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. Author manuscript; available in PMC: 2015 Jan 30.
Published in final edited form as: Cell. 2014 Jan 30;156(3):549–562. doi: 10.1016/j.cell.2013.12.025

Figure 7. Knockdown of NR4A1 inhibits progesterone-mediated permeability.

Figure 7

(A) qPCR analysis of NR4A1 following transfection of HUVECs with either non-targeting (NT) or NR4A1 siRNA. n=3. (B) Baseline HUVEC monolayer resistance following NR4A1 knockdown. (C) HUVEC monolayer resistance after transfection with NR4A1 or NT siRNA in the presence of P4. (D) HUVEC monolayer resistance following adenoviral infection with a NR4A family dominant negative (NR4A DN) and control (GFP) in the presence of P4. (E) HUVECs expressing PR (GFP, green) and transfected with either NT or NR4A1 siRNA were treated with P4 for 24h. PECAM, VE-cadherin, and β–catenin (white) were used to visualize junctions. DAPI (blue) denotes nuclei. Arrowheads indicate a reduction in junctional proteins. Arrows show expanded junctional area. Scale, 50μm. (F) Junctional proteins from GFP or PR infected HUVECs following transfection with either NT or NR4A1 siRNA followed by P4 treatment. GAPDH=loading control. (G) NR4A1 (GFP, green) localization in the vascular endothelium in vivo (PECAM, red). Scale, 25μm (H) PR (blue) and NR4A1 (GFP, green) colocalization in uterine vasculature (PECAM, red). Arrows indicate endothelial cells with NR4A1 and PR. Scale, 20μm (I) qPCR of NR4A1 from the uteri of virgin, ovarectomized (OVX) and PRECKO mice treated with or without P4. n=3. (J) Evans blue content from the uterus and intestine following hormone stimulation of control (wild-type) and NR4A1KO mice. ***p<0.001, ****p<0.001. Error bars=+/− SEM. See also Figure S7.