reports have shown a decline in contrast sensitivity in HD patients, suggesting a loss of outer retinal, or other outer-retina-associated function, but the majority of studies have really focused on inner retinal function; in particular retinal ganglion cells (RGCs) and their axons The authors found a loss of RNFL thickness in the termporal region, but this was a trend, and not significant. They conclude that there are global RNFL changes, but not suitable as a biomarker, and that retinal parameters are not appropriate for monitoring HD disease progression. This is a considerable leap, since what they are really saying is that a) retinal SD-OCT at its present development is not suitable. B) the retinal parameters they looked at are not suitable. C) D-15 color testing is not a good damage marker of retinal parameter changes in HD. All are valuable considerations. I question then why they found changes in total macular volume in HD patients, which does seem to be significant, and which does suggest some associated retinal function besides RNFL. It also suggests that there may be changes in layers other than the RNFL.

In short, the authors need to utilize their data to perform a more detailed analysis of the different layers of the retina. This would make it more comprehensive, and more accurate for the conclusions that they draw (that OCT is useless in identifying HD retinal biomarkers).

**********

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Reviewer #1: No

**********

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PLoS One. 2023 Feb 24;18(2):e0282175. doi: 10.1371/journal.pone.0282175.r002

Author response to Decision Letter 0

18 Dec 2022

Dear reviewers, 

we are pleased to read your comments and insights to our work. You have provided us with key feedback that help us do better research. We had to deal with some issues by stating them in the limitations section. However, we still think that our findings are worth as a publication, since it contradicts older studies and has the largest sample size. We have addressed the comments by making the conclusion more appropriate to our findings. 

Our response to comments is as follows: 

- "The authors evaluate retinal OCT findings as a possible biomarker for HD severity and progression. This is the largest number of patients included in a retinal evaluation of HD, and thus deserves a comprehensive analysis of the different layers, in addition to the RNFL and RGC complex. The temporal RNFL changes they report are small. And there is enough variability that 4 referenced studies were evenly divided in terms of these changes. The expansion repeats (~44) are in agreement with adult onset/complete penetrance, but do not take into consideration more severe/juvenile forms, nor does it consider late disease. However, this is a minor consideration, particularly since the authors are talking about identifying biomarkers for HD."

We added the fact that this study included only adult Huntington's disease patients to the study limitations. 

- "To compare with expansion repeat numbers, age and disease duration, the authors use SD-OCT, contrast sensitivity (for acuity) and D15 color testing. Scotopic/photopic testing was not performed. This is of interest because the authors quote a previous functional study released only as an abstract (Johnson et al, 2012) which suggested inner retinal dysfunction that may be unassociated with RGCs."

We added to the study limitations that the scotopic/photopic testing was not performed. 

- "Previous HD models have shown retinal degeneration, but it is unclear whether the changes in the early models (in the fly and mouse) are true indicators of the disease, since their expansion repeats were huge, and may have contributed to unusual retinal degeneration patterns similar to retinitis pigmentosa. 

Previous reports have shown a decline in contrast sensitivity in HD patients, suggesting a loss of outer retinal, or other outer-retina-associated function, but the majority of studies have really focused on inner retinal function; in particular retinal ganglion cells (RGCs) and their axons The authors found a loss of RNFL thickness in the termporal region, but this was a trend, and not significant. They conclude that there are global RNFL changes, but not suitable as a biomarker, and that retinal parameters are not appropriate for monitoring HD disease progression. This is a considerable leap, since what they are really saying is that a) retinal SD-OCT at its present development is not suitable. B) the retinal parameters they looked at are not suitable. C) D-15 color testing is not a good damage marker of retinal parameter changes in HD. All are valuable considerations."

We have changed the conclusion section accordingly. 

- "I question then why they found changes in total macular volume in HD patients, which does seem to be significant, and which does suggest some associated retinal function besides RNFL. It also suggests that there may be changes in layers other than the RNFL."

We performed a post-hoc analysis showing that the difference in macular volume was a false positive and was appropriately corrected by the false discovery rate correction. We showed that there is no interaction between age and triplet expansion length in the effect on macular volume. The rate of macular volume loss with aging is even (not significantly) steeper in the control group than in the patients group. These analyses were added to the discussion section. 

- "In short, the authors need to utilize their data to perform a more detailed analysis of the different layers of the retina. This would make it more comprehensive, and more accurate for the conclusions that they draw (that OCT is useless in identifying HD retinal biomarkers)."

Analysis of the individual retinal layers of the macula requires a high-quality scan. Examination of the retina using OCT had to be adapted to involuntary movements and the ability to cooperate during the examination in patients with HD. In many patients, the quality of scans does not allow reliable segmentation of the retinal layers in the macula – because of their involuntary movements.  

In any case, we think that the examination that we cannot reliably perform in these patients is not a suitable method for studying the progression of the disease. 

We have changed the limitations section to state these issues and the conclusion section accordingly to make statements about the performed methods only. 

All the changes in the text were highlighted in yellow. 

Additionally, during the review process, funding of our work changed, so we edited the funding sources slightly.  

Thank you for reviewing our work. We look forward to your opinion on our changes. 

Yours faithfully, 

Pavel Dusek

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Decision Letter 1

Alfred S Lewin

1 Feb 2023

PONE-D-22-18702R1

Is Retina Affected in Huntington’s Disease? Is Optical Coherence Tomography a good biomarker?

PLOS ONE

Dear Dr. Dusek,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

Please revise your paper to state that it is difficult to assess Huntington disease patients using current OCT technology, not that OCT cannot be used to evaluate these patients. The expert reviewer has indicated where changes should be made in your paper in the attached "reviewer's comments"

Please submit your revised manuscript by Mar 18 2023 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

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If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

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We look forward to receiving your revised manuscript.

Kind regards,

Alfred S Lewin, Ph.D.

Section Editor

PLOS ONE

Journal Requirements:

Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice.

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Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: All comments have been addressed

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: The authors have replied to my review with some limited changes. My major concern is that it is important to recognize that while the SD-OCT test has wide capabilities, it is limited by the problems of the current ability to examine. I have also evaluated HD patients, and their shake/eye movements make it difficult to get a ‘lock on’ and high quality scans. Thus, it is necessary to distinguish the problems of CURRENT technology, and not simply say that OCT cannot be used (it cannot be used without more refinements). Otherwise, people may simply quote their paper to show ‘it cannot be done’, and suppress future research. Thus the suggested changes:

  1. abstract: in conclusions: ‘Current’, not ‘These

  2. Discussion/third paragraph: add ‘Changes in male maculae are….

  3. Discussion/last paragraph/last sentence: ‘In many patients, due to the limitations of the current technology, the quality of the scans….

  4. Conclusions:

  5. However, it seems that, within the limits of current technology and software,

**********

7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: Yes: Steven L. Bernstein

**********

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PLoS One. 2023 Feb 24;18(2):e0282175. doi: 10.1371/journal.pone.0282175.r004

Author response to Decision Letter 1

7 Feb 2023

Dear prof. Bernstein,

we thank you for your time and for your suggested changes to improve our research. We incorporated them fully into the manuscript:

1. In the Abstract, we changed ‘These retinal parameters are not appropriate for monitoring HD disease progression.’ to ‘Current retinal parameters are not appropriate for monitoring HD disease progression.’

2. In the Discussion, we changed ‘Male maculae are on average greater than female maculae.’ to ‘Changes in male maculae are on average greater than in female maculae.’

3. In the Study Limitations, we changed ‘In many patients, the quality of the scans does not allow reliable segmentation of the retinal layer in the macula.’ to ‘In many patients, due to the limitations of the current technology, the quality of the scans does not allow reliable segmentation of the retinal layer in the macula.’

4. In the Conclusion, we changed ‘However, it seems that the magnitude of these abnormalities is very small and not clinically useful.’ to ‘However, it seems that, within the limits of the current technology and software, the magnitude of these abnormalities is very small and not clinically useful.’

Additionally, we included the Figure 1 description into the manuscript file, as requested by the Editorial Office.

We think that the revised manuscript is now more balanced in terms of study limitations and possible suggestions for future research. We look forward to your opinion on the manuscript.

Yours sincerely,

Pavel Dusek

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Decision Letter 2

Alfred S Lewin

9 Feb 2023

Is Retina Affected in Huntington’s Disease? Is Optical Coherence Tomography a good biomarker?

PONE-D-22-18702R2

Dear Dr. Dusek,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication.

An invoice for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org.

If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org.

Kind regards,

Alfred S Lewin, Ph.D.

Section Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments:

Acceptance letter

Alfred S Lewin

14 Feb 2023

PONE-D-22-18702R2

Is Retina Affected in Huntington’s Disease? Is Optical Coherence Tomography a good biomarker?

Dear Dr. Dusek:

I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department.

If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact onepress@plos.org.

If we can help with anything else, please email us at plosone@plos.org.

Thank you for submitting your work to PLOS ONE and supporting open access.

Kind regards,

PLOS ONE Editorial Office Staff

on behalf of

Dr. Alfred S Lewin

Section Editor

PLOS ONE

Associated Data

    This section collects any data citations, data availability statements, or supplementary materials included in this article.

    Supplementary Materials

    S1 Table. Comparison of published studies and their findings in the main OCT parameters.

    HD–Huntington’s disease patients, HC–healthy controls

    (DOCX)

    Click here for additional data file. (14.2KB, docx)
    Attachment

    Submitted filename: Response to Reviewers.docx

    Click here for additional data file. (15.7KB, docx)
    Attachment

    Submitted filename: REview Jan 2023.docx

    Click here for additional data file. (12.8KB, docx)
    Attachment

    Submitted filename: Response to Reviewers_2023-02-08.docx

    Click here for additional data file. (10.5KB, docx)

    Data Availability Statement

    The data that support the findings of this study are available in the GitHub repository (https://doi.org/10.5061/dryad.ncjsxksxr). Analysis source code may be found in the GitHub repository (https://github.com/PavelDusek/hd-oct).

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