Lipopolysaccharide Activates Calcineurin in Ventricular Myocytes Original Research Article

Objectives We investigated whether lipopolysaccharide (LPS), a proximate cause of inflammation, activates calcineurin in cardiac myocytes and if calcineurin regulates apoptosis in this setting. Background Calcineurin regulates myocardial growth and hypertrophy, but its role in inflammation is unknown. Calcineurin has proapoptotic or antiapoptotic effects depending on the stimuli. Methods Calcineurin 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. Results Lipopolysaccharide increased calcineurin activity in myocytes over 1 to 24 h (t 1/2 = 4.8 h) with an EC50 of 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). Conclusions In 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).