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,§
Departments of
*
Dermatology and
Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia, PA 19104;
The Wistar Institute, Philadelphia, PA 10104; and
§
Department of Microbiology, Albert Szent-Gyorgyi Medical University, Szeged, Hungary
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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.
Depletion of endogenous IL-10, but not of IL-12, abolished the
apoptosis resistance of C. pneumoniae-infected
PBMCs. Furthermore, addition of exogenous IL-10, but not IL-12,
significantly increased the resistance of control inoculum-treated
PBMCs to photoactivated 8-methoxypsoralen- and hypericin-induced
apoptosis. Therefore, we conclude that C. pneumoniae
possesses an antiapoptotic mechanism. The resistance to apoptosis
observed in PBMCs exposed to C. pneumoniae is due, at
least partially, to the IL-10 induced during C.
pneumoniae infection. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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, Fas Ab, and granzyme B/perforin, through blockage of
mitochondrial cytochrome c release and caspase activation.
The role that cytokines play in infection-mediated antiapoptosis and
whether this antiapoptotic effect is unique to C.
trachomatis, however, were not addressed. Furthermore, the
situation of infected cells treated with other clinically relevant
apoptotic inducers such as 8-methoxypsoralen
(8-MOP)3 (11) and hypericin
(12), two light-activated photochemotherapeutic reagents
that have been used in the treatment of lymphoproliferative diseases
and that cause a high level of apoptosis, is also unknown. Chlamydia pneumoniae, the newest member of the chlamydial family, has been established as a common cause of acute and chronic respiratory disease worldwide and implicated in the pathogenesis of coronary artery disease (13, 14). Monocytes/PBMCs have been postulated to serve as a vehicle for systemic dissemination of the infection (15, 16). Like C. trachomatis, C. pneumoniae has a unique intracellular biphasic life cycle. The spore-like elementary bodies facilitate transit between cells. The metabolically active reticulate bodies are responsible for intracellular replication (13, 14). During chlamydial intracellular growth, maintenance of the host cell’s integrity is essential not only for supplying nutrients, but also for shielding the intracellular organisms from host phagocytosis. Therefore, C. pneumoniae may also have evolved mechanisms that allow it to actively interrupt host apoptotic process.
Researchers have shown that C. pneumoniae is a potent
inducer of the proinflammatory cytokines TNF-, IL-1ß, and IL-6 in
human monocytic cells as well as freshly isolated human PBMCs
(17, 18). However, the ability of C.
pneumoniae to induce PBMC secretion of key regulatory factors
in the immune response such as IL-12 and IL-10 has never been
addressed. In this communication, we report that C.
pneumoniae is able to infect human PBMCs and that human PBMCs
respond to infection by secreting the critical immunoregulatory
cytokines IL-12 and IL-10. Furthermore, we demonstrate that PBMCs
treated with C. pneumoniae have increased resistance
to the high level of apoptosis induced by the photoactivated DNA
intercalator, 8-MOP, or hypericin, a naturally occurring photodynamic
compound, as well as to soluble Fas ligand-mediated apoptosis.
Importantly, C. pneumoniae infection-induced IL-10, but
not IL-12, is at least partially responsible for this increased
resistance to apoptosis.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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C. pneumoniae strain TW 183 was purchased from the American Type Culture Collection (ATCC, Rockville, MD) and was propagated in McCoy cells (ATCC) in supplemented DMEM (Life Technologies, Grand Island, NY) in a 35°C, 5% CO2-saturated humidified incubator, as described previously (16). Infected cells were harvested on day 3 or 4 and disrupted by two cycles of freezing/thawing and ultrasonification, and different harvests were pooled. After centrifugation at 1000 rpm for 5 min to remove cell debris, bacteria were concentrated by high speed centrifugation at 25,000 x g for 25 min. Pellets were resuspended in PBS, pH 7.4, mixed with an equal volume of sucrose-phosphate-glutamic acid buffer, aliquoted, and frozen at -70°C until use. A control inoculum was prepared according to the same procedure with uninfected McCoy cells.
Chlamydia titers were determined by immunofluorescence (IF) assay. Briefly, McCoy cells were infected with serial dilutions of bacterial stock, incubated for 72 h, fixed with methanol-acetone (1:1), stained with an antimajor outer membrane protein Ab (IgG3 mouse mAb anti-C. pneumoniae; Dako, Cambridgeshire, U.K.), and followed by FITC-labeled secondary Ab (goat F(ab')2 anti-mouse IgG; Sigma, St. Louis, MO) to identify chlamydial inclusions. After counting inclusions under a fluorescence microscope and correcting for dilution factors, bacterial titers were expressed as inclusion-forming units per ml (ifu/ml). C. pneumoniae infection of human PBMCs/monocytes was also examined by IF assay.
Heat-inactivated (HI) bacteria were prepared by heating a viable bacterial suspension with a known concentration for 45 min at 75°C. Infectivity was abolished, as judged by the undetectable IF titer.
All cultures were free of mycoplasma contamination, as determined by PCR and Hoechst staining (Cell Center, The University of Pennsylvania, Philadelphia, PA).
Human PBMC and monocyte preparations
PBMCs were prepared essentially as previously described (20). First, PBMCs were isolated from blood of healthy human donors through centrifugation on a standard Ficoll gradient (Pharmacia, Piscataway, NJ). The interface containing the mononuclear cell fraction was washed with Dulbecco’s PBS through centrifugation and resuspended in complete medium at the indicated cell concentration. Monocytes were enriched within the fraction of mononuclear cells using their adherence to cell culture dishes (incubation for 2 h at 37°C in RPMI 1640 medium, supplemented with 10% HI FCS). After washing off the nonadherent cells, the monocyte-enriched PBMCs were detached from the plastic surface with cell dissociation solution (Sigma). This procedure yielded monocytes of >90% purity, as confirmed by flow-cytometric analysis with a FACScan (Becton Dickinson, Mountain View, CA). All reagents used in this study were free of endotoxin contamination, as determined by the Limulus amebocyte assay.
Human PBMC infection and cytokine measurement
PBMCs or monocytes were inoculated with various concentrations of C. pneumoniae (viable or HI) and then cultured at 37°C. Supernatants were collected at various times during the culture and stored at -80°C for the cytokine assays. In some experiments, cells were inoculated with the bacteria in the presence or absence of certain cytokine-neutralizing Abs or were treated with certain cytokines before their exposure to the inoculum. In all cases, samples were centrifugated for 1 h at 550 x g immediately after Chlamydia inoculation to facilitate contact between cells and the bacteria.
RIAs for human IL-12 p40, IL-12 p70, TNF-, and IL-10 were performed
as previously described (21, 22, 23, 24), using mAb pairs
C11.79/C8.6, 12H4/C8.6, B154.9/B154.7, and 9D7/12G8, respectively.
RNase protection assay (RPA)
RPA kits were purchased from PharMingen (San Diego, CA). 32P was used for riboprobe labeling. RNAs were extracted with Ultraspect (Biotecx, Houston, TX), and RNA samples were hybridized with an excess amount of [32P]UTP (New England Nuclear-DuPont, Boston, MA)-labeled probes for 12–16 h at 56°C, according to the suggestions of the maufacturer. The protected fragments were fractionated on 5% polyacrylamide/urea-sequencing gels and detected by phosphor imaging.
Cell culture and photoactivation
Purified mononuclear cells were diluted to a concentration of 2 x 106 cells/ml with complete medium and aliquoted into each well of a 24-well plate. Vehicle and predetermined concentrations of hypericin or 8-MOP were added into their respective set of wells in a darkened tissue culture room. Control plates receiving no exposure to photoactivating wavelengths of light were placed immediately in a 37°C incubator. The remaining plates were exposed to either white light from four fluorescent F15T8CW 15 W bulbs under the tissue culture plates or to UVA light with UVA illumination, as described previously (11, 12). Photoactivation with white light lasted for 30 min and delivered the equivalent of 2 J/cm2; UVA light lasted for 4 min and delivered the equivalent of 2 J/cm2. The plates were then incubated at 37°C for 24–72 h, depending on the assay to be performed. The quantity of light delivered to the medium-containing wells of a tissue culture plate was measured with an IL-700 A research radiometer (International Light, Newburyport, MA).
Apoptosis assay
Apoptosis was assessed using a modification of the TUNEL method
previously described by Sgonc et al. (25), as supplied in
kit form by Boehringer Mannheim (Indianapolis, IN). Apoptotic cell
death is characterized by the internucleosomal cleavage of DNA into
fragments of 
180 bp. The TUNEL assay allows for the incorporation of
fluorescein-labeled dUTP (FITC-dUTP) into the free ends of these
apoptotic DNA fragments. Briefly, cells cultured as described above
were fixed in 4% paraformaldehyde for 30 min at 25°C and
permeabilized in 0.1% Triton-containing 0.1% sodium citrate for 2 min
on ice. Terminal deoxynucleotide transferase and nucleotide mixture
were then added to the cells, and the mixture was incubated for 60 min
at 37°C. Then, using a FACScan flow cytometer (Becton Dickinson), the
percentage of cells (of the 10,000 total acquired) undergoing apoptosis
was determined as: [(number of cells incorporating FITC-dUTP into
fragmented DNA)/(total number of cells)] x 100.
Cytokines, Abs, and other reagents
Chinese hamster ovary cell-derived recombinant NK-stimulatory
factor/IL-12 was a gift from Dr. S. Wolf (Genetics Institute, Boston,
MA). Anti-IL-10 mAb 12G8 was a gift from Dr. A. O’Garra (DNAX, Palo
Alto, CA). rTNF- was a gift from Dr. H. M. Shepard (Genentech,
South San Francisco, CA). rIL-4 was purchased from Genzyme (Cambridge,
MA). The mAb, anti-IL-12 (C8.6), was produced and characterized, as
previously described (26). The control Abs, rat normal Ig,
and mouse IgG1 were purchased from Sigma and R&D Systems (Minneapolis,
MN), respectively. Chemically synthesized hypericin was supplied by
VIMRX Pharmaceuticals (Wilmington, DE). A stock solution was prepared
by dissolving 40 mg hypericin in 40 ml 2% benzyl alcohol (vehicle) to
yield a 2 mM solution, then kept in the dark at 4°C. 8-MOP was
purchased from Sigma, and a stock solution of 462.5 µM was prepared
by dissolving 100 µg 8-MOP in 1 ml 2% benzyl alcohol and kept in the
dark at 4°C. Recombinant Fas ligand with Fas ligand enhancer were
purchased from Alexis (San Diego, CA).
Statistical analysis
The data on cytokine production are presented as means ± SD. An unpaired, two-tailed Student’s t test was performed to determine the statistical significance of the data, and p values < 0.05 were considered significant. The data on cell survival are presented as the percentage of cells that survived under various experimental conditions. Percentages are not normally distributed. Therefore, the percentage data were transformed by the arcsin of the square root of the percentage. The transformed data were then analyzed using an analysis of variance. To ensure that the overall probability of detecting a significant difference between two pairs was maintained at a constant type I error of 0.05, comparisons between treatments were made using the Tukey-Kramer multiple comparison test.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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It has been reported that certain C.
trachomatis-infected host cells (HeLa, L929, and U937) are
profoundly resistant to apoptosis induced by proapoptotic stimuli such
as the kinase inhibitor staurosporine, the DNA-damaging agent
etoposide, and immunological molecules including TNF-, Fas Ab, and
granzyme B/perforin (10). To understand whether this
antiapoptotic effect is unique to C. trachomatis and to
explore how Chlamydia-exposed and nonexposed human PBMCs
responded differently following clinically relevant apoptotic inducer
treatment, we tested another species of Chlamydia, C.
pneumoniae, using photoactivated 8-MOP or hypericin
(18). With IF assays, we observed that C.
pneumoniae is able to infect PBMCs, as evidenced by the positive
immunofluorescent signals detected in bacteria-inoculated cells (data
not shown). We also observed that predetermined suboptimal doses of
8-MOP (100 ng/ml and 2 J/cm2 UVA) or hypericin (1 µM and 2
J/cm2 white light) induce 70–80% apoptosis of
PBMCs 60 h following initial photoactivation (Fig. 1
, b and g). But
cells treated with C. pneumoniae (2000–4000 ifu/5 x
106 host cells) before photoexposure had 18% and
40% reduction in apoptosis (Fig. 1, c and h).
Furthermore, there is a dose-dependent relationship between the amount
of C. pneumoniae inoculated and the magnitude of apoptosis
resistance. For instance, with 1 µM of hypericin (2
J/cm2 white light), 2000 ifu of C.
pneumoniae caused a 40% reduction in host cell apoptosis, as
shown in Fig. 1h, while 200 ifu did not cause any
significant changes (data not shown). Cells treated with C.
pneumoniae alone did not significantly affect the rate of
apoptosis (Fig. 1d).
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To understand the mechanism of C. pneumoniae
antiapoptotic activity, we investigated whether PBMCs are able to
respond to C. pneumoniae infection by inducing cytokine
production. Fig. 2 shows that the first
cytokine detected following Chlamydia infection, among
cytokines measured, was TNF-. TNF- peaked at about 8 h
postinoculation (Fig. 2). IL-12 p40 (Fig. 2) and IL-10 (Fig. 2) peaked
between 15–48 and 24–48 h after inoculation, respectively. Optimal
cytokine production was measured at 10,000 ifu of C.
pneumoniae per ml, a maximum dose used in our study. Below 100
ifu/ml, C. pneumoniae was unable to induce any statistically
significant cytokine production (Table I).
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shows that
compared with the viable C. pneumoniae, HI bacteria are not
a strong inducer of the cytokines measured. With 10,000 ifu, viable
C. pneumoniae induced the production of 10.7 ±
1.2 ng of IL-12 p40, 18.8 ± 4.5 ng of IL-10, and 22.9 ± 4.1
ng of TNF-; HI bacteria, however, induced only 0.3 ± 0.1 ng of
IL-12-p40, 1.7 ± 0.5 of IL-10, and 2.8 ± 0.4 ng of
TNF-.
To examine the relationship between different cytokines and to
facilitate our understanding about the functions of those cytokines
produced during the infection, a number of cultures were set up in the
presence or absence of certain cytokines or cytokine-neutralizing Abs.
Fig. 3A demonstrates the
results from a representative RPA experiment in which PBMCs had been
optimally precultured for 16 h in the presence or absence of
IFN-
, then stimulated with C. pneumoniae for 8 h in
the presence or absence of IL-10-neutralizing Ab. In Fig. 3B, lane 1 shows that C. pneumoniae
was able to induce IL12 p40 and IL-10 mRNA expression. If cells had
been pretreated with IFN-, Chlamydia-induced IL-12 p40
mRNA expression was enhanced (Fig. 3B, lane 2).
Chlamydia were also able to induce IFN--pretreated cells
to express IL-12 p35 mRNA (Fig. 3B, lane 2).
Chlamydia-induced IL-10 mRNA expression, however, was
decreased by such a treatment (Fig. 3B, lane 2).
Interestingly, the presence of IL-10-neutralizing Ab in the culture
greatly enhanced IL-12 p40 mRNA expression and induced the expression
of IL-12 p35 mRNA (Fig. 3B, lane 3). Furthermore,
the presence of IL-10-neutralizing Ab enhanced the expression of IL-10
mRNA (Fig. 3B, lane 3).
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enhanced the C.
pneumoniae-induced production of IL-12 p40, p70, and TNF-, but
inhibited the production of IL-10 protein (Fig. 4A). The presence of
IL-10-neutralizing Ab along with Chlamydia also enhanced the
production of IL-12 p40, p70, and TNF- (Fig. 4B).
Addition of exogenous IL-10, on the other hand, caused a significant
reduction in this infection-induced production of IL-12 and TNF-
(Fig. 4B). C. pneumoniae infection mediated PBMC
production of IFN- as well. But under no circumstance was IL-4
production detectable (data not shown).
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To explore the role of cytokines in the antiapoptotic activity of
C. pneumoniae- treated PBMCs, we performed depletion
experiments. As shown in Fig. 5, in
comparison with control Ab treatment, anti-IL-10 treatment before
C. pneumoniae inoculation significantly decreased the
resistance of the cells to photoactivated 8-MOP- or hypericin-induced
apoptosis (Fig. 5, Ae, Ak, and B). The
presence of IL-12-neutralizing Ab, on the other hand, did not
significantly affect the apoptotic activity of these cells (Fig. 5, Ad, Aj, and B). Furthermore,
supplement of exogenous IL-10, but not IL-12, in the absence of
C. pneumoniae, significantly decreased 8-MOP- or
hypericin-induced apoptosis (Fig. 5, Af, Al, and
B). Similar results were obtained in the system of soluble
Fas ligand-mediated cell death (Fig. 6).
Following culture of C. pneumoniae-infected cells with
soluble recombinant Fas ligand, the level of apoptosis was markedly
augmented by addition of anti-IL-10-neutralizing Ab to the culture
system (Fig. 6). Together, these results strongly indicated that
C. pneumoniae infection-mediated secretion of IL-10, but not
IL-12, is at least partially responsible for the antiapoptotic activity
of C. pneumoniae. Similar experiments were also conducted to
investigate the role of IFN- in the antiapoptotic function of
C. pneumoniae-treated cells. We did not observe any
significant effect of IFN- in our system (data not shown).
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The ability of C. pneumoniae to infect human PBMCs and monocytes has been reported previously. In our study, we observed that there are only a small percentage of PBMCs or purified monocytes that by IF assay appear infected. This is consistent with partial resistance to 8-MOP- or hypericin-induced apoptosis. These results may be due to the small amount of inoculum used in our studies. It is also possible, however, that C. pneumoniae infects and protects only specific subpopulations of PBMCs. As to whether those cells that were positive for C. pneumoniae by IF staining are the same cells that have increased resistance to apoptosis induction is an issue currently under investigation.
Photochemotherapy is a unique type of antitumor therapy. It has been used in the treatment of cutaneous T cell lymphoma and Sezary syndrome. The therapeutic efficacy of psoralen in combination with UVA has been linked to the potent induction of apoptosis (11). Hypericin, although still in clinical trial, has also been reported to induce a high rate of apoptotic death of normal, transformed, and malignant T lymphocytes and, thus, has promise for the treatment of cutaneous lymphoproliferative and inflammatory disorders (12). Recently, Abrams et al. (31) described a C. pneumoniae-associated peptide identified in the blood of Sezary syndrome patients that is able to sustain the growth of malignant Sezary T cells and to prevent apoptotic death. This observation is relevant to our in vitro findings. Clearly, understanding the mechanisms of apoptotic resistance induced by C. pneumoniae infection is relevant to enhancing our immunotherapeutic approaches to lymphoproliferative disorders in general and to the use of photochemotherapeutic agents in particular.
Cytokines participate in many physiologic processes, including the
regulation of immune and inflammatory responses. Our previous studies
(19) with murine infection models (BALB/c and 129 mice)
have shown that host immune systems are able to respond to C.
pneumoniae infection by producing a number of cytokines, including
IL-12, TNF-, and IL-10. Although IL-12 appears to play an important
role in early stage host defense against this infection, the function
of IL-10 was not addressed. We observed that depletion of IL-10 by
addition of IL-10-neutralizing Ab to the cell culture significantly
increased the infection-mediated production of IL-12 and TNF-, and
supplementation with exogenous IL-10 at the time of infection decreased
such proinflammatory cytokine production. Together, these results
strongly suggest an anti-inflammatory role of IL-10 produced during
C. pneumoniae infection. Interestingly, the presence of
IL-10-neutralizing Ab in the culture significantly reduced IL-10
protein levels, yet it increased IL-10 mRNA expression, confirming
IL-10 as an autocrine regulatory cytokine, as reported previously by
Masood et al. (32).
To further explore the role of IL-12 and IL-10 produced during C. pneumoniae infection in host cell resistance to apoptosis, we performed blockade experiments with specific mAbs to IL-12 and IL-10. We observed that the presence of anti-IL-10-neutralizing Ab in the infected PBMC culture significantly increased the percentage of apoptotic cells following 8-MOP or hypericin and photoexposure. The presence of anti-IL-12, however, did not change the apoptotic rate. Furthermore, we determined that exogenous IL-10 was able to rescue human PBMCs from photoactivated 8-MOP- or hypericin-induced apoptosis. Exogenous IL-12, however, did not affect the apoptosis-inducing efficacy of those reagents. These findings clearly indicate that the resistance of C. pneumoniae-infected human PBMCs to apoptosis is, at least partially, mediated through the IL-10 produced following infection. The ineffectiveness of HI C. pneumoniae in inducing production of cytokines has been reported previously with an in vivo system (19). In this current study, we observed that HI C. pneumoniae is unable to elicit significant cytokine responses in vitro in comparison with viable organisms. Furthermore, human PBMCs inoculated with HI C. pneumoniae did not show resistance to apoptosis (data not shown). These observations indicated a relationship between cytokines produced during C. pneumoniae infection and the antiapoptotic activity of these bacterium-exposed cells.
The effect of cytokines on cell survival has been studied previously.
It has been shown, in different experimental systems, that signals
transduced through TNFR1 (p55) can induce an activation of proteases,
including caspases. Caspases mediate apoptosis by proteolytic cleavage
of the death substrates (26, 33, 34, 35, 36, 37, 38). TNF- also
increases synthesis of NO in different cells, and this molecule has
been extensively associated with induction of DNA damage and apoptosis
(39, 40, 41, 42). Additionally, TNF- may activate sphingomyelin
breakdown into ceramide, which has a recognized role in apoptosis
(43). IL-10, on the other hand, has been reported to have
variable effects on apoptosis, depending upon the cell type and model
systems used (44, 45, 46). IL-10 has been observed to prevent
lymphocyte activation-induced apoptosis and spontaneous death of
germinal center B cells (47, 48). This effect of IL-10 has
been shown to be associated with increased expression of the
antiapoptotic factor Bcl-2. Furthermore, treatment of cells with
anti-IL-10 diminished cell viability and Bcl-2 expression, while
increasing caspase 1 activation and p53 expression. Balcewicz-Sablinska
et al. (49) reported that IL-10 down-regulates apoptosis
in human alveolar macrophages infected with M. tuberculosis
by inducing the release of TNFR2, leading to the formation of nonactive
TNF--TNFR2 complexes. More recently, Rojas et al. (50)
reported that the TNF-/IL-10 ratio might account for the balance
between apoptosis and survival of virulent M.
tuberculosis-infected murine macrophages. In our system, the time
course for optimal IL-10 production following C. pneumoniae
infection appeared to occur at a point when TNF- production had
fallen to baseline. Thus, induction of IL-10 most likely played a role
in the decline of TNF- levels. As to whether C.
pneumoniae infection-mediated production of IL-10 up-regulates
antiapoptotic factors such as Bcl-2 and Fas-associated death
domain-like IL-1-converting enzyme-inhibitory protein expression and
decreases caspase activation are questions deserving further study.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Alain H. Rook, Department of Dermatology, University of Pennsylvania, 415 Curie Boulevard, Philadelphia, PA 19104.
3 Abbreviations used in this paper: 8-MOP, 8-methoxypsoralen; HI, heat-inactivated; IF, immunofluorescence; ifu, inclusion-forming unit; RPA, RNase protection assay.
Received for publication September 3, 1999. Accepted for publication March 9, 2000.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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and interleukin-10. J. Exp. Med. 183:2559. but not CPP32: multiple interleukin 1-ß converting enzyme-related proteases with distinct substrate recognition properties are active in apoptosis. Proc. Natl. Acad. Sci. USA 93:8395.. J. Immunol. 161:2636. and IL-10 modulate the induction of apoptosis by virulent Mycobacterium tuberculosis in murine macrophages. J. Immunol. 162:6122.This article has been cited by other articles:
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J. Yang, W. C. Hooper, D. J. Phillips, M. L. Tondella, and D. F. Talkington Induction of Proinflammatory Cytokines in Human Lung Epithelial Cells during Chlamydia pneumoniae Infection Infect. Immun., February 1, 2003; 71(2): 614 - 620. [Abstract] [Full Text] [PDF]
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 M. V. Kalayoglu, P. Libby, and G. I. Byrne Chlamydia pneumoniae as an Emerging Risk Factor in Cardiovascular Disease JAMA, December 4, 2002; 288(21): 2724 - 2731. [Abstract] [Full Text] [PDF]
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 L. J. Pinderski, M. P. Fischbein, G. Subbanagounder, M. C. Fishbein, N. Kubo, H. Cheroutre, L. K. Curtiss, J. A. Berliner, and W. A. Boisvert Overexpression of Interleukin-10 by Activated T Lymphocytes Inhibits Atherosclerosis in LDL Receptor-Deficient Mice by Altering Lymphocyte and Macrophage Phenotypes Circ. Res., May 31, 2002; 90(10): 1064 - 1071. [Abstract] [Full Text] [PDF]
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H. Yamaguchi, S. Haranaga, R. Widen, H. Friedman, and Y. Yamamoto Chlamydia pneumoniae Infection Induces Differentiation of Monocytes into Macrophages Infect. Immun., May 1, 2002; 70(5): 2392 - 2398. [Abstract] [Full Text] [PDF]
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J.-L. Perfettini, J. C. Reed, N. Israel, J.-C. Martinou, A. Dautry-Varsat, and D. M. Ojcius Role of Bcl-2 Family Members in Caspase-Independent Apoptosis during Chlamydia Infection Infect. Immun., January 1, 2002; 70(1): 55 - 61. [Abstract] [Full Text] [PDF]
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S. Haranaga, H. Yamaguchi, H. Friedman, S.-i. Izumi, and Y. Yamamoto Chlamydia pneumoniae Infects and Multiplies in Lymphocytes In Vitro Infect. Immun., December 1, 2001; 69(12): 7753 - 7759. [Abstract] [Full Text] [PDF]
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K. Rajalingam, H. Al-Younes, A. Muller, T. F. Meyer, A. J. Szczepek, and T. Rudel Epithelial Cells Infected with Chlamydophila pneumoniae (Chlamydia pneumoniae) Are Resistant to Apoptosis Infect. Immun., December 1, 2001; 69(12): 7880 - 7888. [Abstract] [Full Text] [PDF]
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 G. Trinchieri Regulatory Role of T Cells Producing both Interferon {gamma} and Interleukin 10 in Persistent Infection J. Exp. Med., November 19, 2001; 194(10): F53 - F57. [Full Text] [PDF]
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C. Wahl, F. Oswald, U. Simnacher, S. Weiss, R. Marre, and A. Essig Survival of Chlamydia pneumoniae-Infected Mono Mac 6 Cells Is Dependent on NF-kappa B Binding Activity Infect. Immun., November 1, 2001; 69(11): 7039 - 7045. [Abstract] [Full Text] [PDF]
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S. F. Fischer, C. Schwarz, J. Vier, and G. Hacker Characterization of Antiapoptotic Activities of Chlamydia pneumoniae in Human Cells Infect. Immun., November 1, 2001; 69(11): 7121 - 7129. [Abstract] [Full Text] [PDF]
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 C. Kroegel, J. Rodel, B. Mock, K. W. Garey, and I. Rubinstein Chlamydia pneumoniae, Clarithromycin, and Severe Asthma Chest, September 1, 2001; 120(3): 1035 - 1036. [Full Text] [PDF]
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D. Dean and V. C. Powers Persistent Chlamydia trachomatis Infections Resist Apoptotic Stimuli Infect. Immun., April 1, 2001; 69(4): 2442 - 2447. [Abstract] [Full Text] [PDF]
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