It is debated whether human T cells of heterologous specificity, known as bystander T cells, are activated independently of the TCR during in vivo immune responses. Bangs et al. (p. 1962) developed an in vitro system to address the possibility that bystander activation could occur in the absence of TCR cross-reactivity. Human PBMC were stimulated with the superantigen staphylococcal enterotoxin B (SEB), which could directly activate Vβ17+ T cells but not Vβ13.1+ T cells. However, the Vβ13.1+ T cells were activated in these cultures through a mechanism requiring soluble mediators that did not include IL-15 and IL-23. TCR down-regulation was not observed in these bystander T cells, further supporting the idea that their activation did not involve TCR stimulation. Within the Vβ13.1+ T cell population, CD45RO+ memory T cells were preferentially activated following SEB stimulation. These bystander-activated T cells demonstrated a distinct gene expression profile compared with directly activated and resting T cells. Interestingly, while directly activated T cells up-regulated genes involved in the intrinsic pathway of apoptosis, bystander activated T cells specifically up-regulated genes involved in the extrinsic pathway, suggesting that these cells preferentially underwent death receptor-mediated apoptosis. These data indicate that human T cells can undergo bystander activation and that this process may preferentially eliminate memory T cells.

Lactate is elevated in inflamed tissues and in patients with obesity, hypertension, and/or type 2 diabetes and has been shown to induce insulin resistance. Based on previous studies indicating that lactate could enhance LPS-stimulated inflammatory gene expression, Samuvel et al. (p. 2476) hypothesized that lactate might promote TLR4 activation, which has been suggested to play a role in insulin resistance. Lactate and LPS were found to synergistically induce proinflammatory gene expression, NF-κB transcriptional activity, and expression of the TLR4 co-receptor MD-2, but not of TLR4 itself, in human macrophages. This enhancement of MD-2 expression was required for lactate-mediated augmentation of LPS-induced IL-6 expression. Antioxidants and inhibitors of monocarboxylate transporters (MCTs) inhibited the effects of lactate on LPS-mediated IL-6 production, NF-κB activity, and MD-2 expression, suggesting that the lactate-LPS synergism involved reactive oxygen species and required lactate transport through MCTs. This demonstration of a mechanism by which lactate augments TLR4-mediated inflammation has important implications for the understanding of diseases as wide-ranging as diabetes, sepsis, and atherosclerosis.

As γδ T cells have immune modulatory functions, a clear understanding of the requirements for their maintenance in vivo could enhance their utility in immunotherapeutic applications. MHC molecules are required for αβ T cell homeostasis, but the role of MHC in γδ T cells is not well defined. French et al. (p. 1892) compared the homeostatic expansion of different subsets of γδ T cells in a lymphopenic environment and found that CD8+ γδ T cells proliferated rapidly, whereas NK1.1+ γδ T cells were maintained at relatively low numbers. This apparent advantage of CD8+ γδ T cells was not linked to expression of cytokine receptors nor of the antiapoptotic molecule bcl-2 but could be shown to correlate with expression of Vγ4 and Vδ5, suggesting a role for the γδ TCR in homeostasis. Interestingly, γδ T cell homeostatic expansion, particularly that of NK1.1+ γδ T cells, was enhanced in mice lacking β2-microglobulin (β2m), suggesting that β2m-associated molecules inhibited γδ T cell expansion. Negative regulation of the NK1.1+ γδ T cell subset was not mediated by the NK cell receptors NKG2A or Ly49C. Although the mechanism of β2m-mediated inhibition remains to be defined, the data suggested that the proliferative disadvantage of NK1.1+ γδ T cells could be caused by β2m-associated molecules. Thus, CD8+ and NK1.1+ γδ T cells are differentially regulated during homeostatic expansion, which, in contrast to αβ T cell homeostasis, is impaired in the presence of β2m-associated molecules.

Flavocytochrome b558, as the catalytic core of the phagocytic NADPH oxidase, is responsible for generating the superoxide that is vital for effective host defense. Very little is known about the subcellular distribution of this complex in macrophages, so Casbon et al. (p. 2325) fluorescently tagged the subunits that make up flavocytochrome b558, p22phox, and gp92phox, and examined their localization and trafficking in macrophages. Initial experiments in Chinese hamster ovary cells showed that the individual subunits were localized to the endoplasmic reticulum and trafficked to the plasma membrane upon heterodimer formation. Some flavocytochrome b558 was also observed to recycle from the plasma membrane to the Rab11-expressing perinuclear endocytic recycling compartment. This localization pattern was also observed in primary macrophages and in a macrophage cell line, where flavocytochrome b558 was also found in Rab5-expressing early endosomes. Analysis of flavocytochrome b558 trafficking during phagocytosis suggested that the recycling endosomes might transport flavocytochrome b558 between intracellular membranes and nascent phagosomes. This delineation of flavocytochrome b558 localization may be useful for future studies aimed at the regulation of superoxide production during infection.

Superantigens from Streptococcus pyogenes, including streptococcal pyrogenic exotoxin A (SPE A), and Staphylococcus aureus, including toxic shock syndrome toxin-1 (TSST-1), are involved in the development of inflammatory diseases such as toxic shock syndrome. To better understand the disruption of mucosal tissues that occurs in streptococcal and staphylococcal infections, Brosnahan et al. (p. 2364) analyzed the abilities of these superantigens, together with cytolysins from the same bacterial species, to cross mucosal barriers and directly affect epithelial cell cytokine production. Both cytolysins analyzed, α toxin from S. aureus and streptolysin O (SLO) from S. pyogenes, damaged the vaginal mucosal surface and promoted superantigen penetration of vaginal mucosa. TSST-1, α toxin, and SPE A also induced strong proinflammatory cytokine responses from vaginal epithelial cells, whereas SLO did not. However, SLO directly and rapidly killed vaginal epithelial cells and promoted penetration of S. pyogenes through the epithelium, whereas α toxin did not promote penetration of S. aureus. The authors propose that cytolysins from different species may promote superantigen penetration of mucosal epithelia either by direct damage to the mucosa or by indirect disruption of the mucosal epithelium through the induction of proinflammatory cytokines.

Heme oxygenase-1 (HO-1) can protect against immune activation in transplantation and autoimmunity and has previously been shown to inhibit dendritic cell (DC) maturation and proinflammatory functions. To address the mechanism by which HO-1 might inhibit DC activity, Rémy et al. (p. 1877) assessed the effects of the heme degradation end product carbon monoxide (CO) on DC function. CO, but no other products of HO-1 activity, inhibited the LPS-induced maturation, proinflammatory cytokine production, and allostimulatory capacity of human DCs but did not affect their IL-10 production. HO-1- or CO-mediated inhibition of DC maturation was independent of p38 MAPK, ERK, and JNK signaling. However, HO-1 and CO inhibited LPS-mediated IFN regulatory factor (IRF)-3 activation in human DCs, and HO-1 also inhibited LPS-mediated NF-κB activation. In a model of diabetes induction requiring DC adoptive transfer, transferred DCs treated with CO or induced to express HO-1 were unable to induce diabetes development, indicating that HO-1 and CO impaired the in vivo immunogenicity of DCs. Taken together, these data provide insight into the HO-1-mediated modulation of DC immunogenicity and suggest that CO may promote the development of tolerogenic DCs.

Type I IFN production is important in the host response to both RNA and DNA virus infections and is induced by the activation of IFN regulatory factor (IRF) 3 and IRF7. These transcription factors are phosphorylated by kinases including TANK-binding kinase 1 (TBK1) and inducible IκB kinase (IKK-i), which have been shown to play redundant roles in RNA virus infection. To determine the relative roles of TBK1 and IKK-i in DNA virus infection, Miyahira et al. (p. 2248) examined macrophages and mice lacking TBK1. In vitro experiments demonstrated that TBK1 was required for IFN responses to DNA virus infections, intracellular B-DNA, and TLR4 stimulation. In TBK1−/−IKK-i−/− fibroblasts, reconstitution of TBK1 rescued DNA-induced STAT1 phosphorylation to a greater degree than did IKK-i reconstitution, leading the authors to suggest that these two kinases may have functionally distinct roles in the IFN response to DNA virus infection. IFN production in response to DNA virus infection was further shown to be TBK1/IRF3-dependent in macrophages but TBK1-independent in plasmacytoid dendritic cells. In vivo infection of IRF3−/− and TBK1−/− mice with murine gammaherpesvirus-68 further supported a requirement for the TBK1-IRF3 signaling pathway in host resistance to DNA viruses. Taken together, these data indicate a critical requirement for TBK1 in the macrophage response to DNA virus infection and may provide future avenues for therapeutic manipulation of the host IFN response.

The calcium-binding proteins S100A8 and S100A9 are proposed to act as damage-associated molecular pattern molecules, and their expression is associated with the progression of HIV and other viral infections. Endoh et al. (p. 2258) examined the mechanism of S100 protein up-regulation in viral infection and found that synthetic dsRNA induced S100A8 expression in a murine macrophage cell line and induced both S100A8 and S100A9 in human monocytes and macrophages. This up-regulation occurred via a novel IL-10- and protein kinase R-dependent pathway and was enhanced by IFN-β. S100A8 induction by dsRNA, like its induction by LPS, required signaling via p38 and ERK. Analysis of transcriptional regulation of the gene encoding S100A8 determined that the transcription factor C/EBPβ bound to the promoter region following dsRNA stimulation. Finally, in vivo infection with influenza A virus induced S100A8 expression in the lung, which peaked at day 8 of infection and then declined as the mice recovered. These data help explain the up-regulation of S100 proteins in human viral infection and, as IL-10 is required for this up-regulation and is associated with viral persistence, suggest that S100A8 and S100A9 may contribute to the persistence of RNA viruses.

Summaries written by Jennifer Hartt Meyers, Ph.D.