Fig. 2

MiR-142a-3p is essential for HSPC emergence. (a) 142^T−/− embryos showed decreased expression of HSPC marker runx1 at 26 and 36 hpf by WISH. (b) 142^T−/− embryos displayed decreased expression of HSC marker runx1 at 26 and 36 hpf using qPCR. The expression of runx1 was normalized to β-actin (mean±s.d., n=3, **P<0.05). (c) Runx1 was decreased in 142^T−/− embryos at 26 and 36 hpf by western blot analysis (left panel) and the quantification (right panel). The protein level of Runx1 was normalized to β-Actin. (d) The number of EGFP-labeled HSPCs (red arrowheads) in Tg (cmyb:EGFP) embryos was decreased in the AGM region in 142^T−/− embryos at 36 hpf (mean±s.d., n=5, *P<0.05). White arrowheads indicate cmyb-positive cells in the pronephric duct. cmyb-labeled pronephric duct was counted as an internal control. (e) 142^T−/− embryos displayed reduction of HSPC marker cmyb from 2 to 3.5 dpf by WISH. (f) Differentiated hematopoietic lineages were decreased in 142^T−/− embryos including erythrocytes (gata1), neutrophils (lyz) and lymphocytes (ikaros and rag1) at 4 dpf by WISH. Red arrowheads mark hematopoietic cells in the caudal hematopoietic tissue (CHT), whereas red circles denote the thymus. (g) Hematoxylin and eosin (HE) staining of the kidney showed that decrease in cell population and the volume of the head kidney greatly in 142^−/− adult fish of 12 weeks.