T cells can mediate viral clearance from ependyma but not from brain parenchyma in a major histocompatibility class I- and perforin-independent manner

Viral infection of the central nervous system can lead to disability and death. Yet the majority of viral infections with central nervous system involvement resolve with only mild clinical manifestations, if any. This is generally attributed to efficient elimination of the infection from the brain coverings, i.e. the meninges, ependyma and chorioplexus, which are the primary targets of haematogeneous viral spread. How the immune system is able to purge these structures from viral infection with only minimal detrimental effects is still poorly understood. In the present work we studied how an attenuated lymphocytic choriomeningitis virus can be cleared from the central nervous system in the absence of overt disease. We show that elimination of the virus from brain ependyma, but not from brain parenchyma, could be achieved by a T cell-dependent mechanism operating independently of major histocompatibility class I antigens and perforin. Considering that cytotoxic T lymphocyte-mediated cytotoxicity is a leading cause of viral immunopathology and tissue damage, our findings may explain why the most common viral intruders of the central nervous system rarely represent a serious threat to our healt

Professional Memory CD4+ T Lymphocytes Preferentially Reside and Rest in the Bone Marrow

SummaryCD4+ T lymphocytes are key to immunological memory. Here we show that in the memory phase of specific immune responses, most of the memory CD4+ T lymphocytes had relocated into the bone marrow (BM) within 3–8 weeks after their generation—a process involving integrin α2. Antigen-specific memory CD4+ T lymphocytes highly expressed Ly-6C, unlike most splenic CD44hiCD62L− CD4+ T lymphocytes. In adult mice, more than 80% of Ly-6ChiCD44hiCD62L− memory CD4+ T lymphocytes were in the BM. In the BM, they associated to IL-7-expressing VCAM-1+ stroma cells. Gene expression and proliferation were downregulated, indicating a resting state. Upon challenge with antigen, they rapidly expressed cytokines and CD154 and efficiently induced the production of high-affinity antibodies by B lymphocytes. Thus, in the memory phase of immunity, memory helper T cells are maintained in BM as resting but highly reactive cells in survival niches defined by IL-7-expressing stroma cells

Short-term memory in gene induction reveals the regulatory principle behind stochastic IL-4 expression

Combining experiments on primary T cells and mathematical modeling, we characterized the stochastic expression of the interleukin-4 cytokine gene in its physiologic context, showing that a two-step model of transcriptional regulation acting on chromatin rearrangement and RNA polymerase recruitment accounts for the level, kinetics, and population variability of expression.A rate-limiting step upstream of transcription initiation, but occurring at the level of an individual allele, controls whether the interleukin-4 gene is expressed during antigenic stimulation, suggesting that the observed stochasticity of expression is linked to the dynamics of chromatin rearrangement.The computational analysis predicts that the probability to re-express an interleukin-4 gene that has been expressed once is transiently increased. In support, we experimentally demonstrate a short-term memory for interleukin-4 expression at the predicted time scale of several days.The model provides a unifying framework that accounts for both graded and binary modes of gene regulation. Graded changes in expression level can be achieved by controlling transcription initiation, whereas binary regulation acts at the level of chromatin rearrangement and is targeted during the differentiation of T cells that specialize in interleukin-4 production

Expression of ICOS In Vivo Defines CD4+ Effector T Cells with High Inflammatory Potential and a Strong Bias for Secretion of Interleukin 10

The studies performed to date analyzed the overall participation of the inducible costimulator (ICOS) in model diseases, but did not yield information on the nature and function of ICOS-expressing T cells in vivo. We examined ICOS+ T cells in the secondary lymphoid organs of nonmanipulated mice, in the context of an “unbiased” immune system shaped by environmental antigens. Using single cell analysis, ICOSlow cells were found to be loosely associated with the early cytokines interleukin (IL)-2, IL-3, IL-6, and interferon (IFN)-γ. ICOSmedium cells, the large majority of ICOS+ T cells in vivo, were very tightly associated with the synthesis of the T helper type 2 (Th2) cytokines IL-4, IL-5, and IL-13, and these cells exhibited potent inflammatory effects in vivo. In contrast, ICOShigh T cells were highly and selectively linked to the anti-inflammatory cytokine IL-10. Overall, these data seem to indicate that ICOS cell surface density serves as a regulatory mechanism for the release of cytokines with different immunological properties. Further in vivo functional experiments with in vitro–activated T cells strongly suggested that the ICOS+ population, although representing in vivo only around 10% of T cells bearing early or late activation markers, nevertheless encompasses virtually all effector T cells, a finding with major diagnostic and therapeutic implications

Dissecting the dynamic transcriptional landscape of early T helper cell differentiation into Th1, Th2, and Th1/2 hybrid cells

Selective differentiation of CD4+ T helper (Th) cells into specialized subsets such as Th1 and Th2 cells is a key element of the adaptive immune system driving appropriate immune responses. Besides those canonical Th-cell lineages, hybrid phenotypes such as Th1/2 cells arise in vivo, and their generation could be reproduced in vitro. While master-regulator transcription factors like T-bet for Th1 and GATA-3 for Th2 cells drive and maintain differentiation into the canonical lineages, the transcriptional architecture of hybrid phenotypes is less well understood. In particular, it has remained unclear whether a hybrid phenotype implies a mixture of the effects of several canonical lineages for each gene, or rather a bimodal behavior across genes. Th-cell differentiation is a dynamic process in which the regulatory factors are modulated over time, but longitudinal studies of Th-cell differentiation are sparse. Here, we present a dynamic transcriptome analysis following Th-cell differentiation into Th1, Th2, and Th1/2 hybrid cells at 3-h time intervals in the first hours after stimulation. We identified an early bifurcation point in gene expression programs, and we found that only a minority of ~20% of Th cell-specific genes showed mixed effects from both Th1 and Th2 cells on Th1/2 hybrid cells. While most genes followed either Th1- or Th2-cell gene expression, another fraction of ~20% of genes followed a Th1 and Th2 cell-independent transcriptional program associated with the transcription factors STAT1 and STAT4. Overall, our results emphasize the key role of high-resolution longitudinal data for the characterization of cellular phenotypes.Peer Reviewe

Long-lived virus-reactive memory T cells generated from purified cytokine-secreting T helper type 1 and type 2 effectors

Many vaccination strategies and immune cell therapies aim at increasing the numbers of memory T cells reactive to protective antigens. However, the differentiation lineage and therefore the optimal generation conditions of CD4 memory cells remain controversial. Linear and divergent differentiation models have been proposed, suggesting CD4 memory T cell development from naive precursors either with or without an effector-stage intermediate, respectively. Here, we address this question by using newly available techniques for the identification and isolation of effector T cells secreting effector cytokines. In adoptive cell transfers into normal, nonlymphopenic mice, we show that long-lived virus-specific memory T cells can efficiently be generated from purified interferon γ–secreting T helper (Th) type 1 and interleukin (IL)-4– or IL-10–secreting Th2 effectors primed in vitro or in vivo. Importantly, such effector-derived memory T cells were functional in viral challenge infections. They proliferated vigorously, rapidly modulated IL-7 receptor expression, exhibited partial stability and flexibility of their cytokine patterns, and exerted differential effects on virus-induced immunopathology. Thus, cytokine-secreting effectors can evade activation-induced cell death and develop into long-lived functional memory cells. These findings demonstrate the efficiency of linear memory T cell differentiation and encourage the design of vaccines and immune cell therapies based on differentiated effector T cells

A buprenorphine depot formulation provides effective sustained post-surgical analgesia for 72 h in mouse femoral fracture models

Adequate pain management is essential for ethical and scientific reasons in animal experiments and should completely cover the period of expected pain without the need for frequent re-application. However, current depot formulations of Buprenorphine are only available in the USA and have limited duration of action. Recently, a new microparticulate Buprenorphine formulation (BUP-Depot) for sustained release has been developed as a potential future alternative to standard formulations available in Europe. Pharmacokinetics indicate a possible effectiveness for about 72 h. Here, we investigated whether the administration of the BUP-Depot ensures continuous and sufficient analgesia in two mouse fracture models (femoral osteotomy) and could, therefore, serve as a potent alternative to the application of Tramadol via the drinking water. Both protocols were examined for analgesic effectiveness, side effects on experimental readout, and effects on fracture healing outcomes in male and female C57BL/6N mice. The BUP-Depot provided effective analgesia for 72 h, comparable to the effectiveness of Tramadol in the drinking water. Fracture healing outcome was not different between analgesic regimes. The availability of a Buprenorphine depot formulation for rodents in Europe would be a beneficial addition for extended pain relief in mice, thereby increasing animal welfare