Intercellular Adhesion Molecule-1 Transcription Research Articles

P53 activates ICAM-1 (CD54) expression in an NF-kappaB-independent manner.

Intercellular adhesion molecule-1 (ICAM-1) is a crucial receptor in the cell-cell interaction, a process central to the reaction to all forms of injury. Its expression is upregulated in response to a variety of inflammatory/immune mediators, including cellular stresses. The NF-kappaB signalling pathway is known to be important for activation of ICAM-1 transcription. Here we demonstrate that ICAM-1 induction represents a new cellular response to p53 activation and that NF-kappaB inhibition does not prevent the effect of p53 on ICAM-1 expression after DNA damage. Induction of ICAM-1 is abolished after treatment with the specific p53 inhibitor pifithrin-alpha and is abrogated in p53-deficient cell lines. Furthermore, we map two functional p53-responsive elements to the introns of the ICAM-1 gene, and show that they confer inducibility to p53 in a fashion similar to other p53 target genes. These results support an NF-kappaB-independent role for p53 in ICAM-1 regulation that may link p53 to ICAM-1 function in various physiological and pathological settings.

Galpha(q) and Gbetagamma regulate PAR-1 signaling of thrombin-induced NF-kappaB activation and ICAM-1 transcription in endothelial cells.

As thrombin binding to the G protein-coupled proteinase activated receptor-1 (PAR-1) induces endothelial adhesivity to leukocytes through NF-kappaB activation and intercellular adhesion molecule-1 (ICAM-1) expression, we determined the signaling pathways mediating the response. Studies showed that the heterotrimeric G proteins, Galpha(q), and the Gbetagamma dimer were key determinants of the PAR-1 agonist peptide (TFLLRNPNDK)-induced NF-kappaB activation and ICAM-1 expression in endothelial cells. Cotransfection of RGS3T, a regulator of G-protein signaling that inhibits Galpha(q), or alpha-transducin (Galpha(t)), a scavenger of the Gbetagamma, markedly decreased NF-kappaB activity induced by PAR-1 activation. We determined the downstream signaling targets activated by Galpha(q) and Gbetagamma that mediate NF-kappaB activation. Expression of the kinase-defective protein kinase C (PKC)-delta mutant inhibited NF-kappaB activation induced by the constitutively active Galpha(q) mutant, but had no effect on NF-kappaB activity induced by Gbeta(1)gamma(2). In related experiments, NF-kappaB as well as ICAM-1 promoter activation induced by Gbeta(1)gamma(2) were inhibited by the expression of the dominant-negative mutant of 85-kDa regulatory subunit of PI 3-kinase; however, the expression of this mutant had no effect on the response induced by activated Galpha(q). Cotransfection of the catalytically inactive Akt mutant inhibited the NF-kappaB activation induced by the constitutively active PI 3-kinase mutant as well as that by the activated forms of Galpha(q) and PKC-delta. These results support a model in which ligation of PAR-1 induces NF-kappaB activation and ICAM-1 transcription by the engagement of parallel Galphaq/PKC-delta and Gbetagamma/PI3-kinase pathways that converge at Akt.

Nitric Oxide Inhibits Endothelial IL-1β-induced ICAM-1 Gene Expression at the Transcriptional Level Decreasing Sp1 and AP-1 Activity

Nitric oxide (NO) has frequently been shown to inhibit leukocyte adherence to activated endothelium thus displaying anti-adhesive and immunosuppressive activities. A molecular mechanism contributing to this effect is described. Primary murine aortic endothelial cells were activated with interleukin (IL)-1beta to express intercellular adhesion molecule-1 (ICAM-1) mRNA in the presence or absence of the physiological spontaneous NO-donor S-nitrosocysteine. Subsequently, semiquantitative RT-PCR and gel shift assays with nuclear extracts were performed to analyse the effects of NO on ICAM-1 mRNA expression and on the activity of transcription factors involved in ICAM-1 transcription. In addition, luciferase reporter gene activity of cytokine-activated cells transiently transfected with an ICAM-1 promoter-luciferase construct and cultured in the presence of the slow-releasing NO-donor DETA/NO was determined. NO at subtoxic concentrations decreases IL-1beta-induced endothelial ICAM-1 mRNA expression. This inhibition occurs at the transcriptional level, as NO affects IL-1b-induced ICAM-1 promoter activity in transiently transfected cells. Using gel-shift assays and double-stranded oligonucleotide consensus sequences of the known transcription factor binding sites of the ICAM-1 promoter, Sp1 and AP-1 were identified as transcriptional activators of IL-1beta-driven ICAM-1 expression. The DNA binding of both of these transcription factors to specific binding sites of the ICAM-1 promoter was decreased in MAEC exposed to NO. Our studies indicate that the anti-adhesive effect of NO concentrations equivalent to high-output NO synthesis is mediated, at least in part, by inhibition of ICAM-1 expression via a concerted action of NO on the redox-sensitive transcriptional activators Sp1 and AP-1. This molecular mechanism may contribute to the anti-inflammatory actions of NO synthesized by the inducible NO synthase.

Expression of Intercellular Adhesion Molecule-1 in the Cortex of Preserved Rat Kidneys

Background. Prolonged cold ischemia has been shown to be an important factor in the development of posttransplant renal dysfunction. The exact mechanisms have not been completely defined. The expression of intercellular adhesion molecule-1 (ICAM-1) (CD 54) in rat kidneys stored in University of Wisconsin (UW) solution was studied in an attempt to correlate ischemia time with immunogenicity of the graft.Methods. Kidneys from male Lewis rats were perfused with UW solution, removed, and bathed in UW solution at 4°C for 4, 12, 24, and 48 h. For the evaluation of expression of ICAM-1, immunohistochemical staining, Western blotting, and semiquantitative reverse transcriptase–polymerase chain reaction (RT-PCR) were performed.Results. Immunohistochemical staining in normal, nonischemic kidneys revealed that glomerular capillaries expressed ICAM-1 but that tubular cells did not. The preserved kidneys were analyzed by immunohistochemistry, Western blotting, and semiquantitative RT-PCR and showed increased transcription and expression of ICAM-1 in the cortex of the kidney. Expression reached a maximum at 24 h and declined at 48 h. The ICAM-1 protein expression in the preserved kidney cortex relative to control kidneys was increased at 4 h (1.68 ± 0.60-fold of control kidneys, P = 0.06), 12 h (2.38 ± 0.90-fold, P = 0.02), 24 h (3.70 ± 1.29-fold, P = 0.01), and 48 h (2.00 ± 0.54-fold, P = 0.01). The messenger RNA expression (the ratio of ICAM-1 to glyceraldehyde-3-phosphate dehydrogenase) in preserved kidneys cortex relative to control kidneys was increased at 4 h (1.19 ± 0.14-fold of control kidneys), 12 h (1.38 ± 0.16-fold), 24 h (1.77 ± 0.29-fold), and 48 h (1.19 ± 0.12-fold) (P < 0.05 for all time points).Conclusions. We conclude that cold preservation of rat kidneys in UW solution induces increasing levels of ICAM-1 cell surface expression and gene transcription. Further study is necessary to determine if this increase in adhesion molecule expression increases the immunogenicity of the allograft and contributes to the development of posttransplant renal dysfunction.

Protein kinase C-zeta mediates TNF-alpha-induced ICAM-1 gene transcription in endothelial cells.

We addressed the role of protein kinase C (PKC) isozymes in mediating tumor necrosis factor-alpha (TNF-alpha)-induced oxidant generation in endothelial cells, a requirement for nuclear factor-kappaB (NF-kappaB) activation and intercellular adhesion molecule-1 (ICAM-1) gene transcription. Depletion of the conventional (c) and novel (n) PKC isozymes following 24 h exposure of human pulmonary artery endothelial (HPAE) cells with the phorbol ester, phorbol 12-myristate 13-acetate (500 nM), failed to prevent TNF-alpha-induced oxidant generation. In contrast, inhibition of PKC-zeta synthesis by the antisense oligonucleotide prevented the oxidant generation following the TNF-alpha stimulation. We observed that PKC-zeta also induced the TNF-alpha-induced NF-kappaB binding to the ICAM-1 promoter and the resultant ICAM-1 gene transcription. We showed that expression of the dominant negative mutant of PKC-zeta prevented the TNF-alpha-induced ICAM-1 promoter activity, whereas overexpression of the wild-type PKC-zeta augmented the response. These data imply a critical role for the PKC-zeta isozyme in regulating TNF-alpha-induced oxidant generation and in signaling the activation of NF-kappaB and ICAM-1 transcription in endothelial cells.

Thyrotropin modulates interferon-gamma-mediated intercellular adhesion molecule-1 gene expression by inhibiting Janus kinase-1 and signal transducer and activator of transcription-1 activation in thyroid cells.

TSH is known as an important hormone that plays the major role not only in the maintenance of normal physiology but also in the regulation of immunomodulatory gene expression in thyrocytes. The adhesion molecule intercellular adhesion molecule-1 (ICAM-1) was identified as one of the proteins that are abnormally expressed in the thyroid gland during autoimmune thyroid diseases. In this study we found that TSH inhibits interferon-gamma (IFNgamma)-mediated expression of the ICAM-1 gene, and we investigated the involved mechanisms in rat FRTL-5 thyroid cells. After exposure to IFNgamma, ICAM-1 expression is positively regulated at the level of transcription. This effect occurs via the IFNgamma-activated site (GAS) element in the ICAM-1 promoter as a consequence of the activation of STAT1 (signal transducer and activator of transcription-1), but not of STAT3. On the other hand, after exposure to TSH plus IFNgamma, ICAM-1 transcription is negatively modulated. We found that this inhibitory effect of TSH also occurs via the GAS element. Electrophoretic mobility shift assays confirmed that the IFNgamma-induced DNA-binding activities of STAT1 were reduced by TSH. Furthermore, our results showed that the inhibitory effect of TSH on IFNgamma signaling is caused by inhibition of tyrosine phosphorylation on STAT1, Janus kinase-1 (Jak1), and IFNgamma receptor a, but not Jak2. In conclusion, we have identified a novel mechanism in which TSH modulates the IFNgamma-mediated Jak/STAT signaling pathway through the inhibition of Jak1 and STAT1.

MRNA expression of intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 in acute renal allograft rejection.

The intercellular adhesion molecule-1 (ICAM-1) and the vascular cell adhesion molecule-1 (VCAM-1) show a form of complementary distribution in normal and grafted kidneys. The molecular mechanism by which ICAM-1 and VCAM-1 are increased or induced on vascular cells during acute renal allograft rejection has not been clearly defined. We examined ICAM-1 and VCAM-1 mRNA expression in 17 renal allograft biopsies with (n=12) and without (n=5) features of acute rejection, and four control renal biopsies with no detectable abnormalities by RNA in situ hybridization. The expression of ICAM-1 and VCAM-1 protein was also assessed by immunohistochemical staining of frozen sections. In controls and nonrejecting graft biopsies, the signals of the ICAM-1 and VCAM-1 transcripts in vascular cells were almost negligible. Specific signals of ICAM-1 and VCAM-1 mRNAs were detected on the endothelial cells of small muscular arteries in most cases with acute renal allograft rejection. The messages for ICAM-1 and VCAM-1 were also detected on arterial smooth muscle cells in all the five cases with severe type III rejection. These results suggest that the induced appearance of ICAM-1 and VCAM-1 on the vascular cells of acutely rejecting renal transplants was related to actual cellular synthesis and that both adhesion molecules could act together in the rejection process. They also suggest that the expression of ICAM-1 and VCAM-1 genes by arterial smooth muscle cells may be an important cause of transmural arteritis in severe acute renal allograft rejection.

26] Gene regulation by ultraviolet A radiation and singlet oxygen

Human skin is increasigly exposed to long-wave ultraviolet (UV) radiation that is UVA radiation in the range of 340-400 nm. Sources for UVA radiation exposure are solar radiation and artificial sources used for cosmetic or phototherapeutic purposes. In order to understand the effects of UVA radiation on human skin, it is critical to study the mechanisms by which UVA radiation affects gene expression in human skin cells, particularly epidermal keratinocytes, which constitute the primary cellular target for UVA radiation. By focusing on the regulation of the human intercellular adhesion molecule-1 (ICAM-1) gene in primary human epidermal keratinocytes, it has been demonstrated that UVA radiation-induced gene expression depends critically on the activation of transcription factor AP-2. Interestingly, UVA radiation-induced AP-2 activation and subsequent induction of ICAM-1 transcription were found to be mediated through the generation of singlet oxygen, thus implying that UVA radiation and singlet oxygen-induced gene expression in human epithelial cells may share a common signal transduction pathway. These studies have been made possible by the availability of a variety of techniques that allow the study of gene expression in UVA-irradiated cultured human keratinocytes at different levels of expression, including transcription factor activation, mRNA, and protein expression under various conditions that allow the monitoring of the role of singlet oxygen in this system. These techniques and assay system are discussed along with their relevance for studying UVA radiation-induced gene expression in human keratinocytes.

Insulin like Growth Factor-1 Activates Nuclear Factor-κB and Increases Transcription of the Intercellular Adhesion Molecule-1 Gene in Endothelial Cells

A critical early event in the pathogenesis of occlusive vascular disease is the adhesion of monocytes to endothelial cells. The authors have previously reported that insulin-like growth factor-1 increases monocyte-endothelial cell adhesion and increases the expression of intercellular adhesion molecule-1. In this study, it is hypothesized that the upregulation of intercellular adhesion molecule-1 expression after treatment with insulin-like growth factor-1 is caused by an increase in the transcription of intercellular adhesion molecule-1 in endothelial cells, and that this transcription is regulated, at least in part, by activation of nuclear factor-κB. Adherence cell assays were performed using insulin-like growth factor-1 treated human umbilical vein endothelial cells and human monocytes. To determine the role of nuclear factor-κB. Western blotting using the anti-p65 (activated portion of nuclear factor-κB) was performed on cell lysate of human umbilical vein endothelial cells treated with insulin-like growth factor-1. RT-PCR was performed on RNA extracted from insulin-like growth factor-1-treated human umbilical vein endothelial cells. Intercellular adhesion molecule-1 antibody attenuated the increase in monocyte-endothelial cell adhesion of endothelial cells exposed to insulin-like growth factor-1. We observed an increase in expression of the activated nuclear factor-κB p65 protein in response to insulin-like growth factor-1 treatment. Peak increase occurred at 30 min. This effect was sensitive to pretreatment of human umbilical vein endothelial cells with the insulin-like growth factor-1 receptor antibody. Human umbilical vein endothelial cells treated with insulin-like growth factor-1 for 2 and 4 h revealed a significant increase in intercellular adhesion molecule-1 mRNA as compared with untreated human umbilical vein endothelial cells. Tumor necrosis factor-α produced a larger increase in intercellular adhesion molecule-1 mRNA expression. These results suggest that insulin-like growth factor-1 enhances intercellular adhesion molecule-1 transcription and activates nuclear factor-κB in endothelial cells. The intracellular pathways that increase cell adhesion molecule expression may provide a common link to understanding the monocyte-endothelial cell adhesion that occurs in the early stages of atherosclerosis and restenosis.

Insulin like growth factor-1 activates nuclear factor- κB and increases transcription of the intercellular adhesion molecule-1 gene in endothelial cells

A critical early event in the pathogenesis of occlusive vascular disease is the adhesion of monocytes to endothelial cells. The authors have previously reported that insulin-like growth factor-1 increases monocyte-endothelial cell adhesion and increases the expression of intercellular adhesion molecule-1. In this study, it is hypothesized that the upregulation of intercellular adhesion molecule-1 expression after treatment with insulin-like growth factor-1 is caused by an increase in the transcription of intercellular adhesion molecule-1 in endothelial cells, and that this transcription is regulated, at least in part, by activation of nuclear factor- κB. Adherence cell assays were performed using insulin-like growth factor-1 treated human umbilical vein endothelial cells and human monocytes. To determine the role of nuclear factor- κB, Western blotting using the anti-p65 (activated portion of nuclear factor- κB) was performed on cell lysate of human umbilical vein endothelial cells treated with insulin-like growth factor-1. RT-PCR was performed on RNA extracted from insulin-like growth factor-1-treated human umbilical vein endothelial cells. Intercellular adhesion molecule-1 antibody attenuated the increase in monocyte-endothelial cell adhesion of endothelial cells exposed to insulin-like growth factor-1. We observed an increase in expression of the activated nuclear factor- κB p65 protein in response to insulin-like growth factor-1 treatment. Peak increase occurred at 30 min. This effect was sensitive to pretreatment of human umbilical vein endothelial cells with the insulin-like growth factor-1 receptor antibody. Human umbilical vein endothelial cells treated with insulin-like growth factor-1 for 2 and 4 h revealed a significant increase in intercellular adhesion molecule-1 mRNA as compared with untreated human umbilical vein endothelial cells. Tumor necrosis factor- α produced a larger increase in intercellular adhesion molecule-1 mRNA expression. These results suggest that insulin-like growth factor-1 enhances intercellular adhesion molecule-1 transcription and activates nuclear factor- κB in endothelial cells. The intracellular pathways that increase cell adhesion molecule expression may provide a common link to understanding the monocyte-endothelial cell adhesion that occurs in the early stages of atherosclerosis and restenosis.

NF-κB/Rel Family Members Regulating the ICAM-1 Promoter in Monocytic THP-1 Cells

A κB-site was identified in the promoter of the intercellular adhesion molecule-1 (ICAM-1)gene, which is involved in regulation of ICAM-1 expression by tumor necrosis factor α (TNF-α) and glucocorticoids. We now report on the transcription factors which bind and transactivate this enhancer sequence. In vitro, the ICAM-1 κB site appeared to bind RelA and c-Rel homodimers as well as heterodimers with NF-κB1, but weakly NF-κB1 homodimers. In addition, both RelA and c-Rel, but not NF-κB1, were shown to transactivate an ICAM-1 κB-reporter construct. In monocytic THP-1 cells TNF-α induced two nuclear complexes which in vitro bound to the ICAM-1 κB site. Using antibodies in an electrophoretic mobility supershift assay, one of these complexes was shown to contain NF-κB1 and RelA, and to bind with higher affinity to the consensus κB site in the HIV long terminal repeat. The second complex contained RelA, and exhibited higher affinity towards the ICAM-1 κB than to the HIV-κB site. The glucocorticoid receptor was shown to repress activity of both the RelA homodimer and the NF-κB1/RelA heterodimer. We argue that in vivo RelA homodimers are likely to play a dominant role in TNF-α-induced ICAM-1 transcription in monocytic cells.

Hypoxia induces intercellular adhesion molecule-1 on cultured human tubular cells

The adverse effects of acute renal ischemia are partly mediated through an infiltration of inflammatory cells into the tubulointerstitium. The expression of adhesion molecules such as intercellular adhesion molecule-1 (ICAM-1) by resident renal cells (endothelial cells and tubular cells) may facilitate this process. We investigated whether hypoxia stimulates the expression of ICAM-1 by cultured human proximal tubular cells (HPTC). Hypoxic culture conditions (PO2 < 4 kPa) stimulated the expression of ICAM-1 by HPTC in a time-dependent manner (P < 0.0001) as demonstrated by quantitative flow cytometry analysis. Quantitative PCR demonstrated an increase in ICAM-1 transcription. Re-oxygenation of tubular cells did not increase ICAM-1 expression further. TNF alpha concentration in culture supernatants increased with hypoxia, but blocking experiments demonstrated that TNF alpha was not implicated in hypoxia-induced expression of ICAM-1. Furthermore, the cytokines IL-6 and IL-1 beta were not involved, but the effect of hypoxia was blocked by PDTC, an antioxidant that may inhibit the activation of the transcription factor NF-kappa B. These data demonstrate that hypoxia is a stimulus that induces the synthesis and expression of the adhesion molecule ICAM-1, presumably via the activation of NF-kappa B.

H2O2 and Tumor Necrosis Factor-α Activate Intercellular Adhesion Molecule 1 (ICAM-1) Gene Transcription through Distinct cis-Regulatory Elements within the ICAM-1 Promoter

We investigated the mechanisms by which H2O2 increases intercellular adhesion molecule 1 (ICAM-1; CD54) expression in endothelial cells. The H2O2-induced increase in ICAM-1 mRNA was inhibited by actinomycin D, by the antioxidant N-acetylcysteine, and by 3-amino-benzamide (which blocks oxidant-induced AP-1 activity), but not by pyrrolidine dithiocarbamate (which blocks oxidant-induced NF-kappa B activity). Nuclear run-on and transient transfections of ICAM-1 promoter constructs indicated that H2O2 stimulated ICAM-1 gene transcription by activation of a distinct region of the ICAM-1 promoter. The H2O2-responsive element was localized to sequences between -981 and -769 (relative to start codon). Located within this region are two 16-base pair repeats, each containing binding sites for the transcription factors AP-1 and Ets. A similar composite AP-1/Ets element isolated from the macrophage scavenger receptor gene conferred H2O2 responsiveness to a minimal promoter. Mutation of the 16-base pair repeats within the ICAM-1 promoter prevented H2O2-induced DNA binding activity, and their deletion abrogated the H2O2-induced transcriptional activity. In contrast, TNF alpha induced ICAM-1 transcription via activation of promoter sequences between -393 and -176, a region with C/EBP and NF-kappa B binding sites. The results indicate that H2O2 activates ICAM-1 transcription through AP-1/Ets elements within the ICAM-1 promoter, which are distinct from NF-kappa B-mediated ICAM-1 expression induced by TNF alpha.

Expression of Intercellular Adhesion Molecule-1 Is Regulated by the Actin Network in Epidermoid Carcinoma Cells

Expression of the cell surface protein intercellular adhesion molecule-1 (ICAM-1) is a prerequisite for the interaction of a large variety of cells with leukocytes. Constitutive expression of ICAM-1 in human epidermoid carcinoma cells is low, but is inducible through inflammatory cytokines including interferon γ (INFγ). Disruption of the actin cytoskeleton with dehydrocytochalasin B (CB) increased constitutive and potentiated INFγ-induced ICAM-1 cell surface expression, but did not alter formation of soluble ICAM-1. Actinomycin D inhibited CB-induced ICAM-1 surface expression and CB increased the steady-state levels of ICAM-1 transcripts. However, CB did not alter the glyceralaldehyde-3-phosphate dehydrogenase mRNA levels and the ICAM-1 mRNA half-life. These studies indicate that in human epidermoid carcinoma cells the actin cytoskeleton regulates ICAM-1 transcription and cell surface expression.

Regulatory elements and transcription factors controlling basal and cytokine-induced expression of the gene encoding intercellular adhesion molecule 1.

The gene encoding intercellular adhesion molecule 1 (ICAM-1) is transcriptionally induced in response to inflammatory and immunomodulatory cytokines. To investigate the mechanisms controlling ICAM-1 gene expression, we have identified regulatory DNA sequences responsible for maintaining basal and mediating induced transcription in response to tumor necrosis factor alpha (TNF-alpha) and interferon gamma (IFN-gamma). Regulatory elements centered 115, 60, and 40 bp upstream from the ICAM-1 transcription start site were implicated in cytokine-independent gene expression. Regulatory elements dedicated to TNF-alpha and IFN-gamma were identified 190 and 90 bp, respectively, upstream from the ICAM-1 transcription start site. A combination of mutagenesis and DNA-binding assays revealed that the TNF-alpha response element is composite, consisting of binding sites for both C/EBP and NF-kappa B. The IFN-gamma response element behaved as a simple regulatory element that selectively binds to an IFN-gamma-inducible activity composed, at least in part, of p91. These observations provide a framework for understanding how extracellular signals dynamically regulate the adhesive properties of mammalian cells.

Up-regulation of intercellular adhesion molecule 1 transcription by hepatitis B virus X protein.

Intercellular adhesion molecule 1 (ICAM-1), a counter-receptor for lymphocyte function-associated antigen 1 on T cells, is critically important to a wide variety of adhesion-dependent leukocyte functions, including antigen presentation and target cell lysis. ICAM-1 expression by hepatocytes is increased in areas of inflammation and necrosis during chronic hepatitis B. Whether induction of ICAM-1 is due to the effect of inflammatory cytokines or involves a direct effect of the hepatitis B virus (HBV) remains unknown. In the present study, transfection of the HBV genome into human hepatoma cell lines resulted in enhanced expression of ICAM-1 protein and RNA in the absence of inflammation. Results of subgenomic transfections indicated that the HBV X protein (pX) induced ICAM-1 expression. Nuclear run-on assays showed that pX induced the ICAM-1 gene by increasing its rate of transcription. Although both pX and interferon gamma induced transcription of ICAM-1, addition of interferon gamma to cells expressing pX did not show an additive or synergistic effect. These results indicate that pX can directly regulate expression of ICAM-1 and may participate in the immunopathogenesis of HBV infection.