Release of tumour necrosis factor alpha (TNFalpha) by TNFalpha cleaving enzyme (TACE) in response to septic stimuli in vitro.
BACKGROUND
Tumour necrosis factor alpha (TNFalpha), in its soluble form (solTNF), has been well described as an important cytokine in inflammatory states including sepsis. The transmembrane precursor of solTNF, membrane-bound TNFalpha (memTNF), is cleaved by TNFalpha cleaving enzyme (TACE), the regulation of which is poorly understood. We hypothesized that the diversity of clinical features seen with sepsis caused by different organisms may be a result of differential regulation of TACE. Therefore, we measured these proteins in models of sepsis using flow cytometric methods that we have developed.


METHODS
Surface protein expression of memTNF and TACE, and TACE catalytic activity were measured in human monocytes by flow cytometry following cell stimulation by lipopolysaccharide (LPS), zymosan (a yeast cell wall product) or heat-inactivated Neisseria meninigitidis.


RESULTS
Unstimulated human monocytes express memTNF on the cell surface (mean fluorescence intensity, MFI 131) and this is down-regulated initially in response to LPS (MFI 57) but then recovers to exceed the resting protein expression (MFI 614). TACE protein is also present on the surface of resting cells (MFI 389) but is catalytically inactive until cell stimulation. Stimulation of monocytes with LPS, zymosan and Neisseria meningitidis produced different patterns of TACE activation with time.


CONCLUSIONS
The regulation of memTNF by TACE on monocytes is an important regulatory event in the pro-inflammatory cytokine cascade. As monocytes are important in the inflammatory cascade, we suggest that the control of memTNF and TACE activity on monocytes may play a role in the pathophysiology of sepsis.
Robertshaw
H J
HJ
Anaesthesia and Intensive Care Medicine, St George's Hospital Medical School, Cranmer Terrace, London SW17 0RE, UK. hroberts@sghms.ac.uk
Brennan
F M
FM
D051722:ADAM Proteins; D000072198:ADAM17 Protein; D000942:Antigens, Bacterial / Q000494:pharmacology; D002384:Catalysis; D002478:Cells, Cultured; D005434:Flow Cytometry; D006801:Humans; D008070:Lipopolysaccharides / Q000494:pharmacology; D008666:Metalloendopeptidases / Q000378:metabolism / Q000502:physiology*; D018613:Microscopy, Confocal; D009000:Monocytes / Q000187:drug effects / Q000378:metabolism*; D009345:Neisseria meningitidis / Q000276:immunology; D018805:Sepsis / Q000378:metabolism*; D012995:Solubility; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*; D015054:Zymosan / Q000494:pharmacology
D000942:Antigens, Bacterial; D008070:Lipopolysaccharides; D014409:Tumor Necrosis Factor-alpha; D015054:Zymosan; D051722:ADAM Proteins; D008666:Metalloendopeptidases; D000072198:ADAM17 Protein; C000606462:ADAM17 protein, human