Promoting remyelination: utilizing a viral model of demyelination to assess cell-based therapies

Abstract

Multiple sclerosis (MS) is a chronic inflammatory disease of the CNS. While a broad range of therapeutics effectively reduce the incidence of focal white matter inflammation and plaque formation for patients with relapse-remitting forms of MS, a challenge within the field is to develop therapies that allow for axonal protection and remyelination. In the last decade, growing interest has focused on utilizing neural precursor cells (NPCs) to promote remyelination. To understand how NPCs function in chronic demyelinating environments, several excellent pre-clinical mouse models have been developed. One well accepted model is infection of susceptible mice with neurotropic variants of mouse hepatitis virus (MHV) that undergo chronic demyelination exhibiting clinical and histopathologic similarities to MS patients. Combined with the possibility that an environmental agent such as a virus could trigger MS, the MHV model of demyelination presents a relevant mouse model to assess the therapeutic potential of NPCs transplanted into an environment in which inflammatory-mediated demyelination is established.

Keywords: demyelination; multiple sclerosis; neural precursor cells; neural progenitor cells; remyelination; virus.

Figure 1. Histological characteristics of JHMV-induced neurologic disease

(A) A representative spinal cord was taken from an infected mouse at 5 weeks postinfection and stained with luxol fast blue to determine the extent of demyelination within the white matter tracts. Extensive demyelination is observed throughout the anterior and lateral regions of the white matter. (B) Electron micrograph images (1200×) showing demyelinated axons (red arrows) and remyelinated axons (blue arrow) at 5 weeks postinfection. Reproduced with permission from [119].

Figure 2. Axonal protection and remyelination transplantation of mouse neural precursor cells into JHMV-infected mice

Transplanted NPCs expressing cytoplasmic GFP driven by the myelin proteolipid protein promoter wrap around damaged axons. (A) Representative image showing colocalization between PLP-GFP (green) and damaged axons (yellow) in the JHMV-infected Thy1-YFP spinal cord 15 days post-transfer. Panels on the right show enlarged time lapse images of a PLP-GFP-positive cell wrapping around an axon (minutes:seconds). The top panels depict x–y sections, the lower panels x–z sections. (B) Analysis of the change in volume of PLP-GFP fluorescence of three different NPCs during wrapping as determined by a time lapse 3D reconstruction of the 2P data and the Imaris ‘Surfaces’ tool. (C) Analysis of the change in Thy1-YFP fluorescence intensity during PLP-GFP wrapping presented as arbitrary fluorescent units (AU). (D) Immunostaining in a transverse section of a JHMV-infected Thy1-YFP spinal cord 21 days after GFP-NPC transplantation. MBP (red), YFP⁺ axons (yellow) and colocalization between overlapping GFP-NPC (green) and MBP fluorescence was determined using the Imaris colocalization tool (white). Merging of all three channels is shown on the right. Scale bar = 4 μm. NPC: Neural precursor cell. Reproduced with permission from [107].

Figure 3. Intraspinal transplantation of hNPCs into JHMV-infected mice

(A) Improved (p < 0.05) clinical recovery in hNPC-transplanted JHMV-infected mice was sustained out to 168 days post-transplantation (p.t.) compared with infected mice treated with vehicle alone. (B) Daily IVIS® imaging of luciferase-labeled hNPCs revealed that following intraspinal transplantation, cells are reduced to below the level of detection by day 8 post-transplantation; representative mice are shown. IVIS imaging was performed on vehicle-transplanted mice as a control. (C) Quantification of Treg numbers in spinal cords of mice indicated a significant (p < 0.05) increase in numbers of Tregs in hNPC-transplanted mice versus controls between 8 and 10 days post-transplantation. Data are representative of three independent experiments with a minimum of three mice per group; data are presented as average ± SEM. Mann-Whitney t tests were used to determine the p-values. (D) hNPC-transplanted mice receiving anti-CD25 antibody (purple line) did not display recovery in motor skills compared with either hNPC-treated mice (red line), hNPC-treated mice receiving isotype-matched control antibody (green line) or vehicle control mice (blue line). Reproduced with permission from [119].