T cell subsets

T cell subsets A synopsis of the lecture by Dr. Steve Cobbold for the FHS Physiology Immunology Option

1/ Definitions and relationships of different T cell subsets There are many different, and sometimes confusingly named, T cell and related cell subsets, although much of the confusions arises where subsets have been separately defined on the basis of molecular markers (eg. CD4 T cell) and functional properties (eg. cytotoxic T cell) and the two definitions "overlap" (eg. most cytotoxic T cells are CD8, but CD4 T cells can be cytotoxic in appropriate assays too). The only absolute definition is that a T cell must express either the gd or ab T cell receptor (as this is a genetically irreversible decision) - all other subset definitions are somewhat dependent on the assay used to measure them! 2/ Subsets of T helper cells (Th1 and Th2) defined by cytokines The original observation that mouse CD4+ T cell clones could be divided into two different sets based on their pattern of cytokine expression has become the paradigm for heterogenenity within the T cell response in vitro and in vivo. Th1 and Th2 cells are thought to derive from a non-polarised, naive Th0 precursor that makes a wide range of cytokines, that can differentiate after activation in the presence of IL-12 and IL-18 (from DCs) into Th1 cells that secrete IL-2, IFN-g and lymphotoxin (LT) or in the presence of IL-4 (from B cells or lymphoid DCs?) into Th2 cells that secrete IL-4, IL-5 and IL-10. This initial polarisation of the response towards Th1 or Th2 is then self perpetuating as Th1 cytokines enhance further Th1 responses and down regulate Th2 cytokines, and vise versa. 3/ Antigen presentation for Th1 versus Th2 responses It is still not clear how the initial polarisation towards either Th1 or Th2 is controlled, but it is thought to involve a number of factors including the route of immunisation and the type of APC (eg. skin -> LC -> Th1 or i.v. -> B cell -> Th2), the antigen density/affinity (v. high or v. low antigen dose -> Th2), and the innate immune response (NK cells -> IFN-g -> Th1 or NKT cells -> IL-4 -> Th2). With respect to T cells interacting with the APC it seems that CD40 -> CD40L promotes Th1 responses, while B7 (CD80, CD86) -> CD28 is more important as costimulation for Th2 cells. Once a Th1 or Th2 response has been polarised and established the antigen may be presented directly or indirectly for activation of different effector cells - macrophages, neutrophils and cytotoxic T cells for Th1 responses and eosinophils, mast cells and B cells for the Th2 response. B cells are also dependent on cytokines to promote maturation and isotype switching, with IL-2 and IFN-g promoting IgG2, IL-4 promoting IgG1 and IgE, and IL-5 promoting IgA.

4/ Phenotypic and functional markers for Th1 and Th2 cell subsets Th1 and Th2 clones were defined by their cytokine production pattern in vitro, and there has been much effort to find good markers to identify Th1 or Th2 cells in vivo, but with overall little success. The CD4-like molecule LAG-3 was thought to be a surface marker for Th1 cells, while CD30 (TNF-R family) and ST2L (IL-1R family) were thought to be specific for Th2, but all these markers probably more accurately reflect IFN-g or IL-4 responsive cells respectively, as does the expression of the cytokine receptor signalling molecules Stat-4 and Stat-6. The loss of the IL-12Rb chain is thought to be a marker for Th2 commitment, while the transcription factor GATA-3 is lost on Th1 cells. It was originally thought that chemokine receptor expression would reliably distinguish the different subsets, and although there is some functional division (CCR1, CCR5 and CXCR3 on Th1 while Th1 express CCR3 and CCR4), it is becoming clear that this association is less well defined in vivo. The best way to identify Th1 and Th2 cells is therefore immunofluorescent staining for the appropriate cytokines on fixed and permeabilised cells, after a brief activation step in vitro in the presence of Brefeldin or Monensin that amplify staining by holding the cytokines in the golgi. 5/ Th1 and Th2 responses in disease The Th1/Th2 paradigm has been used to explain a wide variety of disease and pathological conditions, both in expreimental rodent models and in man. The classical example is that of Leishmania infection of C57Bl/6 mice that induces a protective Th1 response, compared to BALB/c mice that insetad make an ineffective Th2 response and die from a progressive infection (the genetic basis is still not clear). A similarly classical example in man is that of Tuberculoid Th1 lesions compared to lepromatous Th2 disease. Generally speaking, Th1 responses protect from invasive bacterial, protozoal and viral infections, and cause autoimmune diseases, while Th2 responses protect from extracellular parasites, helminths, and cause allergic responses in atopic individuals. 6/ Th1 and Th2 in transplantation and pregnancy The Th1/Th2 paradigm has also been applied to transplantation, with Th1 responses being implicated in most forms of acute rejection and graft versus host disease, while Th2 responses have been variably associated with either protection or chronic rejection. However, cloned Th1 or Th2 cells have a similar capacity to reject skin grafts in experimental models, and Tr1/Treg cells are now being implicated in protection and tolerance induction. The foetus is also analogous to an allograft, and Th2 or Treg responses are thought to be protective, while Th1 may lead to resorption or spontaneous abortion.

T cell subsets - selected references The primary literature on Th1, Th2 and other T cell subsets is massive, and I have therefore provided mainly review articles for this lecture, and these will obviously contain original references to those particular aspects of the field that may interest you further. Mosmann, T.R., and Sad, S. The expanding universe of T-cell subsets: Th1, Th2 and more. Immunology Today 17: 138-146 (1996) Fallon, P. Immunopathologyof schistosomiasis: a cautionary tale of mice and men. Immunology Today 21: 29-35 (2000) Murphy, K.M. T lymphocyte differentiation in the periphery. Current Opinion in Immunology 10: 226-232 (1998) Louis, J., Himmelreich, H., et al. Regulation of protective immunity against Leishmania major in mice. Current Opinion in Immunology 10: 459-464 (1998) Abbas, A.K., Murphy, K.M., Sher, A. Functional diversity of helper T lymphocytes. Nature 383: 787-793 (1996) Sallusto, F., Lanzavecchia, A., and Mackay, C.R. Chemokines and chemokine receptors in T cell priming and Th1/Th2-mediated responses. Immunology Today 19: 568-574 (1998) Zelenika, D., Mellor, A., Simpson, E., Stockinger, B., Adams, E., Waldmann, H., & Cobbold, S. Rejection of H-Y disparate skin grafts by monospecific CD4+ T helper 1 (Th1) and T helper 2 (Th2) cells: no requirement for CD8+ T cells or B cells. J. Immunol., 161: 1868-1874 (1998) Hammond, KJ., Pelikan, SB., et al. NKT cells are phenotypically and functionally diverse. Eur. J. Immunol. 29: 3768-81 (1999) Carter, LL., Swain, SL. Single cell analyses of cytokine production. Current Opinion in Immunology 9: 177-82 (1997) Groux-H ; O'Garra-A; Bigler-M; Rouleau-M; Antonenko-S; de-Vries-JE; Roncarolo-MG A CD4+ T-cell subset inhibits antigen-specific T-cell responses and prevents colitis. Nature 389: 737-42 (1997)

Lymphocyte Subset:

CD4 T-cell

Defined by:

CD4 (L3/T4)

MHC-class II restricted

Function:

CD8 T-cell

Defined by:

CD8 (Lyt-2/3) CD3, CD2, TCR (usually ab)

Also expresses:

CD3, CD2, TCR (usually ab )

Also expresses:

Lymphocyte Subset:

MHC-class I restricted

Function:

T-cell responses

T-cell responses Ag

Ag TCR

TCR MHC-II T-cell

T-cell

APC

APC CD8

CD4

Lymphocyte Subset:

Helper T- cell

Lymphocyte Subset: Defined by:

Defined by:

Functions

Also expresses:

CD3, CD2, TCR, CD4 (usually)

Lymphokines Ag

TCR MHC-II

Killing

Ag

APC

TCR

(Macrophage, dendritic cell, or B-cell)

T-cell

Killing of virus infected cells, (and rejection of foreign tissue).

FasL -> Fas Perforin Granzymes

(IL-2, IL-4 etc)

Helper

CD3, CD2, TCR , CD4 or CD8

Function:

Help for T-cells (lymphokines)

Cytotoxic T-cell MHC restricted killing

Also expresses:

Help for antibody (B- cells) Function:

MHC-I

MHC Target

T-cell

cell

CD4 (or CD8/MHC-I for Tc2?)


IL-2 receptor beta chain (p75)

Defined by:

High affinity IL-2 receptor

+

B

Activated T-cell

B IL-2 etc

Memory T-cell

"Secondary responses in vitro"

Function:

CD45RA + CD45RB

IL-2 receptor beta chain (p75)

Immunity to previous Ag encounter

_ CD45RA + CD45RB

Naive T-cell

+ CD45R0 _ _ (RA ,RB )

CDw49 CD29

or

'Th1'

L-selectin

(Th1, Th2 and cytotoxic T-cells)

CD3, CD2, CD4 or CD8, TCR CD45R0 (man), Pgp-1 (mouse CD44) CD29 (VLA antigens), increased integrins CD45RB low and L-Selectin low

Also expresses:

Proliferation in response to antigen

CD25 Antigen

Lymphocyte Subset:

CD2R, CD26, CD71 (Tf-rec) CD3, CD4 or CD8, TCR also MHC-II in man, rat, NOT mouse

B Naive T-cell

Memory T-cells

Activated T-cell

Activation markers eg. CD25 (IL-2 receptor) but note may also be expressed on Treg

Also expresses:

Function:

CD4 or CD8

A

'Th2'

Antigen

IL-2 etc

Activated T-cell

?Time?

T-cell

Long-Term Memory T-cell

CD44

T- regulatory cell

Lymphocyte Subset:

Also expresses:

CD45RA CD45RB

B

Memory

CD44

Defined by:

+ CD45R0 _ _ (RA ,RB )

Lymphocyte Subset:

Poorly defined as yet! Th3 = TGF b , Tr1 = IL-10, Treg = CD25/anergy CD3, CD2, TCR, CD4 (usually) CD45RB low, CD38, CD25, CTLA4?

Down regulation of Th1 (and Th2?) Ag presentation? Functions: Control of autoreactivity Oral tolerance to food and gut flora Transplantation tolerance Anti-inflammatory cytokines

"Suppression" of responses in vitro or transfer in vivo

Defined by: Also expresses:

CD8 (or CD4), CD3, CD45RA?

Function:

Immuno-regulation? Existence controversial a) Anti-idiotype? TCR

TCR

(IL-10, TGF-beta)

Antigen"Suppressor"

Ag TCR MHC-II Type 3 or T reg 1 T-cell

Suppressor T-cell

APC

(B cells, dendritic cell?)

or

specific

T-cell

T-cell MHC-I CD8

b) Anti-TCR CDR peptides + MHC

CD4

Ag-MHC


NK- cell

Killing of K562 (human) YAC-1 (mouse) Killer inhibitory receptors (KIRs) HNK-1 (CD57), NKH-1 (CD56 / NCAM) CD2, CD3+/-, CD8+/-, CD16+/-

Also expresses:

Innate response/Th1 amplification Killing of syngeneic tumours?

Function:


MHC-I HLA-E

or

KIR

Defined by:

CD16 (FcR-III)

Also expresses:

CD2, CD3+/-, CD8+/Antibody dependent cell mediated cytotoxicity (ADCC)

Function:

CD16

K-cell

inhibits

Target

NK- cell

cell K562

CD2

K- cell

Killing

CD16 Crosslinking activates

Lymphocyte Subset:

Ag Target

Lymphocyte or Macrophage

cell Ag

Adhesion molecules (LFA-1->ICAM-1 etc)

Antibody

Interferon g

Lymphocyte Subset:

NKT - cell

Defined by:

NK1.1, CD4 low (mouse) CD3, CD2, limited TCR Va 14 (Ja 281)/V b7 (mouse) Va 24 (Ja Q)/V b 11 (man)

Also expresses:

Amplification of Th2 T-cell responses? Glycolipid Ag?

Function:

TCR


Gamma/delta T-cell

TCR gamma/delta expression CD3, CD2, CD7 usually CD4 negative, CD8+/-

Also expresses:

Non-classical MHC antigen presentation Responses to Mycobacterial Glycolipids

Function:

Ag (glycolipid)

CD1

ab

TCR

NK T-cell

CD1

gd

APC T-cell

APC

CD4 High levels of Cytokines (esp. IL-4)

Lymphocyte Subset:

Helper T- cell (Th1)

Cytokine secretion IFN- g , LT (TNF- b ), IL-2

Defined by:

CD3, CD2, TCR, CD4 (usually), LAG-3

Also expresses:

Activation of Type I effectors (cytotoxic T cells, macrophages NK and K cells, neutrophils)

Functions:

(IFN-g etc)

Cytokines

Ag

CD3, CD2, TCR, CD4 (usually), CD30, ST2L Activation of Type 2 effectors (B cells [esp. IgG1, IgE, IgA] eosinophils, mast cells) Down regulation of Th1 response

Functions:

Cytokines

(IL-4, IL-5, IL-13 etc)

Ag

Stat-4

APC (Macrophage, dendritic cell) CD4

CD40L

Also expresses:

Helper T- cell (Th2)

Cytokine secretion IL-4, IL-5, IL-10

Defined by:

IFN-g receptor

TCR MHC-II Type 1 Helper T-cell

Lymphocyte Subset:

IL-4 receptor

Stat-6

TCR MHC-II Type 2 Helper T-cell

APC (B cells, dendritic cell) CD4

CD40

"Positive Feedback" Amplification of Cytokine Driven T-cell Responses

CD28


CD80/86

Type 2 Cytotoxic T-cell (Tc2) CD8 plus Type 2 cytokines CD3, CD2, TCR

Also expresses: Function:

Killing of infected cells in ongoing Th2 response?


Ag TCR

Killing

MHC-I Target

Tc2 T-cell

cell CD8

IL-4, IL-5, IL-10

Help for B cells?

Regulation of Th1 versus Th2 responses

IL-2

Secondary Lymphoid Chemokine (SLC)

Th1

Macrophage Inflammatory Protein-1 (MIP-1 a,b ) IP-10

GATA-3 (irreversible?)

Activated

CD62L

To Node

Th0

TGF- b

GATA-3

CXCR4

IL-4

DC-CK1R

IL-4

Other sources eg. NKT cells

GATA-3 Stat-6

Dendritic cell chemokine 1

IL-4R

Stromal cell Derived Factor (SDF-1)

RANTES

CCR1 CCR5 CXCR3

Loss of

CCR7

Th0

LAG-3 Stat-4

IL-12

IFN- g IL-12R ab

NK cells IFN- g IL-18 Activated macrophages and dendritic cells

Microbial factors eg. LPS

CD30 ST2L

Th2

Eotaxin

CCR3 CCR4 Macrophage derived Chemokine (MDC)

Loss of IL-12R b (irreversible?)

Th1 versus Th2 responses in Disease Bacterial infection

eg. Leishmania

Macrophage

IFN- g Viral infection

eg. Influenza HIV

IL-12

Th1

IL-2

IgG2 iNOS

g IFN-Macrophage

CTL

iNOS IFN-g IgG2

Neutrophil

Th0

Protection from: Leishmania (C57Bl/6 mice) Listeria HIV (early), Influenza Tuberculosis, Candida

Delayed Hypersensitivity Pathology generated: Psoriasis, IDDM, RA Tuberculoid Leprosy Lesions Hepatosplenic Schistosomiasis Graft rejection, foetal resorption?

Activated

Th0

IL-4 Allergens

eg. Derp-1 (dust mite) Aspf-1 (aspergiillus) Helminths

eg. Schistomsomes Nippostrongylus

IL-4 Mucosal epithelium

Th2

IgE B cell

Pathology generated: Leishmania (BALB/c mice) HIV (late) Allergy: Asthma, Airway Disease Lepromatous Leprosy Lesions Chronic graft rejection

Eosinophil

IgA Mast cells

Immediate Hypersensitivity Protection from: Schistosomiasis, Brugia etc Malaria (late) Autoimmunity: EAE, EAU ?

CD4-PE

Monospecific female (A1xRAG-/-) anti-HY+H-2Ek TCR Tg mice generate both Th1 and Th2 responses as a consequence of rejecting male skin grafts

CD8-QR

anti-mIg-FITC

IFN-g

CD44-QR

IFNg-FITC IL-4

IL-2

IL2-FITC

IL4-FITC

-/-

Rejection of male (but not female) skin by A1(M)xRAG-1 Th1 and Th2 lines (by transfer into "empty" ATX-depleted CBA/Ca female mice)

l

1 R2.2 Th1 clone 2 R2.4 Th2 clone 3 D1 Treg clone

MST = 11 days MST = 13 days MST > 25 days

m

All female grafts

% male skin graft survival

lm

Treg m

Th1

l

Th2

m

m