Figure 7.

WmOPCs are more sensitive to TNFα- and IFNγ-mediated inhibition of proliferation than gmOPCs. Oligodendrocyte progenitor cells (OPCs) isolated from the cortex (gmOPCs) and non-cortex (wmOPCs) of neonatal rat forebrains were left untreated or treated with 10 ng/ml TNFα, 500 U/ml IFNγ, or a combination of TNFα and IFNγ for 48 hours in the presence of PDGF-AA and FGF-2. (a,c) OPC proliferation was determined by immunocytochemistry for the proliferation marker ki67. Representative images are shown in (a); quantitative analysis of the number of ki67-positive of total DAPI-stained cells in (c) (n = 4, at least 150 cells analysed per independent experiment). (b) OPC migration towards a 10 ng/ml PDGF-AA gradient (4 hours) was determined using a transwell assay (n = 5). Grey bars represent gmOPCs white bars represent wmOPCs (b,c). Note that both exposure to TNFα and IFNγ decreased wmOPC proliferation, while IFNγ, but not TNFα, decreased gmOPC proliferation. (d) mRNA expression levels of Tnfrs1a, Tnfrs1b, Ifngr1 and Ifngr2. Hmbs was used as reference gene; the reference gene Eef1a1 showed similar results (data not shown). Note that Ifngr1 expression levels are elevated in wmOPCs compared to gmOPCs. Bars represent mean relative to their respective untreated control, which was set at 1 for each independent experiment (horizontal line). Error bars show the standard error of the mean. Statistical analyses were performed using a one-sample t-test (*p < 0.05) to test for differences between treatments and their respective control and a one-way ANOVA with a Šidák post-test was used to test whether the response to TNFα, IFNγ and TNFα and IFNγ combined differed between gmOPCs and wmOPCs (not significant). Scale bar is 50 µm.