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Regular Articles |
From the Department of Pathology and Skin Cancer Research
Laboratories,*
Cardinal Bernardin Cancer Center, Loyola
University Medical Center, Maywood, Illinois; and the Department of
Microbiology, Molecular Genetics and
Immunology,
University of Kansas Medical
Center, Kansas City, Kansas
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Abstract |
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2-herpesvirus consistently
identified in Kaposi’s sarcoma (KS), primary effusion
lymphoma, and multicentric Castleman’s disease. Although HHV-8
infection appears to be necessary, it may not be sufficient for
development of KS without the involvement of other cofactors. One
potentially important cofactor is HIV-1. HIV-1-infected cells produce
HIV-1-related proteins and cytokines, both of which have been
shown to promote growth of KS cells in vitro. Though
HIV-1 is not absolutely necessary for KS development, KS is the
most frequent neoplasm in AIDS patients, and AIDS-KS is
recognized as a particularly aggressive form of the disease. To
determine whether HIV-1 could participate in the pathogenesis of KS by
modulating HHV-8 replication (rather than by inducing
immunodeficiency), HIV-1-infected T cells were cocultured with
the HHV-8-infected cell line, BCBL-1. The results demonstrate
soluble factors produced by or in response to HIV-1-infected T cells
induced HHV-8 replication, as determined by production of lytic
phase mRNA transcripts, viral proteins, and detection
of progeny virions. By focusing on cytokines produced in the coculture
system, several cytokines known to be important in growth and
proliferation of KS cells in vitro, particularly
Oncostatin M, hepatocyte growth factor/scatter factor,
and interferon-, were found to induce HHV-8 lytic
replication when added individually to BCBL-1 cells. These results
suggest specific cytokines can play an important role in the initiation
and progression of KS through reactivation of HHV-8. Thus,
HIV-1 may participate more directly than previously recognized in KS by
promoting HHV-8 replication and, hence, increasing
local HHV-8 viral load.
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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2-herpesviruses (genus Rhadinovirus) to infect
humans.1,2
This virus has been consistently associated
with Kaposi’s sarcoma (KS), primary effusion lymphomas (PEL, also
termed body cavity-based lymphomas or BCBL), and multicentric
Castleman’s disease.3
Epidemiological evidence strongly
implicates HHV-8 as the etiological agent of KS. This conclusion is
based on the presence of HHV-8 in all four clinical forms of KS and in
virtually all KS lesions,4-7
the presence of
HHV-8-specific antibodies in a majority of KS patients (over 80%), but
a minor proportion of normal healthy blood donors
(<10%),8-11
and the presence of HHV-8-specific
antibodies in patients before the development of KS
lesions.10 All herpesviruses, including HHV-8, share the ability to establish latent infections in their natural host cells.12 In latent infection, the viral genome persists extrachromosomally as a circular episome, viral gene expression is severely restricted, and viral progeny are not produced.13 Previous studies have demonstrated that HHV-8 viral replication in PEL and KS tumor cells is tightly regulated, with the virus being held predominantly in a latent state.14,15 However, treatment of PEL cells with either phorbol esters (TPA) or n-butyrate induces HHV-8 reactivation. During reactivation, HHV-8-infected cells express a variety of lytic cycle genes and produce viral progeny, which ultimately results in the destruction of the host cell.16-19 Regulation of viral replication is critical to disease progression as the proportion of virally infected cells undergoing lytic replication increases, so does the degree of tissue damage and progression of the infection. Indeed, studies have shown that HHV-8 viral load is higher in KS patients than in HHV-8-infected individuals without KS, and HHV-8 viral load also increases during progression of KS to the late (nodular) stage of the disease.13,20,21
Although HIV-1 infection is neither necessary nor sufficient for the development of KS, AIDS-KS is known to be more aggressive, disseminated, and resistant to treatment than other forms of KS, including forms also associated with immunosuppression.22-24 Immunosuppression clearly plays a role in the development of KS, but immunosuppression alone cannot explain the overwhelming prevalence of KS in AIDS patients (relative risk of developing KS in AIDS patients is 70-fold higher than in immunosuppressed transplant patients),25,26 the frequent presentation of KS in the early stages of AIDS before the onset of severe immunosuppression,27,28 and the overwhelming association of KS in patients infected with HIV-1 but not HIV-2.29 Recent studies by Ariyoshi et al demonstrate that KS developed almost exclusively in HIV-1-positive, not HIV-2-positive, patients in Gambia, West Africa, despite essentially equivalent seroprevalance for HHV-8 and severity of immunosuppression in both groups.29
In an attempt to explain the more aggressive nature of KS in
HIV-1-positive patients, HHV-8 replication was examined in the
persistently infected PEL cell line, BCBL-1, after coculture with an
HIV-1-infected CD4+ T cell line. The results demonstrate that soluble
factors produced by HIV-1-infected CEM cells induce lytic cycle
replication of HHV-8. In the present study, we focused on inflammatory
cytokines and found that cytokines important in the growth and
proliferation of KS tumor cells in vitro, particularly
Oncostatin M (OSM), hepatocyte growth factor/scatter factor (HGF/SF),
and interferon- (IFN-), induce HHV-8 lytic cycle replication.
Previous studies from our group and others have demonstrated the
presence of HIV-1 in KS skin lesions in proximity to the cells that
harbor HHV-8. These studies identified HIV-1 transcripts in dermal
dendritic cells, T cells, and basal keratinocytes in
KS.30,31
Similarly, HHV-8 was identified in basal
keratinocytes, eccrine epithelial cells, KS tumor cells, and
endothelial cells.32,33
In addition, recent studies have
found that dendritic cells, keratinocytes, and macrophages can also be
infected by HHV-8 in vitro.34-36
Therefore, we
propose that the HIV-1-mediated enhancement of HHV-8 lytic replication
may increase the local viral load and, hence, increase the likelihood
of tumor formation. Such a mechanism would point to a more direct
relationship between HIV-1 and HHV-8 than previously postulated by
other investigators studying KS.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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CEM-SS cells (kindly provided by Drs. Clive Woffendin and Gary Nabel, University of Michigan, Ann Arbor, MI) were maintained in RPMI-1640 (BioWhittaker, Walkersville, MD) containing 10% heat-inactivated fetal bovine serum (FBS), 2 mmol/L glutamine, 100 U/ml penicillin, and 100 µg/ml streptomycin (RPMI + 10% FBS). The BCBL-1 cell line, originally isolated by Drs. Michael McGrath and Don Ganem, was obtained through the AIDS Research and Reference Reagent Program, Division of AIDS, National Institute of Allergy and Infectious Diseases, National Institutes of Health.18 BCBL-1 cells, an EBV-negative and HHV-8-positive PEL cell line, were cultured in RPMI 1640 supplemented with 5% FBS, 5% normal human AB serum (BioWhittaker) and 2 mmol/L L-glutamine (BCBL-1 media) at 37°C in a humidified, 5% CO2 atmosphere. BC-1 cells (an HHV-8- and EBV-positive PEL cell line) and BC-3 cells (an EBV-negative and HHV-8-positive PEL cell line) were obtained from ATCC and were grown in RPMI + 10% FBS or RPMI + 20% FBS, respectively.37 For experiments using normal human peripheral blood mononuclear cells (PBMC), human whole blood was separated on a Ficoll-Hypaque gradient and PBMCs isolated from the interface. The cells were washed three times in RPMI 1640 and resuspended in RPMI + 10% FBS containing 20 U/ml interleukin-2 (IL-2) before infection with HIV-1.
In a majority of the coculture experiments, Transwell dishes (CoStar)
were used to prevent cell-cell interactions while allowing passage of
soluble factors, such as cytokines and viral particles, between the two
cultures. Several experiments required supplementation of the culture
media with recombinant cytokines. The cytokines used were purchased
from R&D Systems (Minneapolis, MN) and included IFN-
(50–1000 U/ml), tumor necrosis factor-
(TNF-; 1000 U/ml),
IL-6 (50 ng/ml), HGF/SF (100 ng/ml), OSM (10 ng/ml), and IL-2 (20
U/ml).
High-titer stocks of HIV-1IIIB (kindly provided by Drs. Clive Woffendin and Gary Nabel) and HIV-1SF2 (AIDS Research and Reference Reagent Program, Division of AIDS, National Institute of Allergy and Infectious Diseases) were prepared by passaging the viruses on CEM cells. The TCID50 titer (106/ml) was calculated as terminal dilution producing syncytia. CEM cells were incubated with 0.01 multiplicity of infection of HIV-1 for 4 hours at 37°C. After the incubation, cells were washed with RPMI-1640 and resuspended at 3 x 105 cells/ml in RPMI + 10% FBS. Production of HIV-1 was monitored by measuring reverse transcriptase (RT) activity in tissue culture supernatants (performed in duplicate) as previously described.38
Northern Blot Analysis
Total RNA was extracted from cells using a phenol/ guanidine isothiocyanate/chloroform-based technique (Trizol, Gibco BRL, Gaithersburg, MD) following the manufacturer’s instructions. Twelve micrograms of total RNA was fractionated on a 1% agarose formaldehyde gel and transferred to a nylon membrane (Zetabind, Cuno Inc., Meriden, CT). Even loading of RNA and efficiency of transfer was confirmed by staining of the 18S and 28S bands on the membrane with methylene blue.39 The blot was prehybridized with Church’s hybridization buffer and probed with a [32P]-labeled dCTP probes generated using gel-purified PCR products and a random prime label kit (Roche Molecular Biochemicals, Indianapolis, IN). The blots were washed with sodium phosphate buffers containing sodium dodecyl sulfate, EDTA, and bovine serum albumin and exposed to Kodak film.
Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR)
cDNA was synthesized from isolated RNA using the SuperScript Preamplication system for First Strand cDNA Synthesis (Gibco BRL) following the manufacturer’s instructions. To ensure no DNA contamination of the RNA, which could lead to false positive results, the RNA samples were treated with DNase I (Gibco BRL) before reverse transcription. As an additional control, each sample was also subjected to reverse transcription in the absence of RT. Single-stranded cDNA was then amplified using standard PCR techniques as previously described.40,41 Primers used for analysis included HHV-8 ORF26 primers originally described by Chang et al1 and HGF/SF (5'-TTT GCC TTC GAG CTA TCG GG-3', 5'-GAA TTT GTG CCG GTG TGG TG-3'). To prevent PCR contamination due to previously amplified PCR products, all PCR reactions were set up in a separate location from the area used for amplification and analysis. All PCR-related reagents were stored in this room, including dedicated pipettors, aerosol barrier pipette tips, and reaction tubes. Uracil DNA glycosylase and dUTP-spiked dNTP were also used in the PCR reaction to destroy any previously amplified product, as previously described.42,43
Immunoperoxidase Staining
Cytospin preparations of cultured cells were fixed for 10 minutes in 50:50 acetone:methanol and air-dried. The cells were immunostained to detect a variety of antigens using a highly sensitive avidin-biotin immunoperoxidase technique (Vectastain kit, Vector Laboratories, Burlingame, CA) as previously described.44 The chromogen, 3-amino-4-ethylcarbazole was used, producing a positive red reaction. The panel of monoclonal antibodies (mAb) used included CD4 (mouse IgG; Becton-Dickinson, San Jose, CA), HHV-8 ORF 59 (clone 11D1, mouse IgG), and HHV-8 ORF K8.1 A/B (clone 2A3, mouse IgG). Both ORF 59 and K8.1 mAbs recognize HHV-8 lytic cycle proteins and have been previously described.45-47 To calculate the percentage of positive cells, photographs of at least 10 unique fields were taken of every slide, and the number of positive and negative cells counted separately by two individuals, including one who was blinded to the results. Immunostaining was performed on samples from three separate experiments.
Ribonuclease Protection Assay (RPA)
Expression of mRNA encoding for cytokines was evaluated in BCBL-1, CEM, and HIV-1-infected CEM cells both before and after coculture using RPA kits and the manufacturer’s recommended instructions (RiboQuant RPA kit, Pharmingen, San Diego, CA).
Enzyme-Linked Immunosorbent Assay (ELISA)
Production of IFN-, OSM, and HGF/SF was measured in PBMC, PBMC
+ HIV, CEM, CEM + HIV, and BCBL-1 cells both before and after coculture
using Quantikine ELISA kits (R&D Systems). The assay was performed as
recommended by the manufacturer using undiluted tissue culture
supernatants. Each sample was run in duplicate and the assay repeated a
minimum of two times.
Electron Microscopy (EM)
BCBL-1 cells were stimulated for 7 days with HGF/SF as described. The cells were collected, washed once with PBS, and fixed with EM-grade glutaraldehyde in cacodylate buffer. Cells were processed, sectioned, and stained by the Loyola University Core Imaging facility. Analysis and interpretation of electron micrographs was performed by an expert electron microscopist, Dr. Raoul Fresco (Loyola University, Maywood, IL).
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Results |
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To test the hypothesis that HIV-1 may play a role in regulation of HHV-8 replication, BCBL-1 cells were cultured in direct contact with HIV-1IIIB infected CEM cells (CEM + HIV-1IIIB). Northern blot analysis for T0.7 mRNA at 24, 48, 72, and 96 hours after coculture demonstrated an increase in HHV-8 mRNA (data not shown). Expression of T0.7 mRNA increased over time in the cocultured cells, in comparison with untreated BCBL-1 (twofold increase at 24 hours, rising to a sixfold increase at 96 hours). Analysis of mRNA for the cellular "housekeeping" gene glyceraldehyde-3-phosphate dehydrogenase (GAPDH) on the same membrane indicated that loading of RNA was similar (data not shown).
As the factor(s) responsible for increased HHV-8 replication may be
either soluble or cell-associated, similar coculture experiments were
performed in Transwell dishes to prevent direct contact between the two
cell types. Northern blot analysis for ORF26 mRNA (expressed only
during lytic HHV-8 replication) demonstrated an increase in mRNA
expression, which continued to increase over time (Figure 1)
. Of interest, ORF26 mRNA also
increased after coculture with uninfected CEM cells, but to a lesser
extent than demonstrated with CEM+HIV-1IIIB at
the same time point (Figure 1)
. Analysis of data from 5 independent
experiments demonstrated that, on average, ORF26 expression was
increased 2.6 ± 0.39-fold at 72 hours and 3.25 ± 0.44-fold
at 96 hours when comparing BCBL-1 cells cocultured with uninfected
versus HIV-infected CEM cells. In each experiment, analysis
of GAPDH on the same membrane demonstrated loading of RNA was similar.
The results indicated not only that soluble factors are responsible for
induction of HHV-8, but these factors induce HHV-8 lytic phase
replication; not simply a proliferation of latently infected cells.
|
. The
difference in the results between BCBL-1 and BC-1 cells may be due to
the expression of EBV-regulatory proteins in BC-1 cells, which may
suppress HHV-8 re-activation.
|
. The decrease in HHV-8
ORF26 mRNA likely represented exhaustion of critical factors in the
conditioned media over time which are responsible for induction of the
HHV-8 lytic cycle.
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, top row). Of importance, expression of
ORF59 has been previously shown to occur earlier and more frequently in
the lytic cycle compared with expression of ORF K8.1.48
Indeed, after 10 days of coculture with CEM +
HIV-1IIIB cells, 7.9 ± 0.8% of BCBL-1
cells expressed ORF K8.1 compared to 4.6% ± 0.2% of BCBL-1 cells
cultured with uninfected CEM cells and 1.1% ± 0.1% of untreated
BCBL-1 cells (P < 0.005, Figure 4
center row).
Staining for an isotype control mAb, CD4, showed less than 0.4 ±
0.2% reactivity with cocultured BCBL-1 cells; however, 88.6 ±
1.6% of CEM cells were positive for CD4 (Figure 4
, bottom row).
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.
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As HHV-8 lytic replication was induced by soluble factors, we
reasoned that cytokines released either by CEM, CEM +
HIV-1IIIB, or BCBL-1 cells (in response to
culture with HIV-1-infected T cells) were likely candidates to mediate
this response. To identify cytokines modulated in the coculture system,
RPA assays were performed (Table 1)
. Of
particular interest, IFN- mRNA was not detected in CEM or CEM +
HIV-1IIIB cells, but was readily detected in
BCBL-1 cells, particularly after coculture with CEM +
HIV-1IIIB (10-fold increase at 72 hours of
coculture compared to baseline levels). IFN- mRNA was also detected
in BCBL-1 cells cocultured with uninfected CEM cells, but the increase
was significantly less (fivefold increase at 72 hours of coculture)
than in the presence of HIV-1. In addition, OSM mRNA was not expressed
by BCBL-1 cells in any of the assay conditions, but was detected in CEM
and CEM+HIV-1IIIB cells. CEM and
CEM+HIV-1IIIB cells cocultured with BCBL-1 cells
expressed OSM mRNA at modestly increased levels (25% increase compared
to CEM or CEM+HIV-1IIIB alone). As the RPA kits
do not evaluate expression of HGF/SF mRNA, RT-PCR was used to evaluate
production of this mRNA. Results showed HGF/SF mRNA was not detected in
CEM or CEM + HIV, but was readily detected in BCBL-1 cells both before
and after coculture with CEM + HIV (data not shown).
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, OSM, and HGF/SF proteins. IFN- was not detected
in CEM cells, CEM+HIV-1IIIB cells, or BCBL-1
cells, but was produced by BCBL-1 cells cocultured with CEM +
HIV-1IIIB. Levels of IFN- increased over time
starting at undetectable levels at time 0 and reaching 12–20 pg/ml at
8 days of coculture. OSM was not produced by BCBL-1 cells, but was
produced by CEM cells, CEM+HIV-1IIIB cells, and
CEM or CEM+HIV-1IIIB cells cocultured with BCBL-1
cells. The cells produced approximately 10–14 pg/ml of OSM, and there
was no significant difference in expression of OSM between the groups
or over time. HGF/SF was not detected in BCBL-1, CEM or CEM +
HIV-1IIIB cells. However, following coculture of
either CEM or CEM + HIV-1IIIB cells with BCBL-1
cells, HGF/SF could be detected in the culture media. HGF/SF levels
were undetectable at early time points and steadily increased over time
to 90 ± 47 pg/ml (CEM cocultured with BCBL-1), and 206 ± 45
pg/ml (CEM + HIV cocultured with BCBL-1) after 10 days.
As IFN-, OSM, and HGF/SF were all detected at the protein level in
pg/ml concentrations, experiments were designed to determine whether
recombinant cytokines could also induce HHV-8 lytic replication. BCBL-1
cells were stimulated in culture with recombinant cytokines (IFN-,
OSM, HGF/SF, IL-6, TNF-, or IL-2) and HHV-8 replication examined.
Using Northern blot analysis, studies demonstrated an increase in ORF26
mRNA in BCBL-1 cells stimulated with OSM, HGF/SF and IFN-, but not
IL-6, TNF-, or IL-2 at both 3 and 7 days of cytokine stimulation
(Figure 6)
. In addition, as the cytokines
used in this study have all been identified in KS lesions, a cocktail
of all six recombinant cytokines was used to determine whether the
cytokines could act synergistically to promote higher levels of HHV-8
replication. Northern blot analysis demonstrated an increase in ORF26
mRNA levels with the cytokine cocktail that were equivalent or slightly
higher than levels seen with the individual cytokines alone. Induction
of HHV-8 proteins in BCBL-1 cells stimulated with recombinant cytokines
was also demonstrated by immunohistochemical staining (Figure 7)
. The results showed significant
increases in HHV-8 K8.1 expression for OSM, HGF/SF, and IFN-, but
not IL-6, TNF-, or IL-2 at 7 days of stimulation. To confirm that
the stimulated cells were indeed in lytic cycle and producing progeny
virus, EM analysis of cells stimulated with HGF/SF was performed. As
shown in Figure 7
, viral particles were readily identified in cells.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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, HGF/SF,
IL-6, IL-1ß, and TNF-, are produced in KS, both by KS tumor cells
and mononuclear cells, including infected lymphocytes, infiltrating the
lesion in AIDS patients. Many of these cytokines are known to promote
the growth and proliferation of KS tumor cells in
vitro.52,56-63
In addition, cytokines have been show
to induce normal endothelial cells, the likely precursor cell of the KS
tumor cell, to acquire the characteristics typical of KS tumor cells,
including spindle-shaped morphology, production of angiogenic factors,
and expression of activation markers. These results, and others, have
lead to the hypothesis that cytokines play a critical role in the
initiation and development of KS by activating endothelial cells,
promoting production of angiogenic factors, and providing the necessary
growth factors for the tumor cells.51
In the current study, we demonstrated that cytokines produced by
HIV-1-infected cells can induce lytic cycle replication of HHV-8.
Several cytokines known to be important in KS, particularly OSM,
HGF/SF, and IFN-, were able to induce lytic cycle mRNA transcripts
and viral proteins resulting in the production of progeny virions.
These results are consistent with the above mentioned studies
demonstrating the importance of cytokines in KS, and they add
additional support to the hypothesis that cytokines may be involved in
initiation of KS. As HHV-8 is a lymphotropic virus, a potential
mechanism for initiation of KS could involve cytokine stimulation of
latently infected mononuclear cells resulting in induction of lytic
viral replication and production of viral progeny which can go on to
infect endothelial cells. Indeed, recent in situ
hybridization studies by Sturzl et al have shown HHV-8-infected
mononuclear cells adherent to endothelial cells in KS
lesions.64
This direct contact between infected cells and
endothelial cells may be important to infection of the tumor cell
precursor. How infected endothelial cells acquire the malignant
phenotype is currently unknown, but it may involve subsequent
interactions between HHV-8-encoded proteins and host cytokines and/or
growth factors. The end result of this host/virus interaction is
transdifferentiation of the endothelial cell ultimately leading to the
development of full-fledged KS lesions. Indeed, studies have shown that
after opportunistic infections, cytokine levels are increased, and
opportunistic infections frequently precede the development or growth
of KS lesions.65
In addition, early clinical studies
attempting to treat KS with TNF and IFN- were unsuccessful due to
rapid disease progression in many of the patients.66-68
Our results are also consistent with recently published studies by
Monini et al demonstrating that IFN- can induce HHV-8 lytic
replication in KS patient PBMCs resulting in prolonged survival of
HHV-8 in patients PBMCs in culture.69
The data in the
present study confirm and extend the work of Monini et al, indicating
that other cytokines (ie, OSM and HGF/SF) are also able to induce HHV-8
replication, resulting in production of progeny virions. However,
Monini and colleagues concluded that the effect of cytokines on HHV-8
replication were specific for PBMCs, as they found no significant
differences in latent or lytic viral gene expression in BCBL-1 cells
cultured with cytokines.69
The discrepancy between these
reports is likely due to experimental design, including differences
such as cytokine concentration, duration of cytokine stimulation, and
methods used for analysis of the results.
In the coculture system using HIV-1-infected CEM cells and the
HHV-8-infected PEL cell line, BCBL-1, we found that both cells appeared
to contribute to the combination of cytokines which induced HHV-8
replication. Of interest, the BCBL-1 cells, but not the CEM or CEM +
HIV-1 cells, demonstrated expression of IFN- mRNA in RPA assays and
coculture supernatants contained IFN- proteins as demonstrated by
quantitative ELISA. IFN- production is generally attributed to T
cells and natural killer (NK) cells, not to B cells. However, previous
studies have demonstrated the constitutive expression of IFN- mRNA
by B cell lines as well as the production of IFN- protein following
stimulation of the cells with various agents including phorbol esters
and cytokines.70,71
The lack of IFN- production by the
CEM or CEM + HIV-1 cells may be related to this particular T cell line.
Indeed, ELISA assays indicated production of OSM (510 ± 21
pg/ml), HGF/SF (337 ± 33 pg/ml), and IFN- (19 ± 6 pg/ml)
by normal human PBMCs in our coculture system that were equal to or
exceeded levels seen with the CEM cells. Thus, cytokines involved in
the reactivation of HHV-8 would likely be produced under physiological
conditions.
Of course, the results demonstrating the cytokines OSM, HGF/SF, and
IFN-, which are found in KS lesions and are produced either by
HIV-infected cells or in response to HIV-infected cells, can induce
HHV-8 replication does not eliminate the possibility that other
factors, such as HIV-1-related proteins or additional cytokines, play a
role in induction of HHV-8 replication either in vitro or
in vivo. Studies by Ensoli, Gallo and colleagues have
indicated that extracellular HIV-1 Tat protein can induce growth,
migration, invasion and adhesion of both endothelial cells and KS tumor
cells, and stimulation of normal endothelial cells with cytokines
activates the cells so that they become responsive to Tat
protein.51,72,73
An intriguing brief report by Harrington
et al indicates that Tat protein may be able to induce HHV-8
replication; however, this remains to be confirmed.74
In conclusion, the "cross-talk" demonstrated in this study, whereby HIV-1-infected T cells can induce lytic replication of HHV-8 via specific cytokines, provides new opportunities for targeted intervention in AIDS-related KS. Further studies are clearly indicated to examine more carefully the colocalization of HIV-1 and HHV-8 in sites such as cutaneous KS, as previously observed, to determine whether these in vitro findings are relevant to local tissue levels of HHV-8 in vivo.
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Acknowledgements |
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Footnotes |
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Supported by U. S. Public Health Service grants CA76951 (to K. E. F.), CA75893 (to B. J. N.), CA63928 (to B. J. N.), CA75911 (to B. C.), and CA82056 (to B. C.).
Accepted for publication February 9, 2000.
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N. Inoue, J. Winter, R. B. Lal, M. K. Offermann, and S. Koyano Characterization of Entry Mechanisms of Human Herpesvirus 8 by Using an Rta-Dependent Reporter Cell Line J. Virol., July 15, 2003; 77(14): 8147 - 8152. [Abstract] [Full Text] [PDF]
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 L. A. Dourmishev, A. L. Dourmishev, D. Palmeri, R. A. Schwartz, and D. M. Lukac Molecular Genetics of Kaposi's Sarcoma-Associated Herpesvirus (Human Herpesvirus 8) Epidemiology and Pathogenesis Microbiol. Mol. Biol. Rev., June 1, 2003; 67(2): 175 - 212. [Abstract] [Full Text] [PDF]
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S. Pati, J. S. Foulke Jr., O. Barabitskaya, J. Kim, B. C. Nair, D. Hone, J. Smart, R. A. Feldman, and M. Reitz Human Herpesvirus 8-Encoded vGPCR Activates Nuclear Factor of Activated T Cells and Collaborates with Human Immunodeficiency Virus Type 1 Tat J. Virol., May 15, 2003; 77(10): 5759 - 5773. [Abstract] [Full Text] [PDF]
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J. Tang, G. M. Gordon, M. G. Muller, M. Dahiya, and K. E. Foreman Kaposi's Sarcoma-Associated Herpesvirus Latency-Associated Nuclear Antigen Induces Expression of the Helix-Loop-Helix Protein Id-1 in Human Endothelial Cells J. Virol., May 15, 2003; 77(10): 5975 - 5984. [Abstract] [Full Text] [PDF]
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 M. Malnati, F. Broccolo, S. Nozza, L. Sarmati, S. Ghezzi, G. Locatelli, F. Delfanti, B. Capiluppi, A. Careddu, M. Andreoni, et al. Retrospective analysis of HHV-8 viremia and cellular viral load in HIV-seropositive patients receiving interleukin 2 in combination with antiretroviral therapy Blood, August 13, 2002; 100(5): 1575 - 1578. [Abstract] [Full Text] [PDF]
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S. Sakakibara, K. Ueda, J. Chen, T. Okuno, and K. Yamanishi Octamer-Binding Sequence Is a Key Element for the Autoregulation of Kaposi's Sarcoma-Associated Herpesvirus ORF50/Lyta Gene Expression J. Virol., August 1, 2001; 75(15): 6894 - 6900. [Abstract] [Full Text]
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J. P. Zoeteweij, A. V. Moses, A. S. Rinderknecht, D. A. Davis, W. W. Overwijk, R. Yarchoan, J. M. Orenstein, and A. Blauvelt Targeted inhibition of calcineurin signaling blocks calcium-dependent reactivation of Kaposi sarcoma-associated herpesvirus Blood, April 15, 2001; 97(8): 2374 - 2380. [Abstract] [Full Text] [PDF]
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 L.-M. Huang, M.-F. Chao, M.-Y. Chen, H.-m. Shih, Y.-P. Chiang, C.-Y. Chuang, and C.-Y. Lee Reciprocal Regulatory Interaction between Human Herpesvirus 8 and Human Immunodeficiency Virus Type 1 J. Biol. Chem., April 13, 2001; 276(16): 13427 - 13432. [Abstract] [Full Text] [PDF]
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