Journal: The Journal of Clinical Endocrinology and Metabolism

Article Title: The HMGA2-IMP2 Pathway Promotes Granulosa Cell Proliferation in Polycystic Ovary Syndrome

doi: 10.1210/jc.2018-00544

Figure Lengend Snippet: CCND2 and SERBP1 expression in the HMGA2-IMP2 pathway. (A, B) IMP2 binding sites in the CCND2 and SERBP1 genes. IMP2 bound to CCND2 and SERBP1 mRNAs. (C) RIP experiments in HEK293 cells. CCND2 and SERBP1 proteins were pulled down using anti-IMP2 antibodies. IgG and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) were used as negative controls. (D) Knockdown of HMGA2 in KGN cells reduced the CCND2 and SERBP1 protein levels. (E) Knockdown of IMP2 reduced the CCND2 and SERBP1 protein levels in KGN cells. (F, G) Luciferase assays of IMP2 and downstream genes (F, CCND2; G, SERBP1) in KGN cells. In both cases, a dose-dependent increase in luciferase activity of CCND2 and SERBP1 was observed as IMP2 expression increased. *P < 0.05. **P < 0.01. Each value represents the mean of at least three independent experiments. Ctrl, control.

Article Snippet: The primary antibodies for immunoblotting included anti-HMGA2 antibody (ab184616; Abcam, Cambridge, United Kingdom), anti-IMP2 antibody (14672; Cell Signaling, Danvers, MA), anti–cyclin D2 (CCND2) antibody (3741; Cell Signaling, Danvers, MA), anti–glyceraldehyde 3-phosphate dehydrogenase antibody (60004-1-Ig; Proteintech, Wuhan, China), and anti- SERPINE1 mRNA binding protein 1 (SERBP1) antibody (10729-1-AP; Proteintech, Wuhan, China).

Techniques: Expressing, Binding Assay, Luciferase, Activity Assay

Journal: The Journal of Clinical Endocrinology and Metabolism

Article Title: The HMGA2-IMP2 Pathway Promotes Granulosa Cell Proliferation in Polycystic Ovary Syndrome

doi: 10.1210/jc.2018-00544

Figure Lengend Snippet: CCND2 and SERBP1 mRNA and protein expression in control and PCOS GCs. (A, B) CCND2 mRNA and protein expression was higher in PCOS GCs than in control cells. Data were normalized to glyceraldehyde 3-phosphate dehydrogenase (GAPDH). **P < 0.01. (C, D) SERBP1 mRNA and protein expression was higher in PCOS GCs than in control cells. Data were normalized to GAPDH. **P < 0.01. Ctrl, control.

Article Snippet: The primary antibodies for immunoblotting included anti-HMGA2 antibody (ab184616; Abcam, Cambridge, United Kingdom), anti-IMP2 antibody (14672; Cell Signaling, Danvers, MA), anti–cyclin D2 (CCND2) antibody (3741; Cell Signaling, Danvers, MA), anti–glyceraldehyde 3-phosphate dehydrogenase antibody (60004-1-Ig; Proteintech, Wuhan, China), and anti- SERPINE1 mRNA binding protein 1 (SERBP1) antibody (10729-1-AP; Proteintech, Wuhan, China).

Techniques: Expressing

Journal: PLoS ONE

Article Title: MicroRNA-497 Induces Apoptosis and Suppresses Proliferation via the Bcl-2/Bax-Caspase9-Caspase3 Pathway and Cyclin D2 Protein in HUVECs

doi: 10.1371/journal.pone.0167052

Figure Lengend Snippet: (A) Luciferase assay in HUVECs co-transfected with miR-497 or miR-497-mutant and a luciferase reporter containing the Bcl2 3’UTR. (B) Luciferase assay in HUVECs co-transfected with miR-497 or miR-497-mutant and a luciferase reporter containing the CCND2 3’UTR. (C) Bcl2 mRNA was analyzed using qRT-PCR after transfection with miR-497 or the miR-497 inhibitor. (D) CCND2 mRNA was analyzed using qRT-PCR after transfection with miR-497 or the miR-497 inhibitor. (E) The Bcl2/Bax, CCND2, and caspase-3 and -9 proteins were analyzed using western blot analysis after transfection with miR-497. (F) The Bcl2/Bax, CCND2, and caspase-3 and -9 proteins were analyzed using western blot analysis after transfection with miR-497 inhibitor. The data are presented as the mean ± SD from three separate experiments. **P<0.01 versus the control groups.

Article Snippet: The primary antibodies for BCL-2, BAX, CCND-2, caspase-3, and α-Tubulin were all purchased from Cell Signaling Technology.

Techniques: Luciferase, Transfection, Mutagenesis, Quantitative RT-PCR, Western Blot, Control