Manipulation of extracellular matrix mechanical cues to stimulate oligodendrocytes to promote remyelination

The purpose of this project is to test whether extracellular matrix (ECM) mechanical cues can be used to stimulate oligodendrocytes to proliferate quicker and to differentiate into myelinating cells in greater amounts. This was to combat symptoms present with Multiple Sclerosis (MS), a demyelinating disease in which the immune system attacks the myelin sheath. It was hypothesized that the softest gel of the three would have the most significant increase in oligodendrocyte proliferation and differentiation. To culture these cells, a 2D cell coating hydrogel system was implemented, with ratios of 1:1 (the control), 1:3, and 3:1 of gel to dilution solution. To test proliferation both manual cell counts and an MTT assay were performed. The results demonstrated that there was a significant difference in proliferation rates between the control (gel with a ratio of 1:1) and the variable (gels with a ratio of 1:3 and 3:1) samples, with the variable gels having a higher proliferation rate than the control. However, there was not a significant difference between the two variable samples. To test the impact of the differing gels on differentiation of oligodendrocytes into myelinating cells, expression of the PLP and CNPase proteins, which are known to be present during the first couple days of differentiation, was assessed using immunocytochemistry. The results displayed that the cells seeded on the softer gel (1:3 ratio) had higher levels of PLP and CNPase proteins, although to a lesser extent for the latter protein, compared to the cells seeded on the control and other variable gels. Overall, the results showed that manipulating the stiffness of the ECM increased both aspects, and that the softer gels had the greatest impact on differentiation, but not proliferation.