Intrahepatic nuclear factor-κ B activity and α 1-acid glycoprotein transcription do not predict outcome after cecal ligation and puncture in the rat

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Abstract
Sepsis is the leading cause of death in critically ill surgical patients. Septic hepatic dysfunction, an important determinant of outcome, is poorly understood but includes inappropriate transcriptional down-regulation. This may be modulated by proinflammatory cytokines. We hypothesized that intrahepatic changes in tumor necrosis factor (TNF)/interleukin (IL)-1-linked processes, such as the activation of the p50 homodimeric and the p65/p50 heterodimeric isoforms of the transcription factor nuclear factor (NF)-κB or transcription of the acute phase reactant α1-acid glycoprotein (AGP), would correlate with recovery from sepsis. Prospective experimental comparison of sham operation and nonlethal and lethal sepsis in male Sprague-Dawley rats. Nonlethal sepsis was induced by using single-puncture cecal ligation and puncture (CLP). Lethal sepsis was induced via double-puncture CLP. NF-κB DNA binding activity was determined by using electrophoretic mobility shift analysis with differentiation of p50/p50 and p50/p65 isoforms by using appropriate antibodies. AGP transcription was assessed with transcription elongation analysis, intrahepatic IL-1β, and TNF-α abundance by using immunohistochemistry, and serum IL-1β was assessed by using ELISA. Overall NF-κB activity increased equivalently over time after both single- and double-puncture CLP, with a peak occurring 3 hrs after intervention. In single-puncture CLP, there was an increase in the binding of the p50 homodimer form over time. After double-puncture CLP, no such change was observed. AGP transcription was increased equivalently in both models. Intrahepatic IL-1β was detected 16 and 24 hrs after single-puncture CLP and 6 hrs after double-puncture CLP. After double-puncture CLP, intrahepatic TNF-α was detected at 6, 16, and 24 hrs. Serum IL-1β was undetectable after both single- and double-puncture CLP. Although AGP transcription was similar in mild and fulminant sepsis, double-puncture CLP increased the binding activity of the p50 homodimer relative to binding of the p50/p65 NF-κB heterodimer. These results imply that transcriptional activity not linked to acute phase responses is an important determinant of outcome in sepsis.