Bedside to bench to bedside research: Estrogen receptor beta ligand as a candidate neuroprotective treatment for multiple sclerosis

Abstract

Protective effects of pregnancy during MS have led to clinical trials of estriol, the pregnancy estrogen, in MS. Since estriol binds to estrogen receptor (ER) beta, ER beta ligand could represent a "next generation estriol" treatment. Here, ER beta ligand treatment was protective in EAE in both sexes and across genetic backgrounds. Neuroprotection was shown in spinal cord, sparing myelin and axons, and in brain, sparing neurons and synapses. Longitudinal in vivo MRIs showed decreased brain atrophy in cerebral cortex gray matter and cerebellum during EAE. Investigation of ER beta ligand as a neuroprotective treatment for MS is warranted.

Keywords: Estrogen; Experimental autoimmune encephalomyelitis; Multiple sclerosis; Neuroprotection; Pregnancy.

Copyright © 2016 Elsevier B.V. All rights reserved.

Figures

Figure 1. Protective effects of ER beta ligand treatment on female and male mice with chronic EAE

ER beta ligand treatment (30 mg/kg/every other day) or vehicle treatment initiated after EAE onset (day 13) resulted in significant reductions in EAE clinical severity scores (Table 1) and an increase in the number of seconds on the rotarod, p

Figure 2. Whole brain, cerebral cortical and cerebellar atrophy in EAE are reduced by ER beta ligand treatment

(A-C) Anatomic delineations of cerebral cortex (green) and cerebellum (yellow) overlaid onto the coronal, sagittal and horizontal planes of a minimum deformation atlas comprising all images from all groups of mice. (D) Mean whole brain volume over time in healthy controls (black), ER beta ligand-treated mice with EAE (red) and vehicle-treated mice with EAE (blue) at d0, d30 and d60. ER beta ligand-treated EAE mice exhibit less brain atrophy than vehicle-treated EAE mice as early as d30. (E) Mean cerebral cortex volume in the same groups. ER beta ligand-treated EAE mice exhibit less atrophy in the cerebral cortex than vehicle-treated EAE mice by d60. (F) Mean cerebellar volumes in the same groups. ER beta ligand-treated EAE mice exhibit less cerebellar atrophy than vehicle-treated EAE mice by d60.

Figure 3. ER beta ligand treatment of EAE: protective effects in cerebral cortical gray matter

Representative 10X images of cerebral cortical gray matter neurons stained for NeuN in green (A) and synapses stained for the postsynaptic protein PSD-95 in red (B) in healthy control (left), vehicle treated EAE (middle), and ER beta ligand treated EAE (right). Quantification of immunofluorescence staining is shown in bar graphs. Vehicle treated EAE mice (blue bars) had fewer NeuN+ cortical neurons and less PSD-95+ synapses in the cerebral cortex as compared to age-matched healthy controls (black bars). ER beta ligand treated EAE mice (red bars), as compared to vehicle treated EAE mice (blue bars), had higher numbers of NeuN+ cortical neurons and PSD-95+ synapses in the cerebral cortex, with values comparable to age-matched healthy controls. In A, cc indicates corpus callosum adjacent to cerebral cortex. In B, tissues were counter stained with the nuclear stain DAPI (blue). Three to five mice were examined for each treatment group. ** p < 0.005, * p < 0.05, with p-values determined by one-way ANOVA.

Figure 4. ER beta ligand treatment of EAE: protective effects in cerebellar white and gray matter

(A) Representative 10X images of cerebellar white matter immunofluorescence stained with MBP (red, top) and Neurofilament-200 (green, bottom) in healthy control (left), vehicle treated EAE (middle), and ER beta ligand treated EAE (right). Quantification of myelin staining intensity by MBP and axonal densities by NF200 staining is shown by bar graphs. Vehicle treated EAE (blue bars) as compared to healthy controls (black bars) showed reduced myelin staining, while levels of MBP staining in ER beta ligand treated were increased and similar to those in healthy controls. Axon numbers were reduced in vehicle treated EAE compared to healthy controls, wile they were increased in ER beta ligand treated EAE mice. (B) Representative 10X images of cerebellum including the Purkinje layer, stained for Calbindin using chromogen immunohistochemistry (brown, top) and PSD-95 using immunofluorescence (red, bottom) in the same groups of mice. Vehicle treated EAE mice, had fewer Purkinje cells in the Purkinje layer and less PSD-95 staining in the molecular layer of the cerebellum as compared to age-matched healthy controls. ER beta ligand treated EAE mice, as compared to vehicle treated EAE mice, had higher numbers of Purkinje cells and higher levels of PSD-95 staining. IN B, tissues were counter stained with the nuclear stain DAPI (blue). Three to five mice were examined for each treatment group. *** p