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. 2020 Jun 25;15(1):256–273. doi: 10.1016/j.stemcr.2020.06.001

Figure 2.

Figure 2

APP and PS1 Variant HSs Exhibit AD-Related Pathology

(A and B) Characterization of amyloid-β (Aβ) accumulation (intracellular Aβ, a) and secretion (extracellular Aβ, b) in APP variant, PS1 variant, and gender-matched control HSs at DIV 100. The ratio of Aβ42/Aβ40 in HS lysates (A) and the ratio of Aβ42/Aβ40 secreted from HSs into the medium (B) were measured at day 4 after the last medium change. For quantitation, data were normalized to the total protein. Results are presented as mean ± SEM. n = 3 independent differentiations per genotype. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001. Statistical analysis by two-tailed t test. See also Figure S2.

(C and D) Characterization of phosphorylation of tau protein in APP variant, PS1 variant, and gender-matched control HSs. Western blotting analysis of phosphorylation of tau protein in HSs at DIV 100 with actin blot included as a loading control (C). The blots were quantified densitometrically and for quantitation of tau phosphorylation level, data were normalized by the level of actin (D). n = 3 independent differentiations per genotype. ∗p < 0.05. Statistical analysis by two-tailed t test.

(E–G) Characterization of synaptic proteins in APP variant, PS1 variant, and gender-matched control HSs. Confocal images showing representative examples of immunostaining for synaptic markers synaptophysin and drebrin in APP variant, PS1 variant, and gender-matched control HSs at DIV 100 and 3D surface reconstruction of dendritic segment with putative dendritic spines as delineated by drebrin labeling in close proximity to synaptophysin puncta in control HSs (E). Western blotting analysis of synaptic markers synaptophysin, drebrin, and PSD-95 in HSs at DIV 100 with MAP2d blot included as a loading control (F). The blots were quantified densitometrically and for quantitation of synaptic protein levels, data were normalized by the level of MAP2d (G). Results are presented as mean ± SEM. n = 3 independent differentiations per genotype. ∗p < 0.05; ∗∗p < 0.01. Statistical analysis by two-tailed t test. Scale bars, 100 μm (HS confocal images) and 5 μm (3D reconstruction images).