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. 2018 Feb 26;9:816. doi: 10.1038/s41467-018-03105-2

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© The Author(s) 2018

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 1 — Contribution of myocyte-specific hypoxia-inducible factor (HIF) isoforms Hif1a or Hif2a to cardioprotection. a Schematic of breeding approach to generate mice with induced myocyte-specific HIF deletions used in subsequent studies. Hif1a^loxP/loxP or Hif2a^loxP/loxP mice were crossed with Cre-recombinase expressing mice under the control of Myosin-heavy chain promoter (Myosin-Cre+); these mice express cre-recombinase under the control of a tamoxifen-inducer. Control animals (Myosin-Cre+), Hif1a^loxP/loxPMyosin-Cre+ or Hif2a^loxP/loxPMyosin-Cre+ received a daily dose of 1 mg i.p. Tamoxifen five consecutive days to induce the Cre-recombinase activity. After 7 days, animals underwent experimental protocol (60 min of in situ myocardial ischemia followed by 120 min of reperfusion). b HIF1-alpha or HIF2-alpha immunoblot analysis from homogenized myocardial tissue, harvested from male Myosin-Cre+ and Hif1a^loxP/loxPMyosin-Cre+ or Hif2a^loxP/loxPMyosin-Cre+ mice, matched in age and weight. Mice underwent a thoracotomy with no further treatment(-I) or 60 min of myocardial ischemia (+I) followed by 120 min reperfusion; β-Actin (ACTb) served as a loading control. One representative blot out of three experiments is shown. c, d Quantification by densitometry of the HIF-immunoblot results relative to ACTb. Data are expressed as mean fold change ±SD normalized to untreated myocardial tissue from Myosin-Cre+ compared by one-way ANOVA followed by Bonferroni’s multiple comparison test (n = 3 per group; c: F_5,12 = 6.74, p = 0.0033; d: F_5,12 = 21.85, p < 0.0001). e Infarct sizes ±SD in Myosin-Cre+, Hif1a^loxP/loxP Myosin-Cre+ or Hif2a^loxP/loxPMyosin-Cre+ mice, presented as the percentage to the area-at-risk after 60 min of ischemia, followed by 120 min of reperfusion (Myosin-Cre+ n = 5; Hif1a^loxP/loxPMyosin-Cre+ n = 5; Hif2a^loxP/loxPMyosin-Cre+ n = 4; per group mean ± SD; compared by one-way ANOVA followed by Bonferroni’s multiple comparison test; F_2,11 = 7.901; p = 0.0075). f Representative infarct staining from Myosin-Cre+, Hif1a^loxP/loxPMyosin-Cre+ or Hif2a^loxP/loxPMyosin-Cre+. g Troponin serum levels after 60 min ischemia, followed by 120 min of reperfusion in Myosin-Cre+, Hif1a^loxP/loxPMyosin-Cre+ or Hif2a^loxP/loxPMyosin-Cre+ (Myosin-Cre+ n = 5; Hif1a^loxP/loxPMyosin-Cre+ n = 5, and Hif2a^loxP/loxPMyosin-Cre+ n = 4 per group; presented as mean ± SD; compared by one-way ANOVA followed by Bonferroni’s multiple comparison test; F_2,11 = 19.14, p = 0.0003)