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. 2017 Feb;214(2):529–545. doi: 10.1084/jem.20150906

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© 2017 Lv et al.

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Figure 1. — 5-HT promotes the development of HSPCs in vitro and ex vivo. (A) Flow chart of using different chemicals to detect the requirement of 5-HT for HSPCs production by an AGM explants culture system. In brief, E10.0–E10.5 embryo (31–40 sp) AGMs were dissected from the wild-type embryos and cultured on Durapore filters at the air–liquid interface in the presence of 5-HT, fluoxetine, or MT diluted in M5300 long-term culture medium. After 36–48 h, AGMs were used to quantify the mRNA level of hematopoietic related genes (runx1 and gfi1). The remaining AGMs were dissociated with collagenase and used for CFU-C or CFU-S₁₁ assay. (B) CFU-C assay of AGMs treated with 5-HT at 1 mM. 1 ee was used. (C) qPCR analysis showed the expression of runx1 and gfi1 in the AGM treated with 5-HT. (D) CFU-S₁₁ assay with AGMs treated with fluoxetine at 10 µM. The number of spleen colonies in irradiated adult recipients were counted. 1 ee was used. (E) qPCR analysis showed the expression of runx1 and gfi1 in the AGM treated with fluoxetine. (F) CFU-C assay of AGMs treated with MT at 50 µM. 1 ee was used. (G) qPCR analysis showed the expression of runx1 and gfi1 in the AGM treated with MT. The results are presented as mean ± SE. Student’s t test: *, P < 0.05; **, P < 0.01; ***, P < 0.001. n = 3. BFU-E, burst forming unit–erythroid; CFU-GM, CFU-granulomonocyte; Ctrl, control.