isolation-ex-vivo-culture-v1-cd4-t-cells-an-extrathymic-t-cell

Isolation and Ex Vivo Culture of V&#948;1+CD4+&#947;&#948; T Cells, an Extrathymic &#945;&#946;T-cell Progenitor

The thymus, the primary organ for the generation of &#945;&#946; T cells and backbone of the adaptive immune system in vertebrates, has long been ...

Children's University Hospital Tübingen

We calculated the rate of twinning across four captive collections of tufted capuchins (Cebus apella) to be 2.4%. This rate contrast with previous reports that twinning in tufted capuchins is rare. Additionally, we present data on the survival and weight gain of twins in this species as compared to singletons. Twins face their greatest risk of mortality on or before the first day of life, when 45% will die compared to 16% of singletons. After the first day of life, twins and singletons demonstrate comparable survival rates. This, in conjunction with the finding that at no time during the first year of life do twins and singletons differ significantly in their weights, suggests that twinning is a viable reproductive form for these animals, especially in the captive setting where nutritional demands are met.

Twinning in tufted capuchins (Cebus apella): rate, survivorship, and weight gain.

The lifelong generation of αβT cells enables us to continuously build immunity against pathogens and malignancies despite the loss of thymic function with age. Homeostatic proliferation of post-thymic naïve and memory T cells and their transition into effector and long-lived memory cells balance the decreasing output of naïve T cells, and recent research suggests that also αβT-cell development independent from the thymus may occur. However, the sites and mechanisms of extrathymic T-cell development are not yet understood in detail. γδT cells represent a small fraction of the overall T-cell pool, and are endowed with tremendous phenotypic and functional plasticity. γδT cells that express the Vδ1 gene segment are a minor population in human peripheral blood but predominate in epithelial (and inflamed) tissues. Here, we characterize a CD4(+) peripheral Vδ1(+) γδT-cell subpopulation that expresses stem-cell and progenitor markers and is able to develop into functional αβT cells ex vivo in a simple culture system and in vivo. The route taken by this process resembles thymic T-cell development. However, it involves the re-organization of the Vδ1(+) γδTCR into the αβTCR as a consequence of TCR-γ chain downregulation and the expression of surface Vδ1(+)Vβ(+) TCR components, which we believe function as surrogate pre-TCR. This transdifferentiation process is readily detectable in vivo in inflamed tissue. Our study provides a conceptual framework for extrathymic T-cell development and opens up a new vista in immunology that requires adaptive immune responses in infection, autoimmunity, and cancer to be reconsidered.

Human Peripheral CD4(+) Vδ1(+) γδT Cells Can Develop into αβT Cells.

Stimulating the immune system to attack cancer is a promising approach, even for the control of advanced cancers. Several cytokines that promote interferon-γ-dominated immune responses show antitumor activity, with interleukin 12 (IL-12) being of major importance. Here, we used an antibody-IL-12 fusion protein (NHS-IL12) that binds histones of necrotic cells to treat human sarcoma in humanized mice. Following sarcoma engraftment, NHS-IL12 therapy was combined with either engineered IL-7 (FcIL-7) or IL-2 (IL-2MAB602) for continuous cytokine bioavailability. NHS-IL12 strongly induced innate and adaptive antitumor immunity when combined with IL-7 or IL-2. NHS-IL12 therapy significantly improved survival of sarcoma-bearing mice and caused long-term remissions when combined with IL-2. NHS-IL12 induced pronounced cancer cell senescence, as documented by strong expression of senescence-associated p16(INK4a) and nuclear translocation of p-HP1γ, and permanent arrest of cancer cell proliferation. In addition, this cancer immunotherapy initiated the induction of myogenic differentiation, further promoting the hypothesis that efficient antitumor immunity includes mechanisms different from cytotoxicity for efficient cancer control in vivo.

Cancer-targeted IL-12 controls human rhabdomyosarcoma by senescence induction and myogenic differentiation.

Chronic graft-versus-host disease (cGVHD) is a significant hurdle to long-term hematopoietic stem-cell transplantation success. Insights into the pathogenesis and mechanistical investigations of novel therapeutic strategies are limited as appropriate animal models are missing. The immunodeficient NSG mouse - when humanized with human bone marrow, fetal liver and thymus (BLT NSG) - is prone for cGVHD, yet mainly affects the skin. In contrast, the NSG mouse humanized exclusively with CD34(+)-selected, CD3(+)-depleted stem cells (CD34(+)NSG) has neither been described for acute nor chronic GVHD so far. This is the first report about the development of systemic autoimmune cGVHD ≥24 weeks post stem cell receipt involving lung, liver, skin, gingiva and intestine in two NSG cohorts humanized with CD34(+) grafts from different donors. Affected mice presented with sclerodermatous skin, fibrotic lung, severe hepatitis, and massive dental malformation/loss. CD4(+)-dominated, TH2-biased, bulky T-cell infiltrates featured highly skewed T cell receptor (TCR) repertoires, clonal expansions, and autoreactive TCRs. In affected tissues profibrotic IL-13 and -4 dominated over TH1 cytokines IFN-γ and TNF-α. Thus, the time point of manifestation and the phenotype match human systemic pleiotropic sclerodermatous GVHD. The CD34(+)NSG-model's intrinsic deficiency of thymus, thymus-derived regulatory T cells (nTreg) and B cells emphasizes the role of the genetic polymorphism and the cytokines in the pathogenesis of cGVHD. Importantly, the only factor discriminating diseased versus non-diseased CD34(+)NSG cohorts were two risk HLA haplotypes that in human mediate susceptibility for autoimmune disease (psoriasis). Thus, the CD34(+)NSG model may serve as a platform for addressing issues related to the pathophysiology and treatment of human autoimmunity and chronic GVHD.

Chronic graft-versus-host-disease in CD34(+)-humanized NSG mice is associated with human susceptibility HLA haplotypes for autoimmune disease.

GVHD, both acute and chronic, remains the major cause of morbidity and mortality after allogeneic hematopoietic stem cell transplantation. Thus, there is still a great need for therapeutic tools for the prevention and treatment of GVHD. Several biologics have shown promising results in salvage therapies but are attendant on an increased risk for opportunistic infections, lymphoproliferative disorders, and relapse. This is partly due to efficient T cell elimination that neither dissects alloreactive from non-alloreactive T cells nor considers functional and structural distinctiveness of pathogen- and malignancy-reactive γδ and iNKT T cells. A novel, humanized monoclonal antibody, GZ-αβTCR, specific for the human αβ T cell receptor, was evaluated in a xenogeneic GVHD model for its potential to prevent or ameliorate GVHD and prolong survival. We could show that GZ-αβTCR significantly attenuated clinical signs of GVHD and prolonged survival by preferential depletion of CD4 cells and the naïve T cell compartment, the trigger and driver of GVHD. In a regimen that included a preemptive dose, GZ-αβTCR treatment sufficiently abrogated GVHD. Importantly, GZ-αβTCR's specificity spared host cell-mediated immune competence of cell types other than αβT cells: namely γδT cells. GZ-αβTCR's outstanding capacity to prevent GVHD and ameliorate an ongoing GVHD while sparing immune cells other than αβT cells strongly recommends GZ-αβTCR for the prevention and treatment of acute GVHD in clinical settings.

Preemptive administration of human αβ T cell receptor-targeting monoclonal antibody GZ-αβTCR potently abrogates aggressive graft-versus-host disease in vivo.

Article	Year
Twinning in tufted capuchins (Cebus apella): rate, survivorship, and weight gain. Folia primatologica; international journal of primatology\| PubMed ID: 14716148	2004
Human Peripheral CD4(+) Vδ1(+) γδT Cells Can Develop into αβT Cells. Frontiers in immunology\| PubMed ID: 25709606	2014
Cancer-targeted IL-12 controls human rhabdomyosarcoma by senescence induction and myogenic differentiation. Oncoimmunology\| PubMed ID: 26140238	2015
Chronic graft-versus-host-disease in CD34(+)-humanized NSG mice is associated with human susceptibility HLA haplotypes for autoimmune disease. Journal of autoimmunity\| PubMed ID: 26143958	2015
Preemptive administration of human αβ T cell receptor-targeting monoclonal antibody GZ-αβTCR potently abrogates aggressive graft-versus-host disease in vivo. Annals of hematology\| PubMed ID: 26264693	2015

Christian Welker

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