Abstract-Disruption of leptin signaling is associated with obesity, heart failure, and cardiac hypertrophy, but the role of leptin in cardiac myocyte apoptosis is unknown. We tested the hypothesis that apoptosis increases in leptin-deficient ob/ob and leptin-resistant db/db mice and is associated with aging and left ventricular hypertrophy, increased DNA damage, and decreased survival. We studied young (2-to 3-month-old) and old (12-to 14-month-old) ob/ob and db/db mice and wild-type (WT) controls (nϭ2 to 4 per group). As expected, ventricular wall thickness and heart weights were similar among young ob/ob, db/db, and WT mice, but higher in old ob/ob and db/db versus old WT. Young ob/ob and db/db showed markedly elevated apoptosis by TUNEL staining and caspase 3 levels compared with WT. Differences in apoptosis were further accentuated with age. Leptin treatment significantly reduced apoptosis in ob/ob mice both in intact hearts and isolated myocytes. Tissue triglycerides were increased in ob/ob hearts, returning to WT levels after leptin repletion. Furthermore, the DNA damage marker, 8oxoG (8-oxo-7,8-dihydroguanidine), was increased, whereas the DNA repair marker, MYH glycosylase, was decreased in old ob/ob and db/db compared with old WT mice. Both ob/ob and db/db mice had decreased survival compared with WT mice. We conclude that leptin-deficient and leptin-resistant mice demonstrate increased apoptosis, DNA damage, and mortality compared with WT mice, suggesting that normal leptin signaling is necessary to prevent excess age-associated DNA damage and premature mortality. These data offer novel insights into potential mechanisms of myocardial dysfunction and early mortality in obesity.

Mitochondrial generation of reactive oxygen species (ROS) is increased in mice with fatty livers induced by genetic obesity, chronic consumption of ethanol, or methionine/choline-deficient diets. Both nuclear and mitochondrial (mt) DNA are targets for ROS-induced damage and accumulate hydroxylated bases, such as 8-hydroxy-2'-deoxyguanosine (8-oxoG) and base substitution of adenine with 8-oxoG (A*8-oxoG), that introduce mutations that promote cancer as well as cell death. The mammalian homolog of the bacterial DNA mismatch repair enzyme MutY (MYH) removes A*8-oxoG from nuclear and mtDNA, reduces 8-oxoG accumulation, and restores genomic stability after ROS exposure. Cumulative damage to mtDNA occurs as fatty liver disease progresses. Therefore, differences in hepatic MYH activity may influence the severity of fatty liver disease. To evaluate this hypothesis, we compared mtH2O2 production, MYH expression, oxidative DNA damage, and hepatocyte death in healthy mice and different mouse models of fatty liver disease. The results show that diverse causes of steatohepatitis increase mtROS production, limit repair of mtDNA, and oxidatively damage DNA. However, there are important differences in the DNA repair response to oxidant stress among mouse models of fatty liver disease. Independent of the degree of mtROS generation, models with the least MYH exhibit the greatest accumulation of 8-oxoG and the most hepatocyte death. These findings raise the intriguing possibility that inherited or acquired differences in DNA repair enzyme activity may underlie some of the interindividual differences in fatty liver disease outcomes.

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The benefit of the ␤ 2 -adrenergic agonist, clenbuterol, in left ventricular assist device patients with dilated cardiomyopathy has been reported, but its effect on ischemic heart failure (HF) is unknown. We investigated whether clenbuterol improves left ventricular remodeling, myocardial apoptosis and has synergy with a ␤ 1 antagonist, metoprolol, in a model of ischemic HF. Rats were randomized to: 1) HF only; 2) HF ϩ clenbuterol; 3) HF ϩ metoprolol; 4) HF ϩ clenbuterol ϩ metoprolol; and 5) rats with sham surgery. HF was induced by left anterior descending artery (LAD) artery ligation and confirmed by decreased left ventricular fractional shortening, decreased maximum left ventricular dP/dt (dP/dt max ), and elevated left ventricular end-diastolic pressure (LVEDP) compared with sham rats (p Ͻ 0.01). After 9 weeks of oral therapy, echocardiographic, hemodynamic, and ex vivo end-diastolic pressure-volume relationship (EDPVR) measurements were obtained. Immunohistochemistry was performed for myocardial apoptosis and DNA damage markers. Levels of calcium-handling proteins were assessed by Western blot analysis. Clenbuterol-treated HF rats had increased weight gain and heart weights versus HF rats (p Ͻ 0.05). EDPVR curves revealed a leftward shift in clenbuterol rats versus metoprolol and HF rats (p Ͻ 0.05). The metoprololtreated group had a lower LVEDP and higher dP/dt max versus the HF group (p Ͻ 0.05). Clenbuterol and metoprolol groups had decreased myocardial apoptosis and DNA damage markers and increased DNA repair markers versus HF rats (all p Ͻ 0.01). Protein levels of the ryanodine receptor and sarcoplasmic reticulum calcium-ATPase were improved in clenbuterol-, metoprolol-, and clenbuterolϩmetoprolol-treated groups versus HF rats. However, as a combination therapy, there were no synergistic effects of clenbuterolϩmetoprolol treatment. We conclude that clenbuterol ameliorates EDPVR, apoptosis, and calcium homeostasis but does not have synergy with metoprolol in our model of ischemic HF.Clenbuterol is a ␤ 2 -adrenergic receptor agonist first used in the mid-1970s to treat asthma and is approved for this indication in Europe (Salorinne et al., 1975). The drug bears structural similarity to albuterol but has enhanced oral absorption and ␤ 2 -selectivity. Clenbuterol also increases muscle bulk (Choo et al., 1992;Maltin et al., 1993;Carter and Lynch, 1994) because of an anabolic effect that may be mediated via long-term ␤ 2 -activation (MacLennan and Edwards, 1989;Choo et al., 1992). Because of this anabolic action, oral clenbuterol has been abused by athletes to enhance size and strength (Beckett, 1992;Perry, 1993). Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.