SUBSTANCE-P INDUCES TUMOR-NECROSIS-FACTOR IN AN EX-VIVO MODEL SYSTEM

Many studies have shown that tumor necrosis factor (TNF) is a potent s oluble mediator of immunoregulation and inflammation. Neuropeptide sub stance P (SP) has been known to exert significant influence on product ion of certain inflammatory cytokine by immune cells. Immunopathogenic mechanism underlying the effect of neuropeptide substance P (SP) and the specific amino acid sequence of SP that induces TNF has not been c learly studied. Employing ex vivo and in vitro model systems, we inves tigated the direct effect of different sequences of SP on TNF secretio n by whole blood and separated total mononuclear cells. Aliquots of bl ood samples (1 ml) or Ficoll - Hypaque-separated total mononuclear cel ls (1 x 10(6)/ml) were cultured with different concentrations of SP an d its sequences (SP 1-4, SP 4-11) or vasoactive intestinal peptide (VI P) for 24 hr at 37 degrees C. Plasma samples and culture supernatants were assayed for TNF levels in a bioassay using a TNF-sensitive WEHI 1 64 subclone 13 cell line. Plasma from blood samples or lymphocytes tre ated with whole SP and SP 4-11 at 10(-7), 10(-8), and 10(-9) M concent rations induced significant production of TNF compared to negligible l evels, of TNF produced by SP 1-4-treated and untreated cultures. VIP a t all concentrations tested did not induce TNF production and was simi lar to untreated control cultures. Separated mononuclear cells also pr oduced significant levels of TNF in response to SP and SP 4-11. Anti-T NF-alpha antibodies neutralized the TNF induced by SP 4-11 in plasma. These studies suggest that an ex vivo system using whole blood may be an ideal model to study the effects of SP on TNF production. These stu dies also demonstrated that the TNF inducing activity of SP resides in the region containing amino acids 4 to 11. (C) 1995 Academic Press, I nc.