Author Correction: Multidisciplinary study of the secondary immune response in grandparents re-exposed to chickenpox.

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Multidisciplinary study of the secondary immune response in grandparents re-exposed to chickenpox.

Re-exposure to chickenpox may boost varicella-zoster virus (VZV) immunity in the elderly. This secondary immune response is hypothesized to confer protection against herpes zoster. We longitudinally sampled 36 adults over the course of one year after re-exposure to chickenpox. The resulting 183 samples and those of 14 controls were assessed for VZV-specific T-cell immunity and antibody titres. The percentages of VZV-specific CD4+ IL-2-producing T-cells were increased in re-exposed grandparents compared to control participants up to 9 months after re-exposure. Using a longitudinal mixture modelling approach, we found that 25% and 17% of re-exposed grandparents showed a boosting of VZV-specific CD4+ IL-2-producing T-cells and VZV-specific antibodies, respectively. The antibody boosting occurred exclusively in cytomegalovirus (CMV) IgG-positive participants. CMV IgG-positive participants also had higher VZV IE62-specific CD4+ IFN-γ-producing T-cell percentages and VZV-specific antibody titres. The protective effect of re-exposure to chickenpox is likely limited, as boosting only occurred in 17-25% of the VZV re-exposed grandparents and for less than one year.

Brain derived neurotrophic factor in multiple sclerosis: effect of 24 weeks endurance and resistance training.

BACKGROUND AND PURPOSE: Brain derived neurotrophic factor (BDNF) is suggested to play a neuroprotective role in multiple sclerosis (MS). However, the BDNF response to long-term exercise in MS remains unknown. Our objective was to compare resting BDNF profiles of healthy controls (HCs) and persons with relapsing-remitting MS (RRMS) and to investigate the impact of a 24-week exercise intervention on serum BDNF release in MS. METHODS: At baseline, blood BDNF levels were assessed in MS (n = 22, mean Expanded Disability Status Scale 2.6 ± 0.2, mean age 43 ± 2 years) and HCs (n = 19, mean age 47 ± 1 year). Next, persons with MS were randomized to an exercise intervention group (EX, n = 15) or a sedentary control group (SED, n = 7) completing a 24-week randomized controlled trial. In persons with MS, muscle strength, exercise tolerance and body composition were assessed, as compliance measures, at baseline and after 24 weeks. RESULTS: At baseline, the BDNF concentration of persons with RRMS was 21% lower than HCs. Following 24 weeks of intervention, changes in BDNF concentrations differed significantly between EX and SED. In particular, within EX BDNF concentrations increased 13.9% ± 8.8%, whereas it decreased 10.5% ± 4.1% within SED. Furthermore, 24 weeks of exercise induced changes in the compliance measures between EX and SED. In addition, within EX muscle strength, exercise tolerance and lean tissue mass improved, whereas these remained stable within SED. CONCLUSION: In conclusion, BDNF concentration of persons with RRMS was lower compared to HCs and increased after 24 weeks of exercise in persons with MS, compared to the non-exercise MS control group.

Effect of schistosomiasis on CX3CR1-expressing mononuclear phagocytes in the ileum and mesenteric lymph nodes of the mouse.

BACKGROUND: Intestinal dendritic cells (DCs) maintain immune homeostasis, only initiating an active immune response against invading pathogens. However, little information is available on the reaction of mononuclear phagocytes (MNP) to intestinal trematode infection, a reaction equally important in helminth-based therapies. The CD11c(+)  CX3CR1(+)  F4/80(-) DCs in the ileal lamina propria (LP) of the mouse were proven to migrate to the mesenteric lymph nodes (MLNs). We analyzed all MNP subsets present in the mouse LP and MLNs, under steady-state conditions and during acute Schistosoma mansoni-induced inflammation. Furthermore, we studied the uptake of schistosomal antigens by MNP in vivo in the LP and MLNs. METHODS: Using a combination of immunohistochemistry and multiparametric flow cytometry, we investigated distributional changes of the MNP during acute intestinal schistosomiasis. Next, S. mansoni-derived products, i.e., S. mansoni soluble worm proteins (SmSWP) and S. mansoni soluble egg antigens (SmSEA) were intraperitoneally injected into CX3CR1(+/) (GFP) C57BL/6 mice and antigen uptake was analyzed using confocal microscopy. KEY RESULTS: The CD11c(+)  CX3CR1(+)  F4/80(-) DCs significantly increased during intestinal schistosomiasis in the LP and MLNs. Only CX3CR1-expressing DC and MФ subsets, but not other LP DCs, are involved in both SmSWP and SmSEA antigen uptake and processing. CONCLUSIONS & INFERENCES: The significant upregulation of CD11c(+)  CX3CR1(+)  F4/80(-) DCs during intestinal schistosomiasis and the restriction of phagocytosis of parasitic antigens to CX3CR1-expresssing MNP indicate a crucial role for this immune cell niche in response to trematodiasis. These findings add insight into the functional specialization of LP immune cells and add to the understanding of cellular mechanisms behind helminth-based therapies.

Does multiple sclerosis affect glucose tolerance?

Based on current literature, it is not clear if multiple sclerosis (MS) patients are at increased risk to develop impaired glucose tolerance (IGT). Eighty-one MS patients and 45 healthy controls (HC) performed an oral glucose tolerance test. IGT was defined as a fasting glucose concentration of 6.1-6.9 mmol/l and two-hour post-load glucose of 7.8-11.1 mmol/l. The prevalence of impaired fasting glucose concentrations (17% vs 2%) and IGT (11% vs 0%) was higher in MS patients than HC. Accordingly, the areas under the glucose and insulin curves were higher in MS patients. The current study demonstrates an elevated IGT-prevalence in MS.

Dendritic cells in multiple sclerosis: key players in the immunopathogenesis, key players for new cellular immunotherapies?

Many studies have demonstrated the role of the adaptive immune system in the pathogenesis of multiple sclerosis (MS). Recent data suggest that dendritic cells (DCs), which are innate immune cells, also contribute to the pathogenesis of MS. In patients with MS, DCs are abundantly present in brain lesions, and display an altered phenotype and/or function as compared with this in healthy controls. DCs are thus in the position to pathologically influence the effector function of (auto-reactive) T and B cells. Interestingly, current first-line immunomodulating therapies for MS have been shown to restore DC phenotype and function, albeit in a non-specific manner. To date, clinical trials using agents specifically targeting DC function are ongoing. Moreover, several studies worldwide are currently investigating possible strategies to develop tolerogenic DCs. This review focuses on the phenotypic and functional alterations of conventional DCs and plasmacytoid DCs in patients with MS. Furthermore, we discuss how existing immunomodulating therapies for MS patients affect DC function and address future perspectives in the development of immunotherapies specifically targeting DCs.

Efficient mRNA electroporation of peripheral blood mononuclear cells to detect memory T cell responses for immunomonitoring purposes.

To date, the number of immunotherapy vaccines in clinical trials increases exponentially. To evaluate the efficacy of these clinical vaccination trials, we fervently need cellular immunomonitoring tools. In this study, we present a newly developed short-time assay which allows direct ex vivo analysis of multi-epitope antigen-specific T cell immune responses. This novel method is based on mRNA electroporation of isolated peripheral blood mononuclear cells (PBMC). Fresh and cryopreserved PBMC of both healthy volunteers as well as of allogeneic stem cell transplanted patients enrolled in a cytomegalovirus (CMV) dendritic cell vaccination trial were electroporated with CMV pp65-encoding mRNA. Using a direct IFN-gamma EliSPOT and intracellular cytokine flow cytometry we detected significantly higher numbers of CMV pp65-specific IFN-gamma-secreting T cells as compared to the assay with non-treated PBMC and as compared to PBMC electroporated with mRNA coding for an irrelevant protein. Compared to conventional methods to evaluate T cell-mediated immune responses, this method is time-saving and less labor-intensive because it obviates the need for in vitro cultured antigen-presenting cells and because an overnight incubation is sufficient for activation of T cells. Moreover, the use of CMV pp65-encoding mRNA will broaden the immune response because it covers every epitope with potential relevance. This is a major advantage compared to the recognition of a single epitope covered by a HLA-restricted peptides. In summary, we developed a highly efficient mRNA electroporation protocol for fresh and cryopreserved PBMC. This novel method is a rapid and elegant tool and will be convenient for monitoring the cellular immune status of patients in a clinical vaccination setting.

Immunosuppression induced by immature dendritic cells is mediated by TGF-beta/IL-10 double-positive CD4+ regulatory T cells.

Dendritic cells (DC) have important functions in T cell immunity and T cell tolerance. Previously, it was believed that T cell unresponsiveness induced by immature DC (iDC) is caused by the absence of inflammatory signals in steady-state in vivo conditions and by the low expression levels of costimulatory molecules on iDC. However, a growing body of evidence now indicates that iDC can also actively maintain peripheral T cell tolerance by the induction and/or stimulation of regulatory T cell populations. In this study, we investigated the in vitro T cell stimulatory capacity of iDC and mature DC (mDC) and found that both DC types induced a significant increase in the number of transforming growth factor (TGF)-beta and interleukin (IL)-10 double-positive CD4(+) T cells within 1 week of autologous DC/T cell co-cultures. In iDC/T cell cultures, where antigen-specific T cell priming was significantly reduced as compared to mDC/T cell cultures, we demonstrated that the tolerogenic effect of iDC was mediated by soluble TGF-beta and IL-10 secreted by CD4(+)CD25(-)FOXP3(-) T cells. In addition, the suppressive capacity of CD4(+) T cells conditioned by iDC was transferable to already primed antigen-specific CD8(+) T cell cultures. In contrast, addition of CD4(+) T cells conditioned by mDC to primed antigen-specific CD8(+) T cells resulted in enhanced CD8(+) T cell responses, notwithstanding the presence of TGF-beta(+)/IL-10(+) T cells in the transferred fraction. In summary, we hypothesize that DC have an active role in inducing immunosuppressive cytokine-secreting regulatory T cells. We show that iDC-conditioned CD4(+) T cells are globally immunosuppressive, while mDC induce globally immunostimulatory CD4(+) T cells. Furthermore, TGF-beta(+)/IL-10(+) T cells are expanded by DC independent of their maturation status, but their suppressive function is dependent on immaturity of DC.

Proinflammatory response of human leukemic cells to dsRNA transfection linked to activation of dendritic cells.

Leukemic cells exert immunosuppressive effects that interfere with dendritic cell (DC) function and hamper effective antileukemic immune responses. Here, we sought to enhance the immunogenicity of leukemic cells by loading them with the double-stranded (ds) RNA Toll-like receptor 3 (TLR3) ligand polyriboinosinic polyribocytidylic acid (poly(I:C)), mimicking viral infection of the tumor cells. Given the responsiveness of DC to TLR ligands, we hypothesized that the uptake of poly(I:C)-loaded leukemic cells by immature DC (iDC) would lead to DC activation. Primary acute myeloid leukemia (AML) cells and AML cell lines markedly responded to poly(I:C) electroporation by apoptosis, upregulation of TLR3 expression, enhanced expression of major histocompatibility complex (MHC) and costimulatory molecules and by production of type I interferons (IFN). Upon phagocytosis of poly(I:C)-electroporated AML cells, DC maturation and activation were induced as judged by an increased expression of MHC and costimulatory molecules, production of proinflammatory cytokines and an increase of T helper 1 (T(H)1)-polarizing capacity. These immune effects were suboptimal when AML cells were passively pulsed with poly(I:C), indicating the superiority of poly(I:C) transfection over pulsing. Our results demonstrate that poly(I:C) electroporation is a promising strategy to increase the immunogenicity of AML cells and to convert iDC into activated mature DC following the phagocytosis of AML cells.

mRNA-electroporated mature dendritic cells retain transgene expression, phenotypical properties and stimulatory capacity after cryopreservation.

Genetically modified dendritic cells (DC) are increasingly used in vitro to activate cytotoxic T lymphocyte (CTL) immune responses. Because T cell activation protocols consist of multiple restimulation cycles of peripheral blood lymphocytes with antigen-loaded mature DC, continuous generation of DC is needed throughout the experiment. Therefore, cryopreservation of DC loaded with antigen is a valuable alternative for weekly generation and modification of DC. Recently, we described an antigen loading method for DC based on electroporation of defined tumor antigen mRNA. In this study, we demonstrate that mRNA-electroporated DC can efficiently be prepared for cryopreservation. Using an optimized maturation and freezing protocol after mRNA electroporation, we obtained high transgene-expressing viable mature DC. In addition, we showed that these modified cryopreserved DC retain stimulatory capacity in an influenza model system. Therefore, cryopreservation of mature mRNA-electroporated DC is a useful method for continuous availability of antigen-loaded DC throughout T cell activation experiments.