Reperfusion-Activated Akt Kinase Prevents Apoptosis in Transgenic Mouse Hearts Overexpressing Insulin-Like Growth Factor-1

Abstract

Abstract-Hearts of wild-type and insulin-like growth factor-1 overexpressing (Igf-1 ϩ/Ϫ ) transgenic mice were subjected to Langendorff perfusions and progressive periods of ischemia followed by reperfusion. Apoptosis was measured by DNA nucleosomal cleavage and a hairpin probe labeling assay to detect single-base overhang. Transgenic hearts subjected to 20 minutes of ischemia and 4 hours of reperfusion (I/R) sustained a rate of apoptosis of 1.8Ϯ0.3% compared with 4.6Ϯ1.1% for wild-type controls (nϭ4; PϽ0.03). Phosphorylation of the protein kinase Akt/protein kinase B was elevated 6.2-fold in transgenic hearts at baseline and increased another 4.4-fold within 10 minutes of reperfusion, remaining elevated for up to 2 hours. I/R activated Akt in wild-type hearts but to a lesser extent (1.6Ϯ0.3-fold). Pretreatment of transgenic hearts with wortmannin immediately before and during ischemia eliminated reperfusionmediated activation of Akt and neutralized the resistance to apoptosis. The stress-activated kinase p38 was also activated during ischemia and reperfusion in both wild-type and transgenic hearts. Perfusion with the p38 inhibitor SB203580 (10 mol/L) blocked both p38 activation and phosphorylation of Akt and differentially modulated apoptosis in wild-type and transgenic hearts. Pretreatment with SB203580 reduced apoptosis in wild-type hearts but increased apoptosis in transgenic hearts. These results demonstrate that Akt phosphorylation during I/R is modulated by IGF-1 and prevents apoptosis in hearts that overexpress the IGF-1 transgene. (Circ Res. 2001;88:609-614.) Key Words: ischemia Ⅲ hypoxia Ⅲ phosphoinositol-3Ј-kinase Ⅲ p38 mitogen-activated protein kinase Ⅲ SB203580 P rotective and antiapoptotic properties of insulin-like growth factor-1 (IGF-1) have been demonstrated in different models of myocardial ischemia and infarction 1-4 as well as in isolated cardiac myocytes subjected to ischemic or oxidative stress. 5,6 IGF-1 stimulates the phosphoinositol-3Ј (PI3)-kinase pathway, producing phosphoinositides that promote activation of the kinase Akt. 7,8 Activated Akt kinase plays a central role in suppressing apoptosis by modulating the activities of Bcl-2 family proteins, 9 caspase 9, 10 and Fas ligand. 11 Transfer of mutationally activated PI3-kinase and Akt genes has been shown to prevent apoptosis of cardiac myocytes in vitro, 6 and activated Akt delivered by an adenovirus vector reduced apoptosis in the intact heart subjected to ischemia and reperfusion (I/R). 4 We previously reported that Igf-1 ϩ/Ϫ transgenic mice overexpressing IGF-1 had reduced rates of necrosis and apoptosis after myocardial infarction caused by coronary artery ligation. 3 The same hearts were also resistant to necrosis but not apoptosis caused by nonocclusive coronary artery constriction. 12 Other work has shown that the endogenous PI3-kinase pathway is activated in the postinfarcted myocardium, where it may contribute to cell survival and hypertrophy. [13][14][15] Regulation of the endogenous PI3-kinase pathway during I/R has not b...

Adenoviral gene transfer of Akt enhances myocardial contractility and intracellular calcium handling

“…4 The growth-promoting and antiapoptotic properties of IGF-1 activation are indeed partly mediated by Akt activation. 4,6 It has been proposed that myocardial Akt activation may also be involved in the structural and functional changes observed after longterm exposure to GH excess, as typically occurs in the acromegaly. 5,22 In the same model system, we have previously shown an increase of contractility averaging 20% following acute application of graded concentration of IGF-1 in the perfusate, mostly due to augmented Ca 2+ responsiveness of the myofilaments.…”

“…The principal and most extensively investigated physiological actions of Akt consist of growth activation and apoptosis inhibition, both consistently documented in many systems. [4][5][6][7][8][9] However, it appears that Akt may also be involved in the regulation of myocardial contractility. A recently generated transgenic mouse with cardiac-restricted overexpression of constitutively active Akt, the E40K, exhibited a phenotype characterized not only by cardiomyocyte hypertrophy but also by a remarkable increase of myocardial contractility.…”

Celecoxib modulates hypertrophic signalling and prevents load‐induced cardiac dysfunction

“…Analyzing the degree of Akt activation may help to resolve this potential paradox. Whereas in the studies demonstrating protective effects the level of Akt phosphorylation was rather moderate (1.5-to 6-fold) [12,[29][30][31], the level of Akt activity in studies with maladaptive effects was much higher (15-to 80-fold) [14,27,28]. Therefore, it appears that the character of hypertrophy is determined by the degree of Akt activity.…”

“…Accordingly, it appears consistent that a pharmacologically controlled inhibition of Akt activity by celecoxib has cardioprotective effects. However, there are also numerous studies documenting beneficial effects of Akt activation in the heart [12,[29][30][31]. Moreover, physiological hypertrophy in response to exercise training is accompanied by higher levels of activated Akt [12].…”

“…Moreover, physiological hypertrophy in response to exercise training is accompanied by higher levels of activated Akt [12]. Transgenic overexpression of PI3K [30], IGF-1 [31], or the IGF-1 receptor [29], resulting in higher Akt activity, are accompanied by preserved cardiac function. Furthermore, activation of Akt has a pivotal regulatory role for normal postnatal growth of the heart [11].…”