A comparative analysis of cytokine production and tolerance induction by bacterial lipopeptides, lipopolysaccharides and Staphyloccocus aureus in human monocytes

Abstract

SUMMARYMonocytes (MO) and macrophages (MAC ) are important producers of cytokines involved in the pathophysiology of bacterial sepsis. Most studies concentrate on the effects of bacterial lipopolysaccharides (LPS) regarding the induction of cytokine gene expression and secretion in MO/MAC. Here we report that besides LPS, the synthetic lipoprotein analogue lipopeptide N-palmitoyl-S-(2,3-bis(palmitoyl )-(2RS)-propyl )-(R)-cysteinyl-alanyl-glycine (Pam3-Cys-Ala-Gly), another component of the outer membrane of Gram-negative bacteria, as well as heat-killed Staphyloccocus aureus (S. aureus/SAC ) are potent stimuli for cytokines in human MO. For all three investigated stimuli we found an individual pattern of cytokine induction: LPS was most potent in inducing interleukin-6 (IL-6) synthesis, whereas for tumour necrosis factor-a ( TNF-a) secretion SAC was the best stimulus. Comparable amounts of IL-8 were induced by either LPS or Pam3-Cys-AlaGly, with SAC being less effective even at higher concentrations. The addition of serum led to an increase in LPS-, SAC-and Pam3-Cys-Ala-Gly-stimulated TNF-a secretion, indicating that the presence of serum is critical not just for LPS stimulation. Furthermore, as is known for LPS, Pam3-Cys-Ala-Gly and SAC rendered MO refractory to a second bacterial stimulus. Pam3-CysAla-Gly and SAC induced tolerance for itself, but LPS could partially overcome this effect. As the CD14 molecule is discussed as a common receptor for different bacterial components, we investigated whether the TNF-a response of MO could be blocked by anti-CD14 antibodies. MY4, a CD14 antibody, selectively blocked the TNF-a secretion induced by LPS but not by Pam3-Cys-Ala-Gly or SAC. In summary, we conclude that besides LPS, lipopeptide Pam3-CysAla-Gly and SAC are potent stimuli for human MO, while the mechanisms of activation seem to be partially different from LPS.

Synergy and Cross-Tolerance Between Toll-Like Receptor (TLR) 2- and TLR4-Mediated Signaling Pathways

“…Although the molecular mechanism of the synergistic action of MALP-2 and LPS is unknown, our present study indicates that the simultaneous activation of different TLRs could exert the synergistic effects. Indeed, bacterial lipopeptides and lipoprotein have been shown to induce inflammatory cytokine production in synergy with LPS (4,28). The mycoplasmal lipopeptide MALP-2 is structurally related to bacterial lipopeptides and lipoprotein, all of which have been shown recognized by TLR2 (29 -33).…”

“…These include bacterial lipopeptides, peptidoglycan, muramyl dipeptide from Gram-positive bacteria, and bacterial DNA containing unmethylated CpG motif. These bacterial components have also been shown to induce tolerance to the subsequent stimulation (2)(3)(4).…”

Toll-Like Receptor 2 (TLR2) and TLR9 Signaling Results in HIV-Long Terminal RepeatTrans-Activation and HIV Replication in HIV-1 Transgenic Mouse Spleen Cells: Implications of Simultaneous Activation of TLRs on HIV Replication

“…In in vitro and in vivo systems, costimulation with multiple PAMP is known to induce NF-B activation and inflammatory cytokine production synergistically (18,19). Costimulation of HIV-infected monocytic cell lines with TNF-␣ and IL-6 induces HIV expression in a synergistic manner (20 -24).…”

Coordinate activation of endogenous p38a, β, γ, and δ by inflammatory stimuli

“…Our results demonstrate that activation of the p38 pathway by HKSA is similar to that obtained with LPS, indicating the likelihood of the involvement of p38 in responses to both gram-positive as well as gram-negative bacteria. The role of CD14 in mediating activation of macrophages by gram-positive bacteria is still under debate [31][32][33][34][35]. The data presented herein provide additional support for CD14-dependent cell activation by HKSA.…”

“…Of the four agonists examined, HKSA induced the highest levels of p38 activation. It has been shown previously that HKSA is a potent stimulus for induction of cytokine synthesis in human monocytes [31][32][33] but the signal transduction pathways involved in this induction have not been well studied. Our results demonstrate that activation of the p38 pathway by HKSA is similar to that obtained with LPS, indicating the likelihood of the involvement of p38 in responses to both gram-positive as well as gram-negative bacteria.…”