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,
*
Microbiology and Tumor Biology Center, Karolinska Institute, Stockholm, Sweden;
Department of Virology, Swedish Institute for Infectious Disease Control, Stockholm, Sweden; and
Department of Laboratory Medicine, Karolinska Hospital, Stockholm, Sweden
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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instead
prevails as a dominating effector function in the liver
(8). Despite several lines of evidence demonstrating a protective role for NK cells in CMV infection, the mechanism by which NK cells can recognize virus-infected cells is unclear. The control of NK lysis in general is the result of a fine tuning between positive and negative signals (9). Adhesion molecules and other triggering receptors at both the target and the effector cell level can affect lysis by NK cells, probably by contributing to the delivery of a positive signal. In this context, some reports suggest that hCMV infection can increase the expression of ICAM-1 and LFA-3 on infected cells, leading to an increased susceptibility to NK cell killing (10, 11). On the other hand, engagement of HLA class I (HLA-I) molecules on target cells by specific inhibitory receptors is a well-defined negative signal for NK cells (9, 12). hCMV is able to reduce cell surface expression of HLA-I by a variety of mechanisms mediated by viral gene products, such as US2, US3, US6, and US11 (13). This may lead to a CTL failure of hCMV-infected cell recognition. On the other hand, there is a possibility that this reduced HLA-I expression instead leads to NK cell recognition of hCMV-infected target cells (12, 14).
Despite this suggested complementary role between CTL and NK cells in the scrutiny of hCMV-infected cells, it has recently been proposed that hCMV can evade both surveillance mechanisms by a simultaneous use of the US and UL18 genes (14).
The UL18 gene product binds ß2-microglobulin
(ß2m) and shares 
25% of homology in 
1, 2, 3
domains with HLA-I molecules (15), and it may present
endogenous peptides (16). Recently, the possible role of
the MHC class I homologue in NK cell inhibition was investigated both
in mice and humans (17, 18, 19). These studies indicate that
the CMV MHC class I homologue may be involved in inhibiting NK cell
functions. However, another recent report instead suggests an
NK-activating role for this molecule and an increased NK cell-mediated
killing of fibroblasts infected with a laboratory strain of hCMV
(10). Thus, there is no general agreement on the role of
this pseudo class I Ag in the regulation of NK cells. Recently, a
possible mechanism of action of UL18 has been suggested by Cosman et
al. (20), who demonstrated that UL18 is specifically
recognized by the leukocyte Ig-like receptor-1 (LIR-1). This is an
inhibitory receptor of the Ig superfamily, capable of binding classical
and nonclassical HLA class I molecules (for a review see Refs.
21 and 22). It is predominantly expressed on
monocytes, B cells, and a minor subset of T and NK cells
(23, 24, 25). However, there is still a lack of evidence
supporting a role for UL18 in inhibiting NK cell functions.
All the above mentioned studies (10, 17) deal with target cells infected with hCMV laboratory strains, such as the widely used AD169. Laboratory strains may however differ from freshly isolated, clinical hCMV strains (26). We therefore investigated the susceptibility to NK killing of target cells infected both with highly passaged hCMV laboratory strains and with clinical isolates that have been propagated in vitro for <20 passages. Our data show that all the three hCMV clinical strains confer a strong NK resistance, whereas the same target cells infected in parallel with laboratory strains yielded only marginal or nonreproducible effects with respect to NK recognition. Moreover, all the clinical strains studied inhibit the NK cytotoxicity in an HLA-I- and LIR-1-independent way.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Human foreskin fibroblasts (HFFs, HS27) were obtained from American Type Culture Collection (ATCC, Manassas, VA), and human embryonic lung fibroblasts (HLs) were purchased from the Department of Microbiology, Sahlgrenska Hospital, Gothenburg, Sweden. They were used between passages 21 and 37 and 14 and 19, respectively. All fibroblasts were grown in Dulbecco’s modified medium supplemented with 5% FCS, 2 mM L-glutamine, 100 U/ml penicillin, 100 µg/ml streptomycin, and 0.01% Fungizone (Life Technologies, Stockholm, Sweden).
The NK leukemia cell line NKL was obtained from Dr. M. J. Robertson (Bone Marrow Transplantation Program, Indiana University, Indianapolis, IN), and it was grown in RPMI supplemented with 10% FCS and 180 IU/ml IL-2.
The T2 cell line (27) and the HLA-I-deficient EBV-transformed B lymphoblastoid cell line 721.221 were maintained in RPMI 1640 supplemented with 10% FCS, 2 mM L-glutamine, 100 U/ml penicillin, 100 µg/ml streptomycin, nonessential amino acids, and 1 mM sodium pyruvate.
To generate the 721.221-transfectant cell line DT361, a chimeric cDNA containing the leader segment of HLA-B*5801 and the extracellular, transmembrane, and cytoplasmic domains of HLA-G1 was generated by PCR using the following oligonucleotide primers: sense primer 1: 5'-GCGCTCGAGATGCGGGTCACGGCG CCCCGA-3'; antisense primer 1: 5'-CATGGAGTGGGAGCCGGCCCAGGTCTCGGT-3'; sense primer 2: 5'-GGCTCCCACTCCATGAGGTAT-3'; and antisense primer 2: 5'-GCGCAAGCTTTCAATCTGAGTTCTTC-3'. PCR was performed using a wild-type HLA-B*5801 cDNA as a template with primer set 1 and using wild-type HLA-G1 cDNA as a template with primer set 2. Products from these PCR reactions were mixed and used as templates for a subsequent reaction with sense primer 1 and antisense primer 2. The product was digested with XhoI and HindIII and ligated into the pBJneo vector. 721.221 cells were transfected with the chimeric cDNA and selected in 1 mg/ml G418, and transfected cells expressing high levels of HLA-I were sorted by flow cytometry.
PBMC were isolated by centrifugation on Lymphoprep gradients (Nycomed Pharma, Oslo, Norway) from buffy coats of normal donors obtained from the Blood Bank of the Karolinska Hospital (Stockholm, Sweden). After isolation, the PBMC were washed and incubated in serum-free medium in horizontally placed flasks for 1 h at 37°C to remove adherent cells. Nonadherent cells were then collected, washed, and resuspended in RPMI 1640 medium supplemented with 10% FCS, 2 mM L-glutamine, 100 U/ml penicillin, 100 µg/ml streptomycin, nonessential amino acids, and 1 mM sodium pyruvate. For the generation of IL-2-activated killer cells (LAK), 1 x 106/ml nonadherent cells were cultured in the presence of 1000 U/ml IL-2 for 2–4 days and then used in cytotoxicity assays. Polyclonal NK cell cultures were obtained by coculturing nonadherent cells (4 x 105 cells/ml) with irradiated (3000 rad) RPMI 8866 (105 cells/ml) for 10 days as previously described (28). On day 10, the cell population contained 80–90% CD56+CD3- cells as assessed by FACS analysis. For the generation of NK cell clones, polyclonal NK cell populations were depleted of CD3+ cells by magnetic beads (Dynal, Oslo, Norway) coated with anti-CD3 mAb. More than 95% of the remaining cells were CD56+ CD3- as assessed by FACS analysis.
Clones were generated by limiting dilution of the CD3-depleted cells in 96-well plates in the presence of irradiated (3000 rad) allogeneic PBMC and RPMI 8866, supplemented with 0.5 µg/ml PHA and 1000 U/ml IL-2. All cell lines were grown at 37°C in 7.5% CO2 in air.
Virus
The following strains of hCMV were used: the laboratory strains AD169 (continuously passaged from 1988) and Towne, obtained from V. A. Sundqvist (Karolinska Institute) and ATCC (VR-977) respectively; the clinical isolates 4636 (passages 12 or 18, from a bone-marrow transplanted child), 109B (passage 14 or 20, from a kidney transplant recipient), and 13B (passage 14 or 20, from an AIDS patient).
All strains had been propagated on HLs. Virus stocks were harvested from the supernatants of hCMV-infected HLs at 10–14 days postinfection and stored at -70°C. Virus stocks were titrated using a plaque assay and used at a multiplicity of infection (MOI) of 1 or 2. Virus stocks were confirmed to be Mycoplasma free (Mycoplasma T.C. II Rapid Detection System, Gen-Probe, DPC, Skafte, Sweden).
Infection of cells with hCMV
Cells were either treated with medium alone or infected with hCMV at a MOI of 1 or 2. After adsorption of the virus for 1 h at 37°C, the inoculum was removed, and medium containing 5% FCS was added. In all experiments, performed at day 8 (with a MOI of 1) or at day 4 (with a MOI of 2), a 95–100% cytopathogenic effect was obtained, as determined by microscope analysis.
Cytotoxicity assay
Cell-mediated cytotoxicity was assessed in 4-h 51Cr release assays. In these assays, different concentrations of effector cells were incubated with 5 x 103 target cells in V-bottom, 96-well microtiter plates at 37°C in 7.5% CO2 in air. In the blocking experiments, cells were incubated with saturating amounts of specific or isotype control Abs for 15 min at room temperature. Cells were then washed once and used in the assays. Percentage of lysis was determined by counting a 100-µl aliquot of supernatant and using the formula: % specific lysis = 100 x [(sample release - spontaneous release)/(total release - spontaneous release)].
Monoclonal Abs
PE-labeled mAb anti-CD56 was purchased from Becton Dickinson (San Jose, CA). Anti-ICAM-1 (clone LB2) was kindly provided by Dr. Manuel Patarroyo (Microbiology and Tumor Biology Center, Karolinska Institute). Cychrome-labeled anti-CD3 (clone UCHT1) was obtained from PharMingen (San Diego, CA).
Anti-LFA-3 Ab (clone BRIC-5) was purchased from Serotec (Oxford, England). FITC-conjugated rabbit anti-human ß2m Ab and purified mouse IgM were purchased from Dako (Glostrup, Denmark). Control IgG conjugated with different fluorochromes were obtained from Caltag (Burlingame, CA). Anti-HLA-I supernatant from the hybridoma A6-136 (IgM isotype) was kindly provided by Dr. Lorenzo Moretta (Centro Biotecnologie Avanzate, Genoa, Italy). Purified anti-CD56 Ab (clone C218, IgG1 isotype) was obtained from Immunotech (Marseille, France). Purified anti-LIR-1 Ab (clone M405, IgG1 isotype) was kindly provided by Dr. D. Cosman (Immunex, Seattle, WA) and has been previously described (29).
Cells were labeled and analyzed on a FACscan flow cytometer (Becton Dickinson).
Statistical analysis
Statistical analysis of the data was performed using a paired Student t test. A value of p < 0.05 was considered statistically significant.
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Results |
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To assess whether hCMV infection affects NK recognition, HFFs were
infected with the clinical strain 4636 (derived from a bone marrow
transplant recipient) and tested for sensitivity to NK cell-mediated
cytotoxicity 8 days later. The sensitivity of 4636-infected HFFs to a
polyclonal NK cell population (see Materials and Methods)
was much lower than that of uninfected HFFs (Fig. 1
A). The same result was
obtained with 12 of 12 donors tested (Fig. 2 and data not shown), thus excluding the
possibility that this recognition pattern was limited to a single
donor-target cell combination. To exclude also the possibility that
this reactivity was due to the particular effector cell type, we
generated short term LAK cells from the same donor as in Fig. 1A, and NK clones from a different donor. Also these types
of effector cells showed reduced killing of 4636-infected HFFs (Fig. 1B and data not shown). The reduction in the killing was
considerable, ranging between 48 and 92% at an E:T ratio of
12.5:1.
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). The differences in lysis that sometimes could be observed between
HFFs infected with different viral isolates were not due to different
levels of infection, because a similar infection efficiency (95–100%)
was obtained with all strains (data not shown). Inhibition of the
killing ranged between 43 and 87% for 13B and between 15 and 82% for
109B, at an E:T ratio of 10:1.
To verify whether the clinical hCMV strains can confer protection from
lysis by NK cells to fibroblasts other than HFFs, we used the HL cell
line, originated from human embryonic lung fibroblasts. Similar results
were found with polyclonal and LAK cells (Fig. 3), as well as NK clones (data not shown)
as effectors, as we could always observe a reduction in the lysis of
infected HLs. The inhibition ranged between 20 and 72% for 4636,
between 44 and 87% for 13B, and between 30 and 78% for 109B, at an
E:T ratio of 10:1.
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Comparison between NK cell-mediated killing of fibroblasts infected with clinical isolates and AD169 or Towne laboratory strains
It has recently been shown that infection of fibroblasts with the
laboratory strain AD169 increases the susceptibility to NK
cell-mediated killing (10, 11). To compare NK
cell-mediated cytotoxicity of fibroblasts infected with clinical
isolates vs fibroblasts infected with the laboratory strains Towne or
AD169 of hCMV, we performed cytotoxicity assays using HFFs or HLs
target cells and polyclonal NK cultures as effector cells (Fig. 4). Whereas fibroblasts infected with the
clinical strains always showed a consistent reduction in the
susceptibility to killing, AD169-infection resulted in a variable
pattern of reactivity. In some experiments, AD169 infection increased
the susceptibility to NK killing (Fig. 4, A and
B). However, in our experimental conditions, this pattern
was not always observed; in some experiments there was a decrease in
the killing of AD169-infected cells (Fig. 4C). HLs and HFFs
infected with the Towne strain were on the average less susceptible to
lysis compared with the uninfected counterpart. However, the reduced
recognition conferred by Towne infection was weaker than that detected
with clinical strain-infected fibroblasts (ranging between 11 and
25%), and in some experiments no protection was detectable.
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). Also in this experimental
condition, the clinical isolates were able to inhibit NK cell-mediated
killing, and infection with AD169 still resulted in a variable
pattern of NK recognition, being able to increase (Fig. 5) or decrease
(data not shown) the susceptibility to NK killing.
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Lack of correlation between hCMV-induced cell surface adhesion molecules and HLA-I modulation on fibroblasts target cells and their susceptibility to NK cell-mediated lysis
It has been proposed that the susceptibility of hCMV-infected
cells to NK lysis correlates with changes in cell surface expression of
the adhesion molecules LFA-3 and ICAM-1 (10, 11). We
therefore investigated whether the differences observed between cells
infected with hCMV clinical or laboratory strains could be due to a
virally induced modulation of the expression of these adhesion
molecules. HFFs were infected with different strains of hCMV at a MOI 2
(Fig. 6) or MOI 1 (data not shown) and
stained for ICAM-1 and LFA-3 expression at 4 and 8 days postinfection,
respectively. As previously reported for AD169 (10, 11)
and for other clinical isolates not included in this study
(11), there was an increased expression of ICAM-1 upon CMV
infection, with a similar pattern observed for all the virus strains
tested (Fig. 6A). In contrast, infection of fibroblasts
either with AD169 or with clinical strains did not affect LFA-3
expression (Fig. 6B). The same pattern was observed when the
cells were infected with a MOI of 1 and stained 8 days later (data not
shown). In Fig. 6C, the T2 cell line was used as a positive
control for LFA-3 and ICAM-1 staining.
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and 8).
There was a low basal level of HLA-I expression on uninfected cells. A
down-modulation of HLA-I on HFFs infected either with clinical isolates
or with laboratory strains was observed (Fig. 8). Down-modulation of
HLA-I molecules in AD169- and Towne-infected cells was statistically
significant, although this was not the case with clinical
strain-infected cells. Comparable results were obtained with HLs (data
not shown).
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), thus arguing against a role for HLA-I
molecules in the decreased susceptibility to killing. On uninfected
cells, an increase in the cytotoxicity was observed (Fig. 9),
suggesting that on these cells HLA-I partially inhibited NK
cell-mediated killing. Similar results were obtained with HLs (data not
shown).
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Cosman et al. (20) have described that a soluble form
of UL18 is able to bind to LIR-1, an inhibitory receptor belonging to
the Ig-like superfamily. Therefore, one possible mechanism was that
UL18 could protect cells infected with clinical strains from NK lysis
through the engagement and triggering of the inhibitory function of
LIR-1. To address this issue, NK cells were preincubated with
saturating concentrations of anti-LIR 1 (M405) or anti-CD56
(C218) mAb and then used as effectors against HFFs infected with
different hCMV strains (Fig. 10). The
susceptibility to NK lysis was not affected by the anti-LIR-1
treatment. In particular, the inhibition of the killing observed with
cells infected with clinical isolates could not be reverted by blocking
the LIR-1 receptor on the effectors (Fig. 10, A and
B). No substantial changes were observed with the control
anti-CD56 mAb. In these experiments, 10% (Fig. 10A) and
15% (Fig. 10B) of the effector cells expressed LIR-1 (data
not shown).
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, C and D). As previously shown
(30), NKL cells were able to kill 721.221 target cells
(Fig. 10C), whereas they were unable to kill the HLA-G1
expressing transfectant (Fig. 10D). Pretreatment of NKL with
the anti-LIR1 Ab reconstituted the capability to kill DT361 at
levels comparable to untransfected cells, suggesting that LIR-1 was
involved in the recognition of HLA-G1 and that M405 Ab was able to
interfere with LIR-1 function. No effect was seen with the
anti-CD56 control Ab. These results indicate that the inhibition of NK lysis observed with fibroblasts infected with the clinical isolates is not dependent on LIR-1.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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). The inhibition ranged
between 50 and 90%. Similar findings were obtained with fibroblasts
infected with two other clinical isolates, but not when they were
infected with laboratory strains AD169 and Towne (
Figs. 2–5). Thus,
the capability to prevent NK recognition of HFF- and HL-infected cells
may be a general feature of hCMV clinical strains. The level of
infection of target cells ranged between 95 and 100%, thus excluding
any possible influence of uninfected cells, and further supporting the
idea that the hCMV infection itself provided the protection from lysis.
The inhibition of NK cell killing by hCMV clinical strains could be
observed with different NK cell preparations (i.e., polyclonal
cultures, LAK cells, and NK clones) and with two different conditions
of infection (MOI 1 and MOI 2). Whereas the clinical strains
reproducibly inhibited NK recognition, only a marginal effect was
detected on target cells infected with the Towne laboratory strain.
Infection with the viral strain AD169 resulted either in an inhibition
or in an increase of NK susceptibility, and in some experiments no
substantial changes were found. Contradictory data have previously been
reported by other authors concerning the effect of AD169 infection on
NK sensitivity (10, 11, 31, 32). Thus, clinical vs laboratory strains of hCMV differed greatly in their capability to affect NK susceptibility of infected cells. The reason for this difference is at present unknown; genetic differences between the viral strains is one obvious possibility, and a recent report indicates a large DNA sequence difference between clinical isolates and laboratory strains of hCMV (26). The data revealed large scale deletions up to 15 kbp of DNA in laboratory strains that have been extensively passaged in cell culture (i.e., AD169 and Towne). These deletions were not found in low passaged strains, such as Toledo and the clinical strains tested in the study (26). Although at present the functions of the putative proteins encoded in this region are mostly unknown, they may play an important role for the replication, latency, or pathobiology of the virus in the human host. In fact, Penfold et al. (33) have recently shown that one of the genes deleted in laboratory strains encodes for a chemokine. Clinical trials based on immunization of healthy volunteers with AD169, Towne, or Toledo strains indicate that the low passaged Toledo strain exhibits a significantly higher virulence, being able to cause clinically apparent disease (34), compared with other two highly passaged strains (35, 36, 37, 38). It is thus possible that gene products involved in the immunoescape and survival of the virus in the host are deleted in the laboratory strains (33). In the in vitro conditions, there may be no need to escape from the immune system of the host; it is therefore likely that a high number of passages in vitro can lead to the loss of genes that are not essential for hCMV growth and survival in cultured cells.
In an attempt to analyze the possible molecular mechanisms used by the
clinical strains to prevent NK killing, we analyzed the modulation of
the expression of HLA-I molecules after hCMV infection. We observed a
statistically significant down-regulation of HLA-I levels only after
infection with AD169 or Towne laboratory strains compared with
noninfected fibroblasts. Infection with clinical strains also resulted
in HLA-I down-modulation, but always at a lower degree compared with
the effect observed with the laboratory strains. These data argued
against a major role of HLA-I molecules in relation to the difference
in NK sensitivity between infected and noninfected fibroblasts.
However, it was still possible that HLA-I played an important role in
the difference between cells infected with different viral strains.
There were at least two different possibilities: 1) the inhibition of
the killing could be the result of a cooperation between the effect of
HLA-I and a viral factor; 2) HLA-I could play no role at all, and the
inhibition would be due to the viral factor alone. Masking HLA-I
molecules on the surface of target cells with a specific mAb in the
cytotoxicity assays should reconstitute NK recognition if the first
hypothesis was correct. The blocking experiments showed no
reconstitution of cytotoxicity (Fig. 9 and data not shown). Thus,
although fibroblasts infected with clinical strains have higher levels
of HLA-I molecules than fibroblasts infected with laboratory strains,
HLA-I molecules do not appear to play a role in the induction of the
NK-resistant phenotype. Our data are in agreement with previous reports
where no correlation was found between cell surface HLA-I expression
and NK capability to sense hCMV-infected cells (10, 11, 31). While our study was in progress, a report by Fletcher et
al. (11) proposed that an increasing NK cytotoxicity
against hCMV infected targets can be due to an increase of the adhesion
molecules LFA-3 or ICAM-1. Although, as previously reported, we could
observe an increase in ICAM-1 expression on the surface of the cells
infected with different hCMV strains (10, 11), we were
unable to detect any correlation between LFA-3 expression and
susceptibility to NK lysis in both experimental conditions (Fig. 6 and
data not shown).
Another possible mechanism involved in the protection of cells infected
with the clinical isolates could be mediated by the binding of UL18 to
LIR-1-expressing NK cells. LIR-1 is an inhibitory receptor structurally
related to killer inhibitory receptors, having four extracellular
Ig-like domains and delivering an inhibitory signal after engagement
with the ligand (for a review, see Refs. 21 and
22). It is expressed on B cells, on monocytes, and on a
subpopulation of T and NK cells, and it has a broad specificity, being
able to recognize classical and nonclassical HLA class I molecules, as
well as the viral-encoded MHC-I molecule UL18 (20, 23, 24, 25, 30, 39). Thus, in our system, one possibility was that UL18,
expressed on the cell surface of infected cells, was able to engage
LIR-1 on NK cells, thus inhibiting NK lysis. The blocking experiments
argue against this possibility, as we could not restore the killing by
blocking the LIR-1 receptor on NK cells (Fig. 10, A and
B). The range of LIR-1 expression in the polyclonal NK
cultures used in these experiments was 10–15% (data not shown). We
cannot exclude the possibility that the use of effector cells with a
higher and homogeneous LIR-1 expression could lead to different
conclusions. On the other hand, our blocking experiments clearly
suggest that the inhibition is detectable even in the presence of a low
percentage of LIR-1 expressing effector cells, and it is not affected
by anti-LIR-1 mAb.
If we can exclude a role for HLA-I, for adhesion molecules (at least ICAM-1 and LFA-3) and for LIR-1, which is then a possible molecular mechanism involved in the protection of cells infected with the clinical strains? There are at least two different possibilities. First, the inhibition could be mediated by a soluble host cell factor released from infected cells during the cytotoxicity assay, able to act either at the target or at the effector cell level. In this context, IFN-ß, TGF-ß, or soluble HLA-I could play a role in the inhibition of the killing (31, 40, 41, 42). The second possibility still lies at the level of interaction between molecules on effector and target cells. In this context, hCMV clinical strains may induce on the cell surface of infected cells the expression of different types or levels of UL18 or of other molecules involved in the NK recognition.
The majority of the data available in the literature concerning hCMV interactions with NK cells were generated with laboratory strains. Our data suggest that caution should be applied in the interpretation of these data, and this is further emphasized by reports showing that large genomic variations can exist between laboratory and clinical isolates (26).
Our hCMV clinical strains were generated from either transplant recipients during graft rejection or HIV patients. We speculate that during these conditions the restoration of efficient NK activities would contribute to control the overwhelming hCMV infection, as suggested also by observations in mice infected with mCMV (3).
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Cristina Cerboni, Microbiology and Tumor Biology Center, Karolinska Institutet, Doktorsringen 13, Box 280, S-171 77 Stockholm, Sweden.
3 Abbreviations used in this paper: hCMV, human CMV; mCMV, murine CMV; HLA-I, HLA class I; HFFs, human foreskin fibroblasts; HLs, human lung fibroblasts; MOI, multiplicity of infection; LAK, IL-2-activated killer cells; ß2m, ß2-microglobulin; LIR-1, leukocyte Ig-like receptor-1.
Received for publication May 18, 1999. Accepted for publication February 16, 2000.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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chemokine. Proc. Natl. Acad. Sci. USA 96:9839.This article has been cited by other articles:
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M. R. Wills, O. Ashiru, M. B. Reeves, G. Okecha, J. Trowsdale, P. Tomasec, G. W. G. Wilkinson, J. Sinclair, and J. G. P. Sissons Human Cytomegalovirus Encodes an MHC Class I-Like Molecule (UL142) That Functions to Inhibit NK Cell Lysis J. Immunol., December 1, 2005; 175(11): 7457 - 7465. [Abstract] [Full Text] [PDF]
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A.-C. Iversen, P. S. Norris, C. F. Ware, and C. A. Benedict Human NK Cells Inhibit Cytomegalovirus Replication through a Noncytolytic Mechanism Involving Lymphotoxin-Dependent Induction of IFN-{beta} J. Immunol., December 1, 2005; 175(11): 7568 - 7574. [Abstract] [Full Text] [PDF]
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A. Rolle, M. Mousavi-Jazi, M. Eriksson, J. Odeberg, C. Soderberg-Naucler, D. Cosman, K. Karre, and C. Cerboni Effects of Human Cytomegalovirus Infection on Ligands for the Activating NKG2D Receptor of NK Cells: Up-Regulation of UL16-Binding Protein (ULBP)1 and ULBP2 Is Counteracted by the Viral UL16 Protein J. Immunol., July 15, 2003; 171(2): 902 - 908. [Abstract] [Full Text] [PDF]
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J. Odeberg, H. Browne, S. Metkar, C. J. Froelich, L. Branden, D. Cosman, and C. Soderberg-Naucler The Human Cytomegalovirus Protein UL16 Mediates Increased Resistance to Natural Killer Cell Cytotoxicity through Resistance to Cytolytic Proteins J. Virol., April 15, 2003; 77(8): 4539 - 4545. [Abstract] [Full Text] [PDF]
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L. Rasmussen, A. Geissler, C. Cowan, A. Chase, and M. Winters The Genes Encoding the gCIII Complex of Human Cytomegalovirus Exist in Highly Diverse Combinations in Clinical Isolates J. Virol., October 2, 2002; 76(21): 10841 - 10848. [Abstract] [Full Text] [PDF]
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C. S. Falk, M. Mach, D. J. Schendel, E. H. Weiss, I. Hilgert, and G. Hahn NK Cell Activity During Human Cytomegalovirus Infection Is Dominated by US2-11-Mediated HLA Class I Down-Regulation J. Immunol., September 15, 2002; 169(6): 3257 - 3266. [Abstract] [Full Text] [PDF]
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E. C. Y. Wang, B. McSharry, C. Retiere, P. Tomasec, S. Williams, L. K. Borysiewicz, V. M. Braud, and G. W. G. Wilkinson From the Cover: UL40-mediated NK evasion during productive infection with human cytomegalovirus PNAS, May 28, 2002; 99(11): 7570 - 7575. [Abstract] [Full Text] [PDF]
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J. Odeberg and C. Söderberg-Nauclér Reduced Expression of HLA Class II Molecules and Interleukin-10- and Transforming Growth Factor {beta}1-Independent Suppression of T-Cell Proliferation in Human Cytomegalovirus-Infected Macrophage Cultures J. Virol., June 1, 2001; 75(11): 5174 - 5181. [Abstract] [Full Text]
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) at a MOI of 1. The susceptibility of these cells to NK
cell-mediated lysis was then measured on day 8 postinfection in a 4-h
51Cr release assay. A, A polyclonal NK cell
culture was used as effector cells. B, LAK cells from
the same donor were used as effectors.
) or 109B
(
). HFFs were infected at a MOI of 1 and used 8 days after infection
in a 4-h 51Cr release assay. Two independent experiments
are shown, in which NK cells derived from donor 41 (A
and C) or donor 42 were used (B and
D).
);
AD169 and Towne (
); 4636, 13B, and 109B (
). Lysis at an E:T ratio
of 20:1 from three independent experiments is shown. *, Not tested.
A and B, HFFs as target cells;
C, HLs as target cells.
