FIG. 4.

Effect of insulin on SLC29A1 (hENT1) promoter activity. A: Luciferase (Luc) reporter constructs containing two truncations of SLC29A1 promoter (−3198 and −1670 bp from ATG) were transfected in HUVECs from normal (□) or GDM (■) pregnancies, along with Renilla reporter plasmid, and assayed for Firefly and Renilla luciferase activity, respectively. Results depict the ratio of Firefly/Renilla luciferase activity in cells from normal or GDM pregnancies. After 36 h of transfection, cells were incubated for a further 12 h in the absence of insulin (control) or for the last 8 h of the 12-h incubation period in the presence of 1 nmol/L insulin (white [normal] and black [GDM] dashed bars, respectively). Cells were also transfected with the empty pGL3-basic vector or pGL3-control vector (SV40 pGL3) as negative or positive controls, respectively (see research design and methods). *P < 0.05 vs. normal pregnancies in the absence of insulin. †P < 0.05 vs. normal pregnancies in the absence of insulin or GDM pregnancies in the presence of insulin, transfected with the pGL3-hENT1⁻³¹⁹⁸ construct. B: Reporter construct pGL3-hENT1⁻³¹⁹⁸ of SLC29A1 promoter transfected in cells from normal (□) or GDM (■) pregnancies as in A in the absence (–) or presence (+) of l-NAME. C: Reporter construct pGL3-hENT1⁻¹⁶⁷⁰ of SLC29A1 promoter assayed as in B. In B and C, *P < 0.05 vs. all other corresponding values in normal or GDM pregnancies. †P < 0.05 vs. corresponding values in normal or GDM pregnancies in the presence of insulin. Error bars in the graphs designate the SEM.