Lymphocyte distribution and intrahepatic compartmentalization during HCV infection: a main role for...

A�

rchivum Immunologiae et Therapiae Experimentalis, 2002, 5�

0,� 307–316P�

L ISSN 0004-069X

Review

Lymphocyte Distribution and IntrahepaticCompartmentalization during HCV Infection: a Main Role for MHC-Unrestricted T CellsA

�grati C. et al.: MHC-Unrestricted Cells in HCV Infection

CHIARA AGRATI* , CARLA NISII, ALESSANDRA OLIVA, GIANPIERO D’OFFIZI, CARLA MONTESANO,LEOPOLDO PAOLO PUCILLO and FABRIZIO POCCIA

N�

ational Institute for Infectious DiseaseS “L. Spallanzani”, Via Portuense 292, 00149 Rome, Italy

A�

bstract. Hepatitis C virus (HCV) infection induces an acute and chronic liver inflammation through an immun� e--mediated pathway that may lead to cirrhosis and liver failure. Indeed, HCV-related hepatitis is characterized bya� dramatic lymphocyte infiltrate into the liver which is mainly composed by HCV non-specific c ells. Several dataindicated that interferon (IFN)-γ secretion by intrahepatic lymphocytes (IHL) may drive non-specific cell homingt

�o the liver, inducing interferon inducible protein-10 (IP-10) production. An interesti

�ng hallmark of these IHL

is the recruitment of lymphocytes associated with mechanisms of innate immunity, such as natural killer (NK),n� atural killer T (NKT) and γ δ T lymphocytes. CD81 triggering on NK cell surface by the HCV envelopeg lycoprotein E2 was recently shown to inhibit NK cell function in the liver of HCV-infected p� ersons, resultingin a possible mechanism contributing to the lack of virus clearance and to the establishment of chronic infection.In contrast, intrahepatic NKT cells restricted to CD1d molecules expressed on the hepatocyte� surface mayc ontribute to a large extent to liver damage. Finally, an increased frequency of T cells expr� essing the γ δ T cellreceptor (TCR) was observed in HCV-infected liver and recent observations indicate that int

�rahepatic γδ T cell

a� ctivation could be directly induced by the HCV/E2 particle through CD81 triggering. These c ells are not HCVspecific, are able to kill target cells including primary hepatocytes and their ability to produce T helper (Th)1 cy-t

�okines is associated with a higher degree of liver disease. Together, CD1d/NKT and/or E2/CD81 interactionsmay play a major role in the establishment of HCV immunopathogenesis. In the absence of virus clearance, thec hemokine-driven recruitment of lymphocytes with an innate cytotoxic behavior in the liver of HCV-infectedp� atients may boost itself, leading to necroinflammatory and fibrotic liver disease.

Key words: hepatitis C; liver; intrahepatic lymphocytes; NK cells; NKT cells; NT cells; γ δ T lymphocytes; HLA;C

�D1; CD81; E2.

Abbreviations used: HBV – hepatitis B virus, HCV – hepatitis C virus, HIV – human immunodeficiency virus, EBV – Epstein-Barrv� irus, NK – natural killer, IP-10 – interferon inducible protein-10, NKT – natural killer T

�, NT – natural T, MHC – major histocompatibility

c� omplex, ConA – concanavalin A, α-GalCer – α� -galactosylceramide.* Correspondence to: Dr. Chiara Agrati, Laboratory of Immunopathology, “Padiglione Del Vecchio” , National Institute for Infectious

D�

iseases (IRCCS) “L. Spallanzani”, Via Portuense 292, 00149 Rome, Italy, tel.: +39 06 551 709 58, fax: +39 06 551 709 04, e-mail:c� [email protected]

T�

he hepatitis C virus (HCV) is a small, positive--stranded RNA virus, identified as the leading causativea� gent of post-transfusional and community-acquiredn� on-A, non-B viral hepatitis23. The majority of HCVinfections result in chronic hepatitis, often progressingt

�o cirrhosis and/or hepatocellular carcinoma. The high

p� ercentage of chronicity may be due to active escapemechanisms of the virus5

�0, 100 or to HCV-induced al-

t�ered immune responses. In contrast, less than 10% of

i�ndividuals infected with hepatitis B virus will developt

�he chronic disease2

�2. This difference may imply

a� diversity in the “survival strategies” of hepatitisv� iruses and/or distinctions in host immune responsesa� gainst HCV versus HBV. The E2-HCV protein seemst

�o be a central component of distinct HCV evasionm� echanisms3

�4, 86. There is no vaccine for HCV infec-

t�ion and the only available therapy, interferon (IFN)-α

a� nd ribavirin, has proven efficacious in less than 50%o� f patients5

�8.

HCV infection is characterized by a dramatic lym-p� hocyte infiltration into the liver7

�, 62. However, the ma-

j�ority of liver-infiltrating lymphocytes are not HCVspe-

c ific. At present, the role played by lymphocytesrecruited to the liver in terms of protection and/or pa-t

�hogenesis of HCV-induced hepatitis is not well under-

stood. Several lines of evidence argue in favor of im-m� une-mediated hepatic damage triggered by infiltratinglymphocytes rather than a direct HCV-mediated cyto-p� athic effect6

, 64, 101. In the acute phase intrahepatic

lymphocytes (IHL) could be critical for the resolutiono� f the disease, whereas in the chronic phase IHL couldb

!e dangerous, damaging infected or uninfected hepato-

c ytes through both the direct killing of liver cells andt

�he release of T helper (Th)1 cytokines. The productiono� f IFN-γ at the site of infection may have two oppositee� ffects: an important antiviral activity or an efficientc ytotoxic activity against hepatocytes4

"0. In this respect,

a� nimal models of experimentally induced hepatitis sup-p� ort the hypothesis that the secretion of Th1 cytokinesis a critical mechanism in inducing hepatic injury6

, 61.

For example, in a model of transgenic mice expressingIFN-γ under the control of a liver-specific promoter, thea� nimals developed chronic hepatitis9

#0. Furthermore,

IFN-γ has been shown to be the principal mediator oft

�he hepatic inflammatory process induced against he-

p� atitis B surface antigen expressed in the liver of trans-g enic mice3

�. Finally, the expression of interleukin

2 (IL-2) and IFN-γ in the liver of HCV+ persons posi-t

�ively correlated with the extent of hepatic fibrosis and

p� ortal inflammation6 4.

C�

ell recruitment from the blood stream requires thec oncerted action of several adhesion molecules: inte-

g rins, immunoglobulin-like molecules, selectins, andg lycoproteins serving as selectin ligands. Selectin-me-d

$iated adhesion represents the first step in the cascade

r� equired for leukocyte recruitment. Specifically, the L-selectin (CD62L) has been shown to be involved inleukocyte rolling, which is a transient adhesion eventd

$uring early inflammation that allows the lymphocytes

t�o migrate to the inflamed tissue. Two compartments

c ould be discriminated at the single cell level on theb

!asis of the expression of adhesion molecules: periph-

e� ral blood lymphocytes (PBL) and IHL (Fig. 1). Naivec ells expressing the CD62L are predominant in the pe-r� ipheral blood. In contrast, IHL do not express this ad-hesion molecule necessary for extravasation and migra-t

�ion into the inflamed tissue. Intrahepatic cells show an

i�ncreased expression of class II MHC molecules com-

p� ared with PBL, indicating an activated phenotype6 8.

The analysis of naive, central and effector memoryT

� cell subsets in the HCV+% liver showed an enrichment

o� f CCR7– cells, suggesting a liver compartmentaliza-t

�ion of activated/effector cells. Using these criteria for

d$iscriminating IHL from PBL contamination, an en-

richment of lymphocytes with a cytotoxic behavior waso� bserved in the liver of HCV-infected persons1, 2, 66.Specifically, an interesting hallmark of the liver of pa-t

�ients experiencing HCV-related hepatitis is the recruit-

m� ent of lymphocytes associated with mechanisms ofinnate immunity6

6. The innate immune system uses

o� nly a small, relatively inflexible, cell population com-p� osed of natural killer (NK) cells, natural killerT

� (NKT) cells and γ δ T cells. This innate activity of the

immune system provides early antimicrobial immunitya� nd is able to determine the nature of the downstreama� daptive immune response.

In the course of HCV infection, the frequency ofintrahepatic NK cells, NKT cells, and T cell receptor(TCR) γδ T cells (Vδ1 and Vδ2

&) was significantly

higher than in the peripheral blood of the same patients(Fig. 2). In contrast to classical MHC-restricted recog-nition of antigens, NK lymphocytes can “see” and killt

�arget cells deficient in the expression of one or more

MHC class I molecules. This cell subset provides ani

�mportant defense line against viruses through a rapida� nd potent cytotoxic activity and the release of antiviralc ytokines. Moreover, the IFN-γ released by NK cells isa� ble to directly inhibit HBV replication and drive theTh cell response to a Th1 profile40. NK function is ne-g atively and positively regulated through a variety ofr� eceptors, most of which are known to interact withMHC class I molecules12, 24, 55, 63, 77. A dynamic andc oordinated balance between activating and inhibitoryreceptors controls NK cell functions and influences the

308 Agrati C. et al.: MHC-Unrestricted Cells in HCV Infection

selective recognition of virus-infected, tumor or alloge-neic cells. In HCV infection, it was shown that both thev� irus and the chronic inflammation modulate NK re-c eptor expression on the liver-infiltrating lymphocytes,regulating their immune response to the infection9

#7. NK

c ells can also modulate the adaptive response by releas-i

�ng IFN-γ and a wide variety of cytokines and immu-

noregulatory mediators7�

8.I

'n humans, an impairment of NK functions has been

a� ssociated with an increased susceptibility to differentv� iral infections, such as herpes simplex virus, Epstein--Barr virus (EBV), cytomegalovirus and human immu-n� odeficiency virus (HIV)2

�, 18, 59. Similar results were

shown also in chronic HCV patients, confirming a criti-c al role of NK-mediated immunity versus viral infec-t

�ions2

�5. Accordingly, in HBV infection, GU

(IDOTTI e� t

a� l.4"

1, showed that early IFN-γ production by NK cellsa� t the site of infection is a critical event in response tot

�he virus. In particular, in acutely HBV-infected chim-

p� anzees, NK cells were able to control HBV replication

b!efore the peak of T cell infiltration. Thus, NK activity

rather than CD3-mediated immunity was an early criti-c al component in the protection from HBV infection.I

'n contrast, in chronic HCV infection, the NK cell func-

t�ion was shown to be significantly decreased compared

w) ith healthy donors25. The mechanism responsible fort

�his HCV-induced NK cell impairment is not well

u* nderstood. Recently, TSENG et al.9#

2 proposed CD81t

�riggering as a possible candidate to inhibit NK cell

f+unctions through a novel negative signaling mechan-

ism2�

6. HCV interacts with NK cells via E2-CD81 inter-a� ctions, resulting in a direct inhibition of NK cell func-t

�ion. Specifically, in vitro ligation of CD81 on NK cells

b!y anti-CD81 or by immobilized E2 (anti-E2-rHCV-E2)

b!locks NK activation, inhibits cytokine production a� fter

e� xposure to different cytokines (IL-2, IL-12, IL-15) or byC

�D16 crosslinking. Moreover, CD81 crosslinking is

a� ble to block the cytotoxic granule release induced byC

�D16 and to reduce IL-2 induced proliferation, sugges-

t�ing that HCV could interact with NK cells through

E,

2-CD81 interaction. This mechanism may influencet

�he innate immune response to HCV infection and,

t�herefore, the kinetics and magnitude of T and B cell

immunity. Accordingly, an imbalance of Th1 versusT

�h2 responses has been proposed as a possible mech-

a� nism responsible for the viral persistence seen inc hronic HCV infection.

Along with classical NK cells, the HCV-infectedliver shows an enrichment of cells expressing both NK--like and T cell-like recognition structures. These T cellsubsets have a highly restricted TCR repertoire (bothαβ and γδ )

-, are frequently double negative (DN) for

C�

D4 and CD8 and may recognize glycolipid antigensin the context of the CD1 molecule. Human CD1 pro-t

�eins are HLA class I-like molecules that can be divided

into two separate groups: group 1 comprising CD1a,C

�D1b and CD1c and group 2 containing the more

d$ivergent CD1d16, 75. Group 1 CD1 proteins are mainly

e� xpressed on many specialized antigen-presenting cells(APC), including Langerhans cells in the epidermis andd

$endritic cells in different organs3

�7, 60, 65, whereas group

2 CD1 molecules are expressed on the gastrointestinale� pithelium and hematopoietic cells9

#, 10. Immunohisto-

c hemical analysis confirmed that HLA class I is poorlye� xpressed on hepatocytes (data not shown), suggestingt

�hat CD8 T cell recognition of viral peptides in the

c ontext of HLA class I molecules may not represent themain cytolytic pathway occurring in the liver. Accord-i

�ngly, immunohistochemical analysis of CD1a, -b, -c

a� nd -d expression in the liver showed that hepatocytese� xpress high levels of CD1d molecule independently ofHCV infection, suggesting that CD1d could be the main

Fig. 2. Lymphocyte distribution in the peripheral blood and in thel.iver of HCV-infected patients. Panel A – PBL. Panel B

/ – IHL

Fig. 1. Schematic representation of liver section. Intrahepatic lympho-cytes (IHL) circulating in liver sinusoid and/or migrating in liver tissue.H – hepatocytes, I – Ito cells, K – Kuppfer cells, – CD62L

Agrati C. et al.: MHC-Unrestricted Cells in HCV Infection 309

r� estriction element in the liver (AGRATI et al., manu-script in preparation).

αβ NKT cells express a TCR composed of a singlei

�nvariant TCR α-chain (Vα14, Jα2

&81 in mice and Vα2

&4,

J0αQ

1 in humans) and a highly skewed TCR β-chain

(Vβ11 in the humans and Vβ8 in the mouse)4, 29, 33, 53.N

2KT cell activation results in a rapid production of

c ytokines (IL-4 and IFN-γ )-, cell proliferation and NK-

like cytotoxicity5�

, 42, 48. Moreover, it was reported thatstimulation of NKT cells is able to rapidly induce acti-v� ation of innate (NK cells) and adaptive (T cells andB cells) immune responses18. This αβ T cell subset ex-p� resses NK-like markers belonging to the NKRP1 fam-ily and recognizes glycolipid antigens presented byC

�D1d molecules7

�6.

I'n the liver of HCV-infected patients an increase of

V3

α24 NKT cells (up to 20-fold in comparison with thep� eripheral blood) was observed6

6. The memory/acti-

v� ated phenotype associated with the oligoclonal expan-sion suggests that these NKT cells may recognize ane� ndogenous ligand or ubiquitous antigen. The endogen-o� us ligand could normally be expressed in the liver ofhealthy persons, or processed and presented in the con-t

�ext of CD1d to Vα24/Vβ11 TCR-expressing cells as

a� consequence of tissue damage. This subset is able top� roduce high amounts of Th1 cytokines rapidly and tok

4ill other cells with an NK-like mechanism. Recent

findings have suggested a pathological role for NKTc ells based on their ability to produce large amounts ofIFN-γ . It has been reported that Vα14 NKT cells playa� critical role in various diseases, including S

5almonella

infections45, autoimmune diabetes102 and systemic scle-r� osis81. In a murine model of hepatitis induced by in-t

�ravenous injection of concanavalin A (ConA), the liverinjury is strictly associated with the presence of lym-p� hocyte infiltrates. The absence of liver injury in at-h

6ymic nude mice or SCID mice indicates the immuno-

p� athological origin of ConA-induced hepatitis87.Several observations demonstrate that only Vα14+ NTc ells are required for the development of ConA-inducedhepatitis4

"6. For example, the antibody depletion of

N2

K1.1+ cells in vivo confers resistance to hepatitis, andt

�his resistance is also present in β2

& microglobulin or

C�

D1 knock-out mice that have dysfunctional NK1.1+

N2

KT cells83, 89. Moreover, it was demonstrated thatN

2KT cells critically contribute to liver damage caused

b!y the generalized Shwartzman reaction6

7, suggesting

a� general involvement of NKT cells in immune-medi-a� ted hepatitis. Although the contribution of NKT cellst

�o HCV-induced hepatitis remains to be determined in

f+urther studies, we speculate that they are involved in

mediating the liver injury observed in HCV+% persons,

n� ot only by itself83, but also by cooperating with con-v� entional T cells49, 95 and macrophages87. Vα14 T cellsseems to explain their effector activity through severalm� echanisms, such as Fas-FasL interactions82, the per-forin-granzyme system9

#9 and IFN-γ release5

�1, 82.

In the HCV+ liver, an increased fraction of T cellse� xpressing γδ T

�CR was also observed1, 66, 97. γδ T cells

represent a minor population of human peripheral lym-p� hocytes (3–6%)27 and express the CD16 NK marker,i

�ndicating their natural T (NT) nature7

�1. The majority

o� f these cells express the Vδ2 TCR variable segmenta� ssociated with the Vγ 9

7 segment6

9. Vγ 9

7Vδ2 NT lym-

p� hocytes recognize phosphorylated nonpeptidic micro-b

!ial metabolites14, 15, 85 and alkylamines7

�1 without the

r� equirement of antigen uptake or processing, or MHCc lass I or class II expression84. Also, γ δ NT cells haveb

!een shown to release high levels of cytokines such as

IFN-γ , TNF-α5�

2, and β c hemokines involved in the re-c ruitment of cells of the monocyte/macrophage lineaged

$uring an inflammatory reaction. NK receptors ex-

p� ressed on Vγ 97Vδ2

& NT cell surfaces sharply regulate

t�heir activity3

�8, 73, 74 and influence γδ responses in anti-

v� iral reactivity, tumor immunity and autoimunity3�

6, 73.In contrast, Vδ1 NT cells represent a minor lymphocytesubpopulation in the peripheral blood, usually express-ing a naive phenotype in healthy donors. Their Vδ1-en-c oded receptor chain is typically co-expressed with Vγ --encoded chains distinct from Vγ 9

7 9#

1. However, Vδ1+%

c ells are the predominant γδ T cell population in thep� ostnatal thymus20 and represent a major T cell popu-l

8ation in the skin, intestinal and pulmonary epithelium.

U9

nlike αβ T cells, which re-circulate extensively, γ δT

� cells in these epithelial tissues seem to remain im-

mobile. The selective expression of TCR V-gene seg-ments in different epithelial tissues is also observed inmice3

�2, 72 and may reflect the possibility that these

T�

cells are specialized in responding to certain types ofa� ntigens expressed at these sites. Specifically, Vδ1T

� cells represent the major T cell subpopulation in the

human intestine2�8, 30, 70, 96, suggesting their possible role

a� s the first line of defense against the invadingmicrobes entering the gastrointestinal tract.

T�

he ligands recognized by Vδ1 NT lymphocytes arestress antigens of cellular origin. Vδ1+ NT cells recog-n� ize and interact with the MICA and the closely relatedMICB glycoproteins that are expressed mainly in theg astrointestinal and thymic cortical epithelium and onhepatocyte-derived cell lines. The MIC genes haveh

6eat-shock response elements in their promoters and

show a low homology with MHC class I molecules3�9.

T�

he stress-induced expression of MICA and MICB,a� nd their recognition by polyclonal Vδ1 NT cells


t�hrough TCR or the NKG2-D natural killer receptor,

may serve as an immune surveillance mechanism ford

$etecting damaged, infected, or transformed intestinal

e� pithelial cells. Moreover, tissue Vδ1 NT lymphocytesw) ere recently shown to recognize nonpolymorphicC

�D1c molecules80. Specifically, Vδ1 NT cells were

f+ound to proliferate and release Th1 cytokines in re-

sponse to CD1+-presenting cells and to lyse CD1c+ tar-g ets. The recognition of CD1c is TCR mediated andd

$ependent on Vδ1 TCR expression. The reactivity of

γδ NT cell lines and clones to CD1c is highly specifica� nd independent of the presence of exogenous antigen.I

'n the course of HCV infection, γδ T cells are recruited

t�o the liver1, 66, 97. In particular, phenotypic analysis of

t�his T cell subset indicated that this increase is due toC

�D3+ cells expressing the Vδ1 chain of the TCR and

results in an inversion of the intrahepatic Vδ2 to Vδ1ratio1. Interestingly, Vδ1 T cells homing to the liverw) ere specifically induced by HCV infection: indeed,t

�he analysis of HCV– persons failed to show any Vδ1

c ompartmentalization in the liver2. The rapid recruit-m� ent of Vδ1 to the liver of HCV-infected persons mayb

!e driven by the recognition of HCV antigens and/or

b!y alterations of the cytokine and chemokine environ-

m� ent. Indeed, in HIV infection, the increase of the Vδ1T cell subset observed in the blood suggests that HIVi

�nfection induces a Vδ1 mobilization from the mucosat

�issue to the periphery under the influence of various

c ytokines and/or chemokines. Accordingly, as a resulto� f viral interference, in HIV/HCV co-infected personst

�his increased fraction of peripheral Vδ1 T cells tends

t�o home to the site of HCV infection, confirming a re-c ruitment to the liver driven by HCV2.

The exact role played by γδ T cells in the protectiona� nd/or pathogenesis in HCV infection is not well under-stood. However, the involvement of γδ T lymphocytesi

�n immunosurveillance has been suggested in several

infections with herpesviruses, including herpes simplexv� irus13 and cytomegalovirus27. An increased number ofγδ T cells has also been observed in animal models ofinfluenza17 and Sendai virus infection4

"4 as well as in

p� atients with EBV and HIV5�

7, 98. Several reports havep� rovided strong evidence of the anti-inflammatory roleo� f γδ T cells through the homeostatic regulation of αβT

� cells3

�2. Thus, the rapid homing of circulating γ δ NT

c ells to the liver may determine the pattern of the adap-t

�ive responses mediated by the subsequent activation of

MHC-restricted αβ T lymphocytes. Interestingly, theV

3δ1 T cells recruited to the liver display an acti-

v� ated/memory phenotype (CD62L–C�

D45RO+C�

D95+)-

suggesting an antigen-mediated stimulation. However,t

�he lack of selective expression of any Vγ -chain indi-

c ates a polyclonal “superantigen-like” activation. In-t

�rahepatic γδ T lymphocytes from HCV+ persons can

b!e expanded in vitro with a cytokine cocktail contain-

i�ng IL-2, IL-4, IL-7 and IL-159

#3. γδ T cell lines ob-

t�ained from HCV-infected livers show a potent MHC-

-unrestricted cytotoxic activity against NK-sensitive(K562), NK cell-resistant (Daudi and Huh7) targetsa� nd, finally, against primary hepatocytes. In contrast,αβ T cell lines from the liver of the same patients failedt

�o kill any of the target cells tested9

#3. Nevertheless,

intrahepatic γδ T cells recruited to the liver under theinfluence of HCV infection were not specific for HCVa� ntigens. Finally, none of the intrahepatic γδ T celllines were able to kill autologous EBV-transformedB

: cells infected with recombinant vaccinia viruses ex-

p� ressing HCV proteins, indicating that this T cell sub-set does not recognize HCV proteins. Proliferative ex-p� eriments demonstrated that γδ T cells were unable tor� espond to structural (Core, E1, E2) and nonstructural(NS3, NS4, NS5) HCV recombinant proteins, confirm-i

�ng that intrahepatic γδ T cells are not specific for HCV

p� roteins or HCV-infected cells9#3. Thus, intrahepatic

V3

δ1 T lymphocytes can contribute to liver pathologyb

!y killing hepatocytes without HCV specificity. An

a� nalogous MHC-unrestricted and viral non-specific cy-t

�otoxic activity exerted by γδ T lymphocytes on cells

i�nfected with HIV and herpes simplex virus has been

reported13, 47, 56.Another important event in the induction of necroin-

flammatory processes in the liver is the release of in-f

+lammatory cytokines6

8. In HCV infection, e; x vivo

stimulation of intrahepatic lymphocytes showed an in-c reased frequency of IFN-γ producing Vδ1 T cellsc ompared with peripheral Vδ1 T lymphocytes from thesame donors. Cytokine analysis of γδ T cell lines ob-t

�ained from HCV+ livers confirmed the ability of this

T�

cell subset to release high levels of Th1 cytokines9#3.

Interestingly, the percentage of IFN-γ -releasing Vδ1T

� cells is higher in the liver of HCV+ persons going

t�hrough the necroinflammatory process, suggesting that

t�he overall role of Vδ1 T lymphocytes in the HCV+%

liver produces pathogenic rather than protective re-sults1. Thus, Vδ1 T lymphocytes homing to the liver ofHCV+ persons may contribute to the immunopathogen-e� sis of HCV-induced hepatitis through two distinctmechanisms: killing infected and uninfected hepato-c ytes without HCV specificity and releasing inflamma-t

�ory cytokines. The possible ligand of Vδ1 T cells are

h6eat shock proteins or stress proteins of cellular

o� rigin3�

4, 39. Possible candidates are MHC-related pro-t

�eins MICA and MICB that could function as self anti-

g ens and are recognized broadly by intestinal epithelial


V3

δ1 T cells through TCR or the NKG2-D natural killerreceptor without Vγ restriction. The recognition ofstress-induced proteins on the surface of hepatocytesc ould be an important mechanism in the immune sur-v� eillance mechanism for the detection of HCV-infectedc ells. The CD1c was shown to be another possible li-g and, but it is expressed on the hepatocyte’ surfaceneither in the uninfected nor in the HCV+ liver. Morerecently, TSENG et al.9

#3, showed that in vitro stimulation

o� f intrahepatic γ δ T cell lines by immobilized anti-CD81induced a release of significant levels of TNF-α andIFN-γ indicating that CD81 may be a crucial moleculei

�n γδ T cell activation in HCV infection.

C�

D81 is a member of the tetraspan superfamily ofp� roteins. CD81 is a 26 kDa protein composed of4

< transmembrane and 2 extracellular domains. It is ex-

p� ressed on most human tissues, and within a singlet

�issue its levels vary during development and in re-

sponse to cellular activation5�

4. A common charac-t

�eristic of tetraspanins, including CD81, is a propensityt

�o associate physically with a variety of other mem-b

!rane proteins to form signal transduction complexes.

Ligation of CD81 with monoclonal antibodies resultsin a costimulatory signal for B and T cells expressingαβ T� CR. CD81 molecule associates CD19, CD21 andLeu-1311, 35, 43 on the B cell surface to form a multi-m� olecular complex that reduces the threshold for B cella� ctivation19, 35. On the T lymphocyte surface, CD81 ise� xpressed in association with the CD4 and CD8 mole-c ules88 and acts as a costimulatory signal for prolifera-t

�ion and cytokine production7

�9, 103. γδ T cells, which

lack CD4 and CD8 coreceptors, also express CD81 ont

�heir surface but respond differently to CD81 crosslink-

ing than do αβ T cells. Indeed, crosslinking of CD81o� n γδ T lymphocytes results in direct cell activationw) ithout the need of CD3 stimulation9

#4. Moreover, re-

c ent data demonstrated a third different pathway ofC

�D81-mediated signaling, inducing inhibition of NK

f+unctions (cytokine production, cytotoxicity and pro-

liferation).Altogether, several data suggest that innate (non-

-specific) immunity plays an important role in the liverp� athology of HCV infection. Intrahepatic production ofTh1 cytokines and chemokines by HCV infection pro-m� otes the recruitment of non-specific lymhocytes.These innate cells are directly recruited to the infectedliver by the inflammatory process, without the need fora� ntigen recognition and clonal expansion in the regionall

8ymph nodes. This process requires the concerted action

o� f several adhesion molecules and results in the enrich-m� ent of lymphocytes with innate cytotoxic behavior. Int

�he absence of viral clearance, this pathway will boost

i�tself, leading to necroinflammatory and fibrotic liver

d$isease. The molecular mechanisms may involve

C�

D1d/NKT and/or E2/CD81 interactions, promotingt

�he intrahepatic recruitment of non-specific T cells re-leasing proinflammatory cytokines. Thus, new strat

�egies

for therapeutic intervention could be directed to inhibitt

�he molecular interactions responsible for the recruit-

ment and non-specific activation of IHL in HCV-infec-t

�ion.

A=

cknowledgment. This study was supported by grants from “Min-istero della Salute” .

R>

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Received June 2002Accepted July 2002


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