The efficiency of a vaccine largely depends on the appropriate targeting of the innate immune system, mainly through prolonged delivery of antigens and immunomodulatory substances to professional antigen-presenting cells in the lymphoid environment. Particulate antigens, such as virus-like particles (VLP) induce potent immune responses. However, little is known about the relative importance of direct drainage of free antigen to lymph nodes (LN) versus cellular transport and the impact of particle size on the process. Here, we show that nanoparticles traffic to the draining LN in a size-dependent manner. Whereas large particles (500-2000 nm) were mostly associated with dendritic cells (DC) from the injection site, small (20-200 nm) nanoparticles and VLP (30 nm) were also found in LN-resident DC and macrophages, suggesting free drainage of these particles to the LN. In vivo imaging studies in mice conditionally depleted of DC confirmed the capacity of small but not large particles to drain freely to the LN and demonstrated that DC are strictly required for transport of large particles from the injection site to the LN. These data provide evidence that particle size determines the mechanism of trafficking to the LN and show that only small nanoparticles can specifically target LN-resident cells.

Defect configuration may have an impact on the clinical outcome following surgical regenerative therapy of peri-implantitis lesions. While Class Ie defects seem to be promising in conjunction with NBM+CM, Class Ib and Class Ic may be considered as unfavourable.

Three major subsets of Ag-experienced CD8+ T cells have been identified according to their expression of CD62L and CD127. These markers are associated with central memory T cells (CD62L+CD127+), effector memory T cells (CD162L−CD127+), and effector T cells (CD62L−CD127−). In this study we characterized the development of these three populations during acute and chronic viral infections and after immunization with virus-like particles and determined their lineage relation and functional and protective properties. We found that the balance between the three subsets was critically regulated by the availability of Ag and time. After initial down-regulation of CD127, the responding CD8+ T cell population down-regulated CD62L and re-expressed CD127. Dependent on Ag availability, the cells then further differentiated into CD62L−CD127− effector cells or, in the absence of Ag, re-expressed CD62L to become central memory T cells. Although all three populations efficiently produced effector cytokines such as IFN-γ, CD62L−CD127− effector cells exhibited the highest ex vivo lytic potential. In contrast, CD62L+CD127+ central memory T cells most efficiently produced IL-2 and proliferated extensively in vitro and in vivo upon antigenic restimulation. Strikingly, only effector and effector memory, but not central memory, T cells were able to protect against peripheral infection with vaccinia virus, whereas central memory T cells were most potent at protecting against systemic infection with lymphocytic choriomeningitis virus, indicating that the antiviral protective capacities of specific CD8+ T cell subsets are closely related to the nature of the challenging pathogen.

DNA rich in nonmethylated CG motifs (CpGs) greatly facilitates induction of immune responses against coadministered Ags. CpGs are therefore among the most promising adjuvants known to date. Nevertheless, CpGs are characterized by two drawbacks. They have unfavorable pharmacokinetics and may exhibit systemic side effects, including splenomegaly. We show in this study that packaging CpGs into virus-like particles (VLPs) derived from the hepatitis B core Ag or the bacteriophage Qβ is a simple and attractive method to reduce these two problems. CpGs packaged into VLPs are resistant to DNase I digestion, enhancing their stability. In addition, and in contrast to free CpGs, packaging CpGs prevents splenomegaly in mice, without affecting their immunostimulatory capacity. In fact, vaccination with CpG-loaded VLPs was able to induce high frequencies of peptide-specific CD8+ T cells (4–14%), protected from infection with recombinant vaccinia viruses, and eradicated established solid fibrosarcoma tumors. Thus, packaging CpGs into VLPs improves both their immunogenicity and pharmacodynamics.

IL‐2 is a cytokine with multiple and even divergent functions; it has been described as a key cytokine for in vitro T cell proliferation but is also essential for down‐regulating T cell responses by inducing activation‐induced cell death as well as regulatory T cells. The in vivo analysis of IL‐2 function in regulating specific T cell responses has been hampered by the fact that mice deficient in IL‐2 or its receptors develop lymphoproliferative diseases and/or autoimmunity. Here we generated chimeric mice harboring both IL‐2R‐competent and IL‐2R‐deficient T cells and assessed CD8+ T cell induction, function and maintenance after acute or persistent viral infections. Induction and maintenance of CD8+ T cells were relatively independent of IL‐2R signaling during acute/resolved viral infection. In marked contrast, IL‐2 was crucial for secondary expansion of memory CD8+ T cells and for the maintenance of virus‐specific CD8+ T cells during persistent viral infections. Thus, depending on the chronicity of antigen exposure, IL‐2R signaling is either essential or largely dispensable for induction and maintenance of virus‐specific CD8+ T cell responses.

Inhibitory molecules derived from CNS myelin and glial scar tissue are major causes for insufficient functional regeneration in the mammalian CNS. A multitude of these molecules signal through the Rho/Rho kinase (ROCK) pathway. We evaluated three inhibitors of ROCK, Y-27632, Fasudil (HA-1077), and Dimethylfasudil (H-1152), in models of neurite outgrowth in vitro. We show, that all three ROCK inhibitors partially restore neurite outgrowth of Ntera-2 neurons on the inhibitory chondroitin sulphate proteoglycan substrate. In the rat optic nerve crush model Y-27632 dose-dependently increased regeneration of retinal ganglion cell axons in vivo. Application of Dimethylfasudil showed a trend towards increased axonal regeneration in an intermediate concentration. We demonstrate that inhibition of ROCK can be an effective therapeutic approach to increase regeneration of CNS neurons. The selection of a suitable inhibitor with a broad therapeutic window, however, is crucial in order to minimize unwanted side effects and to avoid deleterious effects on nerve fiber growth.

The proteasome is critically involved in the production of MHC class I-restricted T cell epitopes. Proteasome activity and epitope production are altered by IFN-γ treatment, which leads to a gradual replacement of constitutive proteasomes by immunoproteasomes in vitro. However, a quantitative analysis of changes in the steady state subunit composition of proteasomes during an immune response against viruses or bacteria in vivo has not been reported. Here we show that the infection of mice with lymphocytic choriomeningitis virus or Listeria monocytogenes leads to an almost complete replacement of constitutive proteasomes by immunoproteasomes in the liver within 7 days. Proteasome replacements were markedly reduced in IFN-γ−/− mice, but were only slightly affected in IFN-αR−/− and perforin−/− mice. The proteasome regulator PA28α/β was up-regulated, whereas PA28γ was reduced in the liver of lymphocytic choriomeningitis virus-infected mice. Proteasome replacements in the liver strongly altered proteasome activity and were unexpected to this extent, since an in vivo half-life of 12 days had been previously assigned to constitutive proteasomes in the liver. Our results suggest that during the peak phase of viral and bacterial elimination the antiviral cytotoxic T lymphocyte response is directed mainly to immunoproteasome-dependent T cell epitopes, which would be a novel parameter for the design of vaccines.

Optimal vaccine strategies must be identified for improving T-cell vaccination against infectious and malignant diseases. MelQbG10 is a virus-like nano-particle loaded with A-type CpG-oligonucleotides (CpG-ODN) and coupled to peptide16–35 derived from Melan-A/MART-1. In this phase IIa clinical study, four groups of stage III-IV melanoma patients were vaccinated with MelQbG10, given (i) with IFA (Montanide) s.c.; (ii) with IFA s.c. and topical Imiquimod; (iii) i.d. with topical Imiquimod; or (iv) as intralymph node injection. In total, 16/21 (76%) patients generated ex vivo detectable Melan-A/MART-1-specific T-cell responses. T-cell frequencies were significantly higher when IFA was used as adjuvant, resulting in detectable T-cell responses in all (11/11) patients, with predominant generation of effector-memory-phenotype cells. In turn, Imiquimod induced higher proportions of central-memory-phenotype cells and increased percentages of CD127+ (IL-7R) T cells. Direct injection of MelQbG10 into lymph nodes resulted in lower T-cell frequencies, associated with lower proportions of memory and effector-phenotype T cells. Swelling of vaccine site draining lymph nodes, and increased glucose uptake at PET/CT was observed in 13/15 (87%) of evaluable patients, reflecting vaccine triggered immune reactions in lymph nodes. We conclude that the simultaneous use of both Imiquimod and CpG-ODN induced combined memory and effector CD8+ T-cell responses.