Ocular pathology in multiple sclerosis: retinal atrophy and inflammation irrespective of disease duration

Abstract

There has been growing interest in the use of retinal imaging for tracking disease progression in multiple sclerosis. However, systematic and detailed pathological descriptions of retinal tissue in multiple sclerosis are lacking. Graded, histological evaluations on eyes from 82 patients with multiple sclerosis and 10 subjects with other neurological diseases, with immunohistochemistry on a subset, were performed and correlated with clinical and pathological findings. Multiple sclerosis cases demonstrated evidence of retinal atrophy and inflammation even in late-stage disease. Retinal ganglion cell loss was significant and remaining neurons appeared shrunken and were partially engulfed by human leukocyte antigen-DR positive cells with the phenotype of microglia in samples subjected to immunohistochemistry. Neurofilament staining revealed variable but prominent degrees of axonal loss and injury. Neuronal loss was noted in the inner nuclear layer with focal reduction in cell density. Foamy-appearing human leukocyte antigen-DR positive cells were evident near vessels and periphlebitis was found in a small but significant number of multiple sclerosis cases. Glial fibrillary acidic protein staining showed extensive astrocyte hypertrophy and proliferation with prominent gliosis in multiple sclerosis cases. Frequent but previously unreported abnormalities in the iris were documented in the majority of chronic multiple sclerosis cases. The injury to both iris and retina could be seen at all stages of disease. Severity of retinal atrophy was correlated with overall brain weight at time of autopsy (P = 0.04) and a trend for increased atrophy was seen with longer disease duration (P = 0.13). This study provides the first large-scale pathological description of retinas in multiple sclerosis, including patients with different subtypes of disease at all stages, and with variable clinical severity. Changes were seen not only in the retinal nerve fibre layer and ganglion cell layer, but also in the inner nuclear layer, suggesting that retinal injury is more widespread than previously appreciated. Furthermore, the human retina is devoid of myelin, but inflammation was demonstrated to be prominent in multiple sclerosis and to persist in the retina at late stages of disease. The prominent gliosis and inflammation surrounding vessels of the inner retina could potentially impact optical coherence tomography evaluations in multiple sclerosis-as standard techniques exploit presumed differences in tissue reflectivity and utilize automated edge detection algorithms to judge axon loss in the nerve fibre layer. Deciphering the relationships between the different types of retinal pathology may aid us in understanding the factors that drive both inflammation and tissue atrophy in multiple sclerosis.

Figure 1

Haematoxylin and eosin and immunoperoxidase labelling of formalin-fixed, paraffin-embedded microtome sections from other neurological disease control or multiple sclerosis eye samples. Layers of retina are as follows: NFL = nerve fibre layer; GCL = ganglion cell layer; IPL = inner plexiform layer; INL = inner nuclear layer; OPL = outer plexiform layer; ONL = outer nuclear layer; IS = inner segments; OS = outer segments; RPE = retinal pigment epithelium; choroid. (A) Control sample: normal distribution of cells in the ganglion cell layer and inner nuclear layer. (B, C) Varying degrees of disruption in the ganglion cell layer and inner nuclear layer in patients with multiple sclerosis; B shows mild and C severe changes. (D) Perivascular cellular infiltration and periphlebitis in the retina (arrows). (E) Mild gliosis (arrow) in the optic nerve head. (F) Severe gliosis in the optic nerve head. (G, H) Inflammation involving the stroma of the iris.

Figure 2

Haematoxylin and eosin, immunoperoxidase and dual immunofluoresence/confocal microscopy labelling of formalin-fixed, paraffin-embedded, microtome sections from multiple sclerosis eyes. (A, B) Contrast in distribution of neurofilaments in the optic nerve head (black arrows) in two patients with multiple sclerosis; A mild and B severe loss. GFAP immunohistochemistry highlights (C) perivenular gliosis in the optic disc and (D) intense glial-mesodermal reaction in the optic nerve. (E) Many of the inflammatory cells surrounding blood vessels expressed HLA-DR (arrow) and HLA-DR positive cells with ramified processes are also present in the inner nuclear layer in multiple sclerosis eye sections (F, arrow). (G) Tight junction expression (ZO1—Alexa-488, green; propidium iodide stained nuclei -red) in a retinal blood vessel. Most of the junction strands appear normal. However, there is evidence of ‘opening’ of tight junction strands between some endothelial cells (arrow). (H) Dual-imunofluoresence labelling for ZO1 (Alexa-568, red) and GFAP (Alexa-488, green), demonstrates loss of tight junction strands in a retinal blood vessel (arrowed) which is associated with a pronounced glial cell reaction.