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. Author manuscript; available in PMC: 2012 Feb 4.

Published in final edited form as: Circ Res. 2010 Dec 23;108(3):305–313. doi: 10.1161/CIRCRESAHA.110.228437

(A) After 24 hours 100 nM Ang II treatment, NRCMs were harvested for immunoblot analyses of total lysates with anti-cyclin D2 antibody. (B) Mice were subjected to continuous infusion of either PBS (control) or a subpressor dose (200 ng/kg/min) of Ang II by osmotic pump for two weeks. Heart homogenates were subjected to immunoblot analyses with anti-cyclin D2 and anti-α-tubulin antibodies. (C) Twenty-four hours after transduction of Ad-sh-sc or Ad-short hairpin-cyclin D2, NRCMs were treated with or without 100 nM Ang II for 48 hours. Cells were stained with anti-α-actinin antibody (red) and DAPI (blue), and relative cell surface area (cell size) was measured. N>3. Values are mean±SEM. P<0.05. (D) Twelve to 24 hours after transduction of Ad-lacZ or Ad-Trx1, NRCMs were treated with or without 100 nM Ang II for 24 hours. Cells were harvested for immunoblot analyses using anti-cyclin D2, anti-Trx1 and anti-α-tubulin antibodies and the results were quantified by densitometry. N>3. Values are mean±SEM. P<0.05. (E) Tg-Trx1 and NTg mice were subjected to continuous infusion of either PBS or a subpressor dose (200 ng/kg/min) of Ang II by osmotic pumps for two weeks. Heart homogenates were subjected to immunoblot analyses with anti-cyclin D2 and anti-Trx1 antibodies and the results were quantified by densitometry. (F, G) Twenty-four hours after transduction of Ad-sh-sc or Ad-miR-98, NRCMs were treated with or without 100 nM Ang II for 24–48 hours. Cells were harvested for immunoblot analyses using anti-cyclin D2 antibody and the results were quantified by densitometry. (H) Ad-miR-sc or Ad-miR-98 was injected into C57 mice hearts with or without continuous infusion of Ang II (200 ng/kg/min) for two weeks. Heart homogenates were subjected to immunoblot analyses with anti-cyclin D2 antibody. (I) Cartoon shows the cyclin D2 3’UTR with miR-98 binding sites and luciferase constructs harboring either wild type or mutated cyclin D2 3’UTR. NRCMs were co-transfected with one of these two luciferase constructs with or without miR-98 overexpression for 72 hours. Cells were harvested for luciferase assay. N>3. Values are mean±SEM. P<0.05.

(A) After 24 hours 100 nM Ang II treatment, NRCMs were harvested for immunoblot analyses of total lysates with anti-cyclin D2 antibody. (B) Mice were subjected to continuous infusion of either PBS (control) or a subpressor dose (200 ng/kg/min) of Ang II by osmotic pump for two weeks. Heart homogenates were subjected to immunoblot analyses with anti-cyclin D2 and anti-α-tubulin antibodies. (C) Twenty-four hours after transduction of Ad-sh-sc or Ad-short hairpin-cyclin D2, NRCMs were treated with or without 100 nM Ang II for 48 hours. Cells were stained with anti-α-actinin antibody (red) and DAPI (blue), and relative cell surface area (cell size) was measured. N>3. Values are mean±SEM. P<0.05. (D) Twelve to 24 hours after transduction of Ad-lacZ or Ad-Trx1, NRCMs were treated with or without 100 nM Ang II for 24 hours. Cells were harvested for immunoblot analyses using anti-cyclin D2, anti-Trx1 and anti-α-tubulin antibodies and the results were quantified by densitometry. N>3. Values are mean±SEM. P<0.05. (E) Tg-Trx1 and NTg mice were subjected to continuous infusion of either PBS or a subpressor dose (200 ng/kg/min) of Ang II by osmotic pumps for two weeks. Heart homogenates were subjected to immunoblot analyses with anti-cyclin D2 and anti-Trx1 antibodies and the results were quantified by densitometry. (F, G) Twenty-four hours after transduction of Ad-sh-sc or Ad-miR-98, NRCMs were treated with or without 100 nM Ang II for 24–48 hours. Cells were harvested for immunoblot analyses using anti-cyclin D2 antibody and the results were quantified by densitometry. (H) Ad-miR-sc or Ad-miR-98 was injected into C57 mice hearts with or without continuous infusion of Ang II (200 ng/kg/min) for two weeks. Heart homogenates were subjected to immunoblot analyses with anti-cyclin D2 antibody. (I) Cartoon shows the cyclin D2 3’UTR with miR-98 binding sites and luciferase constructs harboring either wild type or mutated cyclin D2 3’UTR. NRCMs were co-transfected with one of these two luciferase constructs with or without miR-98 overexpression for 72 hours. Cells were harvested for luciferase assay. N>3. Values are mean±SEM. P<0.05.