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Induced Hypoxia and Screening for Cardioprotection

hypoxia

Illustration of coronary artery blockage-induced hypoxia in cardiac muscle tissue

Myocardial ischemia is a pathological condition characterized by a reduced oxygen supply that can lead to cellular apoptosis/necrosis, arrhythmia, organ injury, and even death. Ironically, returning hypoxic myocardium to normoxic levels exacerbates the pathology, collectively known as myocardial reperfusion injury. A primary cause of myocardial ischemia is coronary artery block, and while conventional therapies are in place to minimize cellular damage caused by the hypoxic condition, morbidity and mortality remain high and additional therapeutic approaches for cardioprotection are in demand1.

iCell® Cardiomyocytes provide an ideal preparation for investigating hypoxia and discovering novel potential therapeutic interventions. These human cardiomyocytes are amenable to hypoxia induction (Figure 1), quantification of cellular endpoints (Figure 2), and screening for cardioprotective molecules and/or conditions (Figure 3).

Additionally, iCell Cardiomyocytes can be induced to exhibit hypertrophic and diabetic cardiomyopathies, thus making simultaneous investigation of common ischemia co-morbidities a possibility.

Contact CDI’s Technical Support for additional information on implementing these protocols and more into your research.

hypoxia_workflow

Figure 1: Inducing Hypoxia in iCell Cardiomyocytes and Monitoring Cellular Effects Is Straightforward iCell Cardiomyocytes can be driven to a hypoxic condition through culturing in low oxygen (1% for 48 hours). A variety of endpoints such as ROS generation, cellular viability, apoptosis, etc. can be monitored easily and used as screening endpoints for cardioprotective effects.

hypoxic_responses

Figure 2: Hypoxic Responses in iCell Cardiomyocytes Can Be Quantified Hypoxia was induced by incubating iCell Cardiomyocytes in 1% O2 for 48 hours, and apoptosis was monitored via the TUNEL assay. (A) Hypoxic conditions induced significant apoptosis. Nuclear morphology and TUNEL staining under (B) normoxia and (C – E) hypoxia showing nuclear fragmentation (C) and condensation (D – E). Results shown are mean ±SEM. * p<0.05.

 

Figure 3: iCell Cardiomyocytes Enable Screens for Cardioprotective Compounds Inhibitors and growth factors were added to the cells immediately prior to induction of hypoxia (1% O2), and caspase-3/7 activity was measured 48 hours later. iCell Cardiomyocytes treated with growth factors and two of the three caspase inhibitors tested displayed significantly lower caspase 3/7 activity suggesting a cardioprotective effect against hypoxia while necrosis inhibitors had minimal effect. Results: mean ± SEM, * p < 0.05; ** p < 0.01, *** p < 0.001

 

  1. Eltzchig HK, Bonney SK, and Eckle T (2013) Attenuating Myocardial Ischemia by Targeting A2B Adenosine Receptors, Trends Mol Med 19(6):345-54.
  2. Carlson C, Koonce C, et al. (2013) Phenotypic Screening with Human iPS Cell-derived Cardiomyocytes: HTS-compatible Assays for Interrogating Cardiac Hypertrophy, J Biomol Screen 18(10):1203-11.
  3. Drawnel FM, Boccardo S, et al. (2014) Disease Modeling and Phenotypic Drug Screening for Diabetic Cardiomyopathy Using Human Induced Pluripotent Stem Cells, Cell Reports 9(3):810-820.