To determine whether patients with amyotrophic lateral sclerosis (ALS) show retinal axon pathology. METHODS. Postmortem eyes from 10 patients with ALS were sectioned and compared with 10 age-matched controls. Retinal sections were evaluated with periodic acid Schiff and phosphorylated (P-NF) and nonphosphorylated (NP-NF) forms of neurofilament with SMI 31 and 32 antibodies. Spheroids identified in the retinal nerve fiber layer were counted and their overall density was calculated in central, peripheral, and peripapillary regions. P-NF intensity was quantified. Morphometric features of ALS cases were compared with age-matched controls using the exact Wilcoxon matched-pairs signed-rank test. RESULTS. Distinct periodic acid Schiff-positive round profiles were identified in the retinal nerve fiber layer of patients with ALS and were most commonly observed in the peripapillary and peripheral retina. The density of periodic acid Schiff-positive spheroids was significantly greater in patients with ALS compared with controls (P = 0.027), with increased density in the peripapillary region (P = 0.047). Spheroids positive for P-NF and NP-NF were detected. P-NF-positive spheroid density was significantly increased in patients with ALS (P = 0.004), while the density of NP-NF spheroids did not differ significantly between ALS and control groups (P > 0.05). P-NF immunoreactivity in the retinal nerve fiber layer was significantly greater in patients with ALS than in controls (P = 0.002). CONCLUSIONS. Retinal spheroids and axon pathology discovered in patients with ALS, similar to hallmark findings in spinal cord motor neurons, point to disrupted axon transport as a shared pathogenesis. Retinal manifestations detected in ALS suggest a novel biomarker detectable by noninvasive retinal imaging to help to diagnose and monitor ALS disease.

Amyotrophic lateral sclerosis (ALS) is an incurable motor neuron disease with no current valid diagnostic imaging biomarkers. The retina is an extension of the central nervous system and axonal transport defects have been documented in various neurodegenerative diseases. This study reports evidence of axonal pathology in the retina of ALS patients using an interdisciplinary approach that includes the neuropathological study of retinal sections in ALS patients expanded to the optical characteristics of the whole retina preparations using eye imaging technology. The histopathological examination of retina sections revealed round profiles in the retinal nerve fibre layer in 10 out 10 ALS patients and in 4 out of 10 age-matched control patients. All 10 ALS patients showed increased phosphorylated neurofilament immunoreactivity in the retinal nerve fibre layer compared to all 10 control patients. Retinal imaging of whole globes and retina flat-mounts by blue reflectance retinal funduscopy and optical coherence tomography revealed hyper-reflective profiles in the retinal nerve fibre layer. For the first time, approximately 1µm retinal ganglion cells axons were visualized in immunofluorescence stained retina flat-mounts using near-infrared retina fundus imaging and Image Mapping Spectrometer. These findings suggest axonal pathology in retinal ganglion cells and its potential use as a novel non-invasive ocular imaging biomarker for ALS.

Amyotrophic lateral sclerosis (ALS) is an incurable motor neuron disease with no current valid diagnostic imaging biomarkers. The retina is an extension of the central nervous system and axonal transport defects have been documented in various neurodegenerative diseases. This study reports evidence of axonal pathology in the retina of ALS patients using an interdisciplinary approach that includes the neuropathological study of retinal sections in ALS patients expanded to the optical characteristics of the whole retina preparations using eye imaging technology. The histopathological examination of retina sections revealed round profiles in the retinal nerve fibre layer in 10 out 10 ALS patients and in 4 out of 10 age-matched control patients. All 10 ALS patients showed increased phosphorylated neurofilament immunoreactivity in the retinal nerve fibre layer compared to all 10 control patients. Retinal imaging of whole globes and retina flat-mounts by blue reflectance retinal funduscopy and optical coherence tomography revealed hyper-reflective profiles in the retinal nerve fibre layer. For the first time, approximately 1µm retinal ganglion cells axons were visualized in immunofluorescence stained retina flat-mounts using near-infrared retina fundus imaging and Image Mapping Spectrometer. These findings suggest axonal pathology in retinal ganglion cells and its potential use as a novel non-invasive ocular imaging biomarker for ALS.