|
|||||||||
|
|
|
|
|
|
|
|
|
|
|
Prepublished online as a Blood First Edition Paper on December 27, 2002; DOI 10.1182/blood-2002-08-2574.
IMMUNOBIOLOGY
From the Departments of Medicine and Immunology, Mayo
Clinic, Rochester, MN.
CD28 is the quintessential costimulatory molecule expressed on
CD4+ and CD8+ T cells. During chronic
infections and the normal aging process, CD28 expression is lost,
compromising the functional activity of T cells. CD28 loss is promoted
by replicative stress, particularly in the presence of tumor necrosis
factor- T-cell activation requires specific antigen
recognition by the T-cell receptor (TCR) and a second signal from
costimulatory molecules. The classic costimulatory molecule is
CD28.1 CD28 provides signals that promote T-cell survival,
interleukin-2 (IL-2) production, metabolic activity, and T-cell clonal
expansion.2-5 This central role of CD28 suggests that
modulation of the levels of CD28 expression profoundly alters T-cell
function. Indeed, targeted deletion of the CD28 gene in mice results in
an immunocompromised phenotype, typified by reduced helper T-cell
activity and impaired immunoglobulin production.6
Although CD28 is constitutively expressed on the surface of human T
cells, its expression can be down-regulated. A progressive increase in
the percentage of T cells that lack CD28 expression is common with
increasing age in healthy individuals7,8 and in patients
with chronic infections.9
CD4+CD28null T cells can comprise up to 50% of
the total CD4+ T-cell compartment in some individuals older
than 65 years.10-12 CD28null T cells have a
memory (CD45RO+) phenotype, are long lived in vivo, and
form large oligoclonal populations.8,13 In vitro,
successive lymphocyte replication is accompanied by CD28 loss and
telomeric shortening.11,12,14,15 CD28null T
cells have reduced division potential and delayed cell cycle kinetics.
CD4+CD28null T cells are resistant to apoptotic
signals, while the data for CD8+CD28null T
cells are conflicting, and increased and decreased susceptibility to
apoptosis has been described for CD8+ T
cells.12,16 Taken together, CD28null T cells
have features that are hallmarks of an aged immune system. Indeed, an
increased frequency of CD28null T cells has been found to
be the best predictor of humoral incompetence in the
elderly.17
Several additional phenotypic and functional characteristics
distinguish CD4+CD28null T cells from classic
CD4+CD28+ helper T cells. Expression of the
IL-2 receptor (CD25) after activation is short lived compared with
CD4+CD28+ T cells.18
CD4+CD28null T cells also lack CD40 ligand
(CD40L) expression, which renders the cells incapable of promoting
B-cell differentiation and immunoglobulin secretion.19
CD4+CD28null T cells have acquired receptors
that are typically associated with natural killer (NK) cells. In
contrast to CD4+CD28+ T cells, they express
CD8- CD4+ T cells may also completely lose CD28 surface
expression during chronic inflammatory conditions, such as rheumatoid
arthritis (RA),22 Wegener granulomatosis,23
and multiple sclerosis.24 In these diseases,
CD4+CD28null T cells are proinflammatory and
have been implicated in the autoimmune pathogenesis.22,24,25 CD4+CD28null
T cells isolated from patients with RA are autoreactive13
and produce large amounts of interferon- We have hypothesized that modulation of CD28 expression on senescent
CD4+CD28null T cells influences immune
responses. In the context of an aging immune system, re-expression of
CD28 may be desirable to restore immunocompetence. In contrast,
induction of CD28 on CD28null T cells in autoimmune
diseases may augment the autoimmune response. Mechanisms that restore
CD28 expression are, therefore, potential therapeutic targets.
Here, we describe the IL-12 influence on the phenotype and
function of CD4+CD28null T cells. Our study
shows that CD4+CD28null T cells express a
functional IL-12 receptor and that IL-12 modulates expression of the
IL-12-responsive gene, CD161, on these cells. More importantly, IL-12
induces the surface expression of functional CD28 molecules on
CD4+CD28null T cells.
T-cell cloning
T-cell activation
Flow cytometry Flow cytometry was performed on freshly purified PBMCs and on CD4+ T-cell clones and lines with the following mAbs: mouse anti-CD4 (fluorescein isothiocyanate [FITC]), mouse anti-CD4 (peridinin chlorophyll A protein [PerCP]), and mouse anti-CD28 (phycoerythrin [PE]) (all Becton Dickinson); mouse anti-CD4 (allophycocyanin [APC]), mouse anti-CD25 (APC), mouse anti-CD154 (FITC), mouse anti-interferon-
(IFN-), rat anti-IL-12R 1, and rat anti-IL-12R2 (all
BD-Pharmingen); and mouse antiperforin (Ancell, Bayport, MN).
Mouse myeloma immunoglobulin G1 (IgG1) (ICN Pharmaceuticals, Costa Mesa, CA) and Simultest Control (Becton Dickinson) were used as isotype controls. Secondary antibodies were goat antirat immunoglobulin (APC), goat antimouse immunoglobulin (FITC)
(both Becton Dickinson), and rat antimouse immunoglobulin (PerCP)
(BD-Pharmingen). Samples were analyzed on a FACSCalibur flow cytometer
(Becton Dickinson), and the frequencies of T-cell subsets were
calculated by means of WinMDI software (Joseph Trotter, Scripps
Research Institute, LaJolla, CA).
Reverse-transcribed PCR Total RNA was extracted from T-cell clones by means of TRIzol reagent (Invitrogen Life Technologies, Carlsbad, CA), and cDNA was synthesized by means of oligodeoxythymidine (oligo(dT)) and avian myeloblastosis virus (AMV) reverse transcriptase (Roche Molecular Biochemicals, Indianapolis, IN). CD28 transcripts were amplified by polymerase chain reaction (PCR) (5'-CGCCCATGCTTGTAGCGTACG-3' and 5'-GATAGGCTGCGAAGTCGCGTG-3'), and products were electrophoresed on 2% agarose gels.Electrophoretic mobility shift assays Nuclear extracts were prepared10 from T-cell clones and lines 5 to 8 days after incubation in IL-12 alone, activation with immobilized anti-CD3 mAb alone, or activation with immobilized anti-CD3 mAb with IL-12. Resting T-cell lines were harvested 3 weeks after the last stimulation. Briefly, 5 × 106 to 1 × 107 cells were lysed in cold HEPES (N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid) hypotonic buffer, and the nuclei were isolated by centrifugation. Nuclear proteins were extracted in 50 µL high-salt buffer, and protein concentration was determined by means of a protein assay kit (BioRad, Richmond, CA). Nuclear extracts (20 µg) were combined with 30 µL binding buffer containing 10 µg polydoxyinosine-polydeoxycytidine (poly(dI-dC)) (ICN Pharmaceuticals) and 5 µg nonspecific oligonucleotide. To this mixture, 5 µL wash buffer was added. The total reaction volume was adjusted to 50 µL with binding buffer and incubated on ice for 30 minutes. Approximately 40 fmol radiolabeled probe corresponding to the site of the CD28 initiator element10,26 was added and
incubated for an additional 30 minutes at room temperature. Protein-DNA
complexes were resolved on 6% nondenaturing polyacrylamide gels and
were visualized by autoradiography.
As a system control, parallel DNA-binding assays were performed by means of oligonucleotide probes specific for specificity protein-1 (SP1),10 a ubiquitous transcription factor complex.
Constitutive expression of the IL-12 receptor on CD4+CD28null T cells We have previously shown that CD4+CD28null T cells have phenotypic and functional properties in common with NK cells.20 In particular, CD4+CD28null T cells express CD8-
homodimers; several types of human leukocyte antigen (HLA)
class I-recognizing receptors; and CD161, a C-type lectin receptor. A
critical cytokine in the regulation of NK cells is IL-12, which can
induce IFN- secretion and enhance NK-cell cytotoxicity.
To examine whether CD4+CD28null T cells are
responsive to IL-12, we analyzed PBMCs from donors who had an expansion
of CD4+CD28null T cells (greater than 1% of
the CD4+ population) for the expression of the IL-12
receptor. Antibodies specific for the IL-12R
CD4+CD28null T-cells clones are IL-12 responsive To confirm that CD4+CD28null T cells express a functional IL-12 receptor, we examined the capacity of IL-12 to modulate the expression of CD161, an IL-12-responsive gene.20 CD161 was constitutively expressed on CD4+CD28null T-cell clones and was up-regulated after exposure to IL-12 (Figure 2A). In contrast, CD161 on CD4+CD28+ T-cell clones, which express low levels of the molecule, was not inducible by IL-12 exposure (Figure 2B). Exposure of CD4+CD28null T-cell clones to IL-12 had no effect on the expression of members of the KIR family, such as the CD158b receptor (Figure 2C). The expression of CD4 also did not appreciably change after stimulation with IL-12 (Figure 2D).
IL-12 induces expression of CD28 on CD4+CD28null T-cells CD4+CD28null T cells lack expression of CD28 owing to a transcriptional block of the CD28 gene.10,26 To determine whether IL-12 could modulate expression of CD28, CD4+CD28null T-cell lines and clones were stimulated with IL-12. Exposure to IL-12 alone did not induce CD28 transcription although CD161 was up-regulated (Figure 2A and data not shown). Activation with anti-CD3 mAb in the presence of IL-12 restored CD28 transcription, and CD28 mRNA could be detected in IL-12-stimulated clones (Figure 3A). Cell surface re-expression of the CD28 molecule after IL-12 stimulation was confirmed by flow cytometry. Stimulation of resting CD4+CD28null T-cell clones with anti-CD3 alone did not up-regulate CD28 expression. In contrast, activation with anti-CD3 mAb in the presence of IL-12 markedly enhanced CD28 surface expression on 7 of 15 CD28null T-cell clones tested (Figure 3E-F). CD28 expression on CD28+ T-cell clones was unchanged regardless of the mode of stimulation (Figure 3C). There was no obvious difference between the T-cell clones that re-expressed CD28 and those that did not. The 2 types of clones had similar levels of expression of a functional IL-12 receptor and equivalently induced CD161 transcription after IL-12 stimulation. For some of the clones, IL-12 consistently restored CD28 expression in only a fraction of all cells, indicating clonal heterogeneity (Figure 3E).
To determine whether the results on the T-cell clones were representative of the in vivo situation, polyclonal CD4+CD28null T-cell lines were examined. The results were very similar to what was observed on the T-cell clones. CD28 was reinducible by IL-12 in 5 of 11 CD4+CD28null T-cell lines tested (Figure 3D). These results were reproducible and did not depend on the culture conditions or the duration of the T cells in culture, but they were characteristic for the individuals from whom the line or clone was established. CD28 levels were unaffected on CD4+CD28+ T-cell lines following IL-12 exposure (Figure 3B). The effect of IL-12 on CD28 expression was seen to be variable
at low doses of 1 ng/mL, and optimal induction was achieved by 10 to 20 ng/mL IL-12 (Figure 4A). CD28
re-expression was generally not seen early after T-cell receptor and
IL-12 stimulation but was a delayed response. To determine the time
course for CD28 expression, CD4+CD28null T-cell
clones were stimulated with IL-12 and evaluated by serial flow
cytometric analysis. Results are expressed as the mean fluorescence intensity of CD28 expression (Figure 4B). CD28 expression peaked at day
6 following activation, suggesting that IL-12 had an indirect effect on
the de novo transcription and translation of the CD28 gene. Therefore,
all experiments shown in Figure 3B-F were done at day 5 to 6 after
stimulation with an optimal concentration of 20 ng/mL IL-12.
Restoration of CD28 expression on CD4+CD28null T-cell lines and clones was a temporary phenomenon. Representative results are shown in Figure 4C. CD28 expression was maximal 5 days after T-cell receptor and IL-12 stimulation. At that time, IL-12 was removed from the culture medium, and the clones or lines were maintained in only IL-2. Cells started to lose CD28 again between days 7 and 15 after stimulation and were completely negative by day 20. Activation of CD4+CD28null T cells restores DNA-binding activity to the CD28 initiator region Transcription of the CD28 gene is controlled at the level of the CD28 initiator region (INR).26 Loss of DNA-binding activities to CD28 INR sequences is associated with a transcriptional block that leads to the CD28null phenotype. IL-12 stimulation may reconstitute the initiator protein complex, thereby restoring CD28 expression. Alternatively, IL-12 may directly enhance/up-regulate CD28 transcription. To address this question, electrophoretic mobility shift assays with INR-specific probes were performed on nuclear extracts of CD4+CD28null T-cell lines and T-cell clones activated with anti-CD3 mAb in the presence or absence of IL-12 for 5 to 8 days. As expected, the anti-CD3-stimulated and the IL-12-stimulated CD4+CD28null T-cell clones and lines did not yield any binding activity (Figure 5, upper panel). A full restoration of the DNA-binding activity was seen with anti-CD3 mAb and IL-12 in those CD28null T cells that could be induced to express CD28 on the cell surface. In contrast, T-cell clones that could not be induced to express CD28 did not yield any binding activity (data not shown).
The reinduction of CD28 INR-specific transcription factors by IL-12 was a specific response. The level of INR-binding activities in CD28+ T cells was unaffected by T-cell receptor triggering with or without IL-12 (Figure 5, upper panel). Moreover, DNA-binding activities of the SP1 transcription factor complex were also unaffected (Figure 5, lower panel). IL-12 induction of CD28 expression restores costimulatory function To determine whether IL-12 restores functional competence of CD4+CD28null T cells, costimulatory activity of the re-expressed CD28 was assessed for the up-regulation of the T-cell activation markers, CD25 and CD40L.CD4+CD28null T-cell clones and lines were
cultured with anti-CD3 mAb and IL-12 for 6 days to induce CD28
expression. Cells were harvested and activated for 24 hours with
anti-CD3 mAb in the presence or absence of anti-CD28 mAb. Under these
conditions, at least 50% of all cells expressed CD28. Stimulated cells
were then analyzed by 4-color flow cytometry for the expression of CD25
and CD40L on CD4+CD28null and
CD4+CD28+ T cells. Stimulation with anti-CD3
and anti-CD28 mAbs, but not with anti-CD3 mAb alone, resulted in
increased CD25 and CD40L expression on the fraction of T cells that
expressed CD28 (Figure 6, left panel). No
costimulatory activity was seen on CD4+ T cells that failed
to express CD28 (Figure 6, right panel).
The present studies demonstrate that the function of CD4+CD28null T-cells is profoundly influenced by exposure to IL-12. CD4+CD28null T cells express IL-12 receptors and respond by up-regulating a known IL-12-responsive gene, CD161 (Figures 1-2). More interestingly, activation in the presence of IL-12 results in the restoration of CD28 gene transcription and the cell surface appearance of a functional CD28 molecule (Figures 3-4). CD28 gene transcription is mediated by reassembly of a protein complex binding to the initiator element of the CD28 gene promoter (Figure 5). In cells that re-express CD28, the molecule is functional and is able to provide a costimulatory signal that enhances expression of CD40L and CD25 (Figure 6). The IL-12 receptor is a heterodimeric protein composed of The origin of CD4+CD28null T cells is not known. The resemblance of CD4+CD28null T cells to NK T cells suggests that they derive from a lineage that is separate from that of classic CD4+ T cells. However, CD4+CD28null T cells do not express the invariant TCR that is characteristic of CD1d-restricted NK T cells.22 Additionally, recent work suggests that CD4+CD28null T cells successively acquire NK receptors after completion of TCR rearrangement.30 In vitro, T-cell replicative senescence is associated with down-regulation of CD28 expression,7,11,14 and CD4+CD28null T cells are expanded in the elderly8-10 and in patients with autoimmune diseases.22-25 CD4+CD28null T cells, therefore, are likely to derive from CD28+ precursors. CD28 expression can be influenced by cytokines. In vitro, tumor
necrosis factor- Stimulation with IL-12 alone was insufficient to induce CD28 transcription and required concurrent triggering of the TCR. This was in contrast to the induction of the signal transducer and activator of transcription-4 (STAT-4)-dependent gene, CD161. The CD28 promoter has a putative STAT-4-binding site. It is, therefore, possible that re-expression of CD28 requires the concerted action of several transcription factors, including STAT-4. Alternatively, TCR- and IL-12-triggered signaling may act on the CD28 promoter only indirectly. We have previously shown that CD4+CD28null T cells lack CD28 expression owing to a transcriptional block of the CD28 gene and that these cells uniformly lack binding activity of specific nuclear proteins to the CD28 promoter.10,14 It has also been shown that the expression of these nuclear protein complexes correlates with the cell surface expression of CD28 on T cells.14 In transcription assays, this nuclear protein complex is able to drive a CD28 INR-driven promoter construct, suggesting that CD28 loss is regulated at the level of transcription initiation.26 We now demonstrate that activation of CD4+CD28null T cells through the TCR in the presence of IL-12 results in restoration of this initiator complex and consequently reverses the transcriptional block of the CD28 gene. Recent studies suggest that the assembly of this initiator complex requires the posttranslational modification of the common nuclear proteins nucleolin and heterogeneous nuclear ribonucleoprotein-D (hnRNPD).33 How IL-12 stimulation induces this modification is not known; however, such an indirect mechanism is consistent with the delayed effect of IL-12 expression. Loss of CD28 expression is one of several developmental changes that occur in T cells during replicative senescence.7,11,14 Other changes are the loss of CD40L expression and the expression of several genes that are generally associated with NK cells, such as KIRs, CD161 and other C-type lectin receptors, perforin, and granzyme B.19-21 This raises the question of whether IL-12 can reverse the entire process or whether it selectively reactivates the transcription of the CD28 gene. Indeed, IL-12 restored the ability to express CD40L after optimal stimulation (Figure 6). In some clones and lines, IL-12 also down-regulates perforin expression (data not shown). Although these results indicate a broader action of IL-12, the major effect was on CD28 expression. IL-12 is unable to down-regulate KIR expression, but it induces CD161 expression instead of repressing it (Figure 2). The rather selective activity of IL-12 on CD28 expression is consistent with the finding that the CD28 INR sequence motifs have not been identified in other genes.26 Ultimately, defining the IL-12-mediated modification of the CD28 initiator complex will be necessary to determine whether and how perforin down-regulation is related to CD28 re-expression. Re-expression of CD28 upon IL-12/TCR costimulation is not universal for all CD4+CD28null T cells. Approximately 50% of T-cell clones and T-cell lines are completely refractory to the action of IL-12 although they express a functional IL-12 receptor. Some clones and cell lines were heterogeneous with respect to their ability to re-express CD28. The ability or inability of each T-cell clone to activate the transcription of CD28 is reproducible. These results are consistent with a model in which CD28 loss is a stepwise process that is initially reversible upon IL-12 exposure but is ultimately permanent. Both stages correlate with the formation, and eventually the loss, of the CD28 initiator complex.14,26,31 In CD4+ clones with reversible CD28 expression, the loss of the nuclear CD28 initiator complex is reversible. In T cells with permanent CD28 loss, the CD28 initiator complex cannot be formed. Loss of CD28 expression on CD4+ and CD8+ T cells is generally considered to be a feature of immunosenescence,34 and the frequency of CD28null T cells has been shown to be a predictor of humoral incompetence to vaccination in the elderly.17 The lack of CD28 expression may not only be a surrogate marker for immunosenescence but could be of direct functional importance. Among the several defects of CD4+CD28null T cells, the lack of CD40L expression after TCR stimulation is most striking. As a consequence, CD4+CD28null T cells cannot support the differentiation of B cells to produce immunoglobulins.19 Our results demonstrate that IL-12-mediated restoration of CD28 signaling enhances the up-regulation of CD40L expression. Restoration of the helper cell phenotype in immunosenescent T-cell populations may, in part, reverse the age-related decline in immune function. CD4+CD28null T cells have low and unstable expression of CD25,12,18 which contributes to the decreased replicative potential of senescent T cells and to the prolonged survival and accumulation of CD28null T cells in the aging immune system. The increased resistance of CD4+CD28null T cells to apoptosis-inducing signals has been linked to dysregulation of the FLICE (Fas-association death domain [FADD]-like IL-1-converting enzyme)-like inhibitory protein, which is in part controlled by IL-2-mediated signals.12 CD25 expression is dependent on CD28-mediated signals, and restoration of CD28 expression enhances the expression of CD25. Thus, re-expression of CD28 may restore T-cell homeostasis, reduce the expansion of the CD28null compartment, and generate space for naive T-cells.35 In addition to the beneficial effects of restoring immunocompetence,
re-expression of CD28 induced by IL-12 and TCR costimulation may be
detrimental in some chronic inflammatory diseases by enhancing autoimmune responses.13,24,36
CD4+CD28null T cells expanded in patients with
RA, acute coronary syndromes, multiple sclerosis, and some vasculitides
are potent producers of IFN-
We thank James W. Fulbright for assistance in preparing this manuscript and Linda H. Arneson for secretarial support.
Submitted August 21, 2002; accepted December 15, 2002.
Prepublished online as Blood First Edition Paper, December 27, 2002; DOI 10.1182/ blood-2002-08-2574.
Supported by grants from the National Institutes of Health (R01-AG15043, R21-GM58604, R01-AR42527, and R01-AR41974) and the Mayo Foundation.
K.J.W. and A.N.V. contributed equally and are regarded as co-first authors.
The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked "advertisement" in accordance with 18 U.S.C. section 1734.
Reprints: Jörg J. Goronzy, Guggenheim 401, 200 First St SW, Mayo Clinic, Rochester, MN 55905; e-mail: goronzy.jorg{at}mayo.edu.
1. Sharpe AH, Freeman GJ. The B7-CD28 superfamily. Nature Rev Immunol. 2002;2:116-126[CrossRef][Medline] [Order article via Infotrieve]. 2. Boise LH, Minn AJ, Noel PJ, et al. CD28 costimulation can promote T cell survival by enhancing the expression of Bcl-XL. Immunity. 1995;3:87-98[CrossRef][Medline] [Order article via Infotrieve].
3.
Thompson CB, Lindsten T, Ledbetter JA, et al.
CD28 activation pathway regulates the production of multiple T-cell-derived lymphokines/cytokines.
Proc Natl Acad Sci U S A.
1989;86:1333-1337 4. Sperling AI, Auger JA, Ehst BD, Rulifson IC, Thompson CB, Bluestone JA. CD28/B7 interactions deliver a unique signal to naive T cells that regulates cell survival but not early proliferation. J Immunol. 1996;157:3909-3917[Abstract]. 5. Frauwirth KA, Riley JL, Harris MH, et al. The CD28 signaling pathway regulates glucose metabolism. Immunity. 2002;16:769-777[CrossRef][Medline] [Order article via Infotrieve].
6.
Shahinian A, Pfeffer K, Lee KP, et al.
Differential T cell costimulatory requirements in CD28-deficient mice.
Science.
1993;261:609-612 7. Effros RB, Boucher N, Porter V, et al. Decline in CD28+ T cells in centenarians and in long-term T cell cultures: a possible cause for both in vivo and in vitro immunosenescence. Exp Gerontol. 1994;29:601-609[CrossRef][Medline] [Order article via Infotrieve].
8.
Posnett DN, Sinha R, Kabak S, Russo C.
Clonal populations of T cells in normal elderly humans: the T cell equivalent to "benign monoclonal gammopathy."
J Exp Med.
1994;179:609-618 9. Choremi-Papadopoulou H, Viglis V, Gargalianos P, Kordossis T, Iniotaki-Theodoraki A, Kosmidis J. Downregulation of CD28 surface antigen on CD4+ and CD8+ T lymphocytes during HIV-1 infection. J Acquir Immune Defic Syndr. 1994;7:245-253.
10.
Vallejo AN, Nestel AR, Schirmer M, Weyand CM, Goronzy JJ.
Aging-related deficiency of CD28 expression in CD4+ T cells is associated with the loss of gene-specific nuclear factor binding activity.
J Biol Chem.
1998;273:8119-8129
11.
Posnett DN, Edinger JW, Manavalan JS, Irwin C, Marodon G.
Differentiation of human CD8 T cells: implications for in vivo persistence of CD8+ CD28- cytotoxic effector clones.
Int Immunol.
1999;11:229-241
12.
Vallejo AN, Schirmer M, Weyand CM, Goronzy JJ.
Clonality and longevity of CD4+CD28null T cells are associated with defects in apoptotic pathways.
J Immunol.
2000;165:6301-6307 13. Schmidt D, Goronzy JJ, Weyand CM. CD4+ CD7- CD28- T cells are expanded in rheumatoid arthritis and are characterized by autoreactivity. J Clin Invest. 1996;97:2027-2037[Medline] [Order article via Infotrieve].
14.
Vallejo AN, Brandes JC, Weyand CM, Goronzy JJ.
Modulation of CD28 expression: distinct regulatory pathways during activation and replicative senescence.
J Immunol.
1999;162:6572-6579 15. Monteiro J, Batliwalla F, Ostrer H, Gregersen PK. Shortened telomeres in clonally expanded CD28-CD8+ T cells imply a replicative history that is distinct from their CD28+CD8+ counterparts. J Immunol. 1996;156:3587-3590[Abstract]. 16. Spaulding C, Guo W, Effros RB. Resistance to apoptosis in human CD8+ T cells that reach replicative senescence after multiple rounds of antigen-specific proliferation. Exp Gerontol. 1999;34:633-644[CrossRef][Medline] [Order article via Infotrieve].
17.
Goronzy JJ, Fulbright JW, Crowson CS, Poland GA, O'Fallon WM, Weyand CM.
Value of immunological markers in predicting responsiveness to influenza vaccination in elderly individuals.
J Virol.
2001;75:12182-12187 18. Park W, Weyand CM, Schmidt D, Goronzy JJ. Co-stimulatory pathways controlling activation and peripheral tolerance of human CD4+CD28- T cells. Eur J Immunol. 1997;27:1082-1090[Medline] [Order article via Infotrieve]. 19. Weyand CM, Brandes JC, Schmidt D, Fulbright JW, Goronzy JJ. Functional properties of CD4+ CD28- T cells in the aging immune system. Mech Ageing Dev. 1998;102:131-147[CrossRef][Medline] [Order article via Infotrieve]. 20. Warrington KJ, Takemura S, Goronzy JJ, Weyand CM. CD4+, CD28- T cells in rheumatoid arthritis patients combine features of the innate and adaptive immune systems. Arthritis Rheum. 2001;44:13-20[CrossRef][Medline] [Order article via Infotrieve]. 21. Namekawa T, Wagner UG, Goronzy JJ, Weyand CM. Functional subsets of CD4 T cells in rheumatoid synovitis. Arthritis Rheum. 1998;41:2108-2116[CrossRef][Medline] [Order article via Infotrieve]. 22. Schmidt D, Martens PB, Weyand CM, Goronzy JJ. The repertoire of CD4+ CD28- T cells in rheumatoid arthritis. Mol Med. 1996;2:608-618[Medline] [Order article via Infotrieve]. 23. Moosig F, Csernok E, Wang G, Gross WL. Costimulatory molecules in Wegener's granulomatosis (WG): lack of expression of CD28 and preferential up-regulation of its ligands B7-1 (CD80) and B7-2 (CD86) on T cells. Clin Exp Immunol. 1998;114:113-118[CrossRef][Medline] [Order article via Infotrieve]. 24. Markovic-Plese S, Cortese I, Wandinger KP, McFarland HF, Martin R. CD4+CD28- costimulation-independent T cells in multiple sclerosis. J Clin Invest. 2001;108:1185-1194[CrossRef][Medline] [Order article via Infotrieve]. 25. Martens PB, Goronzy JJ, Schaid D, Weyand CM. Expansion of unusual CD4+ T cells in severe rheumatoid arthritis. Arthritis Rheum. 1997;40:1106-1114[Medline] [Order article via Infotrieve].
26.
Vallejo AN, Weyand CM, Goronzy JJ.
Functional disruption of the CD28 gene transcriptional initiator in senescent T cells.
J Biol Chem.
2001;276:2565-2570 27. Sinigaglia F, D'Ambrosio D, Panina-Bordignon P, Rogge L. Regulation of the IL-12/IL-12R axis: a critical step in T-helper cell differentiation and effector function. Immunol Rev. 1999;170:65-72[CrossRef][Medline] [Order article via Infotrieve]. 28. Poggi A, Costa P, Zocchi MR, Moretta L. Phenotypic and functional analysis of CD4+ NKRP1A+ human T lymphocytes: direct evidence that the NKRP1A molecule is involved in transendothelial migration. Eur J Immunol. 1997;27:2345-2350[Medline] [Order article via Infotrieve].
29.
Poggi A, Zocchi MR, Costa P, et al.
IL-12-mediated NKRP1A up-regulation and consequent enhancement of endothelial transmigration of V delta 2+ TCR gamma delta+ T lymphocytes from healthy donors and multiple sclerosis patients.
J Immunol.
1999;162:4349-4354
30.
Snyder MR, Muegge LO, Offord C, et al.
Formation of the killer Ig-like receptor repertoire on CD4+CD28null T cells.
J Immunol.
2002;168:3839-3846
31.
Bryl E, Vallejo AN, Weyand CM, Goronzy JJ.
Down-regulation of CD28 expression by TNF-alpha.
J Immunol.
2001;167:3231-3238 32. Morita Y, Yamamura M, Nishida K, et al. Expression of interleukin-12 in synovial tissue from patients with rheumatoid arthritis. Arthritis Rheum. 1998;41:306-314[CrossRef][Medline] [Order article via Infotrieve].
33.
Vallejo AN, Bryl E, Klarskov K, Naylor S, Weyand CM, Goronzy JJ.
Molecular basis for the loss of CD28 expression in senescent T cells.
J Biol Chem.
2002;277:46940-46949 34. Globerson A, Effros RB. Ageing of lymphocytes and lymphocytes in the aged. Immunol Today. 2000;21:515-521[CrossRef][Medline] [Order article via Infotrieve]. 35. Theofilopoulos AN, Dummer W, Kono DH. T cell homeostasis and systemic autoimmunity. J Clin Invest. 2001;108:335-340[CrossRef][Medline] [Order article via Infotrieve].
36.
Liuzzo G, Kopecky SL, Frye RL, et al.
Perturbation of the T-cell repertoire in patients with unstable angina.
Circulation.
1999;100:2135-2139
© 2003 by The American Society of Hematology.
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
 |
|
 |
|
  I. E. Dumitriu, E. T. Araguas, C. Baboonian, and J. C. Kaski CD4+CD28null T cells in coronary artery disease: when helpers become killers Cardiovasc Res, January 1, 2009; 81(1): 11 - 19. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 G. Liuzzo, L. M. Biasucci, G. Trotta, S. Brugaletta, M. Pinnelli, G. Digianuario, V. Rizzello, A. G. Rebuzzi, C. Rumi, A. Maseri, et al. Unusual CD4+CD28null T Lymphocytes and Recurrence of Acute Coronary Events J. Am. Coll. Cardiol., October 9, 2007; 50(15): 1450 - 1458. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. E. Sadun, S. M. Sachsman, X. Chen, K. W. Christenson, W. Z. Morris, P. Hu, and A. L. Epstein Immune Signatures of Murine and Human Cancers Reveal Unique Mechanisms of Tumor Escape and New Targets for Cancer Immunotherapy Clin. Cancer Res., July 1, 2007; 13(13): 4016 - 4025. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 B. Zal, C. Baboonian, and J. C. Kaski Letter to the Editor: Autoreactive CD4+CD28- T Cells and Acute Coronary Syndromes Arterioscler Thromb Vasc Biol, March 1, 2007; 27(3): E18 - E18. [Full Text] [PDF]
|
|
|
|
 |
|
 W. K. Chiu, M. Fann, and N.-p. Weng Generation and Growth of CD28nullCD8+ Memory T Cells Mediated by IL-15 and Its Induced Cytokines J. Immunol., December 1, 2006; 177(11): 7802 - 7810. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 X. Zhang, A. Niessner, T. Nakajima, W. Ma-Krupa, S. L. Kopecky, R. L. Frye, J. J. Goronzy, and C. M. Weyand Interleukin 12 Induces T-Cell Recruitment Into the Atherosclerotic Plaque Circ. Res., March 3, 2006; 98(4): 524 - 531. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. Amyes, A. J. McMichael, and M. F. C. Callan Human CD4+ T Cells Are Predominantly Distributed among Six Phenotypically and Functionally Distinct Subsets J. Immunol., November 1, 2005; 175(9): 5765 - 5773. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Duftner, R. Seiler, P. Klein-Weigel, H. Gobel, C. Goldberger, C. Ihling, G. Fraedrich, and M. Schirmer High Prevalence of Circulating CD4+CD28- T-Cells in Patients With Small Abdominal Aortic Aneurysms Arterioscler Thromb Vasc Biol, July 1, 2005; 25(7): 1347 - 1352. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. Gniadecki and A. Lukowsky Monoclonal T-Cell Dyscrasia of Undetermined Significance Associated With Recalcitrant Erythroderma Arch Dermatol, March 1, 2005; 141(3): 361 - 367. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. L. Riley and C. H. June The CD28 family: a T-cell rheostat for therapeutic control of T-cell activation Blood, January 1, 2005; 105(1): 13 - 21. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. J. Powell Jr, M. E. Dudley, P. F. Robbins, and S. A. Rosenberg Transition of late-stage effector T cells to CD27+ CD28+ tumor-reactive effector memory T cells in humans after adoptive cell transfer therapy Blood, January 1, 2005; 105(1): 241 - 250. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. Wassink, P. L. Vieira, H. H. Smits, G. A. Kingsbury, A. J. Coyle, M. L. Kapsenberg, and E. A. Wierenga ICOS Expression by Activated Human Th Cells Is Enhanced by IL-12 and IL-23: Increased ICOS Expression Enhances the Effector Function of Both Th1 and Th2 Cells J. Immunol., August 1, 2004; 173(3): 1779 - 1786. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. M. M. van Leeuwen, E. B. M. Remmerswaal, M. T. M. Vossen, A. T. Rowshani, P. M. E. Wertheim-van Dillen, R. A. W. van Lier, and I. J. M. ten Berge Emergence of a CD4+CD28- Granzyme B+, Cytomegalovirus-Specific T Cell Subset after Recovery of Primary Cytomegalovirus Infection J. Immunol., August 1, 2004; 173(3): 1834 - 1841. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. E. Lewis, M. Merched-Sauvage, J. J. Goronzy, C. M. Weyand, and A. N. Vallejo Tumor Necrosis Factor-{alpha} and CD80 Modulate CD28 Expression through a Similar Mechanism of T-cell Receptor-independent Inhibition of Transcription J. Biol. Chem., July 9, 2004; 279(28): 29130 - 29138. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Leslie All Pain, No Gain Sci. Aging Knowl. Environ., July 7, 2004; 2004(27): ns4 - ns4. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. Gerli, G. Schillaci, A. Giordano, E. B. Bocci, O. Bistoni, G. Vaudo, S. Marchesi, M. Pirro, F. Ragni, Y. Shoenfeld, et al. CD4+CD28- T Lymphocytes Contribute to Early Atherosclerotic Damage in Rheumatoid Arthritis Patients Circulation, June 8, 2004; 109(22): 2744 - 2748. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. Li, H.-C. Hsu, C. R. Stockard, P. Yang, J. Zhou, Q. Wu, W. E. Grizzle, and J. D. Mountz IL-12 Inhibits Thymic Involution by Enhancing IL-7- and IL-2-Induced Thymocyte Proliferation J. Immunol., March 1, 2004; 172(5): 2909 - 2916. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
 |
 |
 |
|||||||
 |
 |
 |
 |
 |
 |
 |
 |
||
| Copyright © 2003 by American Society of Hematology Online ISSN: 1528-0020 | |||||||||
Top
Abstract
Introduction
) of IL-12 (10 ng/mL),
and the transcription of CD28 was analyzed by RT-PCR (clones 304I and
210I are shown). (B-F) Re-expression of CD28 was confirmed by flow
cytometry. CD4+CD28null T-cell clones (panels
E-F) and lines (panel D) were stimulated with anti-CD3 mAb,
IL-12, or a combination of anti-CD3 mAb and IL-12. Cells were analyzed
by 4-color flow cytometry after 5 to 6 days; representative histograms
are shown. The combination of anti-CD3 mAb and IL-12 induced CD28
expression in 5 of 11 lines and 6 of 13 clones. In contrast, the levels
of CD28 expression were not affected in
CD4+CD28+ T-cell clones or lines (panels
B-C). In all histograms, the shaded areas represent staining with
an isotype control.
) compared with the
isotype control (
) at each time point. CD28 expression was maximal
after 6 days of culture in IL-12. (C) To examine whether re-expression
of CD28 was transient, T cells were stimulated with anti-CD3 mAb and
IL-12 for 5 days and then maintained in the absence of IL-12. CD28
expression decreased again and was absent at day 20 after initial
stimulation. A representative example of 3 experiments is shown for a
clone (upper panel) and a line (lower panel). In all
histograms, the shaded areas represent staining with an isotype
control.
T-cell clones isolated from patients with multiple sclerosis
have been reported to migrate across an endothelial layer more
effectively than CD161
clones, and this
migratory potential is enhanced by IL-12.