Examples of questions for Cellular Immunology/Cellular Biology and Immunology

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1 Examples of questions for Cellular Immunology/Cellular Biology and Immunology Each student gets a set of 6 questions, so that each set contains different types of questions and that the set of questions covers different chapters of the main lectures (there are no specific questions on the case studies). At the start of the exam, you draw a set of 4 questions for which you get 30 min. written preparation. During the oral exam, the answer to these 4 questions is discussed. You also get 2 additional questions to be directly answered orally. These last 2 questions count for 5 out of 20 marks on the exam. Examples of the 4+2 questions (and the corresponding answer) can be found below (P.S. The full set of 6 questions below is not included in the exam, but some of you may randomly draw a set containing one or a few of the questions below) 1) What are the main types of helper T cells? How do these differ in terms of their main immunological functions and in the defining cytokines and transcription factors involved in their induction and function? (5/20) Th1 Priming and amplifying cytokines: IL-12, IFN-g Polarizing transcription factor: T-bet (induced by IFN-g) Unique cytokine products: IFN-g Main functions: signaling via IFN-g and CD40L induces cellular immunity against intracellular pathogens, by acting on cells of cellular immunity: o macrophage activation o CD8 + CTL activation o class switching in B cells, resulting in opsonizing antibodies (that can interact with Fcg receptors on macrophages) Th2 Priming and amplifying cytokines: IL-25, IL-4 Polarizing transcription factor: GATA-3 (induced by IL-4) Unique cytokine products: IL-4, IL-13, IL-5, IL-10 Main functions: signalling via Th2 cytokines and CD40L induces humoral immunity against extracellular pathogens, by inducing class switching in B cells, resulting in neutralizing antibodies and IgE antibodies interacting with Fce receptors on eosinophils Th17 Priming and amplifying cytokines: TGFb, IL-6, IL-23 Polarizing transcription factor: RORgT (induced by TGFb + IL-6) Unique cytokine products: IL-17A, IL-17F, IL-21, IL-22 Main functions: antimicrobial activity against extracellular bacteria at epithelial surfaces. Hereto, the Th17 cytokines act on epithelial, endothelial, stromal cells. These activated epithelial, endothelial, stromal cells will: o secrete antimicrobial peptides such a b-defensins o secrete inflammatory cytokines and recruit neutrophils

2 2) What are the main stages of B cell development and activation in the spleen? How is this affected by T-cell help? (5/20) Interaction between T helper cells and follicular B cells (dependent on antigen presentation by the B cells on MHCII and on CD40L and cytokine signals from the Th cell) is needed for the germinal centre reaction, allowing: o Class switching (from IgM to IgA/G/E) o Somatic hypermutation (-> allowing affinity maturation of the antibodies, since the B cells that randomly generate the highest affinity antibodies will be able to most effectively take up the antigen through the B cell receptor (surface-bound antibody) and thus successfully compete with other B cells for the T cell help) o Generation of memory B cells (-> T-cell dependent generation of B cell memory)

3 3) Which stimuli can induce maturation and activation of dendritic cells? (2.5/20) 4) What is IL-2? By which cells is it expressed and by which stimuli is its expression regulated? What are its main immunological functions? (2.5/20) o Interleukin-2 = a cytokine o secreted by effector T cells, when these are activated by signalling through the TCR, recognizing peptides presented on MHC molecules. IL-2 production by effector T cells is enhanced when the T cells also receive a costimulatory signal, for example through B7 molecules expressed on antigen presenting cells, binding to CD28 expressed on the T cells. o main growth factor for T cells: important for proliferation and activation of effector T cells in the induction of immunity critical for immune tolerance: by inducing activation-induced cell death in effector T cells as growth factor for Tregs (Tregs need IL-2, but do not secrete IL-2 themselves) (Remark: the function of IL-2 as growth factor for effector T cells can be partly compensated by other growth factors such as IL-7, but not its role in tolerance induction. As a consequence, IL-2 deficient mice develop a lethal lymphoproliferative disease, in which the control of T cell proliferation is lost).

4 Examples of additional questions at the oral exam 5) In which cases can B cell mediated antibody responses be elicited in MHC class II deficient individuals? Why? (2.5/20) MHC class II is needed for positive selection of CD4+ helper T cells during thymic development. MHC class II is also needed for interaction between Thelper cells and follicular B cells in germinal centres. Thus, in MHC class II deficient individuals, there will be no helper T cells and no T cell help for B cell development. As a consequence, you will only get T-cell independent (thymus-independent) antibody responses (by splenic Marginal Zone B-cells and peritoneal B-1 B cells). Two types of thymus-independent (TI) B-cell activation can occur: TI type 1 activation is when a B cell binds to an antigen and receives secondary activation by Toll-like receptors, leading to polyclonal B-cell activation or specific Ab response TI type 2 activation when the antigen is a molecule with multiple repeating subunits which can simultaneously cross link enough B cell receptors to fully activate the B cell (e.g. Ficoll or repetitive polysaccharide antigens on encapsulated bacteria) 6) Which method(s) could be used to study in vivo the role of granzyme B expressed by regulatory T cells in limiting auto-immunity? (2.5/20) If you study the development of autoimmunity in granzyme B deficient mice, all cells (including for example effector CD8+ T cells) will lack granzme B, not only the Tregs. You could isolate Tregs from granzyme B deficient mice and adoptively transfer them into immunodeficient mice (for example athymic nude mice), together with adoptive transfer of effector Tcells from wild-type mice but this may not be a very natural system. To study the role of granzyme B expressed specifically by regulatory T cells, you could use a Treg-specific granzyme B KO mouse, based on the Cre-Lox system. Hereby, you need transgenic mice in which LoxP recombination sites are introduced in the introns of the granzyme B gene. In these floxed mice, the LoxP sites will be spliced out of the granzyme B mrna, resulting in normal granzyme B expression in most cells. However, in cells expressing cre recombinase, recombination of the granzyme B gene will result in a non functional mrna. In addition, you also need another transgenic mouse strain in which the cre recombinase is expressed specifically in Tregs, which could for example be obtained by expressing the cre under control of the Treg-specific FoxP3 promotor (for example introducing the cre

5 gene downstream of the FoxP3 coding region in the genome, with an internal ribosome entry site in between, to allow expression of both FoxP and Cre protein from the same mrna that is under control of the FoxP3 promotor). When breeding the FoxP3-Cre mice with the Granzyme B-LoxP mice, the resulting offspring will contain mice in which Granzyme B is specifically inactivated in cells expressing FoxP3 (i.e. Tregs) and is normal in other cells.

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