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Significance of apoptosis in ischaemia and reoxygenation of the human myocardium
Critical Care volume 6, Article number: 4 (2002)
Apoptosis is triggered by a number of intrinsic and extrinsic factors but its importance in ischaemia-reoxygenation in the human heart is unclear. We quantified apoptosis and necrosis in an in vitro model of human atrial myocardium following various periods of simulated ischaemia (SI) and reoxygenation (R).
Experiments (n = 8 per group) were performed on sections of right atrium subjected to periods of SI (0, 30, 90 and 180 min) followed by R (2, 8 and 24 hours). Cell damage was measured following release of myocyte-specific creatine kinase (CK-MB) and cell viability measured using the vital stain MTT. Cell apoptosis and necrosis were visualised in tissue sections with FITC (TUNEL) and propidium iodide, respectively. Quantification was by confocal microscopy and NIH-image software. Caspase-3-like activity was quantified by fluorometric assay.
CK-MB release and necrosis increased whereas MTT values decreased over the period of SI in a dose-dependent manner. Apoptosis increased (32%) after 90 min SI with 2 hours R and peaked (56%) after 90 min SI with 8 hours R. Apoptosis declined following 180 min R. The smallest induction of apoptosis occurred after 24 hours R. With increasing SI in the 2 hour protocol, there was a progressive rise in the ratio of caspase-3 to total caspase activity, which increased to a maximum of fourfold after 180 min SI and 2 hours R. Caspase-3 levels were similar to fresh tissue after 24 hours.
In this model of SI/R injury of human myocardium, the degree of apoptosis and necrosis varies with the duration of ischaemic insult and the reoxygenation time. After certain periods of ischaemia, apoptosis may be the predominant pathway to cell death.
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Vohra, H., Fowler, A. & Galiñanes, M. Significance of apoptosis in ischaemia and reoxygenation of the human myocardium. Crit Care 6, 4 (2002). https://doi.org/10.1186/cc1808
- Creatine Kinase
- Propidium Iodide
- Fluorometric Assay