Imunologia Molecular

Interesse da Investigação

T lymphocytes play a crucial role in cellular immunity against pathogens, and are also implicated in tumor surveillance. We are interested in the processes of generation, activation and regulation of T lymphocytes. We are currently concentrating on γδ T cells, and their roles in the control of inflammation and tumor surveillance. Using state-of-the-art research tools (human and mouse genome databases, bioinformatics, cDNA microarrays, RNA interference), we envisage the identification of molecules involved in those processes and their integration in cellular mechanisms, to be translated into new treatments for cancer, inflammatory and autoimmune diseases.

Membros do Grupo

Projecto de Investigação

Characterization of regulatory γδ T cell subsets and their mechanisms of action

The phenotype of animal models genetically deprived of γδ T cells clearly suggests the existence of sub-populations of regulatory lymphocytes within that lineage. Thus, in various experimental systems, γδ-deficient mice have higher incidence (relative to controls) of inflammatory lesions, particularly in the skin, gut and brain. Moreover, the pathologic impact of some autoimmune diseases is also exaggerated in animals lacking γδ T cells (for example, in animal models of multiple sclerosis and lupus erythematosus). Although γδ T cells display an aggregate anti-inflammatory phenotype, and express common mediators of T cell suppression (such as TGF-β, IL-10 or CTLA-4), they also constitute a very heterogeneous lineage of lymphocytes, for which we lack the identification of its regulatory subset(s). In this project we aim at characterizing, in terms of surface phenotype, gene expression profile, and mechanisms of action, γδ T cell subsets with immunoregulatory potential. To address this question, we isolate γδ T cells from several mouse tissues and sub-divide them accoridng to the expression of various surface markers, and then analyze their gene expression profiles and their suppressor activity, i.e., the capacity to inhibit the proliferation of other T cells (effector CD4+ CD25- or CD8+ lymphocytes), both in vitro and in vivo. Our main objectives are:
1.To identify murine and human γδ T lymphocyte subsets endowed with the capacity of suppressing αβ T cell responses, namely their proliferation and cytokine secretion, in vitro.
2.To test the novel murine subsets for their ability to prevent inflammation in vivo, in an inflammatory bowel disease mouse model for human Crohn’s disease and ulcerative colitis.
3.To identify molecular mediators of suppression, addressing cell-contact dependent and independent mechanisms. Investigate the role of candidate molecules involved in the suppression, by isolating subsets from genetically modified mice and by using blocking antibodies in vitro and in vivo.
4.To characterize the cellular and molecular mechanisms involved in the generation of the regulatory subsets in the thymus. In partivular, we will address the role of trans-conditioning by CD4+ CD8+ (DP) thymocytes on the development of regulatory T cell subsets (Silva-Santos et al., Science 307: 925-928).

Projecto de Investigação

Molecular mechanisms of γδ T cell recognition and activation in tumor surveillance

Animals lacking γδ T cells are particularly susceptible to the development of tumors. Moreover, γδ T lymphocytes are potent killers of multiple tumor cell lines, including human lymphomas, leukaemias and carcinomas. However, the molecular mechanisms responsible for this activity are unknown: how are tumor cells recognized as targets, and how are the γδ T lymphocytes activated for cytolysis? To answer these questions, we analyze mouse and human tumor cell lines capable of stimulating γδ T lymphocytes. We compare their repertoire of surface proteins, with that of non-activating cells. We also employ bioinformatics tools to identify (based on structural criteria) possible candidates in the murine and human genomes. We are particularly interested in Major Histocompatibility Complex (MHC)-related proteins, such as non-classical class Ib molecules, and their roles in the anti-tumor activity of γδ T lymphocytes. The expression of the selected molecules is studied (by real-time polymerase chain reaction) in a large panel of tumor cell lines and tumor biopsies from patients or animal models. Aspects of the intracellular biology of the candidates are investigated by immunofluorescence and biochemical experiments. These also envisage the identification of molecular partners, for example, receptors on the surface of γδ T lymphocytes that interact with the candidate ligands expressed on tumor cells. Finally, we use RNA-interference to specifically knock-down the expression of the genes of interest in tumor cell lines, and then assess the impact on γδ T cell-mediated tumor cell lysis, either in vitro (against murine and human tumor cell lines) or in vivo (in animal models for tumor development).

Funding

PTDC/BIA-BCM/71663/2006
"Identification of ligands for γδ T cell receptors: a genomics/ transcriptomics approach". Fundação para a Ciência e Tecnologia

PTDC/SAL-MII/71662/2006:
"Molecular mechanisms of tumour antigen recognition and cellular activation of γδ T lymphocytes". Fundação para a Ciência e Tecnologia,

EMBO Young Investigator Programme – project 1440
Installation Grant from the European Molecular Biology Organization

Colaboradores

Queen Mary College, London, United Kingdom
Dr. Daniel Pennington

Instituto de Medicina Molecular, Lisboa, Portugal
Dr. Luís Moita
Dr. João Barata
Dr. Maria Manuel Mota
Dr. Luís Graça

King’s College London, United Kingdom
Prof. Adrian Hayday

The Netherlands Cancer Institute, Amsterdam, The Netherlands
Dr. Jannie Borst

Instituto Gulbenkian de Ciência, Oeiras, Portugal
Dr. Jocelyne Demengeot

Publicações

Gomes, A.Q, Correia, D.V. and Silva Santos, B. (2007). Non-classical Major Histocompatibility Complex proteins as determinants of tumour immunosurveillance. EMBO Reports. in press

Pennington, D.J., Silva Santos, B., Escorcio-Correia, M., Silberzahn, T. and Hayday, A.C. (2006). Early events in the thymus affect the balance of effector and regulatory T cells. Nature, 444, 7122 :1073-7

Graça, L., Silva Santos, B., Coutinho, A. (2006). The blind-spot of regulatory T cells. Eur. J. Immunol. 36, 4 :802-5

Silva Santos, B.*, Pennington, D.J.*, and Hayday, A.C.(* co-first authors) (2005). Lymphotoxin-mediated regulation of role of γδ cell differentiation by αβ T cell progenitors. Science. 307 :925-928

Silva Santos, B.*, Pennington, D.J.*, and Hayday, A.C. (* co-first authors) (2005). γδ T cell development – having the strength to get there. Curr. Opin. Immunol. 17, 2 :108-115. Review

Silva Santos, B.*, Pennington, D.J.*, Shires, J., Theodoridis, E., Pollit, C., Wise, E.L., Tigelaar, R.E., Owen, M.J. and Hayday, A.C. (* co-first authors) (2003). The inter-relatedness and interdependence of mouse T cell receptor γδ+ and αβ+ cells. Nature Immunology. 4, 10 :991-999.