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Transcriptional profile of t follicular helper (Tfh) cells and b cell isotype switching induced by type 1 or type 2 adjuvants

Grant number: 14/19906-2
Support type:Regular Research Grants
Duration: August 01, 2015 - July 31, 2018
Field of knowledge:Biological Sciences - Immunology
Cooperation agreement: Fundação para a Ciência e a Tecnologia (FCT)
Principal Investigator:Momtchilo Russo
Grantee:Momtchilo Russo
Principal investigator abroad: Luis Ricardo Simões da Silva Graça
Institution abroad: Universidade de Lisboa, Portugal
Home Institution: Instituto de Ciências Biomédicas (ICB). Universidade de São Paulo (USP). São Paulo , SP, Brazil
Assoc. researchers:Ana Paula Lepique ; Anderson de Sá Nunes ; Juliana Bortolatto ; Luciana Cristina Mirotti ; Luis Ricardo Simões da Silva Graça

Abstract

A specific subset of CD4 T cells, namely T follicular helper (Tfh) cells, characterized by expression of transcription factor Bcl6, are unique in their ability to access the B cell follicle and drive germinal center (GC) reactions leading to isotype switching, affinity maturation, and production of high affinity antibodies and memory B cells. Therefore, in addition to the influence of "classical" Th1, Th2 (and Th17) cells in driving selection of Ig isotypes, it is critical to understand how Tfh cells (or subsets) participate in this process. The role of Tfh cells in allergic responses remains to be clarified because Bcl6 represses the expression GATA-3, a Th2 of transcription factor. However, GC Tfh cells could produce IL-4 (but not IL-13) in a GATA-3-independent process. Tfh cells also produce IL-21, a cytokine that is important for their differentiation and maintenance. IL-21 was reported to reduce switching to IgE, as mice deficient in IL-21R make high levels of IgE, and intranasal administration of IL-21 reduced IgE response to ovalbumin (OVA) in mice. In addition, it has been shown that generation of IgE-producing plasma and memory B cells can occur in mice, from an extrafollicular origin following sequential class-switch recombination. In humans the follicular vs. extrafollicular origin of IgE was not so thoroughly investigated, although local IgE production on mucosal surfaces, often associated with GC formation has been documented in allergic patients. Our proposal is to determine the transcriptional profile of Tfh cells induced in Th2 or Th1 conditions. We will address two competing hypotheses: One hypothesis is that "help" provided by Tfh cells in the course of a Th2-associated immune response is responsibility of a non-specialized, generic, Tfh cell. Thus, isotype-switch is influenced by contact with generic Tfh cells and cytokines produced in the vicinity by extrafollicular Th2 cells. The alternative model postulates the existence of Tfh subsets specialized in providing Th2-type "help" (a subset we name here Tfh2), but also Th1- and Th17-type of "help" (putative Tfh1 and Tfh17 subsets). The acquisition of such specialized Tfh function may be a consequence of early interaction with dendritic cells (DC), or following interaction with B cells in the T-B border, and it may be influenced by cytokines produced by Th2, Th1 or Th17 cells. Our strategy will involve adoptive cell transfer of TCR transgenic CD4 T cells bearing a congenic marker, followed by immunization with adjuvants leading to type 2 (alum) or type 1 (TLRs agonists) immune responses. This way we will be able to recover antigen-specific cells and analyze the different characteristics acquired by those antigen-specific cells under type 1 or type 2 conditions. Subsequent RNA-Seq studies will allow the identification of signature genes for further validation. The use of gene expression data appears to be the best available method to address a hypothesis (that Tfh subsets are involved in type 1 vs. type 2 responses) and not simply to identify genes associated with a certain phenotype. Our approach will lead to acquisition of information regarding gene expression profiles associated with T cells at different stages of functional specialization: Th1, Th2, pre-Tfh, Tfh, and possibly distinct Tfh subsets. We will, therefore, use this information to identify key molecules for the function of distinct Tfh subsets. In addition, we will use a mouse model of OVA-immunization under type-1 or type-2 conditions and Bcl6-Cre mice to ablate candidate genes on Tfh cells. (AU)