The concept of immune privilege refers to the observation that tissue grafts placed in certain anatomical sites, including the brain and eye, can survive for extended periods of time 1. Immune privilege is thought to reflect an evolutionary adaptation to protect vital structures from damage by inflammatory responses directed against pathogens. It was originally believed that antigens in immune-privileged sites are concealed from the immune system by physical barriers and therefore ignored. However, subsequent studies have shown that antigens do leave immune-privileged sites, that these antigens can induce immune responses, and that immune effector cells can have access to immune-privileged sites. It is now clear that immune privilege is maintained by an active rather than passive process 2,3,4.

The eye is an excellent example of an immune-privileged site. This organ enjoys immune privilege to protect it from destructive inflammation that may impair vision. Immune privilege in the eye is attributed to a variety of mechanisms (for reviews, see references 2–4), including lack of lymphatic drainage, low expression of MHC molecules, increased expression of surface molecules (CD59, membrane cofactor protein, and decay-accelerating factor) that inhibit complement activation, local production of immunosuppressive cytokines such as TGF-β, presence of neuropeptides, and constitutive expression of Fas ligand (FasL). The latter mechanism controls the entry of Fas-expressing lymphoid cells into the eye 5. Additionally, antigen presentation in the eye can elicit regulatory T (Tr) cells that induce immune deviation and suppress inflammatory responses, both locally and systemically (for a review, see reference 2). In this issue, Sonoda et al. 6 demonstrate that immune deviation induced by antigens placed in the anterior chamber (AC) of the eye requires CD1d-reactive natural killer T (NKT) cells. These findings provide insights into the maintenance of immune privilege and indicate that NKT cells play a critical role in the regulation of immune responses.

AC-associated Immune Deviation

It is well established that injection of antigens into the AC of the eye results in a form of immune deviation termed AC-associated immune deviation (ACAID; for a review, see reference 2). This phenomenon was first identified by Kaplan and Streilein 7, who reported that injection of F1 lymphoid cells into the AC of rat eyes alters the recipient's systemic immune response such that rejection of subsequent allografts from the donor strain used for AC priming is impaired. Since then, ACAID has been demonstrated for several antigens, including viruses, corneal allografts, and tumor cells. The hallmarks of ACAID are suppression of delayed-type hypersensitivity (DTH) and inhibition of the synthesis of complement-fixing antibody isotypes.

Antigen introduced into the AC is captured by specialized APCs of the eye that carry the antigen via the blood to the spleen. A deviant form of immunity is then generated which is capable of suppressing Th1-mediated DTH responses and production of complement-fixing antibodies. CD8+ T cells isolated from the spleen of AC-immunized mice can induce ACAID and inhibit DTH responses when transferred into naive syngeneic mice 8. These findings strongly indicate that AC immunization results in the induction of CD8+ Tr cells that inhibit Th1 cells (see Fig. 1). Interestingly, since ACAID in this system is induced by soluble antigen, presentation of antigen to CD8+ Tr cells must follow the poorly characterized exogenous pathway of MHC class I–restricted antigen presentation 4.

There is considerable evidence that the immunosuppressive cytokine TGF-β plays a critical role in the induction and/or effector function of CD8+ Tr cells during ACAID 2. For example, AC injection of antigen induces the generation of T cells in the spleen that produce TGF-β when stimulated with antigen in vitro. Studies by Griffith et al. 9 further indicated that immune tolerance after antigen injection in the eye depends on the death of lymphoid cells by the Fas/FasL system. Although the exact mechanism by which apoptosis of lymphoid cells affects ACAID remains unclear, it was suggested that apoptotic lymphoid cells produce IL-10 which, in turn, induces immune deviation by its effects on APCs 10. The paper from Sonoda et al. 6 provides another piece to this puzzle, demonstrating that NKT cells are required for the induction of CD8+ Tr cells.

NKT Cells

NKT cells represent a relatively novel lymphocyte subset distinct from conventional T cells, B cells, and NK cells. NKT cells share receptor structures with both NK cells and conventional T cells (for reviews, see references 11, 12). NKT cells express typical NK cell markers, including IL-2Rβ, members of the NKR-P1 NK cell–activating receptor family, and members of the Ly49 NK cell–inhibitory receptor family. They also express a TCR with an invariant Vα14-Jα281 chain and a polyclonal Vβ8.2 (and to a lower extent, Vβ7 or Vβ2) chain. Approximately 60% of all NKT cells express CD4, and the remaining cells do not express either CD4 or CD8. These cells have a memory phenotype, expressing high levels of the early activation marker CD69, high levels of CD44 (Pgp-1), and low levels of CD62L (L-selectin). NKT cells are abundant in the thymus and represent a major population of lymphocytes in the spleen, bone marrow, and liver, but are rare in lymph nodes.

Unlike conventional T cells that recognize peptide antigens in the context of self-MHC class I or class II molecules, NKT cells are specific for glycolipid antigens bound with the MHC class I–like molecule, CD1d (for reviews, see references 11, 12). As CD1d expression is required for the development of NKT cells, CD1d-deficient mice have a marked reduction in the number of NKT cells 13,14,15. Autoreactivity appears to be common among NKT cells. Many NKT cell hybridomas were able to react with splenocytes, thymocytes, or CD1d-transfected cells 11. Interestingly, individual hybridomas differed in their reactivity with CD1d-expressing cells, suggesting substantial heterogeneity among NKT cells 16,17. Recent studies have shown that CD1d can bind with a variety of glycolipids, including phosphatidylinositol derivatives 18,19 and glycosylceramides 20. However, the physiological significance of these glycolipids for NKT cell function is unclear.

When activated through their TCR, NKT cells become cytotoxic and quickly produce a variety of cytokines, including large amounts of IL-4, significant amounts of IFN-γ, and some inhibitory cytokines such as TGF-β and IL-10 11,21. Although their physiological functions remain unclear, NKT cells have been implicated in immune responses against infectious agents, bone marrow grafts, tumors, and self-antigens 11,12.

Role of NKT Cells in the Development of ACAID

The paper by Sonoda et al. 6 provides convincing evidence that NKT cells are required for development of ACAID. AC administration of antigen to NKT cell–deficient CD1d knockout mice did not induce ACAID, unless these animals were reconstituted with NKT cells from wild-type mice together with CD1d-expressing APCs. Since anti-CD1d antibodies inhibited development of ACAID in wild-type animals, direct interaction of the invariant Vα14 TCR with CD1d appears to be required. The question then arises as to what NKT cells recognize in the spleen after AC administration of protein antigen. One possibility is that eye APCs express, as a result of exposure to immunosuppressive cytokines such as TGF-β, high levels of CD1d that activate autoreactive NKT cells. Alternatively, introduction of antigen into the AC may cause expression of specific endogenous glycolipids that activate NKT cells.

At what step do NKT cells contribute to the induction of ACAID? Prior studies have shown that CD8+ T cells from the spleen of AC-primed mice can adoptively transfer ACAID to naive animals 8. The studies of Sonoda et al. 6 further extend these findings by demonstrating that NKT cell–depleted spleen cell populations from AC-primed animals can inhibit DTH reactions after adoptive transfer to naive animals. Therefore, these findings indicate that NKT cells are specifically required for the generation of antigen-specific CD8+ Tr cells.

A likely scenario for the induction of ACAID, based on the findings of Sonoda et al. 6, is presented in Fig. 1. Antigen inoculated into the AC of the eye is captured by specialized APCs that carry the antigen to the spleen. These eye APCs may have unique features that are critically important for the induction of ACAID, including increased expression of CD1d (perhaps induced by TGF-β in the AC) and altered antigen-presenting properties (perhaps in response to IL-10 produced by apoptotic lymphoid cells). These eye APCs then activate effector T cells, CD8+ Tr cells, and NKT cells. Activated NKT cells produce cytokines (TGF-β is a likely candidate) that stimulate the generation and expansion of antigen-specific CD8+ Tr cells. In turn, these CD8+ Tr cells can inhibit, by production of cytokines (TGF-β and IL-10 are likely candidates), subsequent Th1-mediated DTH responses to the same antigen.

Implications

There is now compelling evidence that Tr cells participate in many immune responses, including responses against foreign and self-antigens 22. The findings of Sonoda et al. 6 raise the possibility that activation of NKT cells is a general mechanism for the generation of Tr cells. Prior studies have suggested a role for NKT cells in the regulation of immune responses. Various mouse strains with genetic susceptibility for the development of autoimmune disease, including nonobese diabetic (NOD) mice with a propensity for development of type I diabetes, SJL mice with a susceptibility for development of experimental allergic encephalomyelitis, and several lupus-prone strains, were found to have defects in NKT cell development and/or function 23,24,25,26. Reconstitution of NOD mice with NKT cells 27 or transgenic overexpression of NKT cells in these animals 28 was able to inhibit development of diabetes. Further, NKT cells isolated from Vα14-Jα281 transgenic mice either induced or prevented murine lupus in an adoptive transfer model 29, depending on the source and cytokine profiles of the NKT cells that were transferred. Defects in NKT cell development and function were also observed in human patients with systemic sclerosis 30 and in patients with type I diabetes 31. In addition to their role in the regulation of autoimmunity, NKT cells were shown to influence immune responses against tumors 21 and infectious agents 19,32,33,34, and to suppress graft versus host disease 35. In many of these studies it was suggested that NKT cells influence the disease process by production of cytokines. For example, the inhibitory role of NKT cells for the development of Th1-dominated inflammation in several autoimmune diseases was attributed to their production of IL-4, which promotes Th2 immunity. An alternative explanation, not excluded by these studies and supported by the findings of Sonoda et al. 6, is that NKT cells induce the generation of Tr cells that, in turn, inhibit inflammation.

It is unlikely that NKT cells participate in all types of immune regulation controlled by Tr cells. Indeed, Sonoda et al. 6 showed that NKT cells are dispensable for the systemic tolerance induced in response to intravenous injection of antigen. Perhaps other cell types, such as γ/δ T cells and NK cells, can contribute to the regulation of immune responses and induction of self-tolerance. A role for γ/δ T cells as regulators of immune responses has been suggested (for a review, see reference 22).

The realization that NKT cells play a critical role in immune regulation and maintenance of self-tolerance raises the possibility of manipulating these cells to modulate immune responses during prophylaxis and therapy. This should be possible with reagents such as the glycolipid α-galactosylceramide that selectively activates NKT cells 20. Recent studies have shown that administration of this agent to mice can polarize adaptive immune responses towards Th2-dominated immunity 36,37.

Acknowledgments

We thank Andres G. Grandea for critical reading of the manuscript.

L. Van Kaer is an assistant investigator of the Howard Hughes Medical Institute.

References

References
Barker
C.F.
,
Billingham
R.E.
Immunologically privileged sites
Adv. Immunol
25
1977
1
54
[PubMed]
Streilein
J.W.
,
Ksander
B.R.
,
Taylor
A.W.
Immune deviation in relation to ocular immune privilege
J. Immunol
158
1997
3557
3560
[PubMed]
Streilein
J.W.
,
Takeuchi
M.
,
Taylor
A.W.
Immune privilege, T-cell tolerance, and tissue-restricted autoimmunity
Hum. Immunol
52
1997
138
143
[PubMed]
Ferguson
T.A.
,
Griffith
T.S.
A vision of cell deathinsights into immune privilege
Immunol. Rev.
156
1997
167
184
[PubMed]
Griffith
T.S.
,
Brunner
T.
,
Fletcher
S.M.
,
Green
D.R.
,
Ferguson
T.A.
Fas ligand-induced apoptosis as a mechanism of immune privilege
Science
270
1995
1189
1192
[PubMed]
Sonoda
K.
,
Exley
M.
,
Balk
S.
,
Stein-Streilein
J.
CD1-reactive natural killer T cells are required for development of systemic tolerance through an immune-privileged site
J. Exp. Med.
190
1999
1215
1225
[PubMed]
Kaplan
H.J.
,
Streilein
J.W.
Immune response to immunization via the anterior chamber of the eye. I. F1-lymphocyte-induced immune deviation
J. Immunol.
118
1977
809
814
[PubMed]
Wilbanks
G.A.
,
Streilein
J.W.
Characterization of suppressor cells in anterior chamber-associated immune deviation (ACAID) induced by soluble antigen. Evidence of two functionally and phenotypically distinct T-suppressor cell populations
Immunology
71
1990
383
389
[PubMed]
Griffith
T.S.
,
Yu
X.
,
Herndon
J.M.
,
Green
D.R.
,
Ferguson
T.A.
CD95-induced apoptosis of lymphocytes in an immune privileged site induces immunological tolerance
Immunity
5
1996
7
16
[PubMed]
Gao
Y.
,
Herndon
J.M.
,
Zhang
H.
,
Griffith
T.S.
,
Ferguson
T.A.
Antiinflammatory effects of CD95 ligand (FasL)-induced apoptosis
J. Exp. Med.
188
1998
887
896
[PubMed]
Bendelac
A.
,
Rivera
M.N.
,
Park
S.H.
,
Roark
J.H.
Mouse CD1-specific NK1 T cellsdevelopment, specificity, and function
Annu. Rev. Immunol.
15
1997
535
562
[PubMed]
Hong
S.
,
Scherer
D.C.
,
Singh
N.
,
Mendiratta
S.K.
,
Serizawa
I.
,
Koezuka
Y.
,
Van Kaer
L.
Lipid antigen presentation in the immune systemlessons learned from CD1d knockout mice
Immunol. Rev
169
1999
31
44
[PubMed]
Smiley
S.T.
,
Kaplan
M.H.
,
Grusby
M.J.
Immunoglobulin E production in the absence of interleukin-4-secreting CD1-dependent cells
Science.
275
1997
977
979
[PubMed]
Chen
Y.H.
,
Chiu
N.M.
,
Mandal
M.
,
Wang
N.
,
Wang
C.R.
Impaired NK1+ T cell development and early IL-4 production in CD1-deficient mice
Immunity
6
1997
459
467
[PubMed]
Mendiratta
S.K.
,
Martin
W.D.
,
Hong
S.
,
Boesteanu
A.
,
Joyce
S.
,
Van Kaer
L.
CD1d1 mutant mice are deficient in natural T cells that promptly produce IL-4
Immunity.
6
1997
469
477
[PubMed]
Brossay
L.
,
Tangri
S.
,
Bix
M.
,
Cardell
S.
,
Locksley
R.
,
Kronenberg
M.
Mouse CD1-autoreactive T cells have diverse patterns of reactivity to CD1+ targets
J. Immunol.
160
1998
3681
3688
[PubMed]
Park
S.H.
,
Roark
J.H.
,
Bendelac
A.
Tissue-specific recognition of mouse CD1 molecules
J. Immunol.
160
1998
3128
3134
[PubMed]
Joyce
S.
,
Woods
A.S.
,
Yewdell
J.W.
,
Bennink
J.R.
,
De Silva
A.D.
,
Boesteanu
A.
,
Balk
S.P.
,
Cotter
R.J.
,
Brutkiewicz
R.R.
Natural ligand of mouse CD1d1cellular glycosylphosphatidylinositol
Science.
279
1998
1541
1544
[PubMed]
Schofield
L.
,
McConville
M.J.
,
Hansen
D.
,
Campbell
A.S.
,
Fraser-Reid
B.
,
Grusby
M.J.
,
Tachado
S.D.
CD1d-restricted immunoglobulin G formation to GPI-anchored antigens mediated by NKT cells
Science.
283
1999
225
229
[PubMed]
Kawano
T.
,
Cui
J.
,
Koezuka
Y.
,
Toura
I.
,
Kaneko
Y.
,
Motoki
K.
,
Ueno
H.
,
Nakagawa
T.
,
Sato
H.
,
Kondo
E.
CD1d-restricted and TCR-mediated activation of Vα14 NKT cells by glycosylceramides
Science
278
1997
1626
1629
[PubMed]
Tamada
K.
,
Harada
M.
,
Abe
K.
,
Li
T.
,
Tada
H.
,
Onoe
Y.
,
Nomoto
K.
Immunosuppressive activity of cloned natural killer (NK1.1+) T cells established from murine tumor-infiltrating lymphocytes
J. Immunol.
158
1997
4846
4854
[PubMed]
Mason
D.
,
Powrie
F.
Control of immune pathology by regulatory T cells
Curr. Opin. Immunol.
10
1998
649
655
[PubMed]
Gombert
J.M.
,
Herbelin
A.
,
Tancrede-Bohin
E.
,
Dy
M.
,
Carnaud
C.
,
Bach
J.F.
Early quantitative and functional deficiency of NK1+-like thymocytes in the NOD mouse
Eur. J. Immunol
26
1996
2989
2998
[PubMed]
Yoshimoto
T.
,
Bendelac
A.
,
Hu-Li
J.
,
Paul
W.E.
Defective IgE production by SJL mice is linked to the absence of CD4+ NK1.1+ T cells that promptly produce interleukin 4
Proc. Natl. Acad. Sci. USA.
92
1995
11931
11934
[PubMed]
Takeda
K.
,
Dennert
G.
The development of autoimmunity in C57BL/6 lpr mice correlates with the disappearance of natural killer type 1–positive cellsevidence for their suppressive action on bone marrow stem cell proliferation, B cell immunoglobulin secretion, and autoimmune symptoms
J. Exp. Med.
177
1993
155
164
[PubMed]
Mieza
M.A.
,
Itoh
T.
,
Cui
J.Q.
,
Makino
Y.
,
Kawano
T.
,
Tsuchida
K.
,
Koike
T.
,
Shirai
T.
,
Yagita
H.
,
Matsuzawa
A.
Selective reduction of Vα14+ NK T cells associated with disease development in autoimmune-prone mice
J. Immunol
156
1996
4035
4040
[PubMed]
Hammond
K.J.L.
,
Poulton
L.D.
,
Palmisano
L.J.
,
Silveira
P.A.
,
Godfrey
D.I.
,
Baxter
A.G.
α/β-T cell receptor (TCR)+CD4−CD8− (NKT) thymocytes prevent insulin-dependent diabetes mellitus in nonobese diabetic (NOD)/Lt mice by the influence of interleukin (IL)-4 and/or IL-10
J. Exp. Med.
187
1998
1047
1056
[PubMed]
Lehuen
A.
,
Lantz
O.
,
Beaudoin
L.
,
Laloux
V.
,
Carnaud
C.
,
Bendelac
A.
,
Bach
J.F.
,
Monteiro
R.C.
Overexpression of natural killer T cells protects Vα14-Jα281 transgenic nonobese diabetic mice against diabetes
J. Exp. Med.
188
1998
1831
1839
[PubMed]
Zeng
D.
,
Dick
M.
,
Cheng
L.
,
Amano
M.
,
Dejbakhsh-Jones
S.
,
Huie
P.
,
Sibley
R.
,
Strober
S.
Subsets of transgenic T cells that recognize CD1 induce or prevent murine lupusrole of cytokines
J. Exp. Med
187
1998
525
536
[PubMed]
Sumida
T.
,
Sakamoto
A.
,
Murata
H.
,
Makino
Y.
,
Takahashi
H.
,
Yoshida
S.
,
Nishioka
K.
,
Iwamoto
I.
,
Taniguchi
M.
Selective reduction of T cells bearing invariant Vα24JαQ antigen receptor in patients with systemic sclerosis
J. Exp. Med.
182
1995
1163
1168
[PubMed]
Wilson
S.B.
,
Kent
S.C.
,
Patton
K.T.
,
Orban
T.
,
Jackson
R.A.
,
Exley
M.
,
Porcelli
S.
,
Schatz
D.A.
,
Atkinson
M.A.
,
Balk
S.P.
Extreme Th1 bias of invariant Vα24JαQ T cells in type 1 diabetes
Nature
391
1998
177
181
[PubMed]
Denkers
E.Y.
,
Gazzinelli
R.T.
,
Martin
D.
,
Sher
A.
Emergence of NK1.1+ cells as effectors of IFN-γ dependent immunity to Toxoplasma gondii in MHC class I–deficient mice
J. Exp. Med.
178
1993
1465
1472
[PubMed]
Szalay
G.
,
Ladel
C.H.
,
Blum
C.
,
Brossay
L.
,
Kronenberg
M.
,
Kaufmann
S.H.
Cutting edgeanti-CD1 monoclonal antibody treatment reverses the production of TGF-β2 and Th1 cytokines and ameliorates listeriosis in mice
J. Immunol.
162
1999
6955
6958
[PubMed]
Naiki
Y.
,
Nishimura
H.
,
Kawano
T.
,
Tanaka
Y.
,
Itohara
S.
,
Taniguchi
M.
,
Yoshikai
Y.
Regulatory role of peritoneal NK1.1+ αβ T cells in IL-12 production during Salmonella infection
J. Immunol
163
1999
2057
2063
[PubMed]
Zeng
D.
,
Lewis
D.
,
Dejbakhsh-Jones
S.
,
Lan
F.
,
Garcia-Ojeda
M.
,
Sibley
R.
,
Strober
S.
Bone marrow NK1.1− and NK1.1+ T cells reciprocally regulate acute graft versus host disease
J. Exp. Med.
189
1999
1073
1081
[PubMed]
Burdin
N.
,
Brossay
L.
,
Kronenberg
M.
Immunization with α-galactosylceramide polarizes CD1-reactive NK T cells towards Th2 cytokine synthesis
Eur. J. Immunol.
29
1999
2014
2025
[PubMed]
Singh
N.
,
Hong
S.
,
Scherer
D.C.
,
Serizawa
I.
,
Burdin
N.
,
Kronenberg
M.
,
Koezuka
Y.
,
Van Kaer
L.
Activation of NK T cells by CD1d and α-galactosylceramide directs conventional T cells to the acquisition of a Th2 phenotype
J. Immunol.
163
1999
2373
2377
[PubMed]