Microbiota ablation resulted in reduced rotavirus infection/diarrhea and a more durable rotavirus antibody response, suggesting that antibiotic administration before rotavirus vaccination could raise low seroconversion rates that correlate with the vaccine's inefficacy in developing regions.

Activators of innate immunity may have the potential to combat a broad range of infectious agents. We report that treatment with bacterial flagellin prevented rotavirus (RV) infection in mice and cured chronically RV-infected mice. Protection was independent of adaptive immunity and interferon (IFN, type I and II) and required flagellin receptors Toll-like receptor 5 (TLR5) and NOD-like receptor C4 (NLRC4). Flagellin-induced activation of TLR5 on dendritic cells elicited production of the cytokine interleukin-22 (IL-22), which induced a protective gene expression program in intestinal epithelial cells. Flagellin also induced NLRC4-dependent production of IL-18 and immediate elimination of RV-infected cells. Administration of IL-22 and IL-18 to mice fully recapitulated the capacity of flagellin to prevent or eliminate RV infection and thus holds promise as a broad-spectrum antiviral agent.

CD137-mediated signals costimulate T cells and protect them from activation-induced apoptosis; they induce curative antitumor immunity and enhance antiviral immune responses in mice. In contrast, anti-CD137 agonistic mAbs can suppress T-dependent humoral immunity and reverse the course of established autoimmune disease. These results have provided a rationale for assessing the therapeutic potential of CD137 ligands in human clinical trials. In this study, we report that a single 200-μg injection of anti-CD137 given to otherwise naive BALB/c or C57BL/6 mice led to the development of a series of immunological anomalies. These included splenomegaly, lymphadenopathy, hepatomegaly, multifocal hepatitis, anemia, altered trafficking of B cells and CD8 T cells, loss of NK cells, and a 10-fold increase in bone marrow (BM) cells bearing the phenotype of hemopoietic stem cells. These events were dependent on CD8 T cells, TNF-α, IFN-γ, and type I IFNs. BM cells up-regulated Fas, and there was a significant increase in the number of CD8+ T cells that correlated with a loss of CD19+ and Ab-secreting cells in the BM. TCR Vαβ usage was random and polyclonal among liver-infiltrating CD8 T cells, and multifocal CD8+ T cell infiltrates were resolved upon termination of anti-CD137 treatment. Anti-CD137-treated mice developed lymphopenia, thrombocytopenia, and anemia, and had lowered levels of hemoglobin and increased numbers of reticulocytes.

High-risk human papillomaviruses (HPVs) (e.g., HPV-16) cause anogenital and head and neck cancers, and low-risk HPVs (e.g., HPV-6) cause benign hyperproliferative disease. The E7 protein of HPV-16 binds all retinoblastoma tumor suppressor protein (pRB) family members with higher affinity than HPV-6E7. The HPV-16 E7 protein has been reported to target pRB family members for degradation and to immortalize cells. Here we tested the hypothesis that the low-risk E7 protein has an intrinsic ability to decrease expression of pRB family members. First, we introduced a high-affinity pRBbinding site into HPV-6 E7 (6E7G22D) and showed that, in human foreskin keratinocytes, HPV-6 E7G22D decreased the level of pRB protein but not pRB mRNA. Second, we analyzed the ability of wild-type HPV-6 E7 to destabilize the other pRB family members, p107 and p130. HPV-6 E7, like HPV-16 E7, decreased the level of p130 protein. This decrease was inhibited by MG132, a proteasome inhibitor. Binding of HPV-6 E7 to p130 was necessary but not sufficient to decrease the level of p130. Furthermore, the destabilization of p130 correlated with a decrease in the expression of involucrin, a differentiation marker. We suggest that the shared activity of HPV-16 E7 and HPV-6 E7 to destabilize p130 and decrease or delay differentiation may be related to the role of E7 in the HPV life cycle. The added ability of HPV-16 E7 to regulate pRB and p107 may be related to oncogenic activity.keratinocytes ͉ RB family members ͉ human papillomaviruses ͉ differentiation H uman papillomaviruses (HPVs) are nonenveloped viruses that contain an Ϸ8,000-bp circular DNA genome. HPVs infect mucosal and cutaneous stratified squamous epithelia and are divided into high-risk and low-risk viruses based on their pathogenicity (1). The low-risk viruses, e.g., HPV-6 and HPV-11, mainly cause benign hyperproliferative disease, including the most prevalent viral sexually transmitted disease, condyloma acuminata or genital warts. The high-risk viruses, e.g., HPV-16, HPV-18, and HPV-31, are normally found in malignant tumors, including cervical cancer and some head and neck cancers (1). The replication cycle of all HPVs is differentiation-dependent (1, 2). The virus presumably enters through a break in the epithelium and initially infects the basal cells, which are the proliferating epithelial cells (keratinocytes). In these cells, the virus undergoes the nonproductive stage of its life cycle, where it is maintained as a low-copy-number episome. When the infected cell moves to the suprabasal compartment, both the high-risk and low-risk viruses undergo the productive phase of their life cycle: amplifying their DNA, synthesizing structural proteins, and producing infectious virus. Uninfected cells in this suprabasal compartment normally have exited the cell cycle and begun to differentiate. Because HPVs require the host cell DNA replication machinery to replicate, the viral DNA must encode proteins able to generate an intracellular environment within this differentiated compartment approp...

BACKGROUND & AIMS Nanoparticles have been explored as carriers of small interfering RNAs (siRNAs), and might developed to treat inflammatory bowel disease (IBD). Overexpression of CD98 on the surface of colonic epithelial cells and macrophages promotes development and progression of IBD. We developed an orally delivered hydrogel that releases nanoparticles with single-chain CD98 antibodies on their surface (scCD98-functionalized) and loaded with CD98 siRNA (siCD98). We tested the ability of the nanoparticles to reduce levels of CD98 in colons of mice with colitis. METHODS scCD98-functionalized siCD98-loaded nanoparticles were fabricated using a complex coacervation technique. We investigated the cellular uptake and lysosome escape profiles of the nanoparticles in Colon-26 cells and RAW 264.7 macrophages using fluorescence microscopy. Colitis was induced by transfer of CD4+CD45RBhigh T cells to Rag−/− mice or administration of dextran sodium sulfate to C57BL/6 mice. Mice were then given hydrogel (chitosan and alginate) containing scCD98-functionalized nanoparticles loaded with siCD98 or scrambled siRNA (control) via gavage. RESULTS The scCD98-functionalized nanoparticles were approximately 200 nm in size and had high affinity for CD98-overexpressing cells. The scCD98-functionalized siCD98-loaded nanoparticles significantly reduced levels of CD98 in Colon-26 cells and RAW 264.7 macrophages, along with production of inflammatory cytokines (TNFα, IL6, and IL12). In mice with colitis, administration of the scCD98-functionalized siCD98-loaded nanoparticles reduced colon expression of CD98. Importantly, the severity of colitis was also reduced, compared with controls (based on loss of body weight, myeloperoxidase activity, inflammatory cytokine production, and histologic analysis). Approximately 24.1% of colonic macrophages (CD11b+CD11c−F4/80+) in the mice had taken up fluorescently labeled siRNA-loaded nanoparticles within 12 hr of administration. CONCLUSIONS Nanoparticles containing surface CD98 antibody and loaded with siCD98 reduce expression of this protein by colonic epithelial cells and macrophages; oral administration decreases the severity of colitis in mice. This nanoparticle in hydrogel (chitosan/alginate) formulation might be developed to treat patients with IBD.

Highlights d Some mouse colonies developed spontaneous resistance to rotavirus (RV) infection d Fecal microbial transplantation transfers RV resistance d Protection against RV was mediated by segmented filamentous bacteria (SFB) d SFB impeded RV by increasing epithelial cell turnover

Various states of inflammation, including sepsis, are associated with hypoferremia, which limits iron availability to pathogens and reduces iron-mediated oxidative stress. Lipocalin 2 (siderocalin, 24p3) plays a central role in iron transport. Accordingly, Lipocalin 2-deficient mice (Lcn2KO) exhibit elevated intracellular labile iron. Here we report that LPS-induced systemic Lcn2 by 150 fold in WT mice at 24h. Relative to WT littermates, Lcn2KO mice were markedly more sensitive to endotoxemia, exhibiting elevated indices of organ damage (transaminasaemia, LDH) and increased mortality. Such exacerbated endotoxemia was associated with substantially increased caspase 3 cleavage and concomitantly elevated immune cell apoptosis. Further, cells from Lcn2KO mice were hyperresponsive to LPS ex vivo exhibiting elevated cytokine secretion. In addition, Lcn2KO mice exhibited delayed LPS-induced hypoferremia despite normal hepatic hepcidin expression and display decreased levels of the tissue redox state indicators cysteine and glutathione in liver and plasma. Desferroxamine, an iron chelator, significantly protects Lcn2KO mice from LPS-induced toxicity, including mortality, suggesting that Lcn2 may act as an antioxidant in vivo by regulating iron homeostasis. Thus, Lcn2-mediated regulation of labile iron protects the host against sepsis. Its small size and simple structure may make Lcn2 as a deployable treatment for sepsis.

BACKGROUND & AIMS Lipocalin 2 (Lcn2) is a multifunctional innate immune protein whose expression closely correlates with extent of intestinal inflammation. However, whether Lcn2 plays a role in the pathogenesis of gut inflammation is unknown. Herein, we investigated the extent to which Lcn2 regulates inflammation and gut bacterial dysbiosis in mouse models of IBD. METHODS Lcn2 expression was monitored in murine colitis models and upon microbiota ablation/restoration. WT and Lcn2 knockout (Lcn2KO) mice were analyzed for gut bacterial load, composition by 16S rRNA gene pyrosequencing and, their colitogenic potential by co-housing with Il-10KO mice. Acute (dextran sodium sulfate) and chronic (IL-10R neutralization and T-cell adoptive transfer) colitis was induced in WT and Lcn2KO mice with or without antibiotics. RESULTS Lcn2 expression was dramatically induced upon inflammation and was dependent upon presence of a gut microbiota and MyD88 signaling. Use of bone-marrow chimeric mice revealed non-immune cells are the major contributors of circulating Lcn2. Lcn2KO mice exhibited elevated levels of entA-expressing gut bacteria burden and, moreover, a broadly distinct bacterial community relative to WT littermates. Lcn2KO mice developed highly colitogenic T-cells and exhibited exacerbated colitis upon exposure to DSS or neutralization of IL-10. Such exacerbated colitis could be prevented by antibiotic treatment. Moreover, exposure to the microbiota of Lcn2KO mice, via cohousing, resulted in severe colitis in Il-10KO mice. CONCLUSION Lcn2 is a bacterially-induced, MyD88-dependent, protein that play an important role in gut homeostasis and a pivotal role upon challenge. Hence, therapeutic manipulation of Lcn2 levels may provide a strategy to help manage diseases driven by alteration of the gut microbiota.