Figure 6.

STAT3 directly mediated hypoxia-induced miR-211 expression. MiR-211 expression levels were analyzed by polymerase chain reaction (PCR) for both normoxia-treated MSCs (N-MSCs, in black bars) and HP-treated MSCs (HP-MSCs, in white bars) when they were infected with STAT3-over (over, CV as control, shown in A) or STAT3-shRNA (shRNA, NC as control, shown in B), respectively (three separate experiments). The promoter region of miR-211 was divided into seven consecutive segments (C), and ChIP with STAT3 specific antibody was carried out and the pull-down precipitates were analyzed with quantitative PCR, PCR products were run in the ethidium bromide agarose gel, and the image was captured with the grey-scale inverted for better illustration (D), the relative fold of enrichment was normalized to the input (E). Luciferase assay showed that STAT3 can positively regulate the transcription activities specific for the different promoter regions 3, 4, and 5 of miR-211 (F). The experiments were repeated for three times. **Denotes p less than .01 vs. groups that were not infected with vectors containing both STAT3 and specific promoter regions of 3–5 of miR-211. Abbreviations: HP, hypoxic preconditioning; MSCs, mesenchymal stem cells; shRNA, short hairpin ribonucleic acid.