Within overall Th1-like human memory T cell responses, individual T cells may express only some of the characteristic Th1 cytokines when reactivated. In the Th1-oriented memory response to influenza, ...we have tested the contributions of two potential mechanisms for this diversity: variable expression of cytokines by a uniform population during activation, or different stable subsets that consistently expressed subsets of the Th1 cytokine pattern. To test for short-term variability, in vitro-stimulated influenza-specific human memory CD4+ T cells were sorted according to IL-2 and IFNγ expression, cultured briefly in vitro, and cytokine patterns measured after restimulation. Cells that were initially IFNγ+ and either IL-2+ or IL-2- converged rapidly, containing similar proportions of IL-2-IFNγ+ and IL-2+IFNγ+ cells after culture and restimulation. Both phenotypes expressed Tbet, and similar patterns of mRNA. Thus variability of IL-2 expression in IFNγ+ cells appeared to be regulated more by short-term variability than by stable differentiated subsets. In contrast, heterogeneous expression of IFNγ in IL-2+ influenza-specific T cells appeared to be due partly to stable T cell subsets. After sorting, culture and restimulation, influenza-specific IL-2+IFNγ- and IL-2+IFNγ+ cells maintained significantly biased ratios of IFNγ+ and IFNγ- cells. IL-2+IFNγ- cells included both Tbetlo and Tbethi cells, and showed more mRNA expression differences with either of the IFNγ+ populations. To test whether IL-2+IFNγ-Tbetlo cells were Thpp cells (primed but uncommitted memory cells, predominant in responses to protein vaccines), influenza-specific IL-2+IFNγ- and IL-2+IFNγ+ T cells were sorted and cultured in Th1- or Th2-generating conditions. Both cell types yielded IFNγ-secreting cells in Th1 conditions, but only IL-2+IFNγ- cells were able to differentiate into IL-4-producing cells. Thus expression of IL-2 in the anti-influenza response may be regulated mainly by short term variability, whereas different T cell subsets, Th1 and Thpp, may contribute to variability in IFNγ expression.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Background. T-helper (Th) 17 cells are important in the control of Streptococcus pneumoniae. We sought to understand the mechanism of failure of Th17 immunity resulting in S. pneumoniae infections in ...children <2 years old. Methods. Peripheral blood mononuclear cells (PBMCs) from infection-prone (IP) and non-IP (NIP) children 9-18 months old were examined for their responses to heat-killed S. Pneumoniae, using flow cytometry, reverse-transcription polymerase chain reaction, and enzyme-linked immunoassay. We measured cytokine production, proliferation, and differentiation of Th17 cells and the expression of transcription factors in response to S. pneumoniae. Results. PBMCs of IP children stimulated with heat-killed S. pneumoniae had significantly reduced percentages of CD4⁺ Th1 (interleukin2, tumor necrosis factor a) and Th17 (interleukin 17A) cells compared with NIP children. Addition of exogenous Th17promoting cytokines (interleukin 6, 1β, and 23 and transforming growth factor β) restored CD4⁺ Th17 cell function in cells from IP children to levels measured in NIP children. Conclusions. Reduced Th17 responses to S. pneumoniae in PBMCs of IP children can be rescued by addition of Th17-promoting cytokines.
CD73 (5'-ectonucleotidase) is expressed by two distinct mouse CD4 T cell populations: CD25+ (FoxP3+) T regulatory (Treg) cells that suppress T cell proliferation but do not secrete IL-2, and CD25- ...uncommitted primed precursor Th (Thpp) cells that secrete IL-2 but do not suppress in standard Treg suppressor assays. CD73 on both Treg and Thpp cells converted extracellular 5'-AMP to adenosine. Adenosine suppressed proliferation and cytokine secretion of Th1 and Th2 effector cells, even when target cells were activated by anti-CD3 and anti-CD28. This represents an additional suppressive mechanism of Treg cells and a previously unrecognized suppressive activity of Thpp cells. Infiltration of either Treg or Thpp cells at inflammatory sites could potentially convert 5'-AMP generated by neutrophils or dying cells into the anti-inflammatory mediator adenosine, thus dampening excessive immune reactions.
Introduction The HVTN 105 vaccine clinical trial tested four combinations of two immunogens - the DNA vaccine DNA-HIV-PT123, and the protein vaccine AIDSVAX B/E. All combinations induced substantial ...antibody and CD4+ T cell responses in many participants. We have now re-examined the intracellular cytokine staining flow cytometry data using the high-resolution SWIFT clustering algorithm, which is very effective for enumerating rare populations such as antigen-responsive T cells, and also determined correlations between the antibody and T cell responses. Methods Flow cytometry samples across all the analysis batches were registered using the swiftReg registration tool, which reduces batch variation without compromising biological variation. Registered data were clustered using the SWIFT algorithm, and cluster template competition was used to identify clusters of antigen-responsive T cells and to separate these from constitutive cytokine producing cell clusters. Results Registration strongly reduced batch variation among batches analyzed across several months. This in-depth clustering analysis identified a greater proportion of responders than the original analysis. A subset of antigen-responsive clusters producing IL-21 was identified. The cytokine patterns in each vaccine group were related to the type of vaccine – protein antigens tended to induce more cells producing IL-2 but not IFN-γ, whereas DNA vaccines tended to induce more IL-2+ IFN-γ+ CD4 T cells. Several significant correlations were identified between specific antibody responses and antigen-responsive T cell clusters. The best correlations were not necessarily observed with the strongest antibody or T cell responses. Conclusion In the complex HVTN105 dataset, alternative analysis methods increased sensitivity of the detection of antigen-specific T cells; increased the number of identified vaccine responders; identified a small IL-21-producing T cell population; and demonstrated significant correlations between specific T cell populations and serum antibody responses. Multiple analysis strategies may be valuable for extracting the most information from large, complex studies.
Cytokine expression patterns of T cells can be regulated by pre-commitment to stable effector phenotypes, further modification of moderately stable phenotypes, and quantitative changes in cytokine ...production in response to acute signals. We showed previously that the epidermal growth factor family member Amphiregulin is expressed by T cell receptor-activated mouse CD4 T cells, particularly Th2 cells, and helps eliminate helminth infection. Here we report a detailed analysis of the regulation of Amphiregulin expression by human T cell subsets. Signaling through the T cell receptor induced Amphiregulin expression by most or all T cell subsets in human peripheral blood, including naive and memory CD4 and CD8 T cells, Th1 and Th2 in vitro T cell lines, and subsets of memory CD4 T cells expressing several different chemokine receptors and cytokines. In these different T cell types, Amphiregulin synthesis was inhibited by an antagonist of protein kinase A, a downstream component of the cAMP signaling pathway, and enhanced by ligands that increased cAMP or directly activated protein kinase A. Prostaglandin E2 and adenosine, natural ligands that stimulate adenylyl cyclase activity, also enhanced Amphiregulin synthesis while reducing synthesis of most other cytokines. Thus, in contrast to mouse T cells, Amphiregulin synthesis by human T cells is regulated more by acute signals than pre-commitment of T cells to a particular cytokine pattern. This may be appropriate for a cytokine more involved in repair than attack functions during most inflammatory responses.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Human CD4 T cell recall responses to influenza virus are strongly biased towards Type 1 cytokines, producing IFNγ, IL-2 and TNFα. We have now examined the effector phenotypes of CD4 T cells in more ...detail, particularly focusing on differences between recent versus long-term, multiply-boosted responses. Peptides spanning the proteome of temporally distinct influenza viruses were distributed into pools enriched for cross-reactivity to different influenza strains, and used to stimulate antigen-specific CD4 T cells representing recent or long-term memory. In the general population, peptides unique to the long-circulating influenza A/New Caledonia/20/99 (H1N1) induced Th1-like responses biased toward the expression of IFNγ(+)TNFα(+) CD4 T cells. In contrast, peptide pools enriched for non-cross-reactive peptides of the pandemic influenza A/California/04/09 (H1N1) induced more IFNγ(-)IL-2(+)TNFα(+) T cells, similar to the IFNγ(-)IL-2(+) non-polarized, primed precursor T cells (Thpp) that are a predominant response to protein vaccination. These results were confirmed in a second study that compared samples taken before the 2009 pandemic to samples taken one month after PCR-confirmed A/California/04/09 infection. There were striking increases in influenza-specific TNFα(+), IFNγ(+), and IL-2(+) cells in the post-infection samples. Importantly, peptides enriched for non-cross-reactive A/California/04/09 specificities induced a higher proportion of Thpp-like IFNγ(-)IL-2(+)TNFα(+) CD4 T cells than peptide pools cross-reactive with previous influenza strains, which induced more Th1 (IFNγ(+)TNFα(+)) responses. These IFNγ(-)IL-2(+)TNFα(+) CD4 T cells may be an important target population for vaccination regimens, as these cells are induced upon infection, may have high proliferative potential, and may play a role in providing future effector cells during subsequent infections.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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