Influence of deep hypothermia on the tolerance of the isolated cardiomyocyte to ischemia-reperfusion.
The influence of deep hypothermia (4 degrees C) during a substrate-free, hypoxia-reoxygenation treatment was investigated on cardiomyocytes (CM) prepared from newborn rat heart in culture in an in vitro, substrate-free model of ischemia-reperfusion. The transmembranous potentials were recorded with standard microelectrodes. The contractions were monitored photometrically. The RNA messenger (mRNA) and protein expression for protein (HSP70) were analysed by RT-PCR (reverse transcriptase-polymerase chain reaction) and Western blotting, respectively. Simulated ischemia (SI) caused a gradual decrease and then a cessation of the spontaneous electromechanical activity. During the reoxygenation, the CM recovered normal function, provided that SI did not exceed 2.5 h. When SI duration was increased up to 4 h, reoxygenation failed to restore the spontaneous electromechanical activity. Conversely, the exposure of the CM to SI together with deep hypothermia decreased the functional alterations observed, and provided a complete electromechanical recovery after 2.5 h as well as after 4 h of SI. Deep hypothermia alone failed to induce HSP70 mRNA and protein production. On the contrary, HSP70 mRNA production increased after 2.5 and 4 h of deep hypothermia followed by 1 h of rewarming, proportionally to the duration of the cooling period. This augmentation in mRNA was associated with a rise in HSP70 protein content. In summary, it appeared that deep hypothermia exerts a strong cytoprotective action during SI only, whereas cooling CM before SI has no beneficial effect on subsequent SI. Moreover, these results suggested the persistence of a signaling system and/or transduction in deeply cooled, functionally depressed cells. Finally, CM in culture appeared to be a model of interest for studying heart graft protection against ischemia-reperfusion and contributed to clarifying the molecular and cellular mechanisms of deep hypothermia on myocardium.
Bes
S
S
Laboratory of Physiopathology and Pharmacology, Institute of Cardiovascular Research, Dijon, France.
Roussel
P
P
Laubriet
A
A
Vandroux
D
D
Tissier
C
C
Rochette
L
L
Athias
P
P
D000818:Animals; D015153:Blotting, Western; D002478:Cells, Cultured; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000378:metabolism; D007035:Hypothermia; D007511:Ischemia / Q000378:metabolism; D017202:Myocardial Ischemia / Q000517:prevention & control*; D015428:Myocardial Reperfusion Injury / Q000517:prevention & control*; D009206:Myocardium / Q000166:cytology* / Q000378:metabolism; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013696:Temperature; D013997:Time Factors
D018840:HSP70 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D012333:RNA, Messenger; D012313:RNA