TLR2 and TLR4 signaling pathways are required for recombinant Brucella abortus BCSP31-induced cytokine production, functional upregulation of mouse macrophages, and the Th1 immune response in vivo and in vitro

Abstract

Brucella abortus is a zoonotic Gram-negative pathogen that causes brucelosis in ruminants and humans. Toll-like receptors (TLRs) recognize Brucella abortus and initiate antigen-presenting cell activities that affect both innate and adaptive immunity. In this study, we focused on recombinant Brucella cell-surface protein 31 (rBCSP31) to determine its effects on mouse macrophages. Our results demonstrated that rBCSP31 induced TNF-a, IL-6 and IL-12p40 production, which depended on the activation of mitogen-activated protein kinases (MAPKs) by stimulating the rapid phosphorylation of p38 and JNK and the activation of transcription factor NF-kB in macrophages. In addition, continuous exposure (.24 h) of RAW264.7 cells to rBCSP31 significantly enhanced IFN-c-induced expression of MHC-II and the ability to present rBCSP31 peptide to CD4 1 T cells. Furthermore, we found that rBCSP31 could interact with both TLR2 and TLR4. The rBCSP31-induced cytokine production by macrophages from TLR2 2/2 and TLR4 2/2 mice was lower than that from C57BL/6 macrophages, and the activation of NF-kB and MAPKs was attenuated in macrophages from TLR2 2/2 and TLR4 2/2 mice. In addition, CD4 1 T cells from C57BL/6 mice immunized with rBCSP31 produced higher levels of IFN-c and IL-2 compared with CD4 1 T cells from TLR2 2/2 and TLR4 2/2 mice. Macrophages from immunized C57BL/6 mice produced higher levels of IL-12p40 than those from TLR2 2/2 and TLR4 2/2 mice. Furthermore, immunization with rBCSP31 provided better protection in C57BL/6 mice than in TLR2 2/2 and TLR4 2/2 mice after B. abortus 2308 challenge. These results indicate that rBCSP31 is a TLR2 and TLR4 agonist that induces cytokine production, upregulates macrophage function and induces the Th1 immune response.

Leptospira surface adhesin (Lsa21) induces Toll like receptor 2 and 4 mediated inflammatory responses in macrophages

“…Although most lipoproteins signal via TLR2, but our data showed that rLsa21 signals via both TLR2 and TLR4. This finding is consistent with various previous reports demonstrating the production of proinflammatory cytokines and chemokines through TLR2/4 by Leptospira hemolysins23, whole Leptospira or their purified components (LPS, membrane proteins) but also proteins from other bacterial pathogens like Brucella and Mycobacterium 38394041. Our results showed that rLsa21 induced cytokine production mainly through MYD88 dependent signaling pathway as there was a reduced concentration of cytokines (IL-6 and TNF-α) in MyD88 −/− macrophage cell lines (Fig.…”

“…It is important to note that different PAMPs from various pathogens may activate different signaling pathways for production of proinflammatory cytokines. For instance, JNK and NF-κB signaling pathway is activated by Leptospira hemolysins23, p38 and JNK is activated by BCSP31 of Brucella 40 and MPT83 of M. tuberculosis 39, NF-κB by streptococcal hemolysins45 and p38 and NF-κB by streptolysin O and Listeriolysin4647.…”

Cellular and molecular regulation of innate inflammatory responses

“…3,4 TLRs are type I transmembrane molecules which transduce their downstream signaling through the MyD88-dependent pathway or the MyD88-independent but TRIF-dependent pathway, subsequently leading to activation of mitogenactivated protein kinase (MAPK), NF-κB and IRF pathway, inducing the production of proinflammatory cytokines and IFNs (Figure 1). [5][6][7][8] Innate immune activation of phagocytes through TLRs also induces an Mst1-Mst2-Rac signaling axis to activate intracellular microbicidal killing. 9,10 RLRs, a family of cytoplasmic RNA helicases, are essential for innate recognition of viruses and are the key mechanism for the control of viral replication and dissemination.…”

Brucella abortus L7/L12 ve BCSP31 Proteinlerini Birlikte Eksprese Eden Rekombinant Adenovirusların İmmunojenitesi

“…Another promising immunodominant protein specific to Brucella is a 31 kDa protein, BCSP31. Its immune response-inducing properties are attributable to the presence of an immunogenic and protective BCSP fraction, possibly lipopolysaccharide [25] . A study indicated that the recombinant L7/L12-TOmp31 protein elicited stronger humoral and cellular immune responses and provided significant protection level against B. melitensis and B.…”