Fig. 5

C3a-activated astrocytes promote MB cell proliferation through TNF-α secretion. a, b Purified MB cells were cultured in vitro in the presence of recombinant TNF-α, IL-6, or vehicle control (Ctrl) for 48 h and then immunostained for Ki67 (red). DAPI was used to counterstain the cell nuclei (a). The percentage of Ki67 + cells was quantified (b). ***p < 0.001, ns: not significant vs. Ctrl group. c, d Astrocytes were pretreated with 100 nM C3a or vehicle control for 24 h, washed thoroughly, and cultured for another 24 h for CM collection. Then, purified MB cells were cultured with the CM above together with the TNF-α receptor antagonist R-7050 (500 nM) or not for 48 h. Ki67 was immunostained (c), and the percentage of Ki67 + cells was quantified (d). ***p < 0.001, ns: not significant vs. Ctrl CM group, # p < 0.05 vs. (C3a-AS CM + vehicle) group. Images are presented, and values are expressed as the mean ± SEM from three independent experiments