Nicotine inhibits cardiac apoptosis induced by lipopolysaccharide in rats

AbstractObjectivesApoptosis develops in several heart diseases, but the therapeutic options are limited. It was hypothesized that nicotine, which inhibits apoptosis in several cells, inhibits cardiac apoptosis induced by lipopolysaccharide (LPS).BackgroundOver-the-counter nicotine produces sustained levels (10 to 25 ng/ml) that may be antiapoptotic. Low levels of LPS induce apoptosis by activating tissue renin-angiotensin to stimulate angiotensin II, type 1 (AT1) receptors in cardiac myocytes.MethodsAdult Sprague Dawley rats were pretreated with nicotine (6 mg/kg/day) or saline for seven to ten days (miniosmotic pumps). The LPS (1 mg/kg) was injected intravenously. Toll-like receptor 4 (TLR4) and angiotensinogen messenger ribonucleic acid (mRNA) were measured in the heart after 0, 4, 8, 16, and 24 h. Cardiac apoptosis was measured by terminal deoxy-nucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) staining after 24 h. In vitro effects of LPS (10 ng/ml, 24 h) were studied in cardiac myocytes isolated from rats pretreated with nicotine for 7 to 10 days, or after pre-exposing myocytes to nicotine (15 ng/ml) for 1, 4, 16, or 24 h.ResultsNeither nicotine nor LPS affected systolic blood pressure. The LPS increased cardiac apoptosis after 24 h in saline-treated, but not nicotine-treated rats, despite similar increases in cardiac TLR4 and angiotensinogen mRNA over 8 to 16 h. The LPS-induced apoptosis was blocked by pre-exposing myocytes to nicotine for 4 to 24 h (partial inhibition after 1 h). Nicotine did not inhibit apoptosis induced by angiotensin II (100 nM, 24 h).ConclusionsTherapeutic levels of nicotine inhibit LPS-induced cardiac apoptosis. This occurs after LPS increases TLR4 and angiotensinogen mRNA, but proximal to AT1receptor activation. Nicotine may be a novel inhibitor of cardiac apoptosis in conditions associated with circulating LPS (e.g., decompensated heart failure, acute and chronic infections)

Lipopolysaccharide Activates Calcineurin in Ventricular Myocytes

ObjectivesWe investigated whether lipopolysaccharide (LPS), a proximate cause of inflammation, activates calcineurin in cardiac myocytes and if calcineurin regulates apoptosis in this setting.BackgroundCalcineurin regulates myocardial growth and hypertrophy, but its role in inflammation is unknown. Calcineurin has proapoptotic or antiapoptotic effects depending on the stimuli.MethodsCalcineurin activity was measured in left ventricular myocytes from adult Sprague Dawley rats. Cardiac apoptosis was measured by terminal deoxy-nucleotidyl transferase-mediated dUTP nick end-labeling staining and caspase-3 activity after in vitro and in vivo exposure to LPS.ResultsLipopolysaccharide increased calcineurin activity in myocytes over 1 to 24 h (t 1/2 = 4.8 h) with an EC50of 0.80 ng/ml LPS (p < 0.05, n = 4). The LPS (10 ng/ml) effects were mimicked by angiotensin II (Ang II) (100 nmol/l); both increased calcineurin activity and induced apoptosis without additive effects (p < 0.05, n = 5 to 9). Lipopolysaccharide and/or Ang II effects were prevented by 1 h pre-treatment with an Ang II type 1 receptor blocker (losartan, 1 μmol/l), calcineurin inhibitor (cyclosporin A, 0.5 μmol/l), calcium chelator (1,2-Bis(2-amino-5-fluorophenoxy)ethane-N,N,N′,N′-tetraacetic acid tetrakis(acetoxymethyl) ester, 0.1 μmol/l), or by inhibiting sarcoplasmic reticulum (SR) calcium (Ca)-ATPase (thapsigargin, 1 μmol/l) or SR calcium release channel (ryanodine, 1 μmol/l). Left ventricular apoptosis increased from 4 to 24 h after LPS (1 mg/kg intravenously) in vivo, but not in rats pre-treated with cyclosporin A (20 mg/kg/day subcutaneously) for 3 days (p < 0.05, n = 5).ConclusionsIn cardiac myocytes, LPS activates calcineurin in association with apoptosis by Ang II and SR calcium-dependent mechanisms. This expands the paradigm for cardiac calcineurin to be activated by low levels of LPS in inflammation and chronic conditions (e.g., infections, smoking, and heart failure)