Journal: Disease Models & Mechanisms

Article Title: Identification of early indicators of altered metabolism in normal development using a rodent model system

doi: 10.1242/dmm.031815

Figure Lengend Snippet: Regulation of the cell cycle is different in ABW and LBW pups. (A) Western blot analyses of the skeletal muscle of members of the Dnmt1 repressor complex, E2f1, Rb and Hdac1, in ABW and LBW pups at day 1. Gapdh was used as a loading control. (B) Co-IP analysis of the Dnmt1 repressor complex using anti-Dnmt1 antibody, and western blot analysis of the members of the complex E2f1, Rb, Hdac1, in ABW and LBW pups at day 1. Co-IP using IgG was used as a control; 5% of the total extract was used as input. (C) Western blot analysis of skeletal muscle of 1-day-old ABW and LBW pups for phospho Rb (ser 608, ser 780 and ser 795). Gapdh was used as a loading control. (D) Western blot analysis of cyclin D1 and cyclin-dependent kinases Cdk4 and Cdk6, and the G1 phase cell cycle inhibitors P16 and P21, using skeletal muscle of ABW and LBW pups at day 1. Gapdh was used as a loading control.

Article Snippet: Antibodies used were as follows: anti-Dnmt1 (ab92453), anti-Cdk4 (ab131197), anti-Hdac1 (ab7028), anti-cyclin D1 (134175), anti-Glut4 (ab33780), anti-ph-Akt (ab81283), anti-Dnmt3a (ab13888), anti-Dnmt3b ab13604), anti-Gcg (ab92517), anti-ph-IRS1 (ab1194), anti-Akt1/2/3 (ab126811), anti-Pparγ (ab45036) and anti-Rb (ab6075), purchased from Abcam, USA; anti-Pparγ (sc7273) and anti-Rxrα (sc 553), purchased from Santa Cruz Biotechnology, USA; anti-ph-Rb ser780 (RA2133264) and anti-ph-Rb ser 795 (QL2133271), from Thermo Fisher Scientific; anti-Gapdh (2118L), anti-ph-Rb ser 608 (2181s), anti-ph-Akt (4060L), anti-Irs-1 (3407S), anti-Hdac1 (5356s) and anti-P21 (2947s), from Cell Signaling Technology, USA; anti-E2f1 (2651877), from Millipore, USA; anti-AdipoQ (A6354), from Sigma-Aldrich; anti-P16 (10883-1-AP), from Protein Tech, USA; and anti-insulin, from Dako Cytomation, Copenhagen, Denmark.

Techniques: Western Blot, Co-Immunoprecipitation Assay

Journal: Disease Models & Mechanisms

Article Title: Identification of early indicators of altered metabolism in normal development using a rodent model system

doi: 10.1242/dmm.031815

Figure Lengend Snippet: Possible mechanism by which embryonic growth is coordinated and correlated with the maternal environment. Elevated Dnmt1 expression might be increasing methylation in the promoters of genes, resulting in suppression of the transcription of genes required for normal growth. In addition, the formation of the Dnmt1 repressive complex with Rb, E2f1 and Hdac1 might slow down cell cycle progression. These processes, probably working together, could be responsible for the growth reduction observed in the birth weight of LBW pups.

Article Snippet: Antibodies used were as follows: anti-Dnmt1 (ab92453), anti-Cdk4 (ab131197), anti-Hdac1 (ab7028), anti-cyclin D1 (134175), anti-Glut4 (ab33780), anti-ph-Akt (ab81283), anti-Dnmt3a (ab13888), anti-Dnmt3b ab13604), anti-Gcg (ab92517), anti-ph-IRS1 (ab1194), anti-Akt1/2/3 (ab126811), anti-Pparγ (ab45036) and anti-Rb (ab6075), purchased from Abcam, USA; anti-Pparγ (sc7273) and anti-Rxrα (sc 553), purchased from Santa Cruz Biotechnology, USA; anti-ph-Rb ser780 (RA2133264) and anti-ph-Rb ser 795 (QL2133271), from Thermo Fisher Scientific; anti-Gapdh (2118L), anti-ph-Rb ser 608 (2181s), anti-ph-Akt (4060L), anti-Irs-1 (3407S), anti-Hdac1 (5356s) and anti-P21 (2947s), from Cell Signaling Technology, USA; anti-E2f1 (2651877), from Millipore, USA; anti-AdipoQ (A6354), from Sigma-Aldrich; anti-P16 (10883-1-AP), from Protein Tech, USA; and anti-insulin, from Dako Cytomation, Copenhagen, Denmark.

Techniques: Expressing, Methylation