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PubMed1996

Identification of TRAF6, a novel tumor necrosis factor receptor-associated factor protein that mediates signaling from an amino-terminal domain of the CD40 cytoplasmic region.

CD40 signalings play crucial roles in B-cell function. To identify molecules which transduce CD40 signalings, we have utilized the yeast two-hybrid system to clone cDNAs encoding proteins that bind the cytoplasmic tail of CD40. A cDNA encoding a putative signal transducer, designated TRAF6, has been molecularly cloned. TRAF6 has a tumor necrosis factor receptor (TNFR)-associated factor (TRAF) domain in its carboxyl terminus and has a RING finger domain, a cluster of zinc fingers and a coiled-coil domain, which are also present in other TRAF family proteins. TRAF6 does not associate with the cytoplasmic tails of TNFR2, CD30, lymphotoxin-beta receptor, and LMP1 of Epstein-Barr virus. Deletion analysis showed that residues 246-269 of CD40 which are required for its association with TRAF2, TRAF3, and TRAF5 are dispensable for its interaction with TRAF6, whereas residues 230-245 were required. Overexpression of TRAF6 activates transcription factor NFkappaB, and its TRAF-C domain suppresses NFkappaB activation triggered by CD40 lacking residues 246-277. These results suggest that TRAF6 could mediate the CD40 signal that is transduced by the amino-terminal domain (230-245) of the CD40 cytoplasmic region and appears to be independent of other known TRAF family proteins.

Amino Acid Sequence↗
PubMed1999

CD40 signaling of monocyte inflammatory cytokine synthesis through an ERK1/2-dependent pathway. A target of interleukin (il)-4 and il-10 anti-inflammatory action.

Ligation of CD40 on monocytes through its interaction with CD40 ligand (CD154) present on activated T helper cells, results in activation of monocyte inflammatory cytokine synthesis and rescue of monocytes from apoptosis induced through serum deprivation. Both of these consequences of CD40 stimulation have been shown to be dependent on the induction of protein tyrosine kinase activity. CD40-mediated activation of protein tyrosine kinase activity and subsequent inflammatory cytokine production are abrogated by treatment of monocytes with the T helper type 2 cytokines interleukin 4 (IL-4) and interleukin 10 (IL-10). In the current study we demonstrate that stimulation of monocytes through CD40 resulted in the phosphorylation and activation of the extracellular signal-regulated kinases 1 and 2 (ERK1/2) mitogen-activated protein kinases, whereas phosphorylation of mitogen-activated protein kinases family members p38 and c-Jun N-terminal kinase was not observed in response to this stimuli over the time course examined. PD98059, an inhibitor of the upstream activator of ERK1/2, the MAP/ERK kinase MEK1/2, suppressed IL-1beta and tumor necrosis factor-alpha production in a dose-dependent fashion. Pretreatment of monocytes with IL-4 and IL-10 inhibited CD40-mediated activation of ERK1/2 kinase activity when used individually, and are enhanced in effectiveness when used in combination. Together, the data demonstrate that CD40-mediated induction of IL-1beta and tumor necrosis factor-alpha synthesis is dependent on a MEK/ERK pathway which is obstructed by signals generated through the action of IL-4 and IL-10.

CD40 Antigens↗
PubMed2001

The role of Sp1 and NF-kappa B in regulating CD40 gene expression.

CD40 is a member of the tumor necrosis factor receptor superfamily and is a key signaling molecule expressed by antigen-presenting cells of the immune system. In a previous paper, we demonstrated that the expression of CD40 is regulated by both post-transcriptional and post-translational processes. In this paper, we show that basal (constitutive) CD40 gene expression is regulated by a TATA-less promoter, with Sp1 as a key transcription factor. Two Sp1 binding regions were identified in the mouse CD40 promoter at positions -59 to -50 and -74 to -66. Surprisingly, Sp1-mediated CD40 transcription was reduced following lipopolysaccharide stimulation and was associated with a time-dependent reduction in Sp1 DNA binding activity. This reduction seemed to be mediated by phosphorylation of the Sp1 molecule. We also show here that CD40 expression in lipopolysaccharide-stimulated cells is up-regulated by NF-kappaB through two distinct sites. One of these sites (-128 to -119) was shown to bind p50 and p65 members of the NF-kappaB family, while the other site (-562 to -553) bound only p65. Transfectants of p65 were generated using RAW 264 cells, and it was shown that the up-regulation of CD40 mRNA expression was dependent on the presence of the p65 molecule.

Base Sequence↗
PubMed1993

B cell activation via CD40 is required for specific antibody production by antigen-stimulated human B cells.

Costimulatory signals provided by T cells are required for B cells to produce specific antibody (Ab) to T-dependent antigen (Ag) bacteriophage phi x 174. In this study, we demonstrate that if cultured in the presence of anti-CD40, interleukin 10 (IL-10), and Ag, purified B cells can produce antiphage Ab in quantities comparable to those synthesized by B cells cocultured with Ag and T cells. Isotypes produced by B cells in this culture system correspond to those observed in sera of B cell donors. Culture of immunoglobulin (Ig)D- and IgD+ B cells reveals that Ag-induced production of antiphage Ab is restricted to IgD- subset of B cells. In the absence of Ag, anti-CD40/IL-10-stimulated B cells produce only minute amounts of antiphage Ab, indicating that Ag stimulation is indispensable and provides a signal that is synergistic with anti-CD40 and IL-10. Addition of a soluble form of the CD40 ligand (sgp39) to the culture system has a similar effect on specific Ab synthesis as anti-CD40; addition of the soluble construct, CD40 Ig, known to inhibit gp39/CD40 interaction, suppresses in vitro antiphage Ab production by Ag exposed peripheral blood mononuclear cells. Finally, in vivo requirement of gp39/CD40 interaction for specific Ab production was demonstrated by the finding that activated T cells from patients with x-linked hyper IgM syndrome express functionally defective gp39 and respond with depressed Ab titers and fail to switch from IgM to IgG after multiple phage immunizations. These observations illustrate that in vitro and possibly in vivo Ag-specific Ab synthesis requires the presence of Ag and IL-10, and activation signals via CD40.

Antibodies, Viral↗
PubMed1996

CD40 and B cell antigen receptor dual triggering of resting B lymphocytes turns on a partial germinal center phenotype.

Phenotypic alterations occur when resting human B lymphocytes become germinal center (GC) cells. These include the induction of surface CD38, CD95 (FAS/APO-1), and carboxy-peptidase-M (CPM), a recently described GC marker. However, the factors that govern the in vivo induction of these surface molecules on B cells remain unknown. Here, we purified resting (CD38-) human B lymphocytes from tonsils in an attempt to establish culture conditions resulting in the induction of these three GC markers. We show that interferon (IFN) alpha or IFN-gamma, as well as antibodies against the B cell antigen receptor (BCR), could induce CD38 on resting B lymphocytes, a phenomenon further enhanced by CD40 stimulation. Concomitantly, CD95 was upregulated by CD40 ligation and, to a lesser extent, by IFN-gamma. By contrast, CPM expression could be upregulated only through BCR triggering. This CPM induction was specifically enhanced by CD19 or CD40 ligation. CD40 + BCR stimulation of resting B cells with CD40 ligand-transfected fibroblastic cells in the presence of cross-linked anti-BCR monoclonal antibodies resulted in the coexpression of CD38, CD95, and CPM. As GC cells, these cells also expressed CD71, CD80 (B7.1), and CD86 (B7.2), but not CD24. However, CD10+ or CD44- B cells could not be detected in these culture conditions, suggesting that yet other signals are required for the induction of these GC markers. Consistent with a GC phenotype, CD40 + BCR-stimulated cells exhibited reduced viability when cultured for 20 h in the absence of stimulus. These results first demonstrate that cotriggering of resting B cells through BCR and CD40 induces both phenotypic and functional GC features. They also show that IFN and CD19 triggering of resting B cells specifically modulate the expression of GC markers.

ADP-ribosyl Cyclase↗
PubMed1996

High level IL-12 production by murine dendritic cells: upregulation via MHC class II and CD40 molecules and downregulation by IL-4 and IL-10.

We have shown previously that dendritic cells (DC) produce IL-12 upon interaction with CD4+ T cells. Here we ask how this IL-12 production is induced and regulated. Quantitative PCR and in situ hybridization for IL-12 p40 and an ELISA specific for the p70 heterodimer were used to determine IL-12 production. We demonstrate that ligation of either CD40 or MHC class II molecules independently trigger IL-12 production in DC, and that IL-12 production is downregulated by IL-4 and IL-10. The levels of bioactive IL-12 that can be released by triggering with an anti-CD40 mAb or with a T cell hybridoma are high (range 260-4700 pg/ml from 1 X 10(6) DC in 72 h). The CD40-mediated pathway indicates that IL-12 production is induced in DC upon interaction with activated, CD40 ligand-expressing helper T cells, even in the absence of cognate antigen recognition. Side-by-side comparison of IL-12 production, and blocking experiments employing an anti-CD40 ligand mAb, suggest that the CD40-mediated pathway is quantitatively more significant than induction via the MHC class II molecule. The importance of the CD40/CD40 ligand interaction for IL-12 induction in DC likely contributes to the recent finding that mice lacking the CD40 ligand are impaired in mounting Th1 type cell-mediated immune responses.

Animals↗
PubMed1997

CD40 ligation on human cord blood CD34+ hematopoietic progenitors induces their proliferation and differentiation into functional dendritic cells.

Human CD34+ multilineage progenitor cells (CD34HPC) from cord blood and bone marrow express CD40, a member of the tumor necrosis factor-receptor family present on various hematopoietic and nonhematopoietic cells. As hyper-IgM patients with mutated CD40 ligand (CD40L) exhibit neutropenia, no B cell memory, and altered T cell functions leading to severe infections, we investigated the potential role of CD40 on CD34HPC development. CD40-activated cord blood CD34HPC were found to proliferate and differentiate independently of granulocyte/macrophage colony-stimulating factor, into a cell population with prominent dendritic cell (DC) attributes including priming of allogeneic naive T cells. DC generated via the CD40 pathway displayed strong major histocompatibility complex class II DR but lacked detectable CD1a and CD40 expression. These features were shared by a dendritic population identified in situ in tonsillar T cell areas. Taken together, the present data demonstrate that CD40 is functional on CD34HPC and its cross-linking by CD40L+ cells results in the generation of DC that may prime immune reactions during antigen-driven responses to pathogenic invasion, thus providing a link between hematopoiesis, innate, and adaptive immunity.

Animals↗
PubMed1999

Human non-germinal center B cell interleukin (IL)-12 production is primarily regulated by T cell signals CD40 ligand, interferon gamma, and IL-10: role of B cells in the maintenance of T cell responses.

Interleukin (IL)-12 is expressed mainly in antigen-presenting cells after challenge with microbial material or after CD40 activation. Although IL-12 was cloned from human Epstein-Barr virus (EBV)-transformed B cell lines, surprisingly, CD40 ligation on murine B cells did not lead to IL-12 production, suggesting that murine B cells do not produce IL-12. Here we demonstrate that a subset of human tonsillar B cells can be induced to express and secrete bioactive IL-12. The major stimulus to produce IL-12 in human B cells was CD40 ligation. In contrast, B cell receptor cross-linking did not induce IL-12. Expression of IL-12 after CD40 activation was restricted to CD38(-)IgD+/- non-germinal center (non-GC) B cells. CD40 ligation and interferon (IFN)-gamma exhibited synergistic effects on IL-12 production, whereas IL-10 abrogated and IL-4 significantly inhibited IL-12 production by these B cells. In contrast to IL-12, production of IL-6 is conversely regulated, leading to significant increase after CD40 ligation in the presence of the T helper type 2 (Th2) cytokine IL-4. Cord blood T cells skewed towards either a Th1 or a Th2 phenotype maintained their cytokine expression pattern when restimulated with allogeneic resting B cells. Blockade of CD40 and/or IL-12 during T-B interaction significantly reduced IFN-gamma production by the T cells. This suggests a model whereby B cells produce either IL-12 or IL-6 after contact with T cells previously differentiated towards Th1 or Th2. Furthermore, IL-12 and IL-6 might provide a positive feedback during cognate T-B interactions, thereby maintaining T cells' differentiation pattern during amplification of the immune response.

Antigen-Presenting Cells↗
PubMed2001

Differential signaling and tumor necrosis factor receptor-associated factor (TRAF) degradation mediated by CD40 and the Epstein-Barr virus oncoprotein latent membrane protein 1 (LMP1).

Latent membrane protein 1 (LMP1) plays a critical role in B cell transformation by Epstein-Barr virus (EBV) and appears to mimic a constitutively active CD40 receptor. Intracellular tumor necrosis factor (TNF) receptor-associated factor (TRAF) adapter proteins, shown to contribute to signaling by both CD40 and LMP1, were recruited by both molecules to lipid-enriched membrane rafts. However, we found that TRAFs 2 and 3 were subsequently degraded after CD40- but not LMP1-induced signaling. This degradation was proteasome-dependent and required direct TRAF binding by CD40. Using a model system designed to directly compare the signaling potency of the cytoplasmic domains of LMP1 and CD40 in B lymphocytes, we found that LMP1 more potently activates c-Jun kinase and nuclear factor kappaB and induces higher levels of several B cell effector functions than does CD40. This suggests that LMP1 utilizes a modified CD40 signaling pathway. Failure to regulate TRAFs may contribute to the enhanced capacity of LMP1 to activate B cells as well as promote B cell transformation.

Animals↗
PubMed1999

IFN-alpha stimulates proliferation and cytokine secretion of CD40-stimulated B cell chronic lymphocytic leukemia cells in vitro.

Interferon (IFN)-alpha has a therapeutic effect in several B cell malignancies, including low-grade non-Hodgkin's lymphoma (NHL), multiple myeloma, and hairy cell leukemia, whereas its efficacy in the treatment of B cell chronic lymphocytic leukemia (B-CLL) is rather limited. In the present study, we investigated the effect of IFN-alpha on the biologic functions of B-CLL cells, which were stimulated by cross-linking of the CD40 antigen. In cell samples from 16 B-CLL patients, the addition of IFN-alpha to CD40-stimulated purified B-CLL cells caused a significant increase in [3H]thymidine uptake (p < 0.003). In B-CLL cells maximally activated by CD40 cross-linking and interleukin-2 (IL-2)/IL-10, proliferation was not further enhanced or inhibited by IFN-alpha. In contrast, proliferation of normal tonsillar B cells stimulated by the combination CD40/IL-2/IL-10 was significantly inhibited by IFN-alpha. Because B-CLL activation might be enhanced by induction of the autocrine growth factor tumor necrosis factor-alpha (TNF-alpha), we investigated the effect of IFN-alpha on the secretion of B cell tropic cytokines. In fact, IFN-alpha significantly stimulated the secretion of IL-6 and TNF-alpha in CD40-activated B-CLL cells (p < 0.01). Although exogenous addition of TNF-alpha did not influence activation of CD40-stimulated B-CLL cells, neutralization of TNF-alpha by polyclonal antibodies led to a complete inhibition of CD40-mediated proliferation of B-CLL cells, suggesting that enhanced proliferation and increased cytokine production by CD40-activated B-CLL cells are independent IFN-alpha-mediated events. The studies presented here provide evidence that IFN-alpha mediates costimulatory signals in the context of T cell-mediated B-CLL cell activation.

Antineoplastic Agents↗
PubMed2002

CD40 is expressed and functional on neuronal cells.

We show here that CD40 mRNA and protein are expressed by neuronal cells, and are increased in differentiated versus undifferentiated N2a and PC12 cells as measured by RT-PCR, western blotting and immunofluorescence staining. Additionally, immunohistochemistry reveals that neurons from adult mouse and human brain also express CD40 in situ. CD40 ligation results in a time-dependent increase in p44/42 MAPK activation in neuronal cells. Furthermore, ligation of CD40 opposes JNK phosphorylation and activity induced by NGF-beta removal from differentiated PC12 cells or serum withdrawal from primary cultured neurons. Importantly, CD40 ligation also protects neuronal cells from NGF-beta or serum withdrawal-induced injury and affects neuronal differentiation. Finally, adult mice deficient for the CD40 receptor demonstrate neuronal dysfunction as evidenced by decreased neurofilament isoforms, reduced Bcl-x(L):Bax ratio, neuronal morphological change, increased DNA fragmentation, and gross brain abnormality. These changes occur with age, and are clearly evident at 16 months. Taken together, these data demonstrate a role of CD40 in neuronal development, maintenance and protection in vitro and in vivo.

Animals↗
PubMed2002

CD40 regulates the processing of NF-kappaB2 p100 to p52.

The nf-kb2 gene encodes the cytoplasmic NF-kappaB inhibitory protein p100 from which the active p52 NF-kappaB subunit is derived by proteasome-mediated proteolysis. Ligands which stimulate p100 processing to p52 have not been defined. Here, ligation of CD40 on transfected 293 cells is shown to trigger p52 production by stimulating p100 ubiquitylation and subsequent proteasome-mediated proteolysis. CD40-mediated p52 accumulation is dependent on de novo protein synthesis and triggers p52 translocation into the nucleus to generate active NF-kappaB dimers. Endogenous CD40 ligation on primary murine splenic B cells also stimulates p100 processing, which results in the delayed nuclear translocation of p52-RelB dimers. In both 293 cells and primary splenic B cells, the ability of CD40 to trigger p100 processing requires functional NF-kappaB-inducing kinase (NIK). In contrast, NIK activity is not required for CD40 to stimulate the degradation of IkappaBalpha in either cell type. The regulation of p100 processing by CD40 is likely to be important for the transcriptional regulation of CD40 target genes in adaptive immune responses.

Active Transport, Cell Nucleus↗
PubMed2004

Effect of beta2-adrenergic receptor agonists on intercellular adhesion molecule (ICAM)-1, B7, and CD40 expression in mixed lymphocyte reaction.

BACKGROUND: The plasma interleukin (IL)-18 level is elevated in acute rejection after organ transplantation. Although beta2-adrenergic receptor (AR) agonists suppress the rejection of organ and tissue transplants, little is known about their action mechanisms. We examined the effects of endogenous catecholamines and beta2-AR agonists on the expression of intercellular adhesion molecule (ICAM)-1, B7.1, B7.2, CD40, and CD40 ligand (CD40L) in human mixed lymphocyte reaction (MLR) and in an in vitro model of acute rejection in the presence or absence of IL-18. METHODS: ICAM-1, B7.1 B7.2, CD40, and CD40L expression on monocytes was measured by flow cytometry, and the production of interferon (IFN)-gamma and IL-12 was determined by enzyme-linked immunosorbent assay. Lymphocytes proliferation in MLR was measured by [3H]-thymidine uptake. The relevant AR subtypes were characterized using subtype-selective agonists and antagonists. RESULTS: beta2-AR agonists inhibited the expression of ICAM-1 and CD40 during MLR in the absence of IL-18. Among IL-18-induced expression of ICAM-1, B7.1, B7.2, CD40, and CD40L, beta2-AR agonists inhibited ICAM-1 and CD40 expression. beta2-AR agonists prevented the production of IFN-gamma and IL-12 in the presence of IL-18 but had no effect in the absence of IL-18. beta2-AR agonists inhibited lymphocyte proliferation in IL-18-treated MLR. CONCLUSIONS: We found that beta2-AR agonists strongly inhibited the expression of ICAM-1 and CD40, irrespective of the presence or absence of IL-18, which is different from that of histamine and prostaglandin E2.

Adrenergic beta-Agonists↗
PubMed2004

CD40 is necessary for activation of naïve T cells by a dendritic cell line in vivo but not in vitro.

The importance of CD40-CD40L interactions during CD4(+) T-cell activation has been extensively investigated over the years; however, it still remains questionable whether the interaction is a prerequisite for dendritic cell (DC)-mediated antigen-specific priming in vivo. Naïve CD4(+) T cells require two signals for proper activation and induction of differentiation: signal 1 is provided by peptide antigens in the context of the major histocompatibility complex (MHC) class II, while signal 2 is delivered by costimulatory molecules such as CD80 or CD86 present on the antigen-presenting cell (APC). It is well known that the expression of CD80/CD86 is upregulated after interaction between CD40 on APCs and CD40L expressed by at least partly activated T cells. We used a DC line, JawsII, to compare the importance of CD40 expression and downstream signalling in vitro and in vivo. JawsII cells represent pre-immature bone marrow-derived DCs expressing low levels of MHC molecules, low levels of B7 molecules and no CD40. We have previously shown that JawsII cells, despite the lack of CD40 expression, are capable of priming naïve allogeneic T cells in vitro. In correlation with the current literature, we present data showing that constitutive expression of CD40 significantly increases the priming capacity of JawsII cells in vitro. In addition, we show that CD40 expression is required for JawsII cell-dependent T-cell priming in vivo.

Animals↗
PubMed2006

Transfection of CD40 in a human oral squamous cell carcinoma keratinocyte line upregulates immune potency and costimulatory molecules.

BACKGROUND: There is upregulation of class II molecules of the major histocompatibility complex (MHC) by keratinocytes in oral squamous cell carcinoma (OSCC) and inflammatory diseases such as lichen planus. The significance of this expression, or whether it is accompanied by upregulation of membrane-bound costimulatory molecules, is unknown. OBJECTIVES: To compare the expression of CD40, CD80, CD86, MHC class II and intercellular adhesion molecule-1 (ICAM-1), and the ability to induce allogeneic T-lymphocyte proliferation in vitro, of a CD40- OSCC cell line, its CD40+ transfected derivative and null transfectants. METHODS: OSCC cell lines and purified T lymphocytes were cocultured and T cell proliferation recorded. Phenotypes were analysed by flow cytometry. RESULTS: After T lymphocyte proliferation, which all OSCC cell lines were able to induce, there was upregulation of MHC class II and ICAM-1. However, the CD40+ transfectants were the most immunologically potent and were the only cells to show increased expression of CD86 (as well as further upregulation of CD40 and a statistically insignificant rise in CD80). The effects of blocking antibodies on T-cell proliferation were only statistically significant with the CD40+ transfectants. CONCLUSIONS: While not essential, expression of CD40 by OSCC cells is necessary for optimal induction of allogeneic T lymphocytes, possibly because of concurrent upregulation of other membrane-bound costimulatory molecules.

Antigens, CD↗
PubMed1998

Downregulation of the CD95 receptor and defect CD40-mediated signal transduction in B-chronic lymphocytic leukemia cells.

Cross-linking of the CD40 receptors has been shown to induce protein tyrosine kinases (PTK) phosphorylation and prevent apoptosis in Bcl-2 negative germinal center B cells. The expression of CD40 on B chronic lymphocytic leukemia (B-CLL) cells was found to be similar to that of normal B cells. Activation of normal B cells with soluble anti-CD40 monoclonal antibody (mAb) induced tyrosine phosphorylation, prolonged survival and prevented apoptosis. However, activation of CD40 on B-CLL cells using soluble anti-CD40 mAb does not influence survival or apoptosis. Normal B cells entered apoptosis when cultured in the presence of soluble anti-CD95 mAb. This process was independent of PTK activity. On B-CLL cells, the CD95 molecules were downregulated and a transient PTK signal was observed when cross-linking of the receptor by soluble anti-CD95 mAb occurred. Interestingly, B-CLL cells did not enter apoptosis in the presence of anti-CD95 mAb. Our study indicates that survival signals mediated through the CD40 molecule and death signals mediated through the CD95 molecule used different intracellular pathways in control donor B cells. In contrast, B-CLL cells do not respond to these signals. The leukemic B cells showed a defective CD40-mediated signal transduction and downregulated CD95 receptor expression. As a consequence, no apoptosis could be induced in B-CLL cells by a soluble anti-CD95 mAb. The abnormalities of these receptors may contribute to the long-lived status of B-CLL cells.

Apoptosis↗
PubMed2003

Functional caveolae are a prerequisite for CD40 signaling in human renal proximal tubule cells.

The role of caveolae in CD40/CD154 activation of proinflammatory chemokines and their potential role in renal inflammatory disease were explored in primary cultures of human renal proximal tubule epithelial cells. With the use of a cell fractionation assay, caveolin-1 (Cav-1), the defining structural protein of caveolae, was detected exclusively in the cell membrane (detergent insoluble) component of resting and CD40-activated cells. In the unstimulated condition, CD40 was associated with Cav-1, and with activation of the receptor by its cognate ligand CD154, CD40 disassociated from Cav-1. Other previously identified components of the CD40 signaling pathway, namely, SAPK/JNK, p38, and ERK1/2 MAPKs, but not tumor necrosis factor receptor-associated factor 6 (TRAF-6), were also present within caveolae and dissociated from this structure with ligation of the CD40 receptor. Disruption of caveolae with filipin diminished CD40-mediated MAPK activation and blunted downstream monocyte chemoattractant protein-1 (MCP-1) and IL-8 production. Similarly, dislodgment of signaling proteins from their scaffolding with a peptide targeted to the Cav-1 scaffolding domain (CSD) resulted in blunted MAPK activation and augmented IL-8 and MCP-1 production. In contrast, epidermal growth factor (EGF)-mediated tyrosine phosphorylation of the EGF receptor and activation of ERK1/2 were not interrupted by the peptide. We conclude that in human renal proximal tubule epithelial cells, CD40 and its downstream MAPK signaling proteins are located in membrane rafts and that disruption of caveolae or dislodgment of signaling proteins from the CSD diminishes MAPK activation and IL-8 and MCP-1 production in these cells.

Amino Acid Sequence↗
PubMed2003

Soluble CD40 ligand levels indicate lipid accumulation in carotid atheroma: an in vivo study with high-resolution MRI.

OBJECTIVE: The CD40/CD40 ligand pathway mediates inflammatory processes important in atherogenesis and the formation of the intraplaque lipid pool. We tested the hypothesis that plasma levels of soluble CD40 ligand are elevated in patients with evidence of a lipid pool on high-resolution magnetic resonance imaging (MRI) of carotid stenoses. METHODS AND RESULTS: We recruited 49 patients with evidence of carotid atherosclerosis on ultrasonography; 3 patients could not undergo carotid MRI because of claustrophobia. The remaining 46 patients underwent high-resolution MRI of the carotid arteries. Two radiologists blinded to all other data determined the presence or absence of an intraplaque lipid pool based on the loss of signal between the 20-ms echo time (TE20) and the fat-suppressed TE55 fast spin-echo images. Plasma levels of soluble CD40 ligand were determined by ELISA. Baseline levels of soluble CD40 ligand were higher among patients with evidence of intraplaque lipid (n=14) than among those without it (n=32; median, 2.54 ng/mL; interquartile range [IQR], 1.85 to 3.52 vs median, 1.58 ng/mL; IQR, 1.21 to 2.39; P=0.02). In contrast, soluble CD40 ligand levels were not correlated with percent diameter stenosis (r=-0.19; P=0.21). The relative risk for intraplaque lipid associated with soluble CD40 ligand levels above the median was 6.0 (95% confidence interval, 1.15 to 31.23; P=0.03). The magnitude of this predictive effect did not substantially change after adjustment for traditional cardiovascular risk factors (relative risk, 5.12; 95% confidence interval, 0.78 to 33.73; P=0.09). CONCLUSIONS: Plasma levels of soluble CD40 ligand may predict patients with features of high-risk atherosclerotic lesions. These data provide novel insight into the mechanism through which elevated levels of soluble CD40 ligand may reflect cardiovascular risk in humans and illustrate the potential value of interfacing high-resolution MRI with studies of vascular inflammation.

Aged↗