Endogenous signals released from necrotic cells augment inflammatory responses to bacterial endotoxin

Abstract

Stressed cells undergoing necrosis release molecules that acts as endogenous danger signals to alert and activate innate immune cells. Both HMGB1 and HSP70 are induced in activated monocytes/macrophages and also are released from stressed or injured cells. We investigated whether HMGB1 and HSP70 released from necrotic monocytes/macrophages, can act as danger signals to mediate proinflammatory cytokine responses to bacterial endotoxin or lipopolysaccharide (LPS). We show that cell lysate, obtained from necrotic cells directly stimulates the proinflammatory cytokine and chemokine responses in human monocyte/ macrophage cell line, THP-1, as revealed by the induction of TNF-α, IL-6 and IL-8 mRNA expression and protein production. In the presence of LPS, necrotic cell lysate induced a more robust increase in all three proteins. We found that HMGB1 and HSP70 were indeed present in the necrotic cell lysate and were responsible for the significant induction of the proinflammatory cytokine expression, as neutralization with antibodies against both proteins blocked the increase in the cytokine production seen after incubating LPS-stimulated cells with the necrotic cell lysate. We also found that the newly identified triggering receptor expressed on myeloid cells-1 (TREM-1) was involved in mediating the HMGB1-and HSP70-induced cytokine production. Blocking TREM-1 on THP-1 cells with a recombinant chimera prevented the increase in cytokine production, while simultaneous blocking of TLR4 and TREM-1 completely abolished the proinflammatory response, suggesting that TREM-1 synergizes with TLR4 to mediate the effects of such signals from necrotic cells. In addition, blocking HMGB1 or HSP70 simultaneously with TREM-1 did not decrease the cytokine level further, confirming the involvement of TREM-1 in mediating the effect of HMGB1 and HSP70. Although the interaction of HMGB1 and HSP70 with TREM-1 induced IκBα and p38 expression, both of which are required for the inflammatory cytokine expression, blockade of TREM-1 did not affect IκBα expression but markedly reduced p38 activation, as revealed by Western blot analysis. Together, these results demonstrate that HMGB1 and HSP70 released from necrotic cells function as endogenous danger signals to augment the proinflammatory responses in monocytes/macrophage and that TREM-1 relays such signals to the cytokine expression cascade. This mechanism may contribute to the amplification and persistence of the inflammatory response to bacterial infection.

Conditional ablation of HMGB1 in mice reveals its protective function against endotoxemia and bacterial infection

“…There has been a particular focus on HMGB1 in the context of LPS-induced endotoxemia, wherein HMGB1 released by myeloid cells is reported to be capable of activating various innate receptors, especially TLR4, and thereby exacerbating pathogenic inflammatory responses (11,27). We therefore first examined whether and how LysM Cre/+ -Hmgb1 f/f mice respond to LPS.…”

Involvement of the TREM-1/DAP12 pathway in the innate immune responses to Porphyromonas gingivalis

“…TREM-1 activation and expression occurs synergistically with TLRs (Arts et al, 2011;Klesney-Tait and Colonna, 2007;Klesney-Tait et al, 2006), as the TREM family contains both inhibitory and activating receptors capable of modulating the signaling downstream of TLRs (Allcock et al, 2003;Ornatowska et al, 2007). Moreover, TREM-1 has also been associated with NOD-Like Receptors (NLR), responsible for sensing microbial danger signals and amplifying the inflammatory response (El Mezayen et al, 2007;Netea et al, 2006). On the molecular level, TREM-1 regulates immune cell function, by forming an intracellular complex with signalling adapter DNAX activating protein of 12 kDa (DAP12) (Bouchon et al, 2000;Colonna and Facchetti, 2003;Ford and McVicar, 2009;Tomasello and Vivier, 2005).…”

Strong renal expression of heat shock protein 70, high mobility group box 1, inducible nitric oxide synthase, and nitrotyrosine in mice model of severe malaria

“…Thus, it appears that the maximum expression of iNOS and other markers occurs at approximately 3 days after Plasmodium infection. Although no exponential increases were observed in the expression values, it is also possible that the expressions will continue to increase, unlike the degree of parasitemia 29 . Furthermore, the degree of parasitemia is a weak predictor of malaria severity and mortality, as it is not related to the levels of inflammation and organ damage.…”

“…During the inflammatory process, HMGB1 expression is associated with tissue damage (i.e., a damage-associated molecular protein) and is influenced by abiotic stressors, such as heat 28 . In addition, Mezayen et al found that triggering expressed on myeloid cells-1 (TREM-1) is an HMGB-1 mediator that induces cytokine production, which suggests that HMGB-1 is released as an endogenous danger signal that induces the pro-inflammatory response of macrophages and leads to exacerbated kidney damage 29 . However, HSP70 expression increases in the presence of pathogenic stressors, such as heat, ischemia, and elevate levels of pro-inflammatory cytokines that are induced by the expression HMGB1.…”