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*
Department of Medicine, Division of Rheumatology, Center for Rheumatic Diseases in Kangnam St. Mary’s Hospital and
Research Institute of Immunobiology, Catholic Research Institutes of Medical Sciences, Catholic University of Korea, Seoul, Korea; and
Department of Microbiology, Kangwon National University, Chunchon, Korea
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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, and TGF-ß did affect
CD40L-induced VEGF production. Stimulation of FLS with TNF-,
IL-1ß, and TGF-ß increased VEGF production by 1.6-, 2.0-, and
5.2-fold, respectively, and displayed an additive effect on the
production of VEGF by CD40L. VEGF mRNA expression was also up-regulated
by the stimulation of FLS with membranes from the CD40L+ L
cells. Dexamethasone completely abrogated CD40L-induced VEGF
production. In addition, pyrrolidine dithiocarbamate partially
down-regulated CD40L-induced VEGF production, showing that the NF-
B
pathway was partly involved in the signaling of CD40L leading to VEGF
production. Collectively, these results suggest that the interaction
between CD40 on synovial fibroblasts and CD40L expressed on activated T
lymphocytes may be directly involved in the neovascularization in
rheumatoid synovitis by enhancing the production of
VEGF. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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,
TGF-ß, angiogenin, and vascular endothelial growth factor (VEGF) (as
reviewed in Refs. 2, 3). VEGF is a heparin-binding, dimeric glycoprotein that induces endothelial cell proliferation, angiogenesis, and capillary permeability (4, 5). VEGF plays a pivotal role in both normal and pathologic processes such as embryonic development (6), wound healing (7), solid tumor growth, and ascites formation (8). Recently, it has been documented that VEGF may be involved in the pathogenesis of RA. Significantly greater quantities of VEGF are found in the synovial fluid of RA patients than in osteoarthritis or other forms of arthritis (9, 10). VEGF is also highly expressed in the inflamed synovium of RA, where it is produced by synovial fibroblasts and activated macrophages (9, 11). An important stimulus for VEGF release is hypoxia, which up-regulates VEGF protein and mRNA expression in rheumatoid synovial cells (12, 13, 14). In addition, inflammatory mediators which play an important role in the pathogenesis of RA, including PG (15), IL-1, IL-6 (15, 16), and TGF-ß (13, 17), have been described to induce VEGF.
CD40 is a 50-kDa membrane-bound type I glycoprotein described initially
on B lymphocytes, but also expressed on monocytes, thymic epithelium,
dendritic cells, endothelial cells, and fibroblasts (as reviewed in
Refs. 18, 19). CD40 ligand (CD40L), a member of the TNF
superfamily, is a 30- to 33-kDa type II transmembrane protein expressed
on activated T cells, mast cells, basophils, and eosinophils (as
reviewed in Refs. 18, 19). It has been reported that
stimulation with CD40L-expressing cells or purified recombinant CD40L
induces the secretion of proinflammatory cytokines such as IL-1, IL-6,
IL-8, and TNF- from monocytes, dendritic cells, epithelial cells,
and fibroblast, and augments the expression of adhesion molecules and
metalloproteinase (20, 21, 22, 23, 24). However, the effect of CD40L
on the production of VEGF by synovial cells has not been addressed to
date. Before this study, we hypothesized that the interaction of CD40L
on T cells with CD40 on synovial fibroblasts could stimulate
neovascularization at the site of synovitis. To investigate this
hypothesis, we examined whether CD40 ligation could induce VEGF
production from rheumatoid synovial cells. We demonstrate herein that
the ligation of CD40 upon synovial fibroblasts directly enhances VEGF
in both protein and mRNA levels. Moreover, the combined stimulation of
synovial fibroblasts with CD40L and cytokines, including IL-1ß,
TNF-, and TGF-ß, has an additive effect on VEGF production.
Dexamethasone (DEX) abrogates CD40L-induced VEGF production in a
dose-dependent manner and pyrrolidine dithiocarbamate (PDTC) partially
down-regulates CD40L-induced VEGF production, which show that the
NF-B pathway is partly involved in VEGF production by CD40
ligation.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Recombinant TNF- and IL-1ß were purchased from Endogen
(Woburn, MA). Recombinant IL-10 and TGF-ß were purchased from R&D
Systems (Minneapolis, MN). Indomethacin, DEX, and PDTC were obtained
from Sigma (St. Louis, MA). Mouse fibroblastic L cells transfected with
the human CD40L (CD40L+ L cells), as described
previously (25), or synovial fluid (SF) T cells from
patients with RA were used for CD40 activation on cultured synovial
cells. Untransfected (CD40L-) L cells served as
a control. mAb against human CD40 (m3; mouse IgG1) obtained from
Genzyme (Cambridge, MA) was used for the inhibition study. Neutralizing
anti-IL-1ß mAb was purchased from Endogen and anti-TNF-
mAb and anti-TGF-ß mAb was obtained from R&D Systems. All isotype
controls were purchased from Jackson ImmunoResearch (West Grove,
PA).
Isolation and cultures of synoviocytes
Cells were isolated by enzymatic digestion of synovial tissues obtained from RA patients undergoing total joint replacement surgery. Tissues were minced into 2- to 3-mm pieces and treated for 4 h with 4 mg/ml of collagenase (type I; Worthington Biochemical, Freehold, NJ) in DMEM at 37°C in 5% CO2. Dissociated cells were then centrifuged at 500 x g, resuspended in DMEM supplemented with 10% FCS (Life Technologies, Grand Island, NY), 2 mM glutamine, penicillin (100 U/ml), and streptomycin (100 µg/ml), and plated in 75-cm2 flasks. The cultures were kept at 37°C in 5% CO2 and the culture medium was replaced every 3 days. When cells approached confluence, they were passed after diluting 1:3 with fresh medium and recultured until used.
Preparation of L cell membranes
Cell membranes were prepared from L cells as described previously (26). Briefly, cells were washed four times with PBS and suspended at a density of 3 x 107 cells/ml in lysis buffer containing 0.25 M sucrose, 10 mM Tris (pH 7.4), 10 mM NaCl, 0.1 M MgCl2, 1 mM PMSF, and 500 ng/ml polymyxin B. All manipulations were performed at 4°C. Cells were lysed by sonication three times (each burst was 90 W for 8 s) in a Braun sonicator (B. Braum Biotech, Alllentown, PA) with a microprobe tip. Cell lysates were centrifuged at 1000 x g for 15 min, and the resulting supernatants were again centrifuged at 100,000 x g for 30 min. The pellets containing cell membranes were resuspended in RPMI 1640 (Life Technologies) containing 500 ng/ml polymyxin B and stored at -70°C.
VEGF production by CD40L or cytokines
A homogenous population of fibroblast-like synovial cells (FLS)
from passage 4 through 8 were used for each experiment.
CD40L+ or CD40L- L cells
were grown in RPMI 1640 supplemented with 10% FCS and irradiated with
75 Gy before use. FLS were seeded in 24-well plates at 6 x
104 cells/well in 1 ml DMEM/5% FCS and incubated
at 37°C for 24 h, and medium was changed to serum-free DMEM
supplemented with insulin-transferrin-selenium A (Life Technologies).
After another 48-h incubation, the medium was replaced with fresh
DMEM/insulin-transferrin-selenium A, and CD40L+
or CD40L- L cells were added to the wells at
5 x 105 cells/well. In selected wells,
membranes from L cells were added instead of intact cells. As an
inhibition study, anti-human CD40 mAb or unrelated isotype-matched
mouse IgG1 was added to the wells in varying concentrations. Cytokines,
including IL-1ß, TNF-, TGF-ß, and IL-10, were added to the wells
at the onset of culturing. In some experiments, FLS were stimulated
with CD40L+ L cells in the presence or the
absence of 1–50 µg/ml of neutralizing Abs to IL-1ß, TNF-, and
TGF-ß or indomethacin (10-7,
10-6, and 10-5 M) to
determine whether VEGF production was indirectly mediated by IL-1,
TNF-, TGF-ß, and PGE2 produced upon CD40
ligation. Various concentrations of PDTC were added to the wells at
1 h before the stimulation with CD40L, and DEX was added at the
initiation of culture. After 24 h of incubation (unless otherwise
stated), cell-free media were collected and stored at -20°C until
assayed. All cultures were set up in either triplicate or quadruplicate
and the results are expressed as means ± SD.
VEGF production by RA synovial T cells
Synovial fluid mononuclear cells from three patients with RA
were separated by Ficoll-Hypaque (Amersham Pharmacia Biotech, Uppsala,
Sweden) density gradient centrifugation. T cells were separated by
magnetic panning (magnetic cell sorting; Miltenyi Biotec, Sunnyvale,
CA). Purity was 
98% by flow cytometry using anti-CD3 mAb. T
cells were either incubated in a resting state or stimulated with 20
ng/ml of PMA (Sigma) and 5 µg/ml of ionomycin (Sigma) for 6 h.
Some of the stimulated T cells were used to analyze the cell surface
expression of CD40L by flow cytometry using anti-CD40L mAb
(PharMingen, San Diego, CA). Stimulated or unstimulated T cells were
then washed with PBS extensively and fixed in 1% paraformaldehyde for
15 min at room temperature. For VEGF production, FLS were cultured for
24 h with different numbers of fixed T cells in the presence or
absence of anti-CD40 mAb or isotype-matched mouse IgG1.
ELISA of VEGF
VEGF in culture supernatants was measured by sandwich ELISA, as previously described (27), but with minor modification. Ninety-six-well microtiter plates were coated with 100 µl/well of 0.4 µg/ml goat anti-human VEGF165 (R&D Systems) buffered with 50 mM of sodium carbonate (pH 9.6). After incubation overnight at 4°C, the plates were blocked with 1% BSA in PBS for 1 h at room temperature. The human recombinant VEGF165 (R&D Systems) or test samples were added to the wells and then reacted with the plate for 2 h at room temperature. The plates were incubated with 0.2 µg/ml biotinylated goat anti-human VEGF165 (R&D Systems) at room temperature for 2 h. Peroxidase-labeled extravidin (Sigma), diluted 1:1000, was added to react with the plates at room temperature for 1 h. Color reaction was induced by the addition of substrate solution (TMB/H202) and was stopped 30 min later by the addition of 1 M phosphoric acid. An automated microplate reader was used to measure the OD at a wavelength of 450 nm. Between each of these steps, the plates were washed four times with PBS containing 0.1% Tween 20. Human recombinant VEGF165 diluted in culture medium was used as a calibration standard, ranging from 10 to 2000 pg/ml. A standard curve was drawn by plotting OD vs the log of the recombinant VEGF165 concentration.
RNA isolation and Northern blot analysis
Total cellular RNA was isolated from FLS cell culture using
Trizol reagent (Life Technologies). RNA (10 µg) was electrophoresed
through 1% agarose gel containing formaldehyde and the integrity of
RNA was analyzed by ethidium bromide staining. The RNA was then
transferred onto nylon filters (Hybond-N; Amersham Pharmacia Biotech)
and cross-linked by UV light. The 0.6-kb cDNA of human VEGF, consisting
of the entire coding region for the precursor of the 165-residue form
(a generous gift from Dr. Young-Ae Cho, Catholic Research Institute of
Medical Science, Seoul, Korea), was labeled to high specific activity
using [-32P]dCTP and a random primer
labeling kit (Amersham Pharmacia Biotech). After hybridization, the
bands were visualized by autoradiography.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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As shown in Fig. 1
A,
unstimulated FLS constitutively produced VEGF over the 24-h incubation
period (206 ± 25 pg/ml). The levels of VEGF were significantly
increased by the addition of CD40L+ L cells
compared with those on either untreated cultures or cultures with
CD40L- L cells. The VEGF levels from six
separate experiments were 303 ± 54 pg/ml for
CD40L- L cells supernatants and 830 ± 52
pg/ml (4.1-fold over the constitutive levels) for
CD40L+ L cells supernatants.
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B). In addition, VEGF was not detected in
the culture supernatants of L cells without FLS, confirming that the
source of VEGF was FLS, not L cells (Fig. 1A).
The specificity of CD40L in VEGF production was demonstrated by
inhibition studies using anti-CD40 mAb. Treatment of FLS with 5
µg/ml of anti-CD40 mAb for 24 h completely abrogated the
production of VEGF, whereas the equivalent concentration of isotype
control mAb did not (Fig. 1C and data not shown). The
increase of VEGF production by CD40 ligation was evident after only
6 h of culture and persisted up to 96 h (Fig. 2).
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, VEGF production was increased with
increasing numbers of CD40L+ L cells in the FLS
cultures, whereas CD40L- L cells did not
significantly affect the production of VEGF. In all experiments,
CD40L-induced VEGF production from FLS was not attributed to cellular
proliferation, because the number of FLS, determined at 24 h after
incubation, was the same in CD40L+ L cells and
CD40L- L cells cultures (data not shown).
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To investigate the effect of CD40-CD40L interaction on VEGF in a
physiologic condition, SF T cells from three patients with RA, rather
than CD40L+ L cells, were used to induce VEGF
production from FLS. Stimulation of SF T cells with PMA and ionomycin
strongly increased CD40L expression on the cells analyzed by flow
cytometry (
65% after 6 h stimulation; data not shown), which
is consistent with an earlier report (28). When the
stimulated T cells were incubated with FLS for 24 h, they
significantly increased VEGF production from FLS (Fig. 4). The production of VEGF by T cells was
dose dependent, as seen with the CD40L+ L cells
(data not shown). Moreover, anti-CD40 mAb, but not control mAb,
significantly inhibited the ability of SF T cells to produce VEGF.
Together, these data demonstrate that VEGF production by CD40-CD40L
interactions is physiologically relevant and provide further evidence
that CD40L is a major Ag in T cell-mediated VEGF production.
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, TGF-ß, and PG from FLS
CD40L induces the production of IL-1, TNF-, IL-6, and
PGE2 from fibroblasts and endothelial cells
(21, 24, 29, 30), and most of these are able to induce
VEGF. TGF-ß is also known to be a potent inducer of VEGF in synovial
cells and some other cells (13, 17). Thus, a series of
inhibition studies using Abs to IL-1ß, TNF-, and TGF-ß, and
PGE2 synthesis inhibitor was conducted in an
effort to investigate whether VEGF production from FLS was mediated
through endogenous production of these substances. As shown in Table I, the incubation of FLS with these Abs
had no significant effect on the production of VEGF by CD40L, which
demonstrated that the induction of VEGF by CD40L was largely
independent of the production of the respective cytokines. No decrease
of VEGF was also detected in the inhibition study using indomethacin in
the range of 10-7–10-5
M, demonstrating that PGE2 does not play a role
in the production of VEGF by CD40L.
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In the inflamed joints, the resident synoviocytes are exposed to
various proinflammatory and anti-inflammatory cytokines, some of
which are known to promote angiogenesis. An experiment was conducted to
determine the additional effects of cytokines on VEGF production driven
by CD40 ligation. IL-1ß alone (10 ng/ml) increased VEGF production by
about 2.0-fold compared with constitutive VEGF levels with unstimulated
FLS. TNF- (1 ng/ml) also up-regulated VEGF production by 1.6-fold.
TGF-ß (10 ng/ml) remarkably enhanced the production of VEGF by
5.2-fold over constitutive levels. VEGF production induced by IL-1ß,
TNF-, and TGF-ß was further increased when
CD40L+ L cells were coincubated, by factors of
5.9, 5.5, and 9.6, respectively, which indicated that these cytokines
had an additive effect on VEGF production driven by CD40L (Fig. 5). However, IL-10 (0.1–50 ng/ml) did
not affect VEGF production alone or with CD40 stimulation (Fig. 5).
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To determine whether the protein level of VEGF are reflected at
the RNA level, we examined the effect of CD40L on the expression of
VEGF mRNA in FLS using Northern blot analysis. Representative levels of
VEGF mRNA expression in FLS cultured in the presence of membranes of
CD40L+ and CD40L- L cells
are shown in Fig. 6. Unstimulated FLS or
FLS stimulated with membranes of CD40L- L cells
showed a very low constitutive expression of VEGF mRNA
(lanes 1 and 3), whereas stimulation of
FLS with membranes from the CD40L+ L cells
resulted in high levels of the VEGF mRNA (lane
2).
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CD40 ligation results in the activation of transcription factors
NF-B (31, 32), and the inhibitory effect of
glucocorticoids and antioxidant PDTC on NF-B activation are well
documented in other types of cells (33, 34). To verify
whether the NF-B pathway is involved in the FLS production of VEGF
by CD40L, we cocultured FLS and L cells for 24 h with variable
concentrations of DEX and PDTC. As shown in Fig. 7A, DEX inhibited constitutive
and CD40L-induced VEGF production in a dose-dependent manner; the
maximum effect was achieved at a concentration of 2 µM (the highest
dose tested) (Fig. 5A). In addition, the pretreatment of FLS
with 400 µM PDTC 1 h before the addition of
CD40L+ L cells also inhibited VEGF production by
55% (Fig. 5B). The inhibitory effects of DEX or PDTC were
not due to nonspecific toxicity, since the viability of FLS, determined
by MTT assay, was not influenced by DEX (0.1 nM-2 µM) or PDTC
(10–400 µM) (data not shown).
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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production, which provides an additional mechanism capable of
augmenting inflammatory process. T cells expressing CD40L stimulate
synovial fibroblasts and monocytes to generate a series of
proinflammatory cytokines and increase the expression of matrix
metalloproteinase (20, 21, 24). The presence of VEGF in
arthritic synovium strongly suggests its participation in synovitis
(9, 10), probably by promoting angiogenesis, vascular
permeability (4), and microvascular stability in the
synovium (35). Since VEGF appears to be very important for
angiogenesis in vivo, several factors that regulate VEGF production by
synovial cells are of great interest. In this study, we investigated the role of CD40L, displayed by activated T cells, in the production of VEGF by synovial fibroblasts. The production of VEGF was markedly increased by the stimulation of FLS with either CD40L+ L cells or their membrane fractions, which express high levels of CD40L, but was suppressed by anti-CD40 mAb. Moreover, SF T cells of RA patients, stimulated with PMA and ionomycin, also could up-regulate VEGF production. These observations provide strong evidence that CD40L on activated T cells is responsible for the induction of VEGF from FLS and that it induces a novel pathway of RA inflammation. At the site of synovitis, activated T cells are recruited adjacent to the resident synoviocytes by the stimulatory effect of a set of cytokines or chemokines. Consequently, it is possible that through the formation of the CD40L-CD40 bridge, infiltrating T cells induce the proliferation of synovial fibroblasts and up-regulate VEGF, which in turn, could further augment the recruitment of inflammatory cells into the synovium by promoting neovascularization. In this context, CD40L could be responsible for establishing a critical amplification loop, which leads to the persistence of synovitis.
Anti-CD40L mAb treatment is reported to suppress the development of
collagen-induced arthritis, an experimental animal model of RA
(36). Anti-CD40L mAb blocks the development of joint
inflammation, the infiltration of inflammatory cells into synovial
tissue, and the erosion of cartilage and bone. It is also documented
that the prevention of collagen-induced arthritis by anti-CD40L mAb
is possibly mediated by the suppression of circulating Abs to collagen
and by a decrease in the production of inflammatory mediators such as
NO and matrix metalloproteinase by macrophages or synovial cells
(37). In this study, the finding that CD40 ligation
induced VEGF production suggests that anti-CD40L mAb therapy also
may block the interaction between activated T cells and synovial cells
and the subsequent production of VEGF and neovascularization in vivo.
The fact that angiogenesis inhibitors such as integrin
V/ß3 antagonist and
AGM-1470 suppress synovitis in animal models supports this idea
(38, 39).
The production-enhancing effect of CD40 ligation upon inflammatory
cytokines from synovial fibroblasts and monocytes (21, 22, 24, 40) has been well documented. Since IL-1 and TNF- are also
involved in the modulation of VEGF in vivo and in vitro (12, 13, 15, 17, 41), it could be expected that an increase of VEGF by
CD40 ligation would be indirectly enforced by the action of cytokines
(IL-1ß, TNF-) released following CD40 ligation. Given that neither
of the neutralizing Abs to IL-1ß nor TNF- affected the production
of VEGF in the present study, it is unlikely that the up-regulation of
VEGF may be mediated by the indirect effect of these cytokines. It also
seems likely that PGE2 and TGF-ß, potent
inducers of VEGF, are not involved in the production of VEGF by CD40
ligation because indomethacin or anti-TGF-ß Ab did not block the
ability of CD40L to induce VEGF production. Together, these
observations suggest that CD40L induce VEGF, independently of
fibroblast-derived endogenous inducers of VEGF.
Several cytokines are able to modulate the CD40L-dependent activity in
different target cells (40, 42). In the present study,
IL-1ß, TNF-, and TGF-ß increased the secretion of VEGF by
factors of 2.0, 1.6, and 5.2, respectively, compared with the control,
which is consistent with the results of previous studies (12, 13, 15, 41). Notably, the combined effect of
CD40L+ L cells with these cytokines was additive
and not synergistic. When one considers that the sum of stimulatory
effects through independent pathways is usually additive rather than
synergistic, it may be that two kinds of stimuli, CD40L and cytokines,
promote VEGF production via distinct pathways. This possibility is also
supported by our observation that VEGF production by CD40 ligation was
not mediated by IL-1ß, TNF-, and TGF-ß.
CD40L trimer induces clustering of the receptors to initiate signal
transduction. Exactly how signal transduction via CD40 occurs is
unknown, but multiple pathways may be involved (43, 44).
CD40 ligation results in the activation of transcription factors
including NF-B (31, 32), NF-AT (45), and
AP-1 (46, 47). However, the relative importance of these
transcription factors as CD40 effectors is still unclear. In this
study, VEGF mRNA expression was increased by CD40 ligation, indicating
that up-regulated VEGF production by CD40 ligation is attributed to the
transcriptional activation of the VEGF gene. In addition, DEX
completely abrogated up-regulation of VEGF mediated by CD40L. Although
the exact mechanism of glucocorticoid actions remains unclear, it may
be that it blocks the function of NF-B in some way, perhaps by
direct physical association of the glucocorticoid receptor with the
transcription factor (33). Furthermore, DEX also
stimulates the transcription of IB, an inhibitor of NF-B
(34). Our data, along with previous reports, suggest that
CD40 ligation may stimulate NF-B expression in FLS, and NF-B
inhibition by DEX may lead to the abrogation of VEGF production.
However, since glucocorticoid receptors may also interfere with AP-1
and NF-AT (47), we cannot conclude here that a decrease in
the level of VEGF, caused by DEX, can be attributed only to NF-B
inhibition. With this particular possibility in mind, we tested a
single effect of the NF-B blockade on VEGF production using a
dithiocarbamate derivative, PDTC, which inhibits NF-B translocation
in transformed lymphoid cell lines, fibroblasts, and monocytes
(48, 49). The result was that pretreatment with PDTC was
found to abrogate the CD40-mediated up-regulation of VEGF by 55%,
which showed that VEGF production by CD40L was partially mediated by
NF-B activation. However, since the production of VEGF by CD40L was
still evident, notwithstanding the PDTC pretreatment, transcriptional
activation of the VEGF gene by CD40 ligation could not be completely
dependent upon NF-B and is probably also induced by other
transcription factors.
In summary, we observed first that CD40L stimulates VEGF secretion by
synovial fibroblasts. Moreover, IL-1ß, TNF-, and TGF-ß augment
VEGF production by CD40 engagement of synovial cells. Transcription
factor, NF-B, appears to play a some role in the CD40-mediated
activation of VEGF. The present data suggest that the CD40-CD40L
interaction may be one of the major regulating pathways for VEGF
production in rheumatoid FLS. In this context, the strategy to disrupt
the CD40-CD40L conduit could be useful to reduce neovascularization and
inflammation in RA.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Ho-Youn Kim, Department of Medicine, Division of Rheumatology, School of Medicine, Catholic University of Korea, Center for Rheumatic Diseases in Kangnam St. Mary’s Hospital, 505 Banpo-Dong, Seocho-Ku, Seoul, Korea, 137-040.
3 Abbreviations used in this paper: RA, rheumatoid arthritis; VEGF, vascular endothelial growth factor; FLS, fibroblast-like synoviocyte; PDTC, pyrrolidine dithiocarbamate; DEX, dexamethasone; CD40L, CD40 ligand; SF, synovial fluid.
Received for publication September 17, 1999. Accepted for publication March 1, 2000.
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R. M. Andrade, M. Wessendarp, and C. S. Subauste CD154 Activates Macrophage Antimicrobial Activity in the Absence of IFN-{gamma} through a TNF-{alpha}-Dependent Mechanism J. Immunol., December 15, 2003; 171(12): 6750 - 6756. [Abstract] [Full Text] [PDF]
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M. De Bandt, M. H. Ben Mahdi, V. Ollivier, M. Grossin, M. Dupuis, M. Gaudry, P. Bohlen, K. E. Lipson, A. Rice, Y. Wu, et al. Blockade of Vascular Endothelial Growth Factor Receptor I (VEGF-RI), but not VEGF-RII, Suppresses Joint Destruction in the K/BxN Model of Rheumatoid Arthritis J. Immunol., November 1, 2003; 171(9): 4853 - 4859. [Abstract] [Full Text] [PDF]
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A. N. Vallejo, H. Yang, P. A. Klimiuk, C. M. Weyand, and J. J. Goronzy Synoviocyte-Mediated Expansion of Inflammatory T Cells in Rheumatoid Synovitis Is Dependent on CD47-Thrombospondin 1 Interaction J. Immunol., August 15, 2003; 171(4): 1732 - 1740. [Abstract] [Full Text] [PDF]
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M. E. J. Reinders, M. Sho, S. W. Robertson, C. S. Geehan, and D. M. Briscoe Proangiogenic Function of CD40 Ligand-CD40 Interactions J. Immunol., August 1, 2003; 171(3): 1534 - 1541. [Abstract] [Full Text] [PDF]
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