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Departments of
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Pediatrics,
Blood Transfusion,
Parasitology, and
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Obstetrics and Gynecology, Shinshu University School of Medicine, Matsumoto, Japan;
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Nagano Children’s Hospital, Toyoshina, Japan; and
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Department of Laboratory Sciences, School of Health Sciences, Faculty of Medicine, Kanazawa University, Kanazawa, Japan
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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transcripts. Flow cytometric and morphological analyses
demonstrated that the addition of CD70 transfectants to B cell cultures
remarkably promoted differentiation into plasma cells in the presence
of IL-4 and CD40 signaling. Finally, CD27 cross-linking resulted in the
up-regulation of positive regulatory domain I-binding factor-1. Taken
together, our findings indicate that signaling via CD27 on B cells
induces IgE synthesis, in cooperation with IL-4 and CD40 signaling, by
promoting the generation of plasma cells through up-regulation of
positive regulatory domain I-binding factor-1. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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CD27, characterized in detail by van Lier et al. (9, 10), is a type I glycoprotein expressed on T and B cells and in B cell malignancies (11). Recently, we have demonstrated that the interaction between CD27, which belongs to the nerve growth factor receptor/TNF receptor family (12), and CD27 ligand (CD70), which belongs to the TNF family (13) and is expressed not only on activated B but also on T cells, especially activated CD4+ CD45RO T cells (14), can enhance Ig production by B cells (15, 16). In addition, it has been demonstrated that the CD27/CD70 interaction is involved in the differentiation of B cells into plasma cells (17, 18).
In the present study we clarified the role of CD27/CD70 interaction in
B cell IgE synthesis by investigating the effects of CD27 signaling on
B cell proliferation, germline transcripts, differentiation into
plasma cells, and the expression of transcription factors involved in B
cell differentiation such as positive regulatory domain I-binding
factors 1 (PRDI-BF1)2 and B
cell-specific activator protein (BSAP).
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Anti-CD27 mAb (8H5; IgG1), which does not block the ligation of
CD27/CD70 (16, 19), was provided by Dr. T. Morimoto (Dana-Farber Cancer
Institute, Boston, MA). FITC-conjugated anti-CD20 mAb and
PE-conjugated anti-CD20 mAb were purchased from Dako Japan (Tokyo,
Japan). FITC-conjugated anti-CD38 mAb (T16; IgG1), anti-CD40
mAb (MAB89; IgG1), anti-CD70 mAb (HNE51; IgG1), and anti-CD62L
mAb (TQ1; IgG1) were purchased from Immunotech (Marseille, France).
PE-conjugated anti-mouse Ig mAb was obtained from Southern
Biotechnology Associates (Birmingham, AL). Conjugation of biotin to
anti-CD27 mAb (8H5) was performed by the standard technique, using
N-hydroxysuccinimido-biotin (Sigma, St. Louis, MO) in our
laboratory. G418 was purchased from Life Technologies (Grand Island,
NY), and human IL-4 was obtained from Genzyme (Cambridge, MA). The L
cell line expressing murine Fc
II receptor (CD32L) was obtained from
American Type Culture Collection (Manassas, VA). The CD154-CD8 fusion
protein (cell culture supernatant, soluble CD154) was provided by Dr.
A. Aruffo (Bristol-Myers/Squibb Pharmaceutical Research Institute,
Seattle, WA).
Cell preparation
Adult human PBMNCs were isolated from healthy volunteers by Ficoll-Hypaque (Pharmacia, Piscataway, NJ) density gradient centrifugation and were separated with 5% sheep erythrocytes into erythrocyte rosette-positive and -negative (E-) populations (20). After depleting monocytes with silica (IBL, Fujioka, Japan) or by adherence to the plastic surface, E- cells were further purified into B cells by positive selection with anti-CD19 mAb-coated immunomagnetic beads (Dynal, Oslo, Norway), and then anti-CD19 mAb was removed by the use of Detach a Bead (Dynal). Ninety-seven percent of the B cell population was reactive with anti-CD20 mAbs. Proliferation and activation were not observed from the B cells thus obtained. CD20+ CD27- or CD20+ CD27+ B cells were isolated from the monocyte-depleted E- cells by sorting with a FACStarPlus (Becton Dickinson, Mountain View, CA) under sterile conditions. Both populations obtained were >98% pure.
Preparation of transfectants
The preparation of CD70 transfectants or mock transfectants has
been described previously (16). Briefly, total RNA was isolated by the
single-step method (21) from the CD70+ human B cell line
(Daudi cells). After RT-PCR, the amplified cDNA was digested with
EcoRI and XbaI and ligated with the mammalian
expression vector pcDLSR
296 (22). The resulting plasmid or the
vector alone was transfected into the murine pre-B cell line, 300-19
(23) by electroporation. The transfectants were selected by growth in a
culture medium containing a neomycin analogue (G418, Life Technologies)
at 1 mg/ml.
Flow cytometric analysis
Activated purified B cells were stained with anti-CD38-FITC and anti-CD20-PE. Two-color analysis of B cell surface molecules was performed by FACScan (Becton Dickinson). Ab-coated cells were gated on living cells by cell size and granularity, and dead cells were removed by propidium iodide staining and then enumerated by flow cytometric analysis.
Fixation of transfectant cells
The CD70/300-19 or mock/300-19 transfectants were incubated with 1% paraformaldehyde in PBS for 5 min. After washing with PBS three times, the cells were cultured in RPMI 1640 and 10% FCS for 30 min and then used for the analysis.
Ig assay by ELISA
Highly purified peripheral blood B cells were cultured with 50 ng/ml of IL-4 and 1 µg/ml of anti-CD40 in the presence of various concentrations of fixed CD70/300-19 or mock/300-19 with or without 1 µg/ml of anti-CD70 mAb for 12–14 days at 37°C in a humidified atmosphere with 5% CO2. The final cell density was 5–10 x 105/ml in a volume of 200 µl/well. The cultured supernatants were harvested, and the supernatants and the standard human IgE (Chemicon, Temecula, CA) were added to 96-well flat ELISA plates (Nunc, Roskilde, Denmark). The plates were coated with human IgE mAbs (CIA-E-7.12 and CIA-E-4.15, provided by Dr. A. Saxon, Division of Clinical Immunology/Allergy, University of California School of Medicine, Los Angeles, CA). After an overnight culture at 4°C, the supernatants were discarded, and the wells were washed with 0.05% Tween 20 in PBS. Alkaline phosphatase-labeled goat anti-human IgE (Sigma, St. Louis, MO) at a dilution of 1/5000 was added. After 2 h of incubation at room temperature, color detection was performed by 3-[cyclohexylamino]-1-propanesulfonic acid (CAPS) buffer containing p-Nitrophenyl phosphate (pNPP) (Sigma). Calibration was performed with PBS at standard zero levels. The limit of sensitivity for IgE was 5 ng/ml.
B cell proliferation assay
Highly purified unseparated B cells and CD20+ CD27+ and CD20+ CD27- B cells were cultured with fixed mock/300-19 or CD70/300-19 (20% of the B cells added) in the presence of 50 ng/ml IL-4 or IL-4 plus 1 µg/ml anti-CD40 at a final cell density of 2.5–5 x 105/ml in a volume of 200 µl/well in 96-well round-bottom plates (Nunc) for 3 days at 37°C in a humidified atmosphere with 5% CO2. Then the cultures were pulsed with 0.5 µCi of [3H]thymidine. After 18 h of incubation, the cells were harvested with an automatic cell harvester (Packard, Meriden, CT), and [3H]thymidine incorporation was measured on a liquid scintillation analyzer (Packard).
RT-PCR of germline , BSAP, and PRDI-BF1
Highly purified unseparated B cells and CD20+
CD27+ or CD20+ CD27- B cells
(1 x 106 cells) were stimulated with 50 ng/ml IL-4 or
IL-4 plus 1 µg/ml anti-CD40 in the presence of mock transfectants
or CD70 transfectants for 16 h or 4 days. Total RNA was extracted
by the acid-guanidine thiocyanate-phenol-chloroform method using a
RNAzol rapid RNA purification kit (Biotex, Houston, TX). First-strand
cDNA copies were synthesized by using murine Moloney leukemia virus
reverse transcriptase (Life Technologies, Grand Island, NY) with
oligo(dT) (Life Technologies) as a primer in a total volume of 20 µl,
and then PCR was performed. The following oligonucleotide primers were
used for PCR: for C exon, sense primer, 5'-CATGCGGTCCACGACCAAGAC-3';
antisense primer, 5'-CCACTGCACAGCTGGATGGAG-3'; for BSAP,
5'-CAACCAACCAGTCCCAGCTTC-3' and 5'-TCACAATGGGGTAGGACTGCG-3'; and
for PRDI-BF1, 5'-AGCTGACAATGATGAATCTCA-3' and
5'-CTTGGGGTAGTGAGCGTTGTA-3'. Two microliters of cDNA were amplified
in PCR using each primer and Taq DNA polymerase (Life Technologies).
The amplified products were analyzed on a 1.2% agarose gel containing
ethidium bromide and visualized by UV light illumination.
ß2m sense primer 5'-GCTATGTGTCTGGGTTTCAT-3' and antisense
primer 5'-ATCTTCAAACCTCCATGATG-3' were used as controls (24).
Quantitative RT-PCR of germline C
In the germline C transcripts, the amounts of mRNA were
estimated by coamplification of parvo virus DNA for a competitor. PCR
amplification of parvo virus DNA, which includes germline C primers
in both sides, was conducted with flanking primers. The product was 505
bp in length and was separated by PAGE and purified. Each template
contained the same amounts of cDNA from RNA extracted from highly
purified B cells, and one of fivefold dilutions of germline C
transcript competitor from 3.2 x 10-1 to 2.6 x
10-3 attomoles. The equivalent point was determined by
PAGE. To ensure that the amounts of germline C cDNA in the samples
were the same, the amounts of the ß-actin cDNA were evaluated as an
internal mRNA control.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Since triggering via CD27 by CD70 transfectants, which greatly
express CD70 on the surface (Fig. 1
A), induces IgA, IgM, and IgG
synthesis in B cell activation systems with SAC plus IL-2 (15)
and IL-10 (25), we investigated whether CD27/CD70 interaction
participates in B cell IgE synthesis. To clarify the direct effect of
CD27 signaling on IgE synthesis, we highly purified B cells from adult
peripheral blood. As reported previously (26), anti-CD40 or IL-4
alone did not induce IgE synthesis by B cells, whereas in the presence
of anti-CD40 plus IL-4, B cells produced substantial amounts of
IgE. Most significantly, CD70 transfectants greatly augmented IgE
production in the presence of IL-4 plus anti-CD40, which was about
fourfold compared with that after the addition of mock transfectants
(Fig. 1B). The augmentation of IgE synthesis by CD70
transfectants was dose dependent, and the enhancement of IgE production
by CD70 transfectants was completely blocked by the initial addition of
anti-CD70 mAb (Fig. 1B), but not by the control mAb
(data not shown). On the other hand, the addition of IL-4 or
anti-CD40 to CD70 transfectants did not induce detectable levels of
IgE secretion (Fig. 1B). When we stimulated B cells by
cross-linking with anti-CD40 plus CD32 transfectants and by
the addition of soluble CD154 in the presence of IL-4, IgE production
was 32 ± 7 and 21 ± 5 ng/ml, respectively. These data
demonstrate that triggering via CD27 by its ligand, CD70,
synergistically induces B cell IgE synthesis in the presence of IL-4
plus anti-CD40, indicating that the CD27/CD70 interaction is
involved in B cell IgE synthesis by direct cell to cell contact.
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Having demonstrated that peripheral blood B cells can be separated
by CD27 expression, and that CD27+ B cells, but not
CD27- B cells, produce large amounts of IgA, IgM, and IgG
(27), we conducted additional experiments to determine whether B cell
subpopulations separated by CD27 have different functions with regard
to the IgE synthesis. We purified CD20+ CD27+ B
cells and CD20+CD27- B cells by flow
cytometry; the purities of the two cell populations were >98%.
CD27+ B cells produced IgE in the presence of IL-4 plus
anti-CD40, but not in the presence of IL-4 or anti-CD40 alone.
The production of IgE by CD27+ B cells was strongly
enhanced by the addition of CD70 transfectants in the presence of IL-4
plus anti-CD40, whereas mock transfectants did not have such an
enhancing effect. The enhancement by the CD70 transfectants was
specifically blocked by the initial addition of anti-CD70 mAb (Fig. 2). On the other hand, IgE secretion from
CD27- B cells was below measurable levels in the presence
of IL-4 plus anti-CD40, even with the addition of CD70
transfectants (Fig. 2). These data demonstrate that CD27+ B
cells, but not CD27- B cells, have ability to produce IgE
and that signaling via the CD27 molecule participates in the IgE
synthesis.
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We next investigated whether CD27 molecule on B cells contributed
to their proliferation in the presence of IL-4 or IL-4 plus
anti-CD40. IL-4 alone induced low levels of proliferation of
unseparated B cells. The B cell proliferation was enhanced by the
addition of anti-CD40 to IL-4 compared with that with IL-4 alone.
Furthermore, the proliferation of B cells was augmented by the addition
of CD70 transfectants in the presence of IL-4 or IL-4 plus
anti-CD40, whereas mock transfectants did not have such an
enhancing effect. In addition, the augmentation of B cell proliferation
by CD70 transfectants was completely blocked by the initial addition of
anti-CD70 mAb (Table I). B cell
proliferation was greatly increased by IL-4 plus anti-CD40 plus
CD32 transfectants (18,952 ± 2,802 cpm; n = 3) or
IL-4 plus soluble CD154 (15,916 ± 2,156 cpm; n =
3) compared with IL-4 plus anti-CD40 stimulation (2,409 ±
1,067 cpm; n = 7). As the baseline levels of B cell
proliferation were high in these systems with IL-4 plus anti-CD40
plus CD32 transfectants or IL-4 plus soluble CD154 only, the enhancing
effects of CD70 transfectants on B cell proliferation were not
prominent: the values with CD70 transfectants were 21,304 ± 4,012
cpm (n = 3) in the former and 19,185 ± 3,231 cpm
(n = 3) in the latter system. This enhancement of B
cell proliferation by CD70 transfectants was obtained in
CD27+ B cells, but was very mild in CD27- B
cells (Table II). Thus, these results
suggest that the ligation of CD27 on B cells by CD70 transfectants
enhances B cell proliferation in the presence of IL-4 or IL-4 plus
anti-CD40.
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transcripts
Since B cell IgE production was strongly enhanced by the addition
of CD70 transfectants in the presence of IL-4 plus anti-CD40, we
investigated the expression of germline transcripts, which is an
early molecular markers of the isotype switch process, to examine
possible early switch events being induced by CD70 transfectants.
Germline transcripts from highly purified B cells were not obtained
by the addition of medium, CD70 transfectants, or anti-CD40 alone
(data not shown). Representative data by quantitative RT-PCR showed
that germline transcripts were induced by IL-4. In our experiments,
the transcript expression by B cells was not enhanced by the addition
of anti-CD40 to IL-4 (Fig. 3). We
were unable to find differences in the expression of germline by B
cells in CD70 transfectant-treated vs untreated B cells in the presence
of IL-4 or IL-4 plus anti-CD40 (Fig. 3). The addition of CD70
transfectants also had no enhancing effect on transcription in the
presence of IL-4 plus anti-CD40 plus CD32 transfectants
(data not shown). This effect did not change with various culture times
from 16–48 h (data not shown). Although CD27- B cells did
not produce IgE (Fig. 2
), their germline transcripts were obtained
in the presence of IL-4 plus anti-CD40 (Fig. 4). Thus, it appears that the increased
IgE secretion from peripheral blood B cells by CD27/CD70 interaction
does not occur in the process of germline transcripts.
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Recently, we demonstrated that triggering via CD27 by CD70 on
purified peripheral blood B cells yielded an increase in the number of
plasma cells in the presence of IL-10 (17). The finding prompted us to
ascertain whether CD27/CD70 interaction induced differentiation of B
cells into plasma cells secreting IgE in the presence of IL-4 plus
anti-CD40. Accordingly, we studied the effect of CD27 signaling on
peripheral blood B cell differentiation stimulated with IL-4 plus
anti-CD40 by flow cytometric and morphological analyses. Human
peripheral blood B cells expressed high levels of CD20 and CD40 and a
low level of CD38, and plasma cells expressed CD38 strongly. Most
interestingly, B cells stimulated with CD70 transfectants in the
presence of IL-4 plus anti-CD40 for 12–14 days differentiated into
plasma cells. In contrast, IL-4 plus anti-CD40 without CD27
signaling induced a slight increase in the number of plasma cells. The
increase in the differentiation from B cells to plasma cells
disappeared by the blockage between CD27 and CD70 interaction with
anti-CD70 mAb (Fig. 5A).
Morphological analysis showed that CD20-
CD38high cells, which were increased by CD27/CD70
interaction in the presence of IL-4 plus anti-CD40, were typical
plasma cells with a basophilic cytoplasm with a pale Golgi zone and an
eccentric nucleus. In contrast, CD20+ CD38low
cells revealed the morphology of B cells (Fig. 5B). These
results clearly demonstrated that, in agreement with the IgE production
measurement, CD27/CD70 interaction promotes plasma cell differentiation
in the presence of IL-4 plus anti-CD40.
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As a further assessment of the role of CD27 signaling on B cell
differentiation into plasma cells, we finally investigated the effect
of CD27/CD70 interaction on the mRNA expression of PRDI-BF1/Blimp-1,
which has been shown to be involved in multiple steps of B cell
differentiation leading to plasma cell development (28) and the
transcription factor BSAP. As shown in Fig. 6, PRDI-BF1 mRNA expression was
up-regulated by the addition of CD70 transfectants compared with that
with mock transfectants in the presence of IL-4 plus anti-CD40
after both 16 h and 4 days of culture. Up-regulation of BSAP mRNA
expression was observed, although it was not significant, in CD70
transfectant-treated B cells in the presence of IL-4 plus
anti-CD40. The enhancement of PRDI-BF1 and BSAP mRNA levels by CD70
transfectants was completely blocked by the initial addition of
anti-CD70 blocking mAb (data not shown). The murine pre-B cell
lines, mock/300-19 and CD70/300-19, did not express mRNA for human
PRDI-BF1, BSAP, or ß2m (Fig. 6, lanes 1 and
2). These studies indicate that while CD27/CD70 interaction
does not reduce the expression of BSAP, it promotes the generation of
plasma cells at least in part by up-regulating PRDI-BF1 levels.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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It is known that IL-4 has a potent function in the production of IgE by
B cells (34, 35). However, IL-4 alone induces B cell germline
transcripts but not IgE production. By the addition of CD40 signaling
to IL-4, the mature transcripts can be induced, and IgE synthesis
finally occurs (4, 3, 26). In our experiments, CD70 transfectants
significantly augmented IgE production by highly purified B cells
cultured with IL-4 plus anti-CD40. Since the blockage of CD27/CD70
interaction by mAb completely blocks the augmentation of B cell IgE
production by CD70 transfectants (Fig. 1), CD27 signaling evidently
plays an important role in IgE synthesis. When we used highly purified
CD27+ B cells, their IgE production was increased more than
that of unseparated B cells in the presence of IL-4 plus anti-CD40,
whereas CD27- B cells did not entirely produce IgE in the
presence of IL-4 plus anti-CD40 (Fig. 2). These findings support
the view that CD27+ B cells are primed memory B
cells, and CD27- B cells are unprimed naive B cells as we
previously demonstrated in the synthesis of IgA, IgM, and IgG (27).
Several reports indicated that CD70 transfectants induced B cell
proliferation in the presence of SAC plus IL-2 (15, 16, 18). In our
experiment, CD70 transfectants augmented B cell proliferation to about
3 times that found in the presence of IL-4 (Table I). In disagreement
with our data, Jumper et al. reported that stimulation via CD27 had no
effect on B cells in the presence of IL-2 or IL-4, suggesting that the
role of CD70 in regulating B cell growth and differentiation is
questionable (36). The reasons for the discrepancy in B cell
proliferation via CD27 between the reported data (36) and our present
results are not clear, but several possibilities could be postulated.
One possibility is that the degree of the CD70 expression of the
transfectants may be different; CD70 expression of our transfectants is
prominent compared with that of the reported cells (36). Alternatively,
the concentrations of cytokines used in the studies are probably
different between the two studies.
The levels of B cell proliferation in our system using IL-4 plus anti-CD40 were very low compared with the proliferation effects previously reported (36). As cross-linking of anti-CD40 by CD32 transfectants or soluble CD154 in the presence of IL-4 greatly promoted B cell proliferation, the effects of our stimulation systems with IL-4 plus anti-CD40 alone appear to be suboptimal. Enhancement of B cell proliferation by CD70 transfectants were remarkable in the presence of IL-4 plus anti-CD40, but were not prominent in the presence of IL-4 plus anti-CD40 plus CD32 transfectants or IL-4 plus soluble CD154. Thus, our present suboptimal activation system is suitable for observing the enhancing effects of CD27/CD70 interaction on B cell proliferation. On the other hand, IgE synthesis was not as significantly increased in the presence of IL-4 plus anti-CD40 plus CD32 transfectants or IL-4 plus soluble CD154 compared with that in the presence of IL-4 plus anti-CD40. The enhancement of IgE synthesis via CD27 signaling was most significant in the presence of IL-4 plus anti-CD40.
B cell IgE synthesis is preceded by transcription of the germline
gene, and the relationship between this expression and subsequent
switching to C has been reported (37, 38). CD20+
CD27+ and CD20+ CD27- B cells from
healthy adults did not differentiate in their ability to produce
transcripts by exogenous IL-4 (Fig. 4). Therefore, IL-4-induced
transcripts occur similarly in CD20+ CD27-
naive B cell and CD20+ CD27+ B cell
populations. Jumper et al. (36) demonstrated that the addition of CD70
transfectants into EBV-transformed B cells did not augment the germline
expression. Also, triggering via CD27 by CD70 in the present study
exerted no enhancing effect on the expression of the germline in
the presence of IL-4 or IL-4 plus anti-CD40 (Fig. 3). These
findings suggested that CD27/CD70 interaction does not participate in
the IL-4-induced germline expression. However, one possibility
remains that CD27 signaling may induce IgE class switching following
the increase in germline transcripts with IL-4 or IL-4 plus CD40
signaling. To clarify this point, we performed quantitative RT-PCR
using mature primers. Unfortunately, as we could not detect mature
bands using several primer pairs in the presence of IL-4 plus
anti-CD40 plus CD32 transfectants, we failed to estimate the
effects of CD27/CD70 interaction in the IgE class switching.
As reported previously, the CD27 signal resulted in the terminal
differentiation of peripheral blood memory B cells into plasma cells in
B cell activation systems by IL-10 or IL-10 plus IL-2 (17) and SAC or
SAC plus IL-2 (18). In our experiments, B cells also differentiated
into plasma cells by triggering via CD27 in the presence of IL-4 plus
anti-CD40 (Fig. 5), but not in the presence of IL-4 alone (data not
shown). Since CD154 is expressed on T cells soon after the activation,
and CD70 is expressed on T cells later after the activation (17), CD40
triggering probably participates in the early steps of the IgE switch
recombination and then CD27/CD70 interaction may promote the
differentiation of class-switched B cells into IgE-producing plasma
cells (35, 39, 40). CD40/CD154 interaction also induces the expansion
of memory B cells expressing CD27 on the surface. CD27 may be important
in the induction of plasma cells when the expansion of CD27-positive B
cells has already occurred (17, 18). Thus, as we demonstrated here and
previously, CD27 is crucial in controlling the differentiation of all
isotype-committed memory B cells into plasma cells.
PRDI-BF1/Blimp-1, a recently described zinc finger transcription factor, is involved in various steps of B cell terminal differentiation (28), indicating that its expression participates in the generation of plasma cells. Since CD27 cross-linking on B cells enhanced the mRNA expression of PRDI-BF1/Blimp-1, even after the overnight activation, the induction of plasma cells, which was demonstrated by FACS, is probably due to the effect of the expression of PRDI-BF1/Blimp-1 induced by CD27/CD70 interaction. We also investigated the transcription factor BSAP/Pax-5, which is involved in the regulation of B cell proliferation, because it is expressed in all B cell developmental stages except for the plasma cell stage (41). BSAP mRNA expression was unexpectedly up-regulated by triggering via CD27 on B cells. A large majority of the cultured cells in our system was composed of B cells, and plasma cells constituted a small population, even after 8 days of culture. Thus, the remaining B cells may express BSAP. At least, CD27/CD70 interaction on B cells did not directly reduce BSAP expression.
In summary, CD70 transfectants significantly augmented IgE production
from highly purified B cells, especially CD27+ B cells
cultured with IL-4 plus anti-CD40, by enhancing B cell
proliferation and differentiation into plasma cells without any
increase in the expression of germline . Our findings presented in
this paper should be tested in the murine system including
CD27-/- mutant mice (42), although the effects of
CD27/CD70 interaction on B cell Ig synthesis in the murine and human
systems are somewhat different (J. Borst, unpublished
observations). Experiments to examine these aspects are
underway.
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Acknowledgments |
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Footnotes |
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2 Abbreviations used in this paper: PRDI-BF1, positive regulatory domain I-binding factor-1; BSAP, B cell-specific activator protein; PE, phycoerythrin; E-, erythrocyte rosette-negative; SAC, Staphylococcus aureus Cowan strain; Blimp-1, B lymphocyte-induced maturation protein-1.
Received for publication April 3, 1998. Accepted for publication August 26, 1998.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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transcripts in human B cells induced by interleukin 4 to switch to IgE production. J. Exp. Med. 172:463. promoter: an effect and versatile mammalian cDNA expressed system composed of the simian virus 40 early promoter and the R-U5 segment of human T-cell leukemia virus type 1 long terminal repeat. Mol. Cell. Biol. 8:466. transcripts in human B cells stimulated with rIL-4 and EBV. J. Immunol. 145:3468.[Abstract]
-independent and IL-4-dependent differentiation signal directly to human B cells for IgE production. J. Immunol. 146:1836.[Abstract]
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 E. Breitbart, X. Wang, L. S. Leka, G. E. Dallal, S. N. Meydani, and B. D. Stollar Altered Memory B-Cell Homeostasis in Human Aging J. Gerontol. A Biol. Sci. Med. Sci., August 1, 2002; 57(8): B304 - 311. [Abstract] [Full Text] [PDF]
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 K. Tarte, J. De Vos, T. Thykjaer, F. Zhan, G. Fiol, V. Costes, T. Reme, E. Legouffe, J.-F. Rossi, J. Shaughnessy Jr, et al. Generation of polyclonal plasmablasts from peripheral blood B cells: a normal counterpart of malignant plasmablasts Blood, July 30, 2002; 100(4): 1113 - 1122. [Abstract] [Full Text] [PDF]
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 J. Wischhusen, G. Jung, I. Radovanovic, C. Beier, J. P. Steinbach, A. Rimner, H. Huang, J. B. Schulz, H. Ohgaki, A. Aguzzi, et al. Identification of CD70-mediated Apoptosis of Immune Effector Cells as a Novel Immune Escape Pathway of Human Glioblastoma Cancer Res., May 1, 2002; 62(9): 2592 - 2599. [Abstract] [Full Text] [PDF]
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H. Nagumo, K. Agematsu, N. Kobayashi, K. Shinozaki, S. Hokibara, H. Nagase, M. Takamoto, K. Yasui, K. Sugane, and A. Komiyama The different process of class switching and somatic hypermutation; a novel analysis by CD27- naive B cells Blood, January 15, 2002; 99(2): 567 - 575. [Abstract] [Full Text] [PDF]
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V. S. Raman, V. Bal, S. Rath, and A. George Ligation of CD27 on Murine B Cells Responding to T-Dependent and T-Independent Stimuli Inhibits the Generation of Plasma Cells J. Immunol., December 15, 2000; 165(12): 6809 - 6815. [Abstract] [Full Text] [PDF]
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C. Angelin-Duclos, G. Cattoretti, K.-I Lin, and K. Calame Commitment of B Lymphocytes to a Plasma Cell Fate Is Associated with Blimp-1 Expression In Vivo J. Immunol., November 15, 2000; 165(10): 5462 - 5471. [Abstract] [Full Text] [PDF]
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M. Odendahl, A. Jacobi, A. Hansen, E. Feist, F. Hiepe, G. R. Burmester, P. E. Lipsky, A. Radbruch, and T. Dorner Disturbed Peripheral B Lymphocyte Homeostasis in Systemic Lupus Erythematosus J. Immunol., November 15, 2000; 165(10): 5970 - 5979. [Abstract] [Full Text] [PDF]
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S. Morimoto, Y. Kanno, Y. Tanaka, Y. Tokano, H. Hashimoto, S. Jacquot, C. Morimoto, S. F. Schlossman, H. Yagita, K. Okumura, et al. CD134L Engagement Enhances Human B Cell Ig Production: CD154/CD40, CD70/CD27, and CD134/CD134L Interactions Coordinately Regulate T Cell-Dependent B Cell Responses J. Immunol., April 15, 2000; 164(8): 4097 - 4104. [Abstract] [Full Text] [PDF]
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