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Mucosal Dendritic Cells and Immunodeficiency Viruses
Melissa Pope
Laboratory of Cellular Physiology and Immunology, The Rockefeller
University, New York, New York
Dendritic cells [DCs] have been implicated in the pathogenesis of human immunodeficiency
virus type 1 (HIV-1). When skin was used as a model for mucosae, the cutaneous DC–T cell
milieu allowed the growth of HIV-1 and much of the newly produced virus could be detected
in multinucleated DC-T cell syncytia. Such virus replication occurs irrespective of the genetic
subtype, the syncytium- and non–syncytium-inducing capacities of the viruses, and whether
they are classified as T cell– or macrophage-tropic. Similar DC-syncytia have been identified
within the mucosal surfaces of the tonsillar tissue of HIV-1–infected persons. More recently,
it was demonstrated that DC–T cell mixtures from the skin, mucosae, and blood of healthy
macaques similarly support the replication of simian immunodeficiency virus. In both the
human and monkey systems, active virus replication requires the presence of both DCs and
T cells. Further studies using the macaque model are underway to elucidate the role of DCs
in the transmission and spread of HIV infection.
Replication of Human Immunodeficiency Virus Type 1
(HIV-1) Promoted by Cutaneous Dendritic Cells (DCs)
DCs, potent antigen-presenting white blood cells, efficiently
promote the replication of HIV-1 in vitro in collaboration with
CD41 T cells [1–6]. DCs are found throughout the body, including in the blood, lymphoid tissues, and body surfaces (skin
and mucosae) [7]. DCs interact efficiently with both T and B
cells [8] and, therefore, play a major role in controlling immunity to incoming pathogens. As reviewed recently by Banchereau and Steinman [8], although DCs may be involved in the
induction of immune responses to immunodeficiency virus infection, they are in fact exploited by viruses to provide a niche
in which significant virus amplification can occur (see below).
Therefore, understanding the balance between DCs “carrying”
virus and inducing immune responses versus exacerbating infection is critical for the advancement of therapeutic and vaccine strategies.
It has been proposed that DCs at the surface epithelia might
be one of the first cells targeted by the virus and be critical in
All human skin specimens used were authorized for use in research. Animal care operations were in compliance with the regulations detailed under
the Animal Welfare Act and in the Guide for the Care and Use of Laboratory
Animals.
Financial support: NIH (AI-36082, AI-40045, AI-40874, AI-40877;
“Mechanisms of AIDS Pathogenesis,” AI-38573; “Antigenic Variation of
HIV-1 and Related Lentiviruses,” AI-35168); Dorothy Schiff Foundation;
Mellam Family Fund; and Cooperative Agreement No. DAMD17-93-V3004, between the US Army Medical Research and Material Command and
the Henry M. Jackson Foundation for the Advancement of Military
Medicine.
Reprints or correspondence: Dr. Melissa Pope, Laboratory of Cellular
Physiology and Immunology, The Rockefeller University, 1230 York Ave.,
New York, NY 10021 ([email protected]).
The Journal of Infectious Diseases 1999; 179(Suppl 3):S427–30
q 1999 by the Infectious Diseases Society of America. All rights reserved.
0022-1899/99/79S3-0010$02.00
the initiation and subsequent spread of infection to the draining
lymphoid tissue [9]. Because the DC content of the mucosal
epithelia is much like that of the skin (figure 1), we have used
the more accessible skin-derived DCs as a model for the mucosal DCs to study the potential role of these cells in the transmission and spread of HIV-1 infection.
In an organ culture system, DCs and memory T cells migrate
from normal human skin [3, 10]. The DCs have a typical mature
phenotype, expressing many costimulatory and adhesion molecules, high levels of class I and class II major histocompatibility complex proteins, moderate levels of CD83, CD25, and
p55, and a perinuclear spot of CD68. Some of the DCs express
the epidermal DC markers, CD1a and LAG-1, indicating that
there is likely a mixture of dermal- and epidermal-derived DCs
migrating from the skin. The memory T cells in these cultures
consist of both CD41 and CD81 T cells, some of which are
tightly bound to the DCs in DC–T cell conjugates. To investigate the contribution of each cell subset to HIV-1 replication,
the single DCs, single T cells, and the DC–T cell conjugates
can be separated by cell sorting [3, 10].
In our initial studies, we compared the replicative capacity
of macrophage-tropic and T cell line–adapted, non–syncytiuminducing and syncytium-inducing viruses for their capacity to
replicate in this DC–T cell environment. With all viruses tested,
we found that virus growth is dependent on the presence of
both the CD41 T cells and DCs and that most of the replication
occurs in the DC–T cell conjugate fraction [2]. Virus-producing
DC–T cell syncytia are the main source of new virus production
irrespective of whether the virus had been previously characterized as syncytium-inducing or non–syncytium-inducing [2].
Although all viruses required T cells and DCs for virus replication to ensue, the T cells did not need to be actively proliferating, and they did not require an exogenous stimulus [2, 3,
11].
Extending these studies, we have demonstrated that although
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Pope
JID 1999;179 (Suppl 3)
Figure 1. Dendritic cell (DC) localization in skin and mucosal surfaces. Skin/Mucosa: Skin and skin-like epithelia of oral and genital tracts.
DCs are located within stratified squamous epithelium of mucosae and epidermis of skin. DCs (and T cells) also can be found in underlying
lamina propria and dermal regions. DCs and T cells migrate to lymph nodes via afferent lymph. Similar DC–T cell mixtures migrate from skin
and mucosal tissues in organ culture. Tonsillar Mucosa: DCs but few, if any, T cells are present within lumenal surface stratified epithelium.
Lymphoepithelium lining crypts contain DCs and numerous B and T cells positioned within loosely keratinized epithelium. DCs are also found
just beneath epithelium in underlying T cell area of lymphoid tissue. Germinal center DCs (GC DC) and follicular DCs (FDC) are located in B
cell areas.
productive infection is not detectable when DCs are alone, small
numbers of infected DCs (∼100 copies of provirus/5 3 10 4 DCs)
could initiate a vigorous infection upon interaction with syngeneic CD41 T cells for up to 1.5 days after exposure of the
DCs to the virus [3]. Furthermore, with all viruses tested, the
capacity of the DCs to provide infectious virus was affected by
treatment with zidovudine.
Most recently, we have shown that this skin-derived DC–T
cell milieu is supportive of infections with both E and B subtype
viruses [6]. Although there is isolate-dependent variation with
respect to the levels of virus growth, we did not observe any
evidence of subtype-specific replication [6]. Similar observations
that viruses from different genetic subtypes can replicate equally
well in the cutaneous DC environment have also been reported
by Dittmar et al. [12]. Our data and that presented by Dittmar
et al. do not support the notion of subtype-restricted replication
of HIV in the skin-derived DC milieu, which was described in
an earlier study [13], suggesting that additional factors other
than the genetic subtype of the virus are likely involved in
determining transmission.
There still appears to be some contradiction regarding the
infectability of purified DCs [1, 5, 6, 12–16]. The contradiction
may be explained, at least in part, by the method of cell isolation. Furthermore, more recent studies have revealed that the
infectability of DCs may be related to the maturation state of
the DC and the level of expression of relevant chemokine receptors [4, 15–18]. Nevertheless, DCs are clearly capable of
promoting significant levels of virus replication, particularly in
the presence of CD41 memory T cells. However, one must ask
whether similar events occur in vivo, where such cellular locales
may support the active growth of the virus.
Virus-Producing Syncytia in the Tonsillar Mucosae of
HIV-Infected Persons
Numerous DCs are found at many body surfaces [7, 19, 20].
Notably, DCs are abundant within the mucosal epithelia covering the surfaces of the tonsillar tissue as well as the epithelium
that lines the deep invaginations or tonsillar crypts [19, 20]
(figure 1). In addition, there are many B and T lymphocytes
within the epithelium lining the crypts; consequently, it is referred to as a “lymphoepithelium” [19, 20] (figure 1).
Studies of the tonsils and adenoids from several HIV-1–infected persons identified many HIV-positive cells within these
DC–T cell rich areas [19, 20]. Closer examination of the tissue
sections revealed that many of these cells are multinucleated
syncytia expressing high levels of intracellular HIV gag protein
and that they are more prominent in the lymphoepithelia than
in the lumenal surfaces of the tonsils and adenoids. Of interest,
these virus-producing syncytia contain DCs (they stain for proteins known to be expressed by mature DCs). However, the
inclusion of T cells into these syncytia has not been detected.
These findings [19, 20] likely represent an in vivo correlate
of the permissive DC–T cell environment we had described in
our earlier in vitro studies [2, 3, 6] (i.e., an environment in which
DCs and T cells can continuously interact with each other and
provide a niche for virus replication). It is possible that virus
circulating throughout the body (as cell-associated or cell-free
JID 1999;179 (Suppl 3)
HIV and SIV and Dendritic Cells
virus) encounters such a permissive milieu and thereby exploits
this site for further virus replication and spread. This could
happen at similar cellular locales around the body and may be
occurring at both acute and chronic stages of infection.
Macaque DCs and the Replication of Simian
Immunodeficiency Virus (SIV)
To advance our understanding of the interactions between
DCs and immunodeficiency viruses and to determine what role
DCs have in virologic and immunologic aspects of disease, we
have used the SIV-macaque system. This is a reliable system in
which to study immunodeficiency diseases [21].
In our initial studies using the SIV-macaque system, we endeavored to isolate and characterize DCs from the body surfaces of healthy macaques. Using the organ culture method
developed for human skin [10], we can isolate DCs and T cells
from the nasopharyngeal and genital mucosae and from the
skin [22]. The DCs in these preparations exhibit the typical
morphologic and phenotypic characteristics of mature human
DCs (table 1), expressing high levels of major histocompatibility
complex proteins and many costimulatory and adhesion molecules. The percentages of DCs in the suspensions isolated from
the genital mucosae are comparable to those seen in skin-derived suspensions (30%–50%). In contrast, !2% of the leukocytes isolated from the tonsillar tissues are DCs, the majority
being T cells and B cells. In all suspensions, the T cells consist
of both CD41 and CD81 subsets that can be free or bound to
some of the DCs.
As we have described in the human system [2, 3, 6, 11], the
macaque skin–derived DC–T cell mixtures supported SIV replication [22]. Of interest, virus replication also occurred in the
DC–T cell suspensions isolated from the tonsillar and vaginal
mucosae but not in the cervical DC–T cell mixtures. Both
SIVmac251 and SIVmac239 replicated equally well in these
cultures, with much of the new viral protein production occurring in multinucleated syncytia. Using the skin-derived cells
that were separated by cell-sorting techniques into the single
DCs, single T cells, and DC–T cell conjugates, we recently
demonstrated that SIV replication is dependent on the presence
of both the DCs and T cells [23], just as we reported for the
HIV-human skin system [2, 3, 6].
To more readily facilitate studies using macaque DCs, we
adapted a method for the generation of significant numbers of
DCs from monocyte-derived blood precursors [24]. After blood
monocytes are cultured in the presence of the cytokines granulocyte-macrophage colony-stimulating factor and interleukin4, a population of immature DCs can be obtained and further
differentiated into a mature DC population by additional culture in a monocyte-conditioned medium (table 1).
Initial studies on the infectability of these cells have revealed
that, much like the cutaneous DCs, the mature blood-derived
Table 1.
Source
Skin
Mucosae
Blood
Blood
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Isolation and characterization of macaque dendritic cells.
Isolation method
a
Phenotype
Organ culture
Organ culture
GM-CSF, IL-4, MCM
Mature
HLA-DR1111
CD86111
CD2511
CD831
CD681/spot
p5511
GM-CSF, IL-4
Immature
HLA-DR111
CD8611
CD252
CD832
CD68111
p552
NOTE.
GM-CSF 5 granulocyte-macrophage colony-stimulating factor;
IL 5 interleukin; MCM 5 monocyte-conditioned medium.
a
HLA-DR, CD86, CD25, and CD83 expression on cell surface by fluorescence-activated cell sorter analysis (i.e., immature DCs contain intracellular
CD83). CD68 and p55 staining by immunoperoxidase labeling of acetone-fixed
cytospins.
DCs do not support significant levels of virus replication in the
absence of T cells [23]. In fact, syngeneic T cells from various
sites around the body can collaborate with the blood-derived
DCs to support active virus replication. Similar observations
have been made when skin-derived DCs are cultured with syngeneic T cells from skin, blood, lymph node, or spleen. Fluorescence-activated cell sorter analysis of the DC–T cell cocultures also revealed that both skin- and blood-derived DCs can
form DC–T cell conjugates with each the T cell populations
examined [23]. Therefore, the capacity of the mature DC–T cell
milieu to allow virus replication is not unique to the DCs and
T cells isolated from the body surfaces. However, since the T
cells in the skin and mucosae are memory T cells, the possibility
that it is the memory T cell subset that is contributing to the
support of virus replication in the T cell suspensions isolated
from blood, lymph node, and spleen is under investigation.
Concluding Remarks
It has become clear from human and monkey studies that
DCs promote the vigorous replication of HIV [2, 3, 6] and SIV
[22, 23], respectively. In the immune system, DCs are pivotal
in the initiation of antigen-specific T cell immune responses [7].
It is quite possible that DCs play two roles in the pathogenesis
of immunodeficiency virus infection. One role could be during
the onset and spread of infection in acute and (perhaps) chronic
phases of disease. DCs have been implicated, particularly following mucosal exposure to immunodeficiency viruses, as one
of the first cells targeted by the virus, after which they traffic
the virus to the draining nodes where it can be amplified [9].
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A second role would be to induce virus-specific immune responses in the draining lymphoid tissues, which might control
viral replication and subsequent spread. Continued studies investigating the biology of the interaction of immunodeficiency
viruses with DCs are being performed in the monkey system.
Such studies are critical to advance our understanding of the
involvement of DC and other cells in the pathogenesis of
infection.
Pope
JID 1999;179 (Suppl 3)
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