Lymphoid tissue inducer (LTi) cells are recruited to the developing lymph node (LN) anlagen, where they interact with stromal cells to induce LN formation. To gain further insights into LN anlagen-associated stromal cell maturation preceding LTi involvement, Bénézech et al. (p. 4521) investigated the early formation of mouse embryonic inguinal LNs (iLNs). When embryo day (E) 13 and E15 iLNs were microdissected and analyzed, the iLN anlage appeared as a spherical shaped sac formed by endothelial cells and surrounded by layers of mesenchymal cells. Outer and inner cell layers remained separated until E17, when mesenchymally derived stromal cells invaded the inner core and began forming the internal compartments of the iLN anlage. Both iLN- and mesenteric LN-derived E15–E17 cells showed similar development patterns, with stromal cell maturation initiating prior to the completion of LN endothelium differentiation. ICAM-1 and VCAM-1 (I/V) expression levels proved descriptive for stromal cell maturation, as precursor IV mesenchymal cells became immature IintVint stromal cells, which then gave rise to IhighVhighMAdCAM-1+ stromal organizer cells. This final transition required lymphotoxin β receptor signaling and the arrival of LTi cells. These data provide insights into the ontogeny and role of stromal cells in the early stages of LN development.

Viral infection can dangerously exacerbate the symptoms of chronic obstructive pulmonary disease (COPD), a debilitating lung disease strongly correlated with exposure to cigarette smoke (CS). NK cells play an integral role in the antiviral immune response, and Motz et al. (p. 4460) hypothesized that exposure to CS “primes” the NK cell response to viral infection. No intrinsic differences were observed when unstimulated IFN-γ+NKp46+ lung leukocytes from filtered air (FA)- and CS-exposed mice were compared. However, stimulation with viral pathogen-associated molecular patterns (PAMPs) induced a greater percentage of CS-exposed leukocytes to become NKp46+ and produce elevated levels of IFN-γ, compared with FA-exposed leukocytes. These cells also produced greater levels of IFN-γ in response to cytokine stimulation and exhibited an increased capacity to degranulate. Enhanced NK responses were duration dependent, as they were observed only in mice exposed to CS longer than 2 wk and were lost following the cessation of CS exposure. These data reveal that NK cells chronically exposed to CS are “primed” for viral-induced activation and identify NK cells as a potential therapeutic target for the treatment of cigarette-induced COPD.

Single-chain trimer (SCT) DNA vaccine constructs encode single chimeric polypeptide chains that use short amino acid linkers to string together all the components of MHC class I–peptide complexes. The resultant expressed protein complexes typically exhibit more stable MHC–peptide interactions than do those of standard DNA vaccines. To craft a CD8+ T cell-specific SCT DNA vaccine against West Nile virus (WNV), Kim et al. (p. 4423) first identified the clinically relevant WNV envelope protein-derived peptide SVG9 as an immunodominant peptide in WNV-infected HLA-A2 transgenic mice. The SVG9 SCT construct included a short leader sequence, SVG9, human β2-microglobin, and the human class I HLA-A2 H chain. Vaccination of HLA-A2 transgenic mice with SVG9 SCT resulted in 27% of splenic CD8+ T cells being SVG9 specific, and of these cells ~20% produced IFN-γ when stimulated with SVG9 peptide. Of vaccinated mice lethally challenged with strain WNV-NY, only ~30% developed disease symptoms, ~25% became moribund, and the remaining mice appeared disease free. SVG9 SCT-vaccinated mice showed lower viral burdens in the brain than did control SCT-vaccinated mice and developed SVG9-specific memory CD8+ T cells. Taken together, these data provide proof-of-principle that SCT-based vaccines can modulate protection from viral challenge.

Studies have revealed a strong correlation between HIV disease progression and HLA class I genotypes. The population of the United States represents a multitude of genealogies; thus, its HIV-1 patient pool afforded John et al. (p. 4368) the means to investigate whether the frequency of HLA class I polymorphisms differed among racial/ethnic groups. Based on self-identified racial/ethnicity information, the study population was comparable to U.S. HIV prevalence estimates. In 97% of cases, HIV-1 sequences were identified as clade B. HLA diversity varied substantially across racial/ethnic groups: whereas 45% of HLA-A and 18.7% of HLA-B alleles were common to all racial/ethnic groups, the remaining HLA-A/B alleles were associated with defined racial/ethnic groups. To visualize convergences and divergences of HIV-1 adaptations to HLA alleles, a unique “immunoselection profile” was mapped for each individual. Comparative analysis of immunoselection profiles revealed that HLA-associated adaptation patterns of HIV-1 differentially distributed among racial/ethnic groups based on sequence similarities between “epitope-length” intervals of viral proteins. These data reveal that HLA allele frequencies differ significantly among various racial/ethnic groups and provide further evidence that HLA–HIV interactions shape the evolution of viral epitope diversity and pathogenicity.

The lipid lysophosphatidylcholine (LPC) is known to enhance neutrophil microbicidal activities, but the mechanisms of its effects are not defined. To gain further insights, Hong et al. (p. 4401) tested various inhibitors of neutrophil bactericidal activity and identified proteases as essential for LPC’s enhancing effects. In response to Gram-negative bacteria, LPC-treated neutrophils released greater amounts of elastase due to increased azurophil granule–phagosome fusion. Previously, a role for Cl flux through glycine receptors (GlyRs) in neutrophil bactericidal activity had been suggested. Human neutrophils were found to express GlyR subtype α2, and the GlyR antagonist strychnine blocked LPC-mediated enhancement of bacterial clearance. Strychnine treatment abrogated LPC enhancement of azurophil granule–phagosome fusion and LPC-mediated increases in Ca2+ flux as well. The latter was sensitive to the intracellular Cl-dependent regulation of a transient receptor potential channel, TRPM2. Finally, both Ca2+ influx and Cl influx were required for LPC-induced p38 MAPK phosphorylation and subsequent azurophil granule–phagosome fusion. Thus, the bactericidal-enhancing effects of LPC are dependent upon glycine-GlyRα2/TRMP2/p38 MAPK signaling. These findings reveal potential therapeutic targets for enhancing neutrophil bactericidal activity.

The collagen-induced arthritis (CIA) mouse model is commonly used to study the development of autoimmune arthritis. In a search for upregulated genes in inflamed joints during CIA development, Chen et al. (p. 4455) identified the recently described CXC chemokine CXCL14. To study the role of CXCL14 during CIA development, CXL14-overexpressing transgenic (Tg) mice were generated. Compared with wild-type mice, collagen II (CII)-immunized Tg mice showed accelerated onset of CIA and more severe disease. Normal percentages of lymphocytes, dendritic cells, and macrophages were observed in bone marrow, thymus, spleen, and lymph nodes (LNs) of unmanipulated Tg mice, but the draining (inguinal) LNs of CII-immunized Tg mice were markedly enlarged, compared with those of immunized wt mice, due to proportional increases in CD4+ and CD8+ T cell numbers. In response to CII restimulation, draining LN cells of immunized Tg mice exhibited increased proliferative responses and expressed significantly higher levels of IFN-γ than did immunized wild-type mice. Finally, CXCL14 overexpression correlated with increased levels of CII-specific Abs. These data reveal that overexpression of CXCL14 results in increased numbers of autoreactive T cells, greater IFN-γ production, and levels of autoantibodies that together may promote the development of CIA.

In vertebrate cells, proteosomes not only degrade peptides for disposal or to yield functional proteins but also to produce MHC class I peptides. To produce an altered repertoire of peptides, vertebrate cells can also substitute the three catalytic subunits of standard proteosomes with the subunits LMP2, MECL-1, and LMP7 to form immunoproteosomes. Recent reports suggest that immunoproteosomes may also have immune functions unrelated to Ag processing. To explore a possible role for immunoproteosomes in B cell homeostasis, Hensley et al. (p. 4115) compared splenic B cells of immunoproteosome subunit-deficient and wild-type mice. In LMP2−/− mice, B cell numbers were dramatically decreased. This effect appeared to be due to the expression of “mixed” immunoproteosomes, which included the standard proteosome subunit Delta. In response to inactivated influenza A (IAV) immunization, LMP2−/− mice mounted normal IgM responses but exhibited a block in isotype switching. This block was not B cell autonomous, as wild-type B cells adoptively transferred into LMP2−/− mice also failed to properly class switch. Moreover, LMP2−/− innate immune cells expressed subnormal cytokine levels in response to IAV. Although mixed immunoproteosomes appeared to have normal overall protein degradation activity, LMP2−/− B cells exhibited impaired NF-κB activation. Taken together, these data suggest a role for immunoproteosomes in the immune response to IAV that is distinct from Ag processing.

The alarmin high-mobility group box 1 (HMGB1) induces a chemotactic response through binding receptor for advanced glycation end-products (RAGE). Previous studies have implicated the NF-κB signaling pathway in HMGB1/RAGE-induced chemotaxis, but the specific NF-κB–inducing kinases involved remain undefined. In this issue, Penzo et al. (p. 4497) examined the roles of kinases IKKα and IKKβ in the NF-κB response to HMGB1. In response to HMGB1, primary mouse embryo fibroblasts (MEFs) derived from either IKKα or IKKβ conditional knockout mice failed to exhibit chemotaxis, whereas wild-type MEFs were normally responsive. Likewise, macrophages and neutrophils required both kinases to exhibit a chemotactic response. Moreover, in response to i.p. injection of HMGB1, both IKKα and IKKβ conditional knockout mice experienced reduced emigration of neutrophils into the peritoneal cavity, which resulted in less severe peritonitis, compared with wild-type mice. HMGB1/RAGE-induced upregulation of RAGE expression was found to be IKKβ-, but not IKKα-, dependent. In addition, RAGE overexpression rescued IKKβ-null MEFs, but not IKKα-null MEFs, from defective HMGB1-induced migration. Taken together, these results indicate that both IKKα and IKKβ are required for HMGB1-induced chemotaxis; further work is required to elucidate their differential contributions.

Summaries written by Meredith G. Safford, Ph.D.