Anthrax lethal toxin suppresses murine cardiomyocyte contractile function and intracellular Ca2+ handling via a NADPH oxidase-dependent mechanism.
OBJECTIVE
Anthrax infection is associated with devastating cardiovascular sequelae, suggesting unfavorable cardiovascular effects of toxins originated from Bacillus anthracis namely lethal and edema toxins. This study was designed to examine the direct effect of lethal toxins on cardiomyocyte contractile and intracellular Ca(2+) properties.


METHODS
Murine cardiomyocyte contractile function and intracellular Ca(2+) handling were evaluated including peak shortening (PS), maximal velocity of shortening/ relengthening (± dL/dt), time-to-PS (TPS), time-to-90% relengthening (TR(90)), intracellular Ca(2+) rise measured as fura-2 fluorescent intensity (ΔFFI), and intracellular Ca(2+) decay rate. Stress signaling and Ca(2+) regulatory proteins were assessed using Western blot analysis.


RESULTS
In vitro exposure to a lethal toxin (0.05-50 nM) elicited a concentration-dependent depression on cardiomyocyte contractile and intracellular Ca(2+) properties (PS, ± dL/dt, ΔFFI), along with prolonged duration of contraction and intracellular Ca(2+) decay, the effects of which were nullified by the NADPH oxidase inhibitor apocynin. The lethal toxin significantly enhanced superoxide production and cell death, which were reversed by apocynin. In vivo lethal toxin exposure exerted similar time-dependent cardiomyocyte mechanical and intracellular Ca(2+) responses. Stress signaling cascades including MEK1/2, p38, ERK and JNK were unaffected by in vitro lethal toxins whereas they were significantly altered by in vivo lethal toxins. Ca(2+) regulatory proteins SERCA2a and phospholamban were also differentially regulated by in vitro and in vivo lethal toxins. Autophagy was drastically triggered although ER stress was minimally affected following lethal toxin exposure.


CONCLUSIONS
Our findings indicate that lethal toxins directly compromised murine cardiomyocyte contractile function and intracellular Ca(2+) through a NADPH oxidase-dependent mechanism.
Kandadi
Machender R
MR
Center for Cardiovascular Research and Alternative Medicine, University of Wyoming College of Health Sciences, Laramie, Wyoming, United States of America.
Hua
Yinan
Y
Ma
Heng
H
Li
Qun
Q
Kuo
Shu-ru
SR
Frankel
Arthur E
AE
Ren
Jun
J
D000818:Animals; D000942:Antigens, Bacterial / Q000633:toxicity*; D001427:Bacterial Toxins / Q000633:toxicity*; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism*; D002460:Cell Line; D005453:Fluorescence; D006321:Heart / Q000187:drug effects*; D051379:Mice; D009206:Myocardium / Q000166:cytology*; D019255:NADPH Oxidases / Q000378:metabolism*
D000942:Antigens, Bacterial; D001427:Bacterial Toxins; C030325:anthrax toxin; D019255:NADPH Oxidases; D002118:Calcium