Figure 2
Western Blot Analysis of Angiotensin-Converting Enzyme 2 (ACE2) Expression in Human Cardiomyocytes under Normoxia and Hypoxia. A , human cardiomyocytes were stably transfected with wild type (WT) IRS-1 or Arg972 IRS-1, or stably transduced with IRS-1-shRNA. Then the cells were cultured under normoxia or hypoxia for 24 hours in the presence or absence of LY294002 (LY, 50 μM). Lysates were subject to western blot analysis for ACE2 expression (lanes 1–5, normoxia; lanes 6–10, hypoxia). Lysate from human cardiomyocytes stably transfected with empty pcDNA3.1 plasmids was used as a control (VC). Lanes 1 and 6, VC; lanes 2 and 7, cardiomyocytes stably transfected with WT-IRS-1; lanes 3 and 8, cardiomyocytes stably transfected with Arg972-IRS-1; lanes 4 and 9, cardiomyocytes stably transduced with IRS-1-shRNA; lanes 5 and 10, VC cells treated with LY (50 μM) for 24 hours. β-Actin blotting was used as a loading control. B , ACE2 and β-actin blots were measured by densitometry. The density of ACE2 blots was normalized against that of β-actin to obtain a relative blot density, which was expressed as fold changes to that of the VC cells under normoxia (designated as 1). *p<0.05 compared with any group in normoxia; ap<0.05 compared with VC in normoxia; bp<0.05 compared with WT-IRS-1 in normoxia; cp<0.05 compared with Arg972-IRS-1 in normoxia; dp<0.05 compared with IRS-1-shRNA in normoxia; ep<0.05 compared with VC+LY in normoxia; fp<0.05 compared with VC in hypoxia; gp<0.05 compared with WT-IRS-1 in hypoxia; hp<0.05 compared with Arg972-IRS-1 in hypoxia.