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 ariseswhere 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 inappropriate assays too). The only absolute definition is that a T cell must express either the gd or ab T cell receptor (as this is agenetically 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 cytokineexpression has become the paradigm for heterogenenity within the T cell response in vitro and in vivo. Th1 and Th2 cells arethought to derive from a non-polarised, naive Th0 precursor that makes a wide range of cytokines, that can differentiate afteractivation 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 thepresence 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 theresponse towards Th1 or Th2 is then self perpetuating as Th1 cytokines enhance further Th1 responses and down regulate Th2cytokines, 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 factorsincluding 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 antigenmay be presented directly or indirectly for activation of different effector cells - macrophages, neutrophils and cytotoxic T cells forTh1 responses and eosinophils, mast cells and B cells for the Th2 response. B cells are also dependent on cytokines to promotematuration 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 markersto identify Th1 or Th2 cells in vivo, but with overall little success. The CD4-like molecule LAG-3 was thought to be a surfacemarker for Th1 cells, while CD30 (TNF-R family) and ST2L (IL-1R family) were thought to be specific for Th2, but all thesemarkers probably more accurately reflect IFN-g or IL-4 responsive cells respectively, as does the expression of the cytokine receptorsignalling molecules Stat-4 and Stat-6. The loss of the IL-12Rb chain is thought to be a marker for Th2 commitment, while thetranscription factor GATA-3 is lost on Th1 cells. It was originally thought that chemokine receptor expression would reliablydistinguish the different subsets, and although there is some functional division (CCR1, CCR5 and CXCR3 on Th1 while Th1express CCR3 and CCR4), it is becoming clear that this association is less well defined in vivo. The best way to identify Th1 andTh2 cells is therefore immunofluorescent staining for the appropriate cytokines on fixed and permeabilised cells, after a briefactivation 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 rodentmodels 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 isstill 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 acuterejection 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 nowbeing implicated in protection and tolerance induction. The foetus is also analogous to an allograft, and Th2 or Treg responses arethought 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 mainlyreview articles for this lecture, and these will obviously contain original references to those particularaspects 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:

Defined by:

Also expresses:

Function:

CD4 T-cell

CD4 (L3/T4)

CD3, CD2, TCR

MHC-class II restricted

T-cell responses

T-cell APC

CD4

TCR MHC-II

Ag

(usually )ab

Lymphocyte Subset:

Defined by:

Also expresses:

Function:

Helper T- cell

Help for antibody (B- cells)

Help for T-cells (lymphokines)

CD3, CD2, TCR, CD4 (usually)

Functions

T-cell

TCR

Ag

HelperAPC

(Macrophage,dendritic cell,

or B-cell)

Lymphokines (IL-2, IL-4 etc)

MHC-II

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

Lymphocyte Subset:

Defined by:

Also expresses:

Function:

CD3, CD2, TCR

T-cell responses

CD8 T-cell

CD8 (Lyt-2/3)

MHC-class I restricted

T-cell APC

TCR

Ag

MHC-I

CD8

(usually )ab

T-cell

TCR

Ag

Target

cell

CD4 or CD8

MHC

Killing

Lymphocyte Subset:

Defined by:

Also expresses:

Function:

Cytotoxic T-cell

MHC restricted killing

CD3, CD2, TCR, CD4 or CD8

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

FasL -> Fas

GranzymesPerforin

Activated T-cell

eg. CD25 (IL-2 receptor) but note

Lymphocyte Subset:

Defined by:

CD2R, CD26, CD71 (Tf-rec)CD3, CD4 or CD8, TCR

also MHC-II in man, rat, NOT mouse

Proliferation in response to antigen

Also expresses:

Function:

NaiveT-cell

ActivatedT-cell

MemoryT-cell

Antigen IL-2 etc

IL-2 receptorbeta chain (p75)

IL-2 receptorbeta chain (p75)

B B BCD25

High affinityIL-2 receptor

Activation markers

may also be expressed on Treg

Lymphocyte Subset:

Defined by:

Also expresses:

Function:

Memory T-cells

"Secondary responses in vitro"

CD45R0 (man), Pgp-1 (mouse CD44)CD3, CD2, CD4 or CD8, TCR

Immunity to previous Ag encounter

(Th1, Th2 and cytotoxic T-cells)

CD29 (VLA antigens), increased integrinsCD45RB low and L-Selectin low

NaiveT-cell

ActivatedT-cell

Antigen IL-2 etc

CD29

A Bor

CD45RB+

CD45RA+

CDw49 CD45R0+

(RA ,RB )_ _CD45R0

+

(RA ,RB )_ _

CD45RBCD45RA

_

+

'Th1' 'Th2'L-selectin

CD44 CD44

CD45RBCD45RA

MemoryT-cell

Long-Term?Time?Memory

T-cell

Defined by:

Also expresses:

Functions:

Lymphocyte Subset: T- regulatory cell

CD3, CD2, TCR, CD4 (usually)

T-cell

TCR

Ag

or T reg 1

APC

(B cells,dendritic cell?)

cytokines (IL-10, TGF-beta)

MHC-II

CD4

Type 3

Down regulation of Th1 (and Th2?) Ag presentation?

Poorly defined as yet!Th3 = TGF , Tr1 = IL-10, Treg = CD25/anergy b

Control of autoreactivityOral tolerance to food and gut flora

CD45RB low, CD38, CD25, CTLA4?

Transplantation tolerance

Anti-inflammatory

Defined by:

Also expresses:

Function:

Lymphocyte Subset: Suppressor T-cell

"Suppression" of responsesin vitro or transfer in vivo

CD8 (or CD4), CD3, CD45RA?

Immuno-regulation?Existence controversial

"Suppressor"

T-cell

Antigen-

specific

T-cell

Ag-MHC

a) Anti-idiotype?

b) Anti-TCR CDR peptides + MHC

CD8

TCRTCR

MHC-I

or

Lymphocyte Subset:

Defined by:

Also expresses:

Function:

NK- cell

Killing of K562 (human) YAC-1 (mouse)

Killer inhibitory receptors (KIRs)HNK-1 (CD57), NKH-1 (CD56 / NCAM)

Killing of syngeneic tumours?

CD2, CD3+/-, CD8+/-, CD16+/-

Target

cell

KIR

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

NK- cell

K562

CD16

CD2

Cross-linking

activates

KillingFasL -> Fas

GranzymesPerforin

HLA-Einhibits

Interferong

Innate response/Th1 amplification

MHC-Ior

Lymphocyte Subset:

Defined by:

Also expresses:

Function:

K- cell

CD16 (FcR-III)

CD2, CD3+/-, CD8+/-

Antibody dependentcell mediated cytotoxicity

(ADCC)

K-cellTarget

cell

CD16

Antibody

Ag

Ag

Lymphocyteor

Macrophage

Lymphocyte Subset:

Defined by:

Also expresses:

Function:

NKT - cell

NK1.1, CD4 low (mouse)

CD3, CD2, limited TCR

Amplification of Th2T-cell responses?

T-cell APC

CD4

TCR CD1

Glycolipid Ag?

High levels of Cytokines(esp. IL-4)

NKab

V 14 (J 281)/V 7 (mouse)a baV 24 (J Q)/V 11 (man)a ba

Defined by:

Also expresses:

Function:

Lymphocyte Subset: Gamma/delta T-cell

TCR gamma/delta expression

CD3, CD2, CD7usually CD4 negative, CD8+/-

Non-classical MHC antigen presentation

T-cell APC

TCR

Ag (glycolipid)

gdCD1

Responses to Mycobacterial Glycolipids

Defined by:

Also expresses:

Functions:

Lymphocyte Subset: Helper T- cell (Th1)

Activation of Type I effectors

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

Cytokine secretion

T-cell

TCR

Ag

HelperAPC

(Macrophage,dendritic cell)

Cytokines (IFN- etc)

MHC-II

CD4

Type 1

g

(cytotoxic T cells, macrophagesNK and K cells, neutrophils)

Stat-4

IFN- receptorg

CD40CD40L

gIFN- , LT (TNF- ), IL-2bDefined by:

Also expresses:

Functions:

Lymphocyte Subset: Helper T- cell (Th2)

Activation of Type 2 effectors

CD3, CD2, TCR, CD4 (usually), CD30, ST2L

Cytokine secretionIL-4, IL-5, IL-10

(B cells [esp. IgG1, IgE, IgA]eosinophils, mast cells)

Down regulation of Th1 response

T-cell

TCR

Ag

HelperAPC

(B cells,dendritic cell)

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

MHC-II

CD4

Type 2Stat-6

IL-4 receptor

CD80/86CD28

"Positive Feedback" Amplificationof Cytokine Driven T-cell Responses

Defined by:

Also expresses:

Function:

Lymphocyte Subset: Type 2 Cytotoxic T-cell (Tc2)

CD8 plus Type 2 cytokines

CD3, CD2, TCR

T-cell

TCR

Ag

Target

cell

CD8

MHC-I

Killing

FasL -> Fas

GranzymesPerforin

Killing of infected cellsin ongoing Th2 response?

IL-4, IL-5, IL-10

Tc2

Help for B cells?

Th0

Th1

Th2

Th0Activated

IL-2

IL-12

IL-4

IL-4

IFN-g

TGF-b

Other sourceseg. NKT cells

Activated macrophagesand dendritic cells

NK cells

IFN-g IL-18Microbial factors

eg. LPS

IL-1

2Rab

IL-4

R

GATA-3

GATA-3

IL-12RbLoss of

GATA-3Loss of

(irreversible?)

(irreversible?)

Sta

t-4

Sta

t-6

LAG-3

CD30

CCR1CCR5CXCR3

CCR3CCR4

CCR7

CXCR4DC-CK1R

To Node

Secondary LymphoidChemokine (SLC)

Stromal cellDerived Factor

(SDF-1) Dendritic cellchemokine 1 Eotaxin

Macrophage derivedChemokine (MDC)

RANTES

Macrophage InflammatoryProtein-1 (MIP-1 )a,b

IP-10

ST2L

CD62L

Regulation of Th1 versus Th2 responses

Th0

Th1

Th2

Th0Activated

IL-2

IL-12

IL-4

IL-4

IFN-g

Th1 versus Th2 responses in Disease

Eosinophil

Macrophage

Bacterial infectioneg. Leishmania

Viral infectioneg. Influenza

HIV

Allergenseg. Derp-1 (dust mite)

Aspf-1 (aspergiillus)

Macrophage

Mucosal epitheliumHelminths

eg. Schistomsomes

NippostrongylusMast cells

B cell

CTL

Neutrophil

iNOS

IgE

IgA

IFN-g

IFN-giNOS

IgG2

IgG2

DelayedHypersensitivity

ImmediateHypersensitivity

Protection from:Schistosomiasis, Brugia etc

Protection from:Leishmania (C57Bl/6 mice)

HIV (early), InfluenzaTuberculosis, Candida

Listeria

Pathology generated:Psoriasis, IDDM, RA

Graft rejection, foetal resorption?Hepatosplenic SchistosomiasisTuberculoid Leprosy Lesions

Autoimmunity: EAE, EAU ?Malaria (late)

Pathology generated:

Chronic graft rejection

Leishmania (BALB/c mice)HIV (late)

Allergy: Asthma, Airway DiseaseLepromatous Leprosy Lesions

l

l

m

m

m

m

1 R2.2 Th1 clone l MST = 11 days2 R2.4 Th2 clone m MST = 13 days

% m

ale

skin

gra

ft s

urvi

val

Rejection of male (but not female) skin by A1(M)xRAG-1Th1 and Th2 lines

(by transfer into "empty" ATX-depleted CBA/Ca female mice)

-/-

All female grafts

Th2Th1

3 D1 Treg clone MST > 25 days

Treg

IL2-FITC IL4-FITC

IFN -FITCg

IFN-g

CD44-QR

CD8-QR anti-mIg-FITC

CD

4-P

E

IL-4IL-2

Monospecific female (A1xRAG-/-) anti-HY+H-2EkTCR Tg mice generate both Th1 and Th2 responses

as a consequence of rejecting male skin grafts