Micropillar arrays as a high-throughput screening platform for therapeutics in multiple sclerosis

Abstract

Functional screening for compounds that promote remyelination represents a major hurdle in the development of rational therapeutics for multiple sclerosis. Screening for remyelination is problematic, as myelination requires the presence of axons. Standard methods do not resolve cell-autonomous effects and are not suited for high-throughput formats. Here we describe a binary indicant for myelination using micropillar arrays (BIMA). Engineered with conical dimensions, micropillars permit resolution of the extent and length of membrane wrapping from a single two-dimensional image. Confocal imaging acquired from the base to the tip of the pillars allows for detection of concentric wrapping observed as ‘rings’ of myelin. The platform is formatted in 96-well plates, amenable to semiautomated random acquisition and automated detection and quantification. Upon screening 1,000 bioactive molecules, we identified a cluster of antimuscarinic compounds that enhance oligodendrocyte differentiation and remyelination. Our findings demonstrate a new high-throughput screening platform for potential regenerative therapeutics in multiple sclerosis.

Clemastine fumarate as a remyelinating therapy for multiple sclerosis (ReBUILD): a randomised, controlled, double-blind, crossover trial

“…Preclinical work validated the efficacy of clemastine fumarate in vitro and in animal models. 17,19 Additional work confirmed the efficacy of clemastine fumarate in multiple animal models [17][18][19][20][21][22] and showed that this benefit was mediated specifically via remyelination induced from oligo dendrocyte differentiation and not via effects on the immune system. 19 We furthermore showed the capacity of clemastine fumarate to induce oligodendrocyte progenitor cell (OPC) differentiation and myelination with human OPCs (appendix).…”

“…Preclinical data unequivocally showed that clemastine fumarate promotes oligodendrocyte precursor differentiation 17,18 and remyelination [19][20][21][22] without modulating the immune system. 19 The robustness of the findings were documented by the latency improvement observed in both groups of the cohort while on active treatment.…”

“…2,15,16 Whether or not remyelination can be achieved in chronically demyelinated lesions in multiple sclerosis remains an unanswered question. A binary, cell-specific, functional screening method identified clemastine fumarate-a first generation antihistamine-as capable of inducing oligodendrocyte differentiation and myelination due to off-target antimuscarinic effects 17 and was confirmed in a second independent screen. 18 Clemastine fumarate readily crosses the blood-brain barrier and has been available over the counter in the USA since 1992.…”

Experimental Demyelination and Remyelination of Murine Spinal Cord by Focal Injection of Lysolecithin

“…Additionally, the ventral white matter makes up a much larger target area to inject; several hundred microns laterally in the dorsal region would place the capillary outside the column, while the same deviation ventrally would still produce a prominent demyelinating lesion. Some protocols inject lysolecithin into both the dorsal and ventral columns of the same animal 19 . This can increase both the likelihood of proper capillary placement and the number of quantifiable lesions in fewer animals.…”

Model Systems to Define Remyelination Therapies

“…Purification of rodent or murine OPCs either through differential antibody binding (panning) or FACS sorting allows for assessment of the direct response of the cell population to therapeutic exposure. Such approaches have been used recently to identify signaling mechanisms that promote the appearance of mature oligodendrocytes [75][76][77][78]. One concept that has gained significant support in recent years is the notion that the rate-limiting step in remyelination is the differentiation and maturation of oligodendrocytes to myelinating cells.…”