1. No category

From www.bloodjournal.org by guest on November 24, 2014. For personal use only.
1996 88: 3474-3481
Differential regulation of proinflammatory and hematopoietic
cytokines in human macrophages after infection with human
immunodeficiency virus
R Esser, W Glienke, H von Briesen, H Rubsamen-Waigmann and R Andreesen
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Copyright 2011 by The American Society of Hematology; all rights reserved.
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Differential Regulation of Proinflammatory and Hematopoietic Cytokines in
Human Macrophages After Infection With Human Immunodeficiency Virus
By Ruth Esser, Wolfgang Giienke, Hagen von Briesen, Helga Rubsamen-Waigmann, and Reinhard Andreesen
Cells of the macrophage lineage (MAC) play an important
role in human immunodeficiency virus (HIV) infection. However, the knowledge on the extent of macrophage involvement in the pathogenesis of HIV infection is still incomplete.
In this study we examined the secretory repertoire of HIVinfected MAC with respect to the proinflammatory cytokines
tumor necrosis factor-a (TNF-a), interleukin-lp (IL-lp), IL6, IL-8, and the hematopoietic growth factors M-, G- and
granulocyte-macrophage colony stimulating factor (GMCSF). Using a culture system on hydrophobic teflon membranes, blood-derived MO from healthy donors were infected with a monocytotropic HIV-1 isolate (HIV-ID,,71,1).
We
analyzed the constitutive and lipopolysaccharides-stimulated secretion of MO/MAC early after infection as well as
in long-term cultured, virus-replicating cells. The release of
proinflammatory mediators and hematopoietic growth factors were differentially regulated after infection with HIV:
the secretion of TNF-a, IL-Ip, IL-6, IL-8 was upregulated,
whereas a down-regulationof M-, G-, and GM-CSF could be
observed. These results may provide some explanation for
the immunological dysfunction, the hematopoietic failure
and the chronic inflammatory disease occuring in HIV-infected patients.
0 1996 by The American Society of Hematology.
M
therefore contribute to the pathogenesis of HIV-mediated
di~ease.~
TNF-a and IL-IP are produced in response to normal
immune stimuli such as immune complexes, lipopolysaccharides (LPS), and phorbol esters.’ Both molecules are highly
inflammatory, can cause fever and loss of appetite, and contribute to a catabolic state.”.” The main physiological functions of the proinflammatory cytokine interleukin-6 (IL-6)
are the induction of terminal differentiation of activated B
cells into antibody producing cells’* and the induction of the
hepatic acute phase response.I3 IL-8 functions as a chemoattractant for neutrophil^,'^ basophil^,'^ and T lymphocytes,”
affects the adhesion of neutrophils to the endothelium,” and
induces the transendothelial migration of neutrophils.I8 Beside chemotactic activities IL-8 also activates neutrophils.”
The hematopoietic cytokines also regulate the proliferation
and differentiation of hematopoietic progenitor cells. In addition to their proliferative role they contribute in maintaining
cell viability, and functionally stimulate mature macrophages
and granulocytic cells.’’
In the present study we investigated the secretory repertoire of MO/MAC after infection with HIV. Our data show
a differential regulation of proinflammatory cytokines and
hematopoietic growth factors. Although the proinflammatory
cytokines tumor necrosis factor-a (TNF-a), IL-16, IL-6, and
IL-8 are elevated in HIV-infected cultures, MO/MAC downregulate their hematopoietic activity, indicated by a decreased secretion of macrophage colony-stimulating factor
(M-CSF), granulocyte-CSF (G-CSF), and granulocyte macrophage (GM-CSF).
ONOCYTES and macrophages are among the cells to
become infected by human immunodeficiency virus
(HIV) in patients with acquired immune deficiency syndrome (AIDS).’” They not only comprise a reservoir of virus
but also serve as vehicle for viral dissemination throughout
the body.4 The replication of HIV in tissue macrophages has
been associated with clinical manifestations of the disease
including encephalopathy, vacuolar myelopathy, pneumonitis and lymphatic hyperpla~ia.~
The expression of these clinical symptoms is variable among patients as is HIV itself.
HIV-1 and HIV-2 primary isolates differ not only in their
ability to replicate on lymphocytes but also with respect to
their cytopathic effects on MOIMAC.‘ Like on lymphocytes
four distinct replication patterns, defined as a-6 can be observed. Highly replicating HIV-1 strains like HIV- I;, 17111as
well as HIV-2 like HIV-2&, can be found. In contrast to
lymphocytes, macrophages that replicate high amounts of
such strains do not die, but the cultures continue to produce
HIV over many weeks.3 Therefore, in vivo, macrophages are
believed to be a long-term reservoir of HIV.
By secretion of proinflammatory cytokines and hematopoietic growth factors MO/MAC play an important role during an immune response and in hematopoiesis. Pleiotropic
cytokines mediate inflammatory actions and also link the
various components of the immune system. A dysregulated
secretion of these soluble mediators after HIV-infection may
From Georg-Speyer-Haus, Frankfirt am Main; the Department of
Hematology and Oncology, University of Regensburg; Neurological
Institute (Edinger Institute), University of Frankfurt am Main; Buyer
AG, Pharma Research Center, Wuppertal, Germany.
Submitted December 6, 1995; accepted November I , 1996.
The Georg-Speyer-Haus is supported by the Bundesministerium
fur Gesundheit and the Hessische Ministerium fur Wissenschaft und
Kunst. This work was supported by a grant of the Bundesministerium
fur Forschung und Technologie (BGA III-004-89/FVP.l).
Address reprint requests to Reinhard Andreesen, MD, Department
of Hematology and Oncology, Franz-Joosef-Strau Allee 1 I , 93055
Regensburg, Germany
The publication costs of this article were defrayed in part by page
charge puytnent. This article must therefore be hereby marked
“advertisement” in accordance with I8 U.S.C. section 1734 solely to
indicate this fact.
0 1996 by The American Society of Hematology.
0006-4971/96/8809-0032$3.00/0
3474
MATERIALS AND METHODS
Isolation and culture of PBMC. Peripheral blood mononuclear
cells (PBMC) were isolated from healthy donors by density gradient
centrifugation, and cultured in supplemented RPMI 1640 with 5%
heat-inactivated human AB serum cultured in hydrophobic teflon
bags (3 x IO6 PBMC/mL).” The MO-derived MAC were subsequently separated by adherence in 24-well tissue culture plates
(Costar, Cambridge, MA). They were further cultured in WMI 1640
with 5% human AB serum. The adherent cell layer (1 to 2 X lo5
MAClwell) consisted of up to 95% MAC as judged by morphology,
nonspecific esterase staining and expression of CD14 antigen.
Virus isolate and infection of PBMC and MAC. Virus preparations were obtained by propagating the virus in PB T cells and
harvesting the culture at the peak of reverse transcriptase activity.
Blood, Vol 88, No 9 (November 1). 1996: pp 3474-3481
From www.bloodjournal.org by guest on November 24, 2014. For personal use only.
3475
CYTOKINE REGULATION IN HIV-INFECTED MACROPHAGES
The cell suspension (lo6 cells/mL) was stored in aliquots at -70°C
until further use? Only mycoplasma-free virus stocks, tested with a
mycoplasma tissue culture DNA probe assay (Gen-Probe, San Diego,
CA) were used. Mock material was prepared from uninfected T cells
corresponding the protocol for stock virus. The cytokine content of
stock virus and mock material was analyzed: the cytokines IL-ID,
IL-6, IL-8,and the hematopoietic growth factors M-CSF, G-CSF,
GM-CSF could be detected in stock virus as well as in mock material
in similar amounts. TNF-a could not be detected in stock virus nor
in mock material (data not shown). The PBMC and MO-derived
MAC, respectively, were infected with 1 mL stock virus (1 X lo6
PBMC with reverse transcriptase activity of 500,000 to 800,000
cpdmL/90') per 10 mL cell suspension in teflon bags at different
time of culture with the monocytotropic strain HIV-1~~~711,.
This
isolate was derived from a perinatally infected child. It propagates
with a high replication rate on MO/MAC.7 The virus inoculum and
the lymphocytes were removed by washing the adherent cell layer
several times with serum-free medium. The infection of the MO/
MAC cultures were checked by using an HIV antigen enzyme-linked
immunosorbent assay (ELISA) (Organon Teknika, Eppelheim, Germany) and by reverse transcriptase assay.**HIV-antigen and reverse
transcriptase activity was determined in the 24-hour supernatant.
HIV-l-p24 immunostaining. For detection of HIV-1 -p24 antigen the cells were isolated, infected, and cultivated as described
above. Instead of 24-well tissue culture plates the MO-derived MAC
were separated by adherence in plastic chamber slides (Nunc, Wiesbaden, Germany) for later detection of HIV-1 -p24. They were further cultured in RPMI 1640 with 5% human AB serum until they
were fixed for immunocytochemistry at the indicated time points.
After cultivation the cells were washed with phosphate-buffered
saline (PBS), fixed with 4% paraformaldehyde in PBS and stored
in 70% ethanol at -20°C until use. After washing with PBS the
cells were exposed to 10% human serum for 30', followed by incubation with the monoclonal antibody anti HIV-1 -p24 (clon Kal-1,
Laboserv Diagnostica, Giessen, Germany) overnight at 4°C: As negative control (isotype control) IgGl was used (Dianova, Hamburg,
Germany). Afterwards the cells were exposed to peroxidase-conjugated goat antimouse Igs (Dako, Hamburg, Germany) for 30', peroxidase-conjugated rabbit antigoat immunoglobulins (Dako) for 30'
and finally the peroxidase reaction was developed by incubation with
DAB (DAB-kit Vectastain; Camon, Wiesboden, Germany) which
resulted in a brown reaction product. For quantitation positive and
negative cells were counted using a 25X objective. Two hundred
cells were counted two times for each slide and the mean was calculated.
Stimulation of macrophages. At indicated time points medium
was exchanged and MAC were stimulated for 6 and 24 hours with
or without 100 ng/mL LPS (Salmonella abortus q u i , kindly provided by C. Galanos, Max-Planck-Institnt, Freiburg, Germany).
Fig 1. HIV-antigen in HIVlonnll-infectedMO cultures. The
cells were infected with HIVlor17tll
at day 1 after start of CUIture. HIV-antigen concentration
was determined in the 24-hour
supernatant at the indicated
time points. Data presented are
the means ? SEM of a representative experiment done in triplicate. (A) HIV-antigendetermined
with ELISA in the supernatant.
A
Three independent experiments with PBMC from three different
donors were performed, each experiment was done in triplicate.
Cytokine assays. Cell supematant were obtained, filtered
through 0.22 pm membranes (Millipore, Eschbom, Germany), aliquoted and stored at -70°C. The supernatants were analyzed by
ELISAs for IL-ID (R & D Systems, Minneapolis, MN; minimum
detectable dose: 0.3 pg/mL), IL-6 (R & D Systems; minimum detectable dose: 0,35 pg/mL), IL-8 (R & D Systems; minimum detectable
dose: 4,7 pg/mL), G-CSF (R & D Systems, minimum detectable
dose: 7,2 pg/mL), GM-CSF (R & D Systems; minimum detectable
dose: 1,5 pg/mL), TNF-a (Endogen, Boston, MA; minimum detectable dose: 5,O pg/mL) and M-CSF (noncommercial ELISA kindly
provided by E. Alderman, Genetics Institute). The amount cytokine
measured with ELISA was related to the number of adherent MO/
MAC in the culture counted in the well when the supernatant was
harvested. Two hundred cells were counted in three different parts
of the well and the mean was calculated. Variability in cell counting
was insignificant.
RESULTS
Infection of human MO/MAC with HIV-lD117111.In vitro
cultures offer the opportunity to study the interaction of MOI
MAC with HIV. For this investigations we used a culture
system on hydrophobic teflon membranes for blood-borne
MOMAC?' which was adapted for infection of MOIMAC
with HIV in ~ i t r oBoth
. ~ freshly isolated MO as well as MOderived MAC from healthy donors were infected with the
monocytotropic HIV- 1 strain D 117111.' Successful infection
of MOMAC cultures was monitored by assay for viral antigen or reverse transcriptase activity in the supematant. Figure 1 shows a representative time course of infection. Virus
replication reached a maximum after 3 weeks and was constant for several weeks at a high level with approximately
80 ng1mL HIV-antigen produced within a 24-hour period
(24-hour supernatant). A corresponding reverse transcriptase
activity of 5,9 X lo5 cpdmV90' was measured on day 22.
Immunoperoxidase staining methods using anti-HIV- 1-p24
antibodies revealed that early after infection no HIV-1 -p24+
cells were detectable (Fig 1B). After 3 weeks, when a productive infection had been established 70% to 80% of MAC
were HIV-l-p24' (Fig IB). To compensate the decline of
cell counts with time of cultivation, the cytokine concentration was normalized to the cell number determined in the
culture when harvesting the supematant and compared to
mock-infected cells of the same donor.
Secretion of proinjammatory cytokines. When the con-
B
lx)
loo
80
100 -
8o
(KI
60S
40
.'
4-
2o 0
0
2
.-c
20
0
From www.bloodjournal.org by guest on November 24, 2014. For personal use only.
ESSER ET AL
3476
30
[ng/105cells] _
_ _ --___
70
Constitutive secretion
20
-
LPS-stimulated secretion
T
25
[ng/l05 cells]
,
Fig 2. Constitutive and LPSstimulated secretion of proinflammatory cytokines in longterm cultured, virus-replicating
cells at day 22 after start of CUIture. The cells were infected or
mock-infected (uninfected controll with H I V - ~ D ~at~ ~day
I I ~1
after start of culture and stimulated for 24 hours with 100 n g l
mL LPS (S abortus equi) at day
21. Data presented are the
means f SEM of a single representative experiment done in
triplicate out of 3 experiments
performed.
60
i
I
50
I
2o
lo
5
~
0
!
-_11.1-
TNF
I
---..
IL-6
11-6
stitutive and LPS-stimulated secretion of proinflammatory
cytokines was analyzed in long-term cultured, virus-replicating cells at day 22 after start of culture constitutive secretion
could only be detected for 1L-8 (Fig 2). This constitutive
production of IL-8 was increased in the HIV-infected cultures. On LPS-stimulation for 24-hour MAC released high
amounts of TNF-a, IL-6, and further increased the secretion
of IL-8 (Fig 2). Comparing the HIV-infected with the uninfected MO/MAC cultures a marked increase in secretion of
TNF-a, IL-6, and IL-8 could be observed. IL-IO is downregulated during the initial phase of maturation and remains
low in long-term cultured MO/MAC as described.’3 An
upregulated proinflammatory reaction could also be demonstrated early after infection when a productive infection has
not yet been established. Figure 3 shows the LPS-stimulated
secretion of proinflammatory cytokines at day 4. The maximum TNF-a secretion could be observed after 6 hours LPS
stimulation (Fig 3A); the IL-8 amount is similar in the 6
hour and 24 hour supernatant (Fig 3D); IL-6 increased from
6 hours to 24 hours (Fig 3C) and low amounts of IL-lp
were only detectable in the 24-hour supernatant (Fig 3B).
The HIV-infected MO/MAC constantly showed an increased
secretion of all four cytokines. Secretion of proinflammatory
cytokines was analyzed during the course of cultivation and
infection. Figure 4A shows the secretion of TNF-a and HIVantigen production at day 4, 8, and 22 after start of culture
(corresponding to day 3,7, and 21 after infection with HIV).
Early after infection when HIV-antigen production in the
infected cultures could not be detected yet (day 4) or is still
low (day 8) as well as in later stages when a productive
TNFalpha [ng/lOE5 cells1
A
11-8
IL-lbeta [pg/lOE5 cells]
T
5,
_ _
11-8 [ng/lOE5 cells]
11-6 lng/lOE5 cells1
T
Fig 3. LPS-stimulated secretion (24 hours) of proinflammatory cytokines in MO early after
start of culture at day 4. The cells
were infected or mock-infected
(uninfected control) with HIV1D117111 at day 1 and stimulated
with 100 nglmL LPS for 6 hour
and 24 hour. Data presented are
the means f SEM of a single representative experiment done in
triplicate out of 3 experiments
performed.
6 h LPS
24 h LPS
10uninfected MO
6 h LPS
infected MO
I
24 h LPS
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CYTOKINE REGULATION IN HIV-INFECTED MACROPHAGES
3411
G C S F [ng/lOE5 cells]
TNF-alpha [ng/lOE5 cells]
Fig 4. Timecourse analysis of
LPS-stimulated secretion (24
hours) of TNF-CY (A) and G-CSF
(6) in HIV-infected MO.The cells
were infected or mock-infected
(uninfected control) with HIV1ol17111
at day 1 after start of culture. Viral replication in the infected cultures is shown by the
mean of HIV-antigen measured
in the supernatant of three parallel cultures. Data presented are
the means 2 SEM of a single representative experiment done in
triplicate out of 3 experiments
performed.
80
6 1
B
A
0
I
nuninfected MO minfected MO +HIV-antigen
infection has been established (day 22), secretion of TNF-a
was increased in the infected cultures (Fig 4A). Similar results were obtained with IL-6 and IL-8 (data not shown).
When MO-derived MAC were infected at day 6 after start
of culture, again an elevated secretion of TNF-a, IL-6, and
IL-8 in the HIV-infected MAC was observed (Fig 5).
Secretion of hematopoietic growth factors. The stimulated secretion of hematopoietic growth factors was investigated early during MO to MAC maturation as the secretion
of M-CSF is suppressed by LPS beyond day 6 in culture.23
Table 1 shows the secretion of hematopoietic growth factors
by MO on day 2. Only control MO secreted M-CSF constitutively whereas in HIV-infected MO cultures no M-CSF
could be detected without stimulation. When LPS was added,
an increasing response to LPS during the time of stimulation
was observed in the uninfected cultures whereas the HIVinfected MO showed no M-CSF secretion. No constitutive
production of G-CSF and GM-CSF was seen neither in control nor in HIV-infected MO, respectively. On LPS stimulation G-CSF and GM-CSF were secreted by HIV-infected
MO at a significantly lower amount compared to the uninfected control cells. Secretion of hematopoietic growth fac-
.-
days after start of culture
days after start of culture
TNF IIL-6 [ngllO~cells]
1
tors G-CSF and GM-CSF was also analyzed during the
course of cultivation and infection. Figure 4B shows the
secretion of G-CSF and HIV-antigen production at day 4, 8,
and 22 after start of culture (corresponding to day 3, 7,
and 21 after infection with HIV). As the upregulation of
proinflammatory cytokines (shown for TNF-a in Fig 4A)
the downregulation of hematopoietic growth factors (shown
for G-CSF in Fig 4B) after infection with HIV could be
observed early after infection when HIV-antigen production
in the infected cultures could not be detected yet (day 4) or
is still low (day 8) as well as in later stages when a productive
infection has been established (day 22). Similar results were
obtained with GM-CSF (data not shown). The response of
LPS-induced hematopoietic growth factor production was
even more pronounced in mature MAC (Fig 6).
DISCUSSION
We have shown that MO and MAC, which have been
infected with HIV in vitro showed an increased secretion of
proinflammatory cytokines (TNF-a, IL- Ip, IL-6, and IL-8)
in long-term cultured, virus-replicating cells, as well as early
after infection when a productive infection has not yet been
IL-8 [ng/lO%ells]
350
LPS-stimulated secretion
300
250
200
150
100
d-
50
0
TNF
IL-6
IL-8
Fig 5. LPS-stimulatedsecretion (24 hours) of proinflammatory cytokines of HIV-infected MAC at day
13 after start of culture. The cells were infected or
mock-infected[uninfectedcontrol)with HIV-lol171a
at
day 6 after start of culture. Data presented are the
means 2 SEM of a single representative experiment
done in triplicate out of 3 experiments performed.
From www.bloodjournal.org by guest on November 24, 2014. For personal use only.
3478
ESSER ET AL
Table 1. Secretion of Hematopoietic Growth Factors by HIV-Infected MAC*
M-CSF [pg/105Molt
G-CSF [pg/105Molt
Control*
HIV
GM-CSF lpg/105Molt
Control*
HIV
Control*
HIV
With LPS
l h
6h
24 h
Without LPSO
24 h
<20
<20
<IO
79 5 3
1,955 5 181
<20
25 2 1
49 -+ 3
270 t 22
<10
27 2 5
9 2 5 11
110
26 2 3
113 c 7
<IO
13 c 2
41 2 3
444 2 45
<20
<IO
110
110
<10
*After isolation the MO were infected or mock-infected with HIV-1D117111
(day 0 after start of culture), cultured for 2 days and then stimulated
with 100 nglmL LPS ( S abortus equi).
t Data presented are the mean ? SEM of a single representative experiment done in triplicate out of three experiments.
* T h e control MO were mock-infected.
5 For constitutive secretion the medium was changed 24 hours before obtaining the supernatant.
established. Concomitant with this upregulation of proinflammatory cytokines, HIV-infected MO/MAC downregulate their hematopoietic activity, reflected by a decreased
secretion of M-CSF, G-CSF, and GM-CSF.
Proinflammatory cytokines, which may increase host resistence to viral or bacterial infections when released in moderate amounts, are detrimental in the context of AIDS. HIV
replication can be upregulated by proinflammatory cytokines
like TNF-a, IL-lP, and IL-6.24In addition alterations in the
inflammatory gene expression caused by HIV infection could
alter immune responses, cause tissue damage by increasing
oxidant release from leukocytes, inducing expression of
acute-phase proteins, leading to a cachectic state and even
upregulate virus repli~ation.’~
Our results about the secretion of proinflammatory cytokines by HIV-infected MO/MAC add to the controversy that
arises from the conflicting results published. Several reports
have shown induction of proinflammatory cytokines by in
vitro infected MONAC by HIV infe~tion.’~-’~
However, others have presented conflicting data and do not find an altered
mRNA expression or cytokine release in MO infected with
HIV in ~ i t r o . ~ Roy
~ - ~ et
’ al,13 reported a decreased LPSstimulated IL- 1 secretion by HIV-infeced M0.33Several differences in experimental techniques may account for these
discrepancies, eg, culture conditions, different HIV isolates,
the use of cell lines rather than primary MAC, the point of
time for infection as well as for cytokine analysis, etc. In
our system MO and lymphocytes were cultivated together
only for the first days but the virus inocculum and the lymphocytes were removed after the infection. This procedure
has proven to yield very reproducible data comparing cells
of the same donor. We found a variability between primary
cells isolated from different donors concerning viral replication and level of cytokine secretion, but comparison of uninfected and infected cultures from cells of the same blood
donation resulted in reproducible findings concerning the
kind of effect (upregulation of proinflammatory cytokines
and downregulation of hematopoietic growth factors). Furthermore, the infection of primary MO/MAC should yield
responses that are much closer to the natural situation than
studies with cell lines.
Mononuclear phagocytes are key effectors of the inflam-
[pg/lOE5 cells]
3500
3000 -
T
2500
2000
1500
1000
500
n
”
G-CSF
uninfected MAC
OM-CSF
HIV-infected MAC
Fig 6. LPS-stimulated secretion (24 hours) of GCSF and GM-CSF in HIV-infected MAC at day 13 after
start of culture. The cells were infected or mock-infected (uninfected controll with HIV-lo,,7Mat day 6
after start of culture. Data presented are the means
2 SEM of a single repreuntative experiment done
in triplicate out of 3 experiments performed.
From www.bloodjournal.org by guest on November 24, 2014. For personal use only.
CYTOKINE REGULATION IN HIV-INFECTED MACROPHAGES
3479
HIV in vitro the observed cytokine dysregulation could be
matory component of the cellular immune response and,
attributed to HIV-infection.
through cytokine production, interact with all phases of the
Hematologic abnormalities have been documented in all
immune system. During a normal immune response proinstages
of HIV-infection and constitute of anemia, granulocyflammatory cytokines are involved in coordinating activities
of T cells and B cells, as well as in directing cytotoxi~ity.~~topenia, and thrombocytopenia.6' GM-CSF plays a critical
role in pulmonary homeostasis. Pathologic analysis of
Therefore, reduced expression of any of these inflammatory
knockout mice lacking GM-CSF showed a progressive accuproteins could lead to an impaired immune response whereas
mulation of surfactant lipids and proteins in the alveolar
their overexpression may induce fever, cachexia and neurospace,63the characteristic of the idiopathic human disorder
logical symptom^.^^.^^ The fever and wasting syndrome seen
pulmonary alveolar proteinosis also seen in HIV-infected
in AIDS patients may be well related to enhanced production
patients.&2This alveolar proteinosis is secondary to a deof TNF-a and IL-1. In addition, JL- 1 and TNF-a may also
ranged surfactant catabolism or clearance.
be involved in the pathogenic mechanisms of Kaposi sarThe mechanism by which HIV mediates the differential
comae26.37.38
Furthermore the upregulated IL-6 secretion
regulation of cytokine production as described here is not
might explain the chronic B-cell activation and elevated Ig
clear. Some reports showed that productive infection of the
production in HIV-infected patientsz9Gastrointestinal disortarget cells is not necessary to observe an increased cytokine
der like chronic diarrhea and malabsorption syndrome are
production after infection with HIV. Inactivated virus as
often observed in AIDS patients in the absence of any known
well as lymphocytotropic HIV-1 isolate are able to induce
pathogens.39HIV infection of enterochromaffin cells in the
cytokine p r o d ~ c t i o n . ~Blocking
~ ~ ~ ~ "experiments
~
with sCD4
intestinal mucosa might deregulate motility and digestive
could show that the viral binding to the MAC surface is
functions of the intestine.a In addition, the possible involvenecessary and sufficient to account for a signal transduction
ment of MAC-derived cytokines in these disorders in the
pathway resulting in cytokine expression.65Induction of cylamina propria should be considered.26IL-8 as an chemoattokine production by native gpl20 and monoclonal antibody
tractant may contribute to perivascular leucocytic accumulato CD4 as well as their inhibition by neutralizing antibodies
tions observed in various tissues (kidney, heart, gastrointestiagainst gp120 reveal evidence for the important role of bindnal tract, and central nervous system [CNS]) of HIV-infected
ing of HIV-1-gpl20 to the cellular CD4 m o l e c ~ l e . In
~~~'~~~~
patients:' An elevated IL-8 production especially by alveosummary a biphasic impact of the virus on monocyte cytolar macrophages of HIV patients with nonspecific interstitial
kine production appears to occur: initial interaction of the
pneumonitis was reported.42
monocyte-CD4 with HIV-1-gpl20 results in a signal leadConflicting results have also been published on the cytoing to transient monocyte activation and a subsequent upregkine activity in vivo. Elevated serum levels of proinflammaulation during viral replication, possibly mediated by viral
tory cytokines were detected in HIV-infected patients!34 In
Viral proteins other than gp120 also contribute
addition PB cells from HIV-infected patients cultured in
vitro also release high levels of ~ y t o k i n e s ~as~do
, ~tissue
~ - ~ ~ to cytokine induction after HIV infection. Recent reports
showed that, eg, HIV-1 -tat, the viral transactivating protein
macrophage^.^'.^'-^^ However, Jones et al,55reported normal
can not only be released from infected cells and taken up
concentration of cytokines in serum. Also a decreased probut may also induce TNF-a," IL-la,
by uninfected
duction of TNF-a by alveolar MAC" or IL-1 by MOS7from
IL-lp, and IFN-7 in a dose-dependent manner.7' Another
patients with AIDS is reported there.
regulatory protein, HIV-1-nef was shown to induce IL-6
Apart from these conflicting results on the influence of
mRNA in PBL.72
HIV infection on proinflammatory cytokines there are also
Our findings that cytokine production is altered early after
conflicting reports about the regulation of hematopoietic
infection with HIV as well as in long-term cultured, virus
growth factor production during HIV infection. In contrast
replicating cells are consistent with the theory of a biphasic
to our results Molina et alS8described an unchanged GMimpact of the virus on cytokine production described.
CSF production by in vitro HIV-infected MO/MAC. HowTo conclude, our results show the differential regulatory
ever, Molina et a15' analyzed the total cytokine content (celleffects of HIV on proinflammatory cytokines and hematopoiassociated and secreted). Probably the decrease of GM-CSF
etic growth factors. Such effects on cytokines could alter
in the supernatant is based on a dysregulated secretion. Intraoverall immune function as well as virus reactivation and
cellular detection by immunocytochemistry showed that
pathogenesis, including virus-associated immunodeficiency
GM-CSF and G-CSF are produced constitutively but restates.
leased only upon stimulation (Esser et al, 1996, submitted).
The results of Molina et a1 could be due to the unchanged,
ACKNOWLEDGMENT
cell-associated GM-CSF. Conflicting results exist also on
the production of GM-CSF in HIV-infected patients. AlWe gratefully acknowledge Silke Muller and Annegret Rehm for
though some investigatorsS9.@' described low levels of GMexcellent technical assistance, E. Alderman for providing the MCSF production, Hober et a16' found normal or even high
CSF assay, and C. Galanos for LPS.
levels of GM-CSF.6' In contrast to in vitro studies the cause
of cytokine dysregulation in HIV-infected patients could be
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