FIG. 3.

Isoflurane represses the self-renewal of mES cells by upregulating the level of miR-9. (A) Level of miRNA-9 detected by qRT-PCR in mES cells treated with isoflurane for 6 h. Ctrl means mES cells with no isoflurane. Data shown are mean±SD (n=3). **P<0.01. (B) Level of miRNA-9 in mES cells transfected with pre-miR-9 or miR-9 inhibitor and control groups. Ctrl means mES cells transfected with miRNA control or miRNA inhibitor control. miR-9 means mES cells transfected with pre-miR-9. Data shown are mean±SD (n=3). *P<0.05, **P<0.01. (C) Representative pictures of AP detection staining of mES cells transfected with pre-miR-9 or miR-9 inhibitor and control groups. (D) Stemness markers, Oct4, Sox2, Nanog, and Lin28, in mES cells transfected by pre-miR-9 and miRNA control groups detected by qRT-PCR. Data shown are mean±SD (n=3). **P<0.01. (E) Stemness markers in miR-9 inhibitor and miRNA inhibitor control groups detected by qRT-PCR. Data shown are mean±SD (n=3). *P<0.05. (F) AP detection staining of mES cells treated with isoflurane or isoflurane plus miR-9 inhibitor compared with control mES cells. Ctrl means the control group treated with no isoflurane and miRNA control. (G) Detection of stemness markers, Oct4, Sox2, Nanog, and Lin28, in mES cells by qRT-PCR in (F). Data shown are mean±SD (n=3). **P<0.01. (H) Immunostaining of stemness markers, Nanog, Sox2, and SSEA1, of mES cells in (F). The scale bar represents 100 μm. miRNA, microRNA. Color images available online at www.liebertpub.com/scd