J Mol Cell Cardiol 31, 147–158 (1999) Article No. jmcc.1998.0855, available online at http://www.idealibrary.com on Quantification of Cardioprotective Gene Expression in Porcine Short-term Hibernating Myocardium Babett Bartling 1 , Juliane Hoffmann 1,2 , Ju ¨ rgen Holtz 1 , Rainer Schulz 3 , Gerd Heusch 3 and Dorothea Darmer 1 1 Institute of Pathophysiology, Martin Luther University, Halle-Wittenberg, Magdeburger Str. 18, D-06097 Halle; 2 Clinic of Gastroenterology, Otto von Guericke University, Magdeburg, Leipziger Str. 44, D-39120 Magdeburg; and 3 Department of Pathophysiology, University Essen, Hufelandstr. 55, D-45147 Essen, Germany (Received 7 July 1998, accepted in revised form 29 September 1998) B. B, J. H, J. H, R. S, G. H D. D. Quantification of Cardioprotective Gene Expression in Porcine Short-term Hibernating Myocardium. Journal of Molecular and Cellular Cardiology (1999) 31, 147–158. Several cardioprotective proteins are induced during myocardial ischemia, such as heat shock proteins and anti-apoptotic Bcl-2-related proteins which, when experimentally overexpressed, have been shown to prevent ischemia-induced myocyte loss. As this pathophysiological induction is obviously not sufficient to prevent losses of myocytes, we analysed whether it could occur under moderate myocardial ischemia with hibernation, thus potentially contributing to myocyte protection under these conditions. Therefore, using anesthetized pigs with documented myocardial hypoperfusion and short-term hibernation, we investigated the left ventricular mRNA expression of the inducible heat shock protein Hsp70 and of the anti-apoptotic Bcl-x L in comparison with the pro-apoptotic Bak and Fas expression. For transcriptional analyses, the porcine cDNA sequences of Bcl-x L , Bak and Fas were identified by polymerase chain reaction (PCR) or by screening of a porcine heart cDNA library and cloned. Using reverse transcription polymerase chain reaction (RT-PCR), we observed an unchanged mRNA expression of inducible Hsp70, Bcl-x L , Bak and Fas after 85 min of hypoperfusion in the short- term hibernating myocardium, as well as after 30 min of subsequent reperfusion in the stunned myocardium, compared with transcription in a non-hypoperfused control area of the same ventricle. In conclusion, the mRNA expression of inducible Hsp70 and of several apoptosis-modulating proteins is not altered during moderate myocardial ischemia resulting in short-term hibernation of the affected area and during subsequent stunning. 1999 Academic Press K W: Heat shock; Hsp70; Apoptosis; Bcl-x L ; Fas; Bak; Hibernation; Stunning; Myocardium; Pig. 1991). Its intracellular signal transduction domain, Introduction called the ‘‘death domain’’, is activated by extra- cellular binding of the Fas ligand (FasL) which is During total coronary occlusion, myocyte necrosis as well as typical signs indicating apoptotic myocyte either soluble or presented by circulating T lym- phocytes (Suda et al., 1993). death can be detected in the ischemic area (for a review, see Bartling et al., 1998a). In rats, this is In addition to this potentially pro-apoptotic sys- tem, several cardioprotective reactions are activated associated with an increased myocardial expression of the apoptosis-triggering surface receptor Fas during severe ischemia in the myocardium, such as upregulation of heat shock proteins (Currie et (Kajstura et al., 1996), a member of the tumor necrosis factor (TNF) receptor family (Itoh et al., al., 1993), supporting protein folding and trafficking Please address all correspondence to: Dorothea Darmer, Institut fu ¨ r Pathophysiologie, Martin-Luther-Universita ¨t Halle-Wittenberg, Magdeburger Str. 18, 06097 Halle, Germany. 0022–2828/99/010147+12 $30.00/0 1999 Academic Press