Long-term expression of protein kinase C in adult mouse hearts improves postischemic recovery

Abstract

Activation of protein kinase C (PKC) protects the heart from ischemic injury; however, its mechanism of action is unknown, in part because no model for chronic activation of PKC has been available. To test whether chronic, mild elevation of PKC activity in adult mouse hearts results in myocardial protection during ischemia or reperfusion, hearts isolated from transgenic mice expressing a low level of activated PKC␤ throughout adulthood (␤-Tx) were compared with control hearts before ischemia, during 12 or 28 min of no-flow ischemia, and during reperfusion. Left-ventricular-developed pressure in isolated isovolumic hearts, normalized to heart weight, was similar in the two groups at baseline. However, recovery of contractile function was markedly improved in ␤-Tx hearts after either 12 (97 ؎ 3% vs. 69 ؎ 4%) or 28 min of ischemia (76 ؎ 8% vs. 48 ؎ 3%). Chelerythrine, a PKC inhibitor, abolished the difference between the two groups, indicating that the beneficial effect was PKC-mediated. 31 P NMR spectroscopy was used to test whether modification of intracellular pH and͞or preservation of high-energy phosphate levels during ischemia contributed to the cardioprotection in ␤-Tx hearts. No difference in intracellular pH or high-energy phosphate levels was found between the ␤-Tx and control hearts at baseline or during ischemia. Thus, long-term modest increase in PKC activity in adult mouse hearts did not alter baseline function but did lead to improved postischemic recovery. Furthermore, our results suggest that mechanisms other than reduced acidification and preservation of highenergy phosphate levels during ischemia contribute to the improved recovery.A ctivation of protein kinase C (PKC) has been postulated to play an important role in modulating cardiac contractile function and cellular growth (1). Furthermore, there are a number of studies showing that PKC inhibitors block, whereas PKC activators mimic, the cardioprotective effect of ischemic preconditioning in a variety of animal models (2-5) and, importantly, in human myocytes (6). However, studies with PKC activators have two major limitations. First, widely used PKC activators, such as phorbol ester, are nonselective and cause significant changes in contractile function in normoxic hearts. Second, experiments with PKC activators can be performed only in an acute setting; they cannot be used as therapeutic agents because of their divergent effects on multiple organ systems.Recently, by using the binary tetracycline-controlled transactivator (tTA) system, a transgenic mouse line with conditional expression of a constitutively active PKC ␤-isoform (PKC␤*) has been developed (7,8). The transgene is targeted to cardiac myocytes with the rat ␣-myosin heavy-chain promoter. By using this strategy, a modest increase in PKC activity can be achieved in a spatially and temporally restricted fashion (7). If it can be shown that chronic expression of PKC␤* is cardioprotective, we will have an unique model to (i) define the mechanism(s) underlying PKCmediated myocardial pro...

Gender Differences in Cardioprotection against Ischemia/Reperfusion Injury in Adult Rat Hearts: Focus on Akt and Protein Kinase C Signaling

“…However, similar to wortmannin, chelerythrine treatment resulted in a significant reduction of postischemic recovery of left ventricular function and an increase in myocardial infarct size in female but not in male hearts and eliminated the gender differences in the postischemic recovery and myocardial infarct size. In agreement with the present finding, previous studies showed that chelerythrine had no effect on postischemic function and infarct size in male hearts (Tian et al, 1999;Ding et al, 2004). The effect of chelerythrine on female hearts has not been previously examined.…”

“…Control hearts received the vehicle only (0.04% DMSO). The concentrations of wortmannin and chelerythrine were chosen on the basis of reported IC 50 of the inhibitors (5 nM wortmannin; Arcaro and Wymann, 1993;0.66 M chelerythrine;Herbert et al, 1990) and the concentrations used in the previous studies for wortmannin (100 nM) (Fujio et al, 2000;Mockridge et al, 2000;Tong et al, 2000;Bell and Yellon, 2003;Okumura et al, 2004;Tsang et al, 2004) and chelerythrine (2-4 M) (Tian et al, 1999;Mockridge et al, 2000;Stamm et al, 2001;Przyklenk et al, 2003;Ding et al, 2004) in animal models of ischemia and reperfusion injury. In separate experiments, some male hearts were pretreated with 5 M PKC⑀ translocation inhibitor peptide (PKC⑀-TIP; Calbiochem) for 20 min before ischemia and reperfusion with no wash-out period.…”

Cardiac-Specific Expression of Heme Oxygenase-1 Protects Against Ischemia and Reperfusion Injury in Transgenic Mice

“…22 Body weights and heart weights of the mice used in the experiments were similar among all four groups (Table). Hearts were stabilized for 30 minutes, and baseline function was measured.…”

“…Hearts were rapidly excised, arrested in ice-cold buffer, and connected via the aorta to the perfusion cannula as described. 22 Retrograde perfusion was maintained at a constant pressure of 70 mm Hg by gravity. The flow of thebesian veins was drained by a thin polyethylene tube (PE-10) through the apex of the left ventricle (LV).…”

Convergence of Protein Kinase C and JAK-STAT Signaling on Transcription Factor GATA-4

“…Second, although numerous hypertrophic signals were shown to activate different PKC isoforms, the bulk of the evidence argues against an involvement of PKC in initiating and/or maintaining hypertrophy (11,28). In fact, upregulation of PKC␤ in the adult heart has been shown to have beneficial effects (75). Importantly, pharmacologic inhibition of PKC did not block AII-induced hypertrophy (74) but did abrogate AII activation of c-fos transcription (62) and ANF biosynthesis (34).…”