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 PKCbeta throughout adulthood (beta-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 beta-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 and/or preservation of high-energy phosphate levels during ischemia contributed to the cardioprotection in beta-Tx hearts. No difference in intracellular pH or high-energy phosphate levels was found between the beta-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 high-energy phosphate levels during ischemia contribute to the improved recovery.

Figure 1

Percentage changes in LV-developed pressure (Left) and LVEDP (Right) in control (open symbols) and β-Tx (filled symbols) hearts during 12-min ischemia and 24-min reperfusion. Data are shown as means ± SEM. The ischemia period is indicated by the shaded area.

Figure 2

Percentage changes in RPP (Left) and LVEDP (Right) in control and β-Tx hearts during 28-min ischemia and 40-min reperfusion. (Upper) Comparisons among the three genotypes used for controls (open square for −/+; filled square for +/−; open diamond for −/−). (Lower) Comparisons between pooled control (open symbols) and β-Tx (filled symbols) hearts treated with (triangles) or without (circles) CH. Data are shown as means ± SEM. The ischemia period is indicated by the shaded area.

Figure 3

Representative ³¹P NMR spectra for a β-Tx (Left) and a control (Right) heart at baseline (Bottom), at the end of 12-min ischemia (Middle), and at the end of 24-min reperfusion (Top). For each spectrum, from left to right, the peaks represent Pi, PCr, and γ-, α-, and β-phosphates of ATP. The difference in the chemical shift of Pi and PCr is proportional to the pHi. Note the shift of Pi peak during ischemia, indicating intracellular acidosis.

Figure 4

Group means for pHi (Left), [ATP] (Center), and [PCr] and [Pi] (Right) at baseline, during 12-min ischemia and 24-min reperfusion for control (open symbols) and β-Tx (filled symbols) hearts. Data are shown as means ± SEM. The ischemia period is indicated by the shaded area.

Figure 5

Group means for pHi, [Pi], [PCr], and [ATP] at baseline, during 28-min ischemia and 40-min reperfusion for control (open symbols) and β-Tx (filled symbols) hearts treated with (triangles) or without (circles) CH. Data are shown as means ± SEM. The ischemia period is indicated by the shaded area.