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. 2010 Nov 15;107(48):20828–20833. doi: 10.1073/pnas.1008301107

Fig. 1.

Fig. 1.

miR-200b/429, ZEB1, and ZEB2, and contraction-associated genes are coordinately regulated during late gestation. Mature miR-200b and miR-429 are significantly up-regulated (A) and ZEB1 and ZEB2 mRNA are significantly down-regulated (B) across late gestation in the mouse myometrium, beginning at 17.5 dpc. mmu, Mus musculus. (C) ZEB1 protein in nuclear extracts of murine myometrium mirrors the decline in ZEB1 mRNA. β-actin was used as a loading control. The immunoblot shown is representative of three independent gestational series of mice. Densitometry analysis of all three series revealed a significant decrease in ZEB1 protein between 15.5 dpc and labor (Student’s t test, P < 0.05; n = 3 mice per group). (D) Contraction-associated genes CXN-43 and OXTR were significantly up-regulated in the same samples as in A and B, beginning at 18.5 dpc. Expression of each miRNA/mRNA was determined by qRT-PCR, normalized to GAPDH/U6, and expressed as the fold increase over 15.5 dpc. Mean ± SEM values are shown. For one-way ANOVA, miR-200b: F(4,20) = 14.82, P < 0.0001; miR-429: F(4,20) = 17.49, P < 0.0001; ZEB1: F(4,20) = 12.71, P < 0.0001; ZEB2: F(4,20) = 5.37, P = 0.004; CXN-43: F(4,20) = 12.93, P < 0.0001; OXTR: F(4,20) = 11.03, P < 0.0001. (Multiple comparison test compared with 15.5 dpc: *P < 0.05, **P < 0.01; n = 10 mice each for 15.5 dpc and laboring groups, 5 mice each for 16.5–18.5 dpc.) miR-200b/429 were significantly up-regulated (E) and ZEB1 and ZEB2 mRNA were significantly down-regulated (F) in laboring myometrium as compared with nonlaboring controls. hsa, Homo sapiens. (G) ZEB1 protein expression was decreased in nuclear extracts of laboring myometrium. β-actin was used as a loading control. The immunoblot shown is representative of three replicate experiments. Densitometry analysis of all blots comparing laboring myometrium with nonlaboring controls revealed a significant decrease in ZEB1 protein in labor (Student’s t test, P < 0.05; n = 9 per group). (H) CXN-43 and OXTR in the same samples as in E and F were significantly up-regulated in laboring myometrium. Expression of each miRNA/mRNA was determined by qRT-PCR, normalized to U6/h36B4, and expressed as the fold increase over nonlaboring controls. Mean ± SEM values are shown (Student’s t test, *P < 0.05; n = 14 for laboring myometrium, n = 23 for nonlaboring controls).