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. Author manuscript; available in PMC: 2021 Mar 1.
Published in final edited form as: Glia. 2020 Oct 24;69(3):792–811. doi: 10.1002/glia.23930

FIGURE 5.

FIGURE 5

The effects of guanidine compounds on oligodendrocyte precursor cell (OPC) proliferation. (a) Phase contrast images of Oli-neu cells treated for 2 days with dimethyl sulfoxide (DMSO) (control) or 50 μM guanidine compounds 002, 005, 006, 008, or 012. Scale, 100 μm. (b) Flow cytometry of Oli-neu cells treated with DMSO (control) or 50 μM compounds 002 or 006 for 16 hr, stained with Annexin V (horizontal axis) and propidium iodide (vertical axis). (c,d) Quantification of EdU incorporation in OPCs (c) and primary astrocytes (d) after treatment with 50 μM compounds for 2 days. The graphs represent the percentage of EdU+ cells among all DAPI+ cells. * p < .05, *** p < .001. Dunnett’s multiple comparisons test. (e) Proliferation of NIH 3T3 cells measured by Cell Titer-Glo assay in 0, 50, and 250 ng/ml hPDGF AA in the presence of vehicle only or 10 μM 002 or 006. ** p = .0100, *** p = .0005. Compound 006 at 10 μM did not significantly reduce proliferation compared to DMSO alone (p = 0.1478 in 50 ng/ml PDGF AA and 0.0711 in 250 ng/ml PDGF AA). Tukey’s multiple comparisons test. (f,g) Dose–response effects of compounds 002 and 006 on OPC and astrocyte proliferation measured by Cell Titer-Glo assay. **** p < .0001 in 002-treated cells compared with DMSO only in the presence of PDGF AA. #### p < .0001 in 006-treated cells compared with DMSO only in the presence of PDGF AA. Sidak’s multiple comparisons test. (h) Detection of EdU in green fluorescent protein (GFP)+ OPCs in the white matter of slice cultures from the forebrain of P8 NG2cre:Z/EG double transgenic mice. Large panels indicate double labeling for GFP and EdU. Smaller panels are single channel images. Scale 50 μm. Arrowheads indicate EdU+ GFP+ cells. Top left: no PDGF AA. Other panels were incubated with 50 ng/ml human PDGF AA in the absence (control) or presence of 10 μM compound 002 or 006. (i) Quantification of EdU incorporation into OPCs in slice cultures treated with 002 or 006 in the absence or presence of 50 ng/ml human PDGF AA. **** p < .0001. Tukey’s multiple comparisons test