PPAR-gamma activation fails to provide myocardial protection in ischemia and reperfusion in pigs

Abstract

Peroxisome proliferator-activated receptor (PPAR)-gamma modulates substrate metabolism and inflammatory responses. In experimental rats subjected to myocardial ischemia-reperfusion (I/R), thiazolidinedione PPAR-gamma activators reduce infarct size and preserve left ventricular function. Troglitazone is the only PPAR-gamma activator that has been shown to be protective in I/R in large animals. However, because troglitazone contains both alpha-tocopherol and thiazolidinedione moieties, whether PPAR-gamma activation per se is protective in myocardial I/R in large animals remains uncertain. To address this question, 56 pigs were treated orally for 8 wk with troglitazone (75 mg x kg(-1) x day(-1)), rosiglitazone (3 mg x kg(-1) x day(-1)), or alpha-tocopherol (73 mg x kg(-1) x day(-1), equimolar to troglitazone dose) or received no treatment. Pigs were then anesthetized and subjected to 90 min of low-flow regional myocardial ischemia and 90 min of reperfusion. Myocardial expression of PPAR-gamma, determined by ribonuclease protection assay, increased with troglitazone and rosiglitazone compared with no treatment. Rosiglitazone had no significant effect on myocardial contractile function (Frank-Starling relations), substrate uptake, or expression of proinflammatory cytokines during I/R compared with untreated pigs. In contrast, preservation of myocardial contractile function and lactate uptake were greater and cytokine expression was attenuated in pigs treated with troglitazone or alpha-tocopherol compared with untreated pigs. Multivariate analysis indicated that presence of an alpha-tocopherol, but not a thiazolidinedione, moiety in the test compound was significantly related to greater contractile function and lactate uptake and lower cytokine expression during I/R. We conclude that PPAR-gamma activation is not protective in a porcine model of myocardial I/R. Protective effects of troglitazone are attributable to its alpha-tocopherol moiety. These findings, in conjunction with prior rat studies, suggest interspecies differences in the response to PPAR-gamma activation in the heart.

Fig. 1

Instrumentation of the heart. LV, left ventricle; LA, left atrium.

Fig. 2

Myocardial expression of peroxisome proliferator-activated receptor (PPAR)-γ. Ribonuclease protection assay demonstrates expression of PPAR-γ₁ and PPAR-γ₂ mRNA in subendocardial samples from ischemic-reperfused region of 1 representative heart from each experimental group. Subcutaneous fat is a positive control. Quantitative analysis of group data is shown in Table 2.

Fig. 3

Systolic function in the ischemic region. Preload-adjusted regional external work (see text) was utilized as a load-insensitive measure of regional systolic function. Under baseline (preischemic) conditions, regional external work index did not differ among groups (25 ± 2, 26 ± 4, 26 ± 1, and 25 ± 5 mmHg/cm² in untreated, troglitazone, rosiglitazone, and α-tocopherol groups, respectively). Therefore, data during ischemia and reperfusion are expressed as a fraction of baseline. Although each group exhibited contractile dysfunction during ischemia and reperfusion, dysfunction was less severe among troglitazone (n = 6) and α-tocopherol (n = 7) groups than among rosiglitazone (n = 12) and untreated (n = 15) groups. B, baseline; I, after 90 min of low-flow ischemia; R, after 90 min of reperfusion.

Fig. 4

Myocardial cytokine mRNA expression. Representative ribonuclease protection assays show expression of IL-1β, IL-6, and IFN-γ in subendocardial tissue. Lane 1, minimal expression in nonischemic myocardium; lane 2, stimulation of expression of all 3 cytokines by 90 min of low-flow ischemia (Isch) and 90 min of reperfusion (Reperf); lanes 3–5, attenuation of cytokine expression in ischemic-reperfused myocardium by pretreatment with troglitazone and α-tocopherol, but not rosiglitazone.

Fig. 5

Myocardial cytokine mRNA and protein expression. Pooled data are shown for pigs pretreated with troglitazone (n = 6), rosiglitazone (n = 12), or α-tocopherol (n = 6) and untreated pigs (n = 7). Top: cytokine mRNA expression by ribonuclease protection assay, normalized to expression of GAPDH in each heart. Data are medians ± interquartile range. Bottom: cytokine protein expression by enzyme-linked immunosorbent assay. Values are means ± SE. U, untreated; T, troglitazone; R, rosiglitazone; E, α-tocopherol; ND, not detected. *P < 0.05 vs. nonischemic. ‡P < 0.05 vs. untreated.

Fig. 6

Myocardial lactate uptake. Myocardial lactate uptake becomes negative (net release) during ischemia and recovers partially with reperfusion. Over the course of the experiment, lactate uptake was greater in troglitazone and α-tocopherol groups (test compounds with an α-tocopherol moiety) than in rosiglitazone and untreated groups (no α-tocopherol moiety). P value indicates an intergroup difference over the course of the experiment.

Fig. 7

Effect of chronic troglitazone treatment on myocardial free fatty acid (FFA) uptake. Myocardial FFA uptake is plotted as a function of arterial FFA concentration ([FFA]) under nonischemic, ischemic, and reperfusion conditions for 14 untreated pigs and 12 troglitazone-pretreated pigs. In 6 pigs from each group, arterial FFA concentrations were increased by infusion of triglyceride emulsion (10% Liposyn) and heparin. Lines indicate linear regression of data. Chronic troglitazone treatment had no significant effect on myocardial FFA uptake under any condition.