Cardiovascular Pharmacology

Hypoxia reoxygenation induces premature senescence in neonatal SD rat cardiomyocytes

Abstract

Aim:

To investigate whether hypoxia reoxygenation induces premature senescence in neonatal Sprague-Dawley (SD) rat cardiomyocytes.

Methods:

Cardiomyocytes were isolated from neonatal SD rat heart and identified by immunohistochemistry. The control cultures were incubated at 37 °C in a humidified atmosphere of 5% CO2 and 95% air. The hypoxic cultures were incubated in a modular incubator chamber filled with 1% O2, 5% CO2, and balance N2 for 6 h. The reoxygenated cultures were subjected to 1% O2 and 5% CO2 for 6 h, then 21% oxygen for 4, 8, 12, 24, and 48 h, respectively. Cell proliferation was determined using bromodeoxyuridine labeling. The ultrastructure of cardiomyocytes was observed by using an electron microscope. β-Galactosidase activity was determined by using a senescence β-galactosidase Staining Kit. p16INK4a and telomerase reverse transcriptase (TERT) mRN A levels were measured by real time quantitative PCR. TERT protein expression was determined by immunohistochemistry. Telomerase activities were assayed by using the Telo TAGGG Telomerase PCR ELISAplus kit.

Results:

The initial cultures consisted of pure cardiomyocytes identified by immunohistochemistry. The proportion of BrdU positive cells was reduced significantly in the hypoxia reoxygenation-treated group (P<0.01). Under the condition of hypoxia reoxygenation, mitochondrial dehydration appeared; p16INK4a and TERT mRNA levels, β-galactosidase activity, TERT protein expression and telomerase activities were all significantly increased (P<0.01 or P<0.05).

Conclusion:

These data indicate that premature senescence could be induced in neonatal SD rat cardiomyocytes exposed to hypoxia reoxygenation. Although TERT significantly increased, it could not block senescence.

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Correspondence to Ke-jiang Cao.

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Project supported by National Natural Science Foundation of China (No 30671073).

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Zhang, F., Chen, M., Shan, Q. et al. Hypoxia reoxygenation induces premature senescence in neonatal SD rat cardiomyocytes. Acta Pharmacol Sin 28, 44–51 (2007). https://doi.org/10.1111/j.1745-7254.2007.00488.x

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Keywords

  • cardiomyocytes
  • hypoxia reoxygenation
  • p16INK4a
  • telomerase reverse transcriptase
  • β-galactosidase

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