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Leukocyte Differentiation Antigens Cell Adhesion MoleculesNK cellsContents Concept of leukocyte differentiation antigens （白细胞分化抗原）Concept of cluster of differentiation （CD分子，分化群）Cell adhesion molecules NK cellsLeukocyte differentiation antigens Leukocyte differentiation antigens:Cell surface molecules which may appear or disappear from the cell membrane of leukocytes in the different stages of differentiation and activation.（第6版p58）Leukocyte differentiation antigens are also found on cells other than leukocytes,such as erythrocytes,endothelial cells,epithelial cells.Immunoglobulin superfamily Cytokine receptor family C-Type lectin superfamily Integrin family TNF superfamily TNF receptor superfamily Cluster of differentiation(CD):Cell surface molecules can be recognized by particular monoclonal antibodies.All of the monoclonal antibodies that react with a particular membrane molecule are grouped together as a cluster of differentiation(CD).Each new mAb that recognizes a cell surface molecule is analyzed for whether it falls into a recognized CD designation;if not,it is given a new CD designation that reflects a new membrane molecule.CD1CD363,up to 363 most recently(as of 2010)in humanHuman CDGroups CD（example）T cell CD2、CD3、CD4、CD5、CD8、CD28、CD152(CTLA 4)、CD154(CD40L)、CD272(BTLA)、CD278(ICOS)、CD294(CRTH2)B cell CD19、CD20、CD21、CD40、CD79a(Ig)、CD79b(Ig)、CD80(B7-1)、CD86(B7-2)、CD267(TACI)、CD268(BAFFR)、CD269(BCMA)、CD307(IRTA2)Myeloid cell CD14、CD35(CR1)、CD64(Fc RI)、CD256(APRIL)、CD257(BAFF)、CD312(EMR2)Platelet CD36、CD41、CD42a CD42d、CD51、CD61、CD62PNK cell CD16(Fc RIII)、CD56(NCAM-1)、CD94、CD158(KIR)、CD161(NKR-P1A)、CD314(NKG2D)、CD335(NKp46)、CD336(NKp44)、CD337(NKp30)Non-lineage CD30、CD32(Fc RII)、CD45RA、CD45RO、CD46(MCP)、CD55(DAF)、CD59、CD252(OX40L)、CD279(PD1)、CD281 CD284(TLR1-TLR4)、CD289(TLR9)、CD305(LAIR-1)、CD306(LAIR-2)、CD319(CRACC)Human CDGroups CD（example）Adhesion molecules CD11a-CD11c、CD15、CD15s(sLx)、CD18、CD29、CD49a CD49f、CD54(ICAM-1)、CD62E、CD62L、CD324(E-cadherin)、CD325(N-cadherin)、CD326(EpCAM)Cytokine/CD25(IL-2R)、CD95(Fas)、CD116-CDw137、CD178(FasL)、Chemokine receptors CD183(CXCR3)、CD184(CXCR4)、CD195(CCR5)、CD261 CD264(TRAIL-R1 TRAIL-R4)Endothelia CD105(TGF-RIII)、CD106(VCAM-1)、CD140(PDGFR)、CD144、CD299(DCSIGN-related)、CD309(VEGFR2)、JAM1(CD321)、JAM2(CD322)Carbohydrate structures CD15u、CD60a CD60c、CD75、CDw327 CDw329Dendritic cells CD80,CD86,CD85Stem cells CD133、CD243 Erythrocytes CD233 CD242 Stromal cells CD331-334（FGFR1-FGFR4）The CD system is commonly used as cell markersCD8CD8cytotoxicCell types and subtypes defined by CDsB cellT cellCD4+T cellTregs(CD4+CD25+)Type of cell CD markers stem cells CD34+,CD31-all leukocyte groups CD45+Granulocyte CD45+,CD15+Monocyte CD45+,CD14+T lymphocyte CD45+,CD3+T helper cell CD45+,CD3+,CD4+Cytotoxic T cell CD45+,CD3+,CD8+B lymphocyte CD45+,CD19+or CD45+,CD20+Thrombocyte CD45+,CD61+Natural killer cell CD16+,CD56+,CD3-Rules for assigning a CD number at least one Workshopcharacterized antibody and good molecular data.the antigen is expressed on the surface of cells involved in immune reactions.the antibodies react with primary cells(not just transfectant or recombinant protein).the antibody is available.CD acts as Receptors or Ligands.Initiates a signal cascade,altering the behavior of the cell,or has other functions,such as cell adhesion.Functions of CDs1.Mediate cell-cell interaction:Homing,activation,effecter functionsFunctions of CDs2.Receptors recognize exogenous or soluble substance:pathogens,antibodies,cytokinesCDs related to T cell activationTCR signalingCD4 and CD8transmembrane glycoproteinco-receptor of T cell*CD4:first and second domains bind to nonpolymorphic region of MHC molecules*CD8:IgV-like domain of chain binds to nonpolymorphic 3 region of MHCmolecules*for cell adhesion *Involved in TCR-CD3 signal transduction APCT细胞细胞Structure of CD4Structure of CD8CCR51.CD4 is a primary receptor used by HIV-1 to gain entry into host T cells.2.HIV-1 attaches to CD4 with a protein in its viral envelope known as gp120.The binding to CD4 creates a shift in the conformation of gp120 allowing HIV-1 to bind to other surface receptors on the host cell,the chemokine receptors CCR5.Following a structural change in another viral protein(gp41),HIV inserts a into the host cell that allows the outer membrane of the virus to fuse with the cell membrane.3.HIV infection leads to a progressive reduction in the number of T cells possessing CD4 receptors.Therefore,medical professionals refer to the CD4 count to decide when to begin treatment for HIV-infected patients.Normal blood values are 500-1200 x106/L.CD40 and CD40L(CD154)BCR complexco-receptorCDs related to B cell activationCD79/CD79 Heterodimer cytoplasmic domains contain ITAM.Function:1.transfers the signals that lead to B cell activation.2.transports the IgBCR signalingCD19/CD21/CD81CD19/CD21/CD81 interactions with complement associated with antigen play a role in antigen-induced B-cell activation.CD21=CR2,C3dR,EBV RCDs constituting FcR familyDifferent types of human Fc receptorsRole of FcRI(CD64)in the Ab-mediated opsonization of microbesRole of FcRIII(CD16)and FcRI in ADCC(antibody-dependent cell-mediated cytotoxicity)Cell adhesion molecules,CAM A group of proteins involved in adhesion between cells or between cells and extracellular matrix(ECM)CAMs and CD molecules are respectively designated according to the different aspects of these molecules.CAMs are grouped depending on their adhesion functions.CD molecules contain the majority of CAMs that have obtained CD designation.A minority of CAMs have not been given CD designation.Categories of CAM1.IgSF(immunoglobulin superfamily)2.Integrin family3.Selectin family4.Cadherin family5.Others:CD44 IntegrinIntegrin family is so named because the molecules of this family primarily mediate the cellular adherence to extracellular matrix,enabling cells and extracellular matrix to form the integration.Structure:Heterodimeric proteins consisting of two non-covalently bound polypeptides (chain 1 out of 18,and chain 1 out of 8).Components of integrin family 1:VLA(very late activation antigen,12 members),ligand:VCAM-1(vascular cell adhsion molecule-1)T cell activation(co-stimulation and adhesion)2:LFA-1(CD11a,lymphocyte function associated antigen-1,4 members);Mac-1(CD11b)ligand:ICAM-1(intercellular adhesion molecule-1)T cell activation,circulation,inflammation 3:gpIIb ligand:Fg(fibrinogen)Platelet aggregationDistribution Broad distribution on the surface of cells.Selectin family includes L-selectin,P-selectin and E-selectin,which play important roles in leukocyte adhesion to endothelium,inflammation and lymphocyte homing.Selectin family1.Structure Transmembrane glycoproteins,with a number of extracellular domains homologous to those seen in complement control proteins(CCP).The extracellular region also contains a domain related to the EGF receptor and a distal C type lectin-like domain.2.Members of selectin family L-selectin:leukocytes(Cd62L)P-selectin:platelets and megakaryocytes(Cd62P)E-selectin:activated endothelia(Cd62E)3.Ligand:carbohydrates,such as CD15sFunctions of CAM1.Serving as co-receptors and providing co-stimulatory signals in immune cells recognition and activation CD4/CD8-MHC II/I(coreceptor)CD28-B7 (costimulation)2.Adhesion between leukocytes and vascular endothelial cells in inflammation3.Lymphocyte homing and recirculationThe sequence of events in the migration of blood leukocytes to sites of infection.Selectins mediate weak tethering and rolling of blood neutrophils on the endothelium;integrins mediate firm adhesion of neutrophils to the endothelium,and chemokines activate the neutrophils and stimulate their migration through the endothelium to the sites of infection.Role of CAMs in migration of nave and effector T cells.Nave T cells home to lymph node as a result of L-selectin binding to its ligand on HEVs(high-endothelial venules),which are present only in lymph nodes.Activated T cells home to sites of infection in peripheral tissues,and this migration is mediated by E-selectin,P-selectin and integrins.Leucocytes adhesion to blood vesselApplication of CD,CAM and Their mAb 1.Elucidation of pathogenesis CD4 and AIDS,gp120 binds to CD4 2.Application in diagnosis leukemia,AIDS(ratio of CD4+to CD8+T cells)3.Prophylactic and Therapeutic application Graft rejection:Anti-CD3,Anti-CD25 non-Hodgkins lymphoma:Anti-CD20Natural Killer cellsNK cells-What are they?3rd lineage of lymphocytes Function in innate immunity to protect against viruses,bacteria,&tumors Produce cytokines&kill abnormal cellsHuman CD3-,CD56+Mouse CD3-,NKp46+(in B6 mice NK1.1+)NK Cells-Where do they come from?Progenitor in bone marrow Thymus not required nude mice have normal NK cells Do not rearrange,or d-TcR or Ig normal NK cells in scid and Rag-/-miceNK Cells-Where do they live?5-20%peripheral blood lymphocytes 5%lymphocytes in spleen Abundant in liver Low frequency in thymus,bone marrow,uninfected lymph nodes and lymphatics 90%of lymphocytes in decidual tissueNK Cells-What do they do?Cell mediated-cytotoxicity Antibody-dependent cellular cytotoxicity(ADCC)Early-interferon production Secretion of TNF,LT,GM-CSF,IL-5,M-CSF,IL-3,IL-10,IL-13,MIP-1,MIP-1,RANTES,etc.NK cells-How do they kill?Predominantly perforin/granzymesKagi et al Nature 369:31,1994Secreted or membrane TNFDegliantoni et al J Exp Med 162:1512 1985Fas ligandArase et al J Exp Med 181:1235,1995TRAILZamai et al J Exp Med 188:2375,1998NK cells-Cytokine Activation*Interferon-/augments cytolytic activity*IL-15 required for development,induces proliferation,increases cytotoxicityIL-12&IL-18 augments INF productionIL-2 induces proliferation,increases cytotoxicity.physiological relevance?NK cells-what are they good for?Protection from viruses&tumorsWhat are NK cells are good for?Humans lacking NK cells susceptible to Varicella zoster&CMV Biron et al.N Engl J Med 320:1731 1989Mouse NK cells protect against CMV-requires perforin and IFNBukowski et al.J Immunol 131:991 1983,J Exp Med 161:40 1985;Welsh et al.J Exp Med 173:1053,1991;Scalzo et al.J Exp Med 171:1469,1990;Orange et al.J Exp Med 182:1045,1995Mouse NK cells protect against mousepox-requires IFNJacoby et al.Arch.Virol.108:49,1989;Chaudhri et al.PNAS 101:9057,2004;Fan et al.PLoS Pathogen 8:e30,2008 Mouse NK cells protect against Ebola virus-requires perforin,not IFNWarfield et al J Exp 200:169,2004Class I+tumors grow in vivoClass I-tumors are rejectedClass I-tumors in NK-depleted mice grow in vivoNK Cells Reject Tumors Lacking MHC Class IImmune surveillance for Missing SelfNK cells preferentially kill cells that have lost MHC class IProvides protection against cells escaping T cell recognition（tumor and viral-infected cells tend to reduce the expression of MHC I to avoid the attack from T cells）Predicts existence of inhibitory receptors for MHC class I that spare normal cells from NK cell attack Karre et al.Nature 319:675,1986Normal healthy cellsRBC,neural cellsVirus-infected cells,tumorStressed cellsVirus-infected cells,tumorHuman Chr 19q13Killer cell Ig-like Receptors(KIR)Activating&inhibitoryNK receptorsKiller Cell Ig-like Receptors (KIR)Ig superfamily7-12 functional genes on human chromosome 19q13Extensive allelic polymorphism(no rearrangement)Mono-allelic expression possibleInhibitory KIR recognize polymorphic HLA-A,-B,and CActivating receptors have no intrinsic signaling capacity.associate with DAP12 ITAM-adapter proteinExpressed by subsets of NK cells and memory T cells(usually CD8+T cells)Human NK Receptors for HLA class IIg-domainITIMK/RC-lectin domainKIR3DL3KIR2DL3KIR2DL2KIR2DL1?HLA-CS77N80HLA-CS77N80HLA-CN77K80KIR2DL4HLA-G?KIR3DL1HLA-Bw4KIR3DS1?KIR2DL5A/B?KIR2DS3KIR2DS5KIR2DS1KIR2DS2KIR2DS4KIR3DL2HLA-CweaklyHLA-Cweakly?HLA-A?LigandsHuman ReceptorsHLA-EHLA-ECD94/NKG2ACD94/NKG2CLILRB1HLA-A,B,C,E,F,G2DL22DS22DL5B2DS32DL13DP12DL5A2DS42DS52DS13DL23DL12DL42DP12DL33DL3A haplotype2DL12DS43DL23DL12DL42DL33DL3B haplotype2DL22DS22DL12DL5A2DS13DL23DS12DL43DL3Human KIR Haplotypes Differ in Gene Content and Organization 3DS12DS5Generated by gene conversion-exon swappingDifficult to distinguish loci versus allelesUhrberg et al.1997;Wilson et al.2000;Vilches et al.2000;Hsu et al.2002Group A haplotype2DL12DS43DL23DL12DL42DL33DL3#alleles:49113826157553 x 106 possible combinations in the group A haplotypes(Courtesy P.Parham)Mice dont have KIR genesMice have Ly49 genes that do the same jobThe NK Complex on Mouse Chromosome 6 and Human Chromosome 12p12-p13NKG2D,C,ACD94Ly49A-INKR-P1A,B,CNKG2D,F,E,C,ANKR-P1ACD94HumanMouse Ly49LHumans dont have functional Ly49 genesLy49 ReceptorsC-type lectin-like superfamilyPolygenic&polymorphicExtensive allelic polymorphism(no rearrangement)Inhibitory Ly49 recognize polymorphic H-2D and H-2KActivating receptors have no intrinsic signaling capacity.associate with DAP12 ITAM-adapter protein Expressed by subsets of NK cells and memory T cells(usually CD8+T cells)Inhibitory and activating Ly49 receptors+Ly49D,HDAP12-ITIMRecruit phosphatases-inhibit responsesLy49A,C,G,I.Recruit Syk&Zap70kinases-activateLigand=MHC class IITAMNKG2DC-type lectin-like superfamily1 gene,non-polymorphic,conserved mice-humansHomodimer expressed on all NK cells,d T cells,and CD8+T cellsR in transmembrane associates with D in DAP10 transmembraneDAP1010 kd homodimerCytoplasmic YINM recruits Grb2&p85 PI3-kinaseNKG2D ligands in mice and humansMany genes Many allelesNKG2D ligands MHC class I-like dont require peptide or 2-microglobulin Bind with nM affinity to NKG2D Low levels expressed on healthy tissues Induced on virus-infected cells and tumor cells Induced by DNA damage Elevated in autoimmune diseasesWhat is the biological role of the NKG2D ligands?“Danger signals”to alert the immune system to infection or genomic damageNK cells-Influence on Adaptive ImmunityEarly INF production by NK cells may skew CD4+T cells towards Th1INF production by NK cells may cause IgG class switching in B cellsCross-talk with dendritic cells
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