Tumor necrosis factor-alpha-induced caspase activation mediates endotoxin-related cardiac dysfunction.
OBJECTIVE
Sepsis-induced cardiac dysfunction is a serious clinical syndrome characterized by hypotension, decreased systemic vascular resistance, and elevated cardiac index. Although cytokines such as tumor necrosis factor (TNF)-alpha have been shown to play a significant role early in this response, the downstream effects of TNF-alpha signaling on cardiac function, specifically its relationship to apoptosis, have not been fully elucidated.


METHODS
Previous studies from our laboratory have identified endotoxin-induced apoptosis in cardiac cells in vitro. To further determine the role of lipopolysaccharide-induced apoptosis in vivo, mice were injected intraperitoneally with lipopolysaccharide (4 mg/kg), and cardiac apoptosis was detected and inhibited using a broad-spectrum caspase inhibitor.


METHODS
University research laboratory.


METHODS
Adult male wild-type (B6:129PF1/J) and TNF receptor 1/receptor 2 (TNFR-1/2) knockout mice (B6;129S-Tnfrsf1aTnfrsf1b).


METHODS
We sought to determine the dependence of cardiac apoptosis on TNF-alpha signaling and determine the physiologic role of caspase activation on lipopolysaccharide-induced cardiac dysfunction.


RESULTS
Cardiac apoptosis was determined at baseline and at 2, 4, 8, and 24 hrs by detection of capase-3 and -8 activity, cytoplasmic levels of Bax/Bcl-2, cleaved caspase-3 immunohistochemistry, and terminal deoxynucleotidyl transferase UTP nick-end labeling (TUNEL) staining of histologic sections in wild-type and TNFR-1/2 knockout mice. To determine the role of caspase activation in lipopolysaccharide-induced cardiac dysfunction, a broad-spectrum caspase inhibitor Z-Val-Ala-Asp (ome)-FMK (sad) was given, and cardiac function was determined in isolated beating hearts (Langendorff preparation). Our experiments determined that caspase-3-dependent apoptosis was active in cardiac tissue by 2 hrs and that this activation was completely mediated by TNFR-1/2. The Bax/Bcl-2 ratios supported the finding and time course of apoptosis, whereas TUNEL staining of cardiac tissue sections identified sporadic apoptotic ventricular cells. The administration of zVAD significantly inhibited myocardial caspase-3 activity and preserved cardiac physiologic function (Langendorff preparation).


CONCLUSIONS
Endotoxin induces a TNF-alpha-dependent apoptotic cascade in the myocardium, which contributes to the development of cardiac dysfunction.
Carlson
Deborah L
DL
Department of Pediatrics, University of Texas Southwestern, Dallas, TX, USA.
Willis
Monte S
MS
White
D Jean
DJ
Horton
Jureta W
JW
Giroir
Brett P
BP
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis; D053148:Caspase 3; D053181:Caspase 8; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism*; D004731:Endotoxins / Q000633:toxicity; D004789:Enzyme Activation; D006331:Heart Diseases / Q000201:enzymology* / Q000382:microbiology; D020287:In Situ Nick-End Labeling; D008070:Lipopolysaccharides; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D014409:Tumor Necrosis Factor-alpha / Q000502:physiology*
D000590:Amino Acid Chloromethyl Ketones; D061945:Caspase Inhibitors; D004731:Endotoxins; D008070:Lipopolysaccharides; D014409:Tumor Necrosis Factor-alpha; C476093:z-Val-Ala-Asp(Ome)-fluoromethylketone; C505409:Casp3 protein, mouse; C505526:Casp8 protein, mouse; D053148:Caspase 3; D053181:Caspase 8; D020169:Caspases