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. 2020 Mar 13;8:153. doi: 10.3389/fcell.2020.00153

FIGURE 6.

FIGURE 6

FOXD3 regulates RND3 expression, inhibits proliferation and migration, and promotes apoptosis in HTR-8 cells in vitro. (A) CCK-8 assay of FOXD3 knockdown and FOXD3-overexpressing HTR8 cells and their respective control groups. (B,D) Transwell migration assay of FOXD3 knockdown without or with RND3 overexpression and FOXD3-overexpressing without or with siRND3 HTR-8 cells and their respective control groups. Original magnification: × 100. Scale bar = 100 μm. (C) Flow cytometry analysis of the apoptosis rate of siNC, siFOXD3, siFOXD3 + RND3 and Vector, FOXD3, FOXD3 + siRND3 group. Histogram of Annexin V APC+/7-AAD– represents early apoptotic cells. Histogram of Annexin VAPC+/7-AAD+ cells represents late apoptotic cells. The sum of the former two categories equals the total number of apoptotic cells. (E) The mRNA expression level of RND3 and FOXD3 in the villous tissues of patients with RM and HC was determined using qRT-PCR (n = 24). RND3 mRNA expression was correlated with FOXD3 mRNA expression in villous tissues. Correlation analysis was performed using Spearman’s rank correlation test. (F–I) Western blot analysis of FOXD3, RND3, MMP2, Cyclin D1, and BAX expression in HTR-8 cells at 48 h after transfection with siNC and siFOXD3 or control vector and FOXD3 overexpression vector. AT 24 h after transfection with siFOXD3 or FOXD3 overexpression vector, HTR-8 cells were transfected RND3 overexpression vector or siRND3 again. Data represent the means ± SD of three independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 vs. siNC, Vector, siFOXD3 or FOXD3 group.