Cytokines and Thelper subsetsI. Characteristics of Cytokines (CKs)Cytokine NetworkPowerPoint PresentationSlide 5Activation of Cells = Production of CytokinesCytokine Network on a Local LevelII. Measuring CytokinesAntigen Capture ELISA for IL-2III. Actions of CytokinesActions of CytokinesIV. Properties of Cytokines PleiotropySynergy Experimental ExampleSlide 14Slide 15VI. CD4+ T helper Subsets Th1/Th2 Cytokine BiasSlide 17Th0 ---> Th1 or Th2 Original mouse experiments on Th cells (Mosmann et al (DNAX) 1986 J Immunol)Th1/Th2Th1/Th2 Antagonism IL-4 blocks Th1 IFN-g blocks Th2Th1/Th2 Regulation T-bet is a transcription factor that is required for Th1 specific genes such as IL-12RbIL-4 vs IFN-gRole for Th1 vs Th2 in Immune ResponseWhat controls Th1 vs Th2?Influence of APC Subsets on Th1/ Th2 Dendritic cellToll-like receptors (TLRs) Influence of APCs on Th1/ Th2TLR vs IL-12 in Th1/ Th2 developmentMyD88-/- mice fail to control acute Toxoplasma infectionMyD88-/- mice default to Th2 Role for TLR Activation in Th1/Th2Th Cytokine Bias in Disease ExamplesTh Cytokine Bias in Disease:Leprosy Skin disease caused by Mycobacterium lepraeCytokine Bias in LeprosySlide 33VII. Cytokine ReceptorsCytokine Receptor FamiliesIL-2 Receptor Subfamily Shared common g subunitX-Linked SCID Common g chain DeficiencyGM-CSF Receptor Subfamily Hematopoietin ReceptorsSlide 39Cytokine Receptor Signaling JAKs and STATs (Model of Signal)JAKs and STATs Usage by CK ReceptorsSlide 42Cytokines and Thelper subsetsMCB 150, Pr CoscoyI. Characteristics of Cytokines (CKs) CKs are small proteins (<30 kDa) CKs are similar to hormones and growth factors. CKs control the immune system. Cytokines include subsets called lymphokines, monokines, chemokines, etc.Cytokine Network CKs are part of complex system that regulates the immune system. CKs are primarily produced by immune system but many other organs (liver, brain, endocrine glands) make CKs to influence immune response.Cytokine NetworkT helper and Macrophage Interactionhighlighted Effector T cellTh cells control immune system through CKs.Activation of Cells = Production of CytokinesProduction of CKs is usually part of immune cell activation. Cytokines are produced in response to different stimuli (e.g antigen receptor, cytokine/chemokine receptor).T cell IL-2IL-2 geneB cellIL-2 ReceptorIL-4Cytokine Network on a Local LevelIL-4 ReceptorIL-4 geneII. Measuring CytokinesProtein amount by ELISA. Good for in vitro experiments.Protein amount by bioactivity assay using CK dependent cell lines.RNA message by PCR.Antigen Capture ELISA for IL-2III. Actions of CytokinesAutocrine acts on same cell that produced it. IL-2 for T cell activation Paracrine acts on nearby cells. T cell help for B cellsEndocrine acts on cell at a distance (through bloodstream). Inflammatory cytokines.Actions of CytokinesIL-2 forT-cell activationT-cell helpfor B cellsInflammatory cytokinesIV. Properties of CytokinesPleiotropySame cytokine has different effects on cells can be activating or inhibiting. Example: IL-10 can be inhibitory to macrophages and Th1 cells yet activating for Th2 cells and B cells.SynergyExperimental ExampleProliferation of CTLLs(CTLLs are cytokine dependent cell line)Both IL-2 or IL-4 activate CTLLs in vitro. Maximal proliferation obtained when both IL-2 and IL-4 added. Both cytokines need to be blocked in order to inhibit growth.Properties of CytokinesPleiotropyRedundancySynergyAntagonismCK Secreted Primary target HallmarkIL-1 Macs,endotheliumIncreases CAMsActivates macsPro-InflammatoryIL-2 Th cells Th1, Tc, NK cells Th1 cytokineIL-4 Th2 cells Th2 cells, B cells Th2 cytokineIL-6 Macs Th2 cells Th2 cells, B cells ProInflam/Th2IL-7 Bone marrow Stem cells HematopoiesisIL-8 Neutrophils Endothelial cells ChemokineIL-10 Th2 cells, Macs, Th2 and B cells Th2 cytokineIL-12 Macrophages,dendritic cellsTh1, Tc, NK cells Th1 cytokineTNF-α Macs, T, Bendothelialtumor cells, macs,B cells, T cellsPro-InflammatoryTNF-β Th1 cells macrophages, Th1cells,InflammatoryIFN-α/βNeutrophils,endo/epithelialUninfected cells Anti-viralcytokinesIFN γTh1 cells,macrophagesmacrophages, Th1 Th1 cytokineInflammatoryV. FunctionsOf CytokinesVI. CD4+ T helper SubsetsTh1/Th2 Cytokine BiasCD4+ Thelper cells can be divided into subsets based on their cytokine production. Th1 cells produce IL-2, IFN-, TNF- CKs which activate cell mediated immunity Th2 cells activate IL-4, IL-6, IL-10 CKs that activate humoral immunity These Th subsets were originally identified using mouse T cell clones.Mouse Th SubsetCytokine Th1 Th2Table 12-4 from GoldsbyTh0 ---> Th1 or Th2Original mouse experiments on Th cells(Mosmann et al (DNAX) 1986 J Immunol) Antigen specific T cells placed in culture with antigen and APCs to make T cell lines. Spleen cells (Th0) add IL-12  Th1 cells IL-2, IFN-, TNF-Spleen cells (Th0) add IL-4  Th2 cells  IL-4, IL-6,IL-10(Th0 --precursor cell that produces IL-2, IL-4, and IFN-.)Th1/Th2 Naïve Th0 IL-2, IL-4, IFN-IL-4IL-12EffectorTh1 cellIL-2, IFN-EffectorTh2 cellIL-4, IL-6, IL-10Th1/Th2 AntagonismIL-4 blocks Th1 IFN- blocks Th2IL-4IL-12 Th1 cellIL-2, IFN-Th2 cellIL-4, IL-6, IL-10IFN-IL-4Th1/Th2 RegulationT-bet is a transcription factor that is required for Th1 specific genes such as IL-12RIL-12 Th1 cellIL-2, IFN-Th2 cellIL-4, IL-6, IL-10IFN-Enhances T-betIL-4 blocks T-betIL-4 vs IFN-T-bet (Th1 associated) activated by IFN- and turned off by IL-4. Conversely in Th2 transcription factor GATA-3 activated by IL-4 turned off by IFN-.Role for Th1 vs Th2 in Immune ResponseBoth subsets activated in lymph nodes (LN) immune responses to complex antigens. Th1 cells leave LN to find activated endothelium tissue to activate macrophages.Th2 cells can stay in LN to activate B cells.What controls Th1 vs Th2? 1) Amount of antigen. Mouse experiments originally showed high dose for Th1. 2) MHC and TCR affinity. High affinity TCR = Th1. 3) Dendritic cell subsets during activation. APC subsets activate Th1 or Th2 preferentially. 4) Toll-like receptor activation.Influence of APC Subsets on Th1/ Th2Dendritic cellMyeloid-like dendritic cells produce abundant IL-12 and drive Th1. Lymphoid-like dendritic cells produce low levels of IL-12 are permissive for Th2.Toll-like receptors (TLRs) Influence of APCs on