ABSTRACT
OBJECTIVE: Nuclear receptors are known to regulate both immune and barrier functions in the GI tract. The nuclear orphan receptor NR2F6 has been shown to suppress the expression of proinflammatory cytokines in T lymphocytes. NR2F6 gene expression is reduced in patients with IBS or UC, but its functional role and tissue dependency in healthy and inflamed gut have not yet been investigated. DESIGN: Intestinal inflammation was induced in wild-type, Nr2f6-deficient, Rag1-deficient or bone marrow-reconstituted mice by administration of chemical (dextran sodium sulfate (DSS)) and immunogenic (T cell transfer) triggers. Disease phenotypes were investigated by survival, body weight, colon length and analysis of immune cell infiltrates. Additionally, histology, intestinal permeability, tight junction proteins, bacterial fluorescence in situ hybridisation, apoptosis, cell proliferation and mucus production were investigated. RESULTS: Nr2f6-deficient mice were highly susceptible to DSS-induced colitis characterised by enhanced weight loss, increased colonic tissue destruction and immune cell infiltration together with enhanced intestinal permeability and reduced Muc2 expression. T cell transfer colitis and bone marrow reconstitution experiments demonstrated that disease susceptibility was not dependent on the expression of Nr2f6 in the immune compartment but on the protective role of NR2F6 in the intestinal epithelium. Mechanistically, we show that NR2F6 binds to a consensus sequence at -2 kb of the Muc2 promoter and transactivates Muc2 expression. Loss of NR2F6 alters intestinal permeability and results in spontaneous late-onset colitis in Nr2f6-deficient mice. CONCLUSION: We have for the first time identified a fundamental and non-redundant role of NR2F6 in protecting gut barrier homeostasis.
Subject(s)
COUP Transcription Factors/metabolism , Colitis/metabolism , Colitis/pathology , Animals , Colitis/etiology , Dextran Sulfate , Disease Models, Animal , Intestinal Mucosa/metabolism , Intestinal Mucosa/pathology , Mice , Mucin-2/metabolism , Repressor Proteins , Tight Junction Proteins/metabolismABSTRACT
Nuclear receptor subfamily 2, group F, member 6 (NR2F6) is an orphan member of the nuclear receptor superfamily. Here, we show that genetic ablation of Nr2f6 significantly improves survival in the murine transgenic TRAMP prostate cancer model. Furthermore, Nr2f6(-/-) mice spontaneously reject implanted tumors and develop host-protective immunological memory against tumor rechallenge. This is paralleled by increased frequencies of both CD4(+) and CD8(+) T cells and higher expression levels of interleukin 2 and interferon γ at the tumor site. Mechanistically, CD4(+) and CD8(+) T cell-intrinsic NR2F6 acts as a direct repressor of the NFAT/AP-1 complex on both the interleukin 2 and the interferon γ cytokine promoters, attenuating their transcriptional thresholds. Adoptive transfer of Nr2f6-deficient T cells into tumor-bearing immunocompetent mice is sufficient to delay tumor outgrowth. Altogether, this defines NR2F6 as an intracellular immune checkpoint in effector T cells, governing the amplitude of anti-cancer immunity.
Subject(s)
CD4-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/immunology , COUP Transcription Factors/genetics , Immunologic Surveillance , Immunotherapy, Adoptive/methods , Prostatic Neoplasms/therapy , Animals , CD4-Positive T-Lymphocytes/cytology , CD4-Positive T-Lymphocytes/transplantation , CD8-Positive T-Lymphocytes/cytology , CD8-Positive T-Lymphocytes/transplantation , COUP Transcription Factors/deficiency , COUP Transcription Factors/immunology , Cell Line, Tumor , Gene Expression Regulation, Neoplastic , Humans , Immunologic Memory , Interferon-gamma/agonists , Interferon-gamma/genetics , Interferon-gamma/immunology , Interleukin-2/agonists , Interleukin-2/genetics , Interleukin-2/immunology , Lymphocyte Activation , Male , Mice , Mice, Inbred C57BL , Neoplasm Transplantation , Prostatic Neoplasms/genetics , Prostatic Neoplasms/immunology , Prostatic Neoplasms/mortality , Repressor Proteins , Signal Transduction , Survival Analysis , Tumor Necrosis Factor-alpha/agonists , Tumor Necrosis Factor-alpha/genetics , Tumor Necrosis Factor-alpha/immunologyABSTRACT
Aging is associated with an accruing emergence of non-functional tissues. Mesenchymal stem cells (MSC) bring forth progenitors with multi-lineage differentiation potential, yet, they also exhibit anti-inflammatory and tissue-protective properties. Due to aging, altered tissue microenvironments constrict controlled stem cell proliferation and progenitor differentiation, thus diminishing the fitness of MSC. Therefore, deepening our understanding of metabolic, molecular and environmental factors impacting on MSC during human aging as well as providing new vistas on their role in promoting healthy aging and preventing age-associated disease is pivot. It is anticipated that integrative quantification of systemic parameters dominantly impacting on MSC will also enable effective personalized prognosis and provision of effective early medical interventions. Working along this line, it can be envisaged that standards in medical therapies can be individually adjusted by accounting not solely for the patient's chronological age or other physical parameters rather than specific physiological parameters which are believed to functionally shape stem cell niches within the bone marrow.
Subject(s)
Cellular Senescence , Mesenchymal Stem Cells/cytology , Aged , Bone Diseases, Metabolic/genetics , Bone Marrow/pathology , Bone Marrow Cells/cytology , Cell Differentiation , Cell Lineage , Cell Proliferation , Humans , Inflammation , Osteoclasts/cytology , Risk Factors , Stem Cell Niche , Stem Cells/cytologyABSTRACT
Bone-derived stroma cells contain a rare subpopulation, which exhibits enhanced stemness characteristics. Therefore, this particular cell type is often attributed the mesenchymal stem cell (MSC). Due to their high proliferation potential, multipotential differentiation capacity, and immunosuppressive properties, MSCs are now widely appreciated for cell therapeutic applications in a multitude of clinical aspects. In line with this, maintenance of MSC stemness during isolation and culture expansion is considered pivot. Here, we provide step-by-step protocols which allow selection for, and in vitro propagation of high quality MSC from human bone.
