Impact of IFN-γ on TPO- and eltrombopag-induced c-MPL internalization in human HSPCs. (A) Time course of c-MPL internalization upon exposure of human CD34⁺ cells to TPO, with or without IFN-γ, or to IFN-γ alone. P values shown were calculated by comparing TPO with TPO + IFN-γ (upper panel). Representative images of CD34⁺ cells 30 minutes after exposure to TPO, TPO + IFN-γ, or IFN-γ alone (lower panel). (B) Time course of c-MPL internalization upon exposure of human CD34⁺ cells to eltrombopag with or without IFN-γ, or to IFN-γ alone. P values shown were calculated by comparing eltrombopag with eltrombopag + IFN-γ (upper panel). Representative images of CD34⁺ cells 30 minutes after exposure to eltrombopag, eltrombopag + IFN-γ, or IFN-γ alone (lower panel). (C) Quantification of the fold decrease in internalization scores caused by exposure to IFN-γ for CD34⁺ cells cultured with TPO or eltrombopag as a function of time. Internalization scores were calculated as described. Internalization data were obtained using multispectral imaging flow cytometry. Data are mean ± SEM. *P < .05, ***P < .001, ****P < .0001, repeated measures 2-way ANOVA with Sidak’s multiple-comparison test or multiple 2-tailed unpaired Student t tests (n = 3). BF, brightfield; MPL, c-MPL receptor.