Nature Immunology

© 2009 Nature America, Inc. All rights reserved.
ARTICLES
Figure 6 CD103 + DCs present
skin-derived self antigen to
CD8 + T cells. Proliferation of
5 10 4 CFSE-labeled OVAspecific
CD8 + T cells (OT-I)
after 60 h of culture together
with serial dilutions of DC
subsets isolated (as described
in Fig. 4a) from the skindraining
lymph nodes of K5mOVA
mice. Data are pooled
from two individual experiments
(mean ± s.e.m.).
Divided OT-I/well (×10 3 )
LC
dDC
CD8α + CD103
DC
+ DC
after infection (Supplementary Fig. 7 online). In this case,
FITC + CD103 + DCs, which had migrated from the skin after viral
recrudescence, presented viral antigens to CD8 + T cells. Notably,
FITC – CD103 + DCs also efficiently presented viral antigen. Presentation
by this latter population most likely reflected the fact that only a
small proportion of the CD103 + DCs are amenable to labeling
with FITC. That idea was reinforced by the finding that although
the number of CD103 + DCs in the axillary lymph nodes
increased fourfold between day 2 and day 5 (Supplementary Fig. 8
online), only about 6% of the cells were labeled with FITC (Supplementary
Fig. 5e).
Access to viral antigen is not limited to CD103 + DCs
‘Preferential’ MHC class I–restricted presentation of viral antigen by
CD103 + DCs could relate to access to viral antigens rather than a
dominant ability to cross-present. To investigate that possibility, we
explored whether other migratory DCs had access to viral antigen for
MHC class II–restricted presentation. To achieve this, we generated a
new line of T cell antigen receptor–transgenic mice that produce CD4 +
T cells specific for glycoprotein D (gD) of HSV-1 (gDT-II mice;
P.G.W., S.B., G. Davey, W.R.H., F.R.C., A.G.B., unpublished observations).
We used CD4 + gDT-II T cells as responders to examine
presentation in the brachial and axillary lymph nodes on days 2 and
5(Fig. 5). This showed that all DCs could present antigens to virusspecific
CD4 + T cells on day 5 in the axillary lymph nodes (Fig. 5d)
and brachial lymph nodes (Fig. 5c) and all but the Langerhans cells
presented on day 2 in the brachial lymph nodes (Fig. 5a). As expected,
there was no presentation on day 2 in the axillary lymph nodes
(Fig. 5b). These studies support the view that the dominant ability of
CD103 + DCs to present MHC class I–restricted antigens on day 5 is
related to their ability to process viral antigen into this pathway, rather
than their ability to access the antigen. This was especially evident by
comparison of MHC class I– and class II–restricted presentation on
day 5 by dermal DCs and CD103 + DCs. Analysis of cytokines
produced by gDT-II cells on day 5 in response to viral antigens
presented by the various DC subsets showed a bias toward production
of T helper type 1 cytokines (interferon-g, interleukin 2 and tumor
necrosis factor) in amounts that mirrored proliferation but did not
differ among DC subtypes (Supplementary Fig. 9 online).
CD103 + DCs cross-present epidermal antigens
It is difficult to distinguish between cross-presentation and direct
infection of DCs during viral responses. As an alternative approach to
examine the cross-presentation ability of skin-derived migratory DCs,
we used the DC-sorting protocol described above for isolation of
CD103 + DCs to examine the presentation of a membrane-associated
form of ovalbumin (OVA) by DCs from mice expressing OVA in
epidermal keratinocytes under control of the promoter of the gene
10
8
6
4
2
0
DC/well (×10 3 0 1.6 3.1 6.3 12.5 25
)
encoding keratin 5 (K5-mOVA mice). In accordance with our studies
of HSV-1 infection, this showed that CD103 + DCs were the dominant
DC population presenting MHC class I–restricted epidermal skin
antigen in the steady state (Fig. 6). As this antigen was expressed by
keratinocytes but not by DCs 23 , it was apparent that in this case,
CD103 + DCs used cross-presentation.
DISCUSSION
This study has shown that langerin-positive CD103 + dermal DCs are
the dominant migratory DCs of the skin that present MHC class I–
restricted antigens. This dominance most likely relates to their ability
to cross-present, at least for the types of antigens tested here. An
alternative explanation is that presentation of viral antigens by
CD103 + DCs occurs as a consequence of ‘preferential’ infection.
Definitive resolution of this possibility is problematic by most
approaches available, as it is almost impossible to distinguish between
an infected cell and a cell that has captured infected cellular material.
Although some doubt may exist over whether CD103 + DCs use crosspresentation
for HSV-1 antigens, it is likely that this function was
responsible for the presentation of OVA in K5-mOVA mice. The
failure of classical dermal DCs to cross-present OVA in this case might
be explained by limited access to antigen, as OVA is expressed by
epidermal cells. This explanation cannot apply to Langerhans cells,
however, as they reside directly in the region of OVA expression in
K5-mOVA mice 31 . Unfortunately, we have not been able to detect
presentation of OVA on MHC class II by any DC subset in K5-mOVA
mice, which precludes a definitive statement about access by dermal
DCs. However, the ability of all migratory DCs to present viral
antigens on MHC class II during HSV infection of the skin indicates
that all had access to viral antigens, yet only the CD103 + DCs
presented these antigens efficiently on MHC class I.
It is worth considering whether migratory DCs might access viral
antigens in the draining lymph nodes rather than the skin. As mature
DCs are very poor at capturing antigen for presentation 20 , and
migratory DCs have a mature phenotype in the lymph nodes, it is
most likely that viral antigens presented by migratory DCs are
captured in the skin. More importantly, if CD103 + DCs captured
viral antigen in the lymph nodes, then they might be expected to
cross-present it efficiently on both days 2 and 5, as reported for the
CD8a + DCs, yet cross-presentation by CD103 + DCs was absent on day
2. Furthermore, the ability of CD103 + DCs to cross-present skinderived
OVA in the K5-mOVA mice indicated that this is related to
capture in the skin, as capture in the lymph nodes should lead to
similar access and cross-presentation by CD8a + DCs. Although the
possibility that virus was captured in the draining lymph nodes cannot
be completely excluded, the finding that all migratory DCs presented
MHC class II–restricted viral antigens yet only CD103 + DCs presented
these antigens in an MHC class I–restricted way supports the main
point of this report, which is that CD103 + DCs are the dominant
migratory subset that cross-presents viral antigen. Our data are mostly
compatible with published studies of HSV-2 (ref. 32) and HSV-1
(ref. 33) showing that a dermal-like population of DCs provides a
principal antigen-presenting contribution to both CD4 + and CD8 +
T cell antiviral responses. Notably, we specifically separated CD103 +
DCs from classical dermal DCs and identified the CD103 + DCsasthe
key dominant cross-presenting migratory population.
A corollary of the conclusion that CD103 + DCs are the dominant
cross-presenting population of migratory DCs from the skin is that
both Langerhans cells and classical dermal DCs are inefficient at crosspresentation.
Earlier studies have provided evidence that Langerhans
cells do not prime CD8 + T cell responses to HSV 9,16 .Thatdoesnot
492 VOLUME 10 NUMBER 5 MAY 2009 NATURE IMMUNOLOGY