Intercellular Adhesion Molecule-1 mRNA Levels Research Articles

Induction of intercellular adhesion molecule-1 by water-soluble components of Hericium erinaceum in human monocytes

Hericium erinaceum is a medicinal mushroom that has been traditionally used in Asian countries for the treatment of cancers and infectious diseases. Although the immunomodulating activity of H. erinaceum is considered to be responsible for its medicinal activity, its action mechanisms are poorly understood. In the present study, we investigated the capability of water-extracted H. erinaceum (WEHE) to induce the expression of intercellular adhesion molecule-1 (ICAM-1), which regulates the migration of immune cells. THP-1, a human monocytic cell-line, or human peripheral blood mononuclear cells (PBMC) were stimulated with WEHE (0-30 μg/mL) and subsequently analyzed using flow cytometry to examine the surface expression of ICAM-1 protein. Steady-state levels of ICAM-1 mRNA were estimated using real-time reverse transcription-polymerase chain reaction analysis. Electrophoretic mobility shift assay was conducted to examine transcription factors involved in ICAM-1 transcription. WEHE induced ICAM-1 expression at both protein and mRNA levels in THP-1 cells in a dose- and time-dependent fashion. A similar pattern of ICAM-1 induction was also observed in CD14(+) monocytes in human PBMC that were stimulated with WEHE. The ICAM-1 expression on THP-1 cells stimulated with WEHE was suppressed by specific inhibitors for extracellular signal-regulated kinases (ERK) and reactive oxygen species (ROS). Additionally, exposure of THP-1 cells to WEHE increased the DNA binding activities of NF-κB, AP-1, SP-1 and STAT-1 transcription factors, all of which are known to be required for ICAM-1 gene expression. These results suggest that WEHE induces ICAM-1 expression in human monocytes through ERK- and ROS-dependent signaling pathways, resulting in the subsequent activations of NF-κB, AP-1, SP-1, and STAT-1 transcription factors.

Glycyrrhetinic acid inhibits ICAM-1 expression via blocking JNK and NF-κB pathways in TNF-α-activated endothelial cells

To investigate the effects of glycyrrhetinic acid (GA), an active component extracted from the root of Glycyrrhizae glabra, on the expression of intercellular adhesion molecule-1 (ICAM-1) in tumor necrosis factor-alpha (TNF-alpha)-activated human umbilical vein endothelial cells (HUVEC). ICAM-1 mRNA and protein levels were detected using RT-PCR and cell enzyme-linked immunosorbent assays. The adherence of human monocytic THP-1 cells labeled with [(3)H]thymidine to HUVEC was determined by counting radioactivity with a scintillation counter. The activation of mitogen-activated protein kinases as well as the degradation of I kappaB and nuclear factor-kappaB (NF-kappaB) or phospho-c-Jun in the nucleus were detected by western blots. NF-kappaB binding activity was detected using electrophoretic mobility shift assay. GA (50 and 100 micromol/L) significantly inhibits TNF-alpha-induced ICAM-1 mRNA and protein expressions, as well as THP-1 cell adhesiveness in HUVEC. GA selectively inhibited TNF-alpha-activated signal pathway of c-Jun N-terminal kinase (JNK), without affecting extracellular signal-regulated kinase 1/2 and p38. Furthermore, GA apparently inhibited I kappaB/NF-kappaB signaling system by preventing I kappaB degradation, NF-kappaB translocation, and NF-kappaB/DNA binding activity. Finally, pretreatment with GA or the inhibitors of NF-kappaB, JNK, and p38 reduced the ICAM-1 protein expression induced by TNF-alpha. GA inhibits TNF-alpha-stimulated ICAM-1 expression, leading to a decrease in adherent monocytes to HUVEC. This inhibition is attributed to GA interruption of both JNK/c-Jun and I kappaB/NF-kappaB signaling pathways, which decrease activator protein-1 (AP-1) and NF-kappaB mediated ICAM-1 expressions. The results suggest that GA may provide a beneficial effect in treating vascular diseases associated with inflammation, such as atherosclerosis.

The Expression of Intercellular Adhesion Molecule-1 Induced by CD40-CD40L Ligand Signaling in Orbital Fibroblasts in Patients with Graves' Ophthalmopathy

The aim of the present study was to examine the effect of CD40 ligand (CD40L) on intercellular adhesion molecule-1 (ICAM-1) production and involvement of mitogen-activated protein kinases (MAPKs) and nuclear factor-kappaB (NF-kappaB) signaling pathways by CD40-CD40L ligand in orbital fibroblasts (OFs) in patients with Graves' ophthalmopathy (GO). OFs were stimulated with soluble CD40L, and conditioned media and lysed cells were subjected to Western blot and real-time quantitative polymerase chain reaction analyses. Inhibitors specific to various signal transduction pathways were used to determine the signal pathways involved. ICAM-1 protein and mRNA in OFs of GO groups were upregulated by CD40L compared with those in normal OFs. Treatment of OFs with CD40L increased ICAM-1 mRNA levels in a dose- and time-dependent manner. CD40L induced the phosphorylation of p38 MAPK, extracellular-regulated kinase1/2 (ERK1/2), c-Jun NH2-terminal kinase (JNK), and IkappaB and the activation of NF-kappaB. Inhibition of the ERK1/2, p38, JNK, and NF-kappaB pathways blocked CD40L-induced ICAM-1 secretion. Furthermore, the activation of NF-kappaB was significantly affected by ERK1/2 and JNK inhibitors, but not by p38. OF ICAM-1 expression was predominantly p38 MAPK and NF-kappaB dependent. ERK1/2 and JNK were implicated in the NF-kappaB pathway. Analyses of ICAM-1 mRNA synthesis revealed that CD40L-induced ICAM-1 expression was mediated by multiple factors. CD40L can potently induce ICAM-1 expression in OF cells through multiple signal pathways. The p38 MAPKs and NF-kappaB pathways may play an important permissive role in CD40L-induced ICAM-1 expression in OFs.

SO2 Inhalation Contributes to the Development and Progression of Ischemic Stroke in the Brain

Epidemiological literatures show an association between air pollution and ischemic stroke, and effective pollutants may include SO(2), NO(x), O(3), CO, and particulates. However, existing experimental studies lack evidence as to the presence of effects for SO(2), which has been the focus in developing countries with increasing use of coal as the main resource. In the present study, we treated Wistar rats with SO(2) at various concentrations and determined endothelin-1 (ET-1), inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and intercellular adhesion molecule 1 (ICAM-1) messenger RNA (mRNA) and protein expression in the cortex. The results show that SO(2) elevated the levels of ET-1, iNOS, COX-2, and ICAM-1 mRNA and protein in a concentration-dependent manner. Then, we set up rat model of ischemic stroke using middle cerebral artery occlusion (MCAO) and further treated the model rats with filtered air and lower concentration SO(2) for the same period. As expected, elevated expression of ET-1, iNOS, COX-2, and ICAM-1 occurred in the cortex of MCAO model rats exposed to filtered air, followed by increased activation of caspase-3 and cerebral infarct volume. Interestingly, SO(2) inhalation after MCAO significantly amplified above effects. It implies that SO(2) inhalation caused brain injuries similar to that of cerebral ischemia, and its exposure in atmospheric environment contributed to the development and progression of ischemic stroke.

Mechanisms of Toll-Like Receptor 4 (TLR4)-Mediated Inflammation After Cold Ischemia/Reperfusion in the Heart

Toll-Like Receptor 4 (TLR4) signaling mediates early inflammation after cold ischemia-reperfusion (I/R). We hypothesized that the TLR4 coreceptor CD14, the intracellular adaptor proteins myeloid differentiation factor 88 (MyD88) and TIR domain-containing-adaptor inducing IFNbeta (TRIF) would be required for cold I/R induced inflammation. High mobility group box 1 (HMGB1) is a putative endogenous activator of TLR4. Therefore, we also assessed the contribution of HMGB1 in cold I/R induced inflammation. Syngeneic heart transplants were performed in mice deficient in CD14, MyD88, TRIF, or wild-type mice. In other experiments, anti-HMGB1 neutralizing antibody or control IgG was administered at reperfusion. Donor hearts were subjected to 2 hr of cold ischemia and retrieved after 3 hr of reperfusion. After cold I/R, grafts revealed striking translocation of HMGB1 out of the nucleus in cardiac myocytes. Administration of an anti-HMGB1 neutralizing antibody resulted in reduced systemic interleukin (IL)-6, tumor necrosis factor alpha (TNFalpha), and intercellular adhesion molecule-1 (ICAM-1) messenger RNA (mRNA) levels (P< or =0.05). Compared with controls, CD14 knock-out (KO) mice exhibited significantly lower (P< or =0.05) systemic IL-6 and JE/monocyte chemotractant protein-1 levels after cold I/R. Intragraft TNFalpha and IL-1beta mRNA levels were also significantly lower (P< or =0.05) in CD14 KO grafts. MyD88 KO mice exhibited significantly lower (P< or=0.05) systemic IL-6 levels compared with control mice after cold I/R. Intragraft TNFalpha, IL-6, and ICAM-1 mRNA levels were also significantly lower (P< or =0.05) in MyD88 KO grafts. Significantly lower levels (P< or =0.05) of serum IL-6, monocyte chemotractant protein-1 as well as intragraft TNFalpha, IL-6, IL-1beta, and ICAM-1 were observed after cold I/R in TRIF deficient animals compared with controls. CD14, MyD88, TRIF, and HMGB1 contribute to the inflammatory response that occurs after cold I/R. These results provide insight into the mechanisms of TLR4-mediated inflammation after cold I/R.

Stasis Predisposes Ileal Pouch Inflammation in a Rat Model of Ileal Pouch-Anal Anastomosis

While restorative proctocolectomy with ileal pouch-anal anastomosis (IPAA) has become the definitive surgical treatment for patients suffering from chronic ulcerative colitis (CUC), pouchitis still remains a major late complication. Fecal stasis has been implicated in the etiology of ileal inflammation; however, the mechanism(s) remain unclear, in part due to the lack of an animal model. Our goal was to surgically mimic the IPAA procedure in a rat to investigate the hypothesis that stasis leads to biochemical changes that predispose the ileal pouch to inflammation. Thirty-two Sprague-Dawley rats underwent total colectomy with either straight ileorectal (IRA) or IPAA, and 11 nonoperated rats served as controls (Controls). Twenty-one d postoperatively, 48 h serial barium radiographs and 12 h charcoal transit follow-through studies were performed. Following sacrifice, ileal tissue was harvested for the measurement of myeloperoxidase activity (MPO) activity, intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1) mRNA levels, and histology. Serial barium radiographs showed stasis in the ileal pouch compared with IRA animals, and charcoal transit times that were two times longer (P ≤ 0.05) than that in the straight IRA rats. Ileal pouch MPO levels were significantly elevated in the IPAA rats compared with the straight IRA rats. ICAM-1 and VCAM-1 mRNA levels were not associated with neutrophil infiltration. These studies showed that ileal pouch stasis predisposes biochemical and histological evidence of ileal pouch mucosal inflammation. Studies such as this may provide the rationale for novel, adjunct therapies for the management of pouchitis in patients having undergone IPAA for CUC.

Positive effects of the middle layer of Phyllostachys nigra var. henonis (PN), Bambusae Calulis in Taeniam, on asthma murine model through down-regulated ICAM-1 of BMMC (36.12)

Abstract The middle layer of Phyllostachys nigra var. henonis (PN), Bambusae Calulis in Taeniam, has been used for cough and sputum for a long time. Cough is one of the major symptoms of asthma. However, there were few of scientific investigation about PN. To investigate the positive effect of PN on asthma, we measured the production of IL-4 of bronchoalveolar lavage fluid (BalF), Immunoglobulin E (IgE) level of serum and immune cell infiltration at lung tissue by HE stain in ovalbumin (OVA) induced asthma murine model (Balb/C, female, 6 weeks). In asthma model, there was IL-4 of BalF decreased by PN, however, it was not significantly. The IgE induced by OVA in serum significantly decreased by PN (40 mg/kg). Furthermore, we observed the decrease of immune cell infiltration up-regulated by OVA in lung tissue. We focused on decrease of cell infiltration by PN. Intercellular adhesion molecule-1 (ICAM-1) and L-selectin regulate IgE production by controlling mast cell accumulation at sites of inflammation. And then we investigated the mRNA and protein levels of ICAM-1 and L-selectin in bone marrow mast cells (BMMC). BMMC were incubated with 10 ng/ml of TNF-alpha fnand IL-3 plus 1, 10 and 100 £gg/ml of PN for 4 hours. The mRNA and protein level were measeured by using RT-PCR and Fluorescence activated cell sorter (FACS). PN had down-regulated both mRNA and protein of ICAM-1, however, no difference at mRNA and protein of L-selectin in BMMC. Together all things, PN has anti allergy effects on asthma, based on down-regulation of ICAM-1. Acknowledgements: This study was supported by a grant from Seoul R&amp;BD Program, Republic of Korea.

INSULIN REGULATES CYTOKINES AND INTERCELLULAR ADHESION MOLECULE-1 GENE EXPRESSION THROUGH NUCLEAR FACTOR-κB ACTIVATION IN LPS-INDUCED ACUTE LUNG INJURY IN RATS

Diabetic patients have increased susceptibility to infection, which may be related to impaired inflammatory response observed in experimental models of diabetes, and restored by insulin treatment. The goal of this study was to investigate whether insulin regulates transcription of cytokines and intercellular adhesion molecule 1 (ICAM-1) via nuclear factor-kappaB (NF-kappaB) signaling pathway in Escherichia coli LPS-induced lung inflammation. Diabetic male Wistar rats (alloxan, 42 mg/kg, i.v., 10 days) and controls were instilled intratracheally with saline containing LPS (750 microg/0.4 mL) or saline only. Some diabetic rats were given neutral protamine Hagedorn insulin (4 IU, s.c.) 2 h before LPS. Analyses performed 6 h after LPS included: (a) lung and mesenteric lymph node IL-1 beta, TNF-alpha, IL-10, and ICAM-1 messenger RNA (mRNA) were quantified by real-time reverse transcriptase-polymerase chain reaction; (b) number of neutrophils in the bronchoalveolar lavage (BAL) fluid, and concentrations of IL-1 beta, TNF-alpha, and IL-10 in the BAL were determined by the enzyme-linked immunosorbent assay; and (c) activation of NF-kappaB p65 subunit and phosphorylation of I-kappaB alpha were quantified by Western blot analysis. Relative to controls, diabetic rats exhibited a reduction in lung and mesenteric lymph node IL-1 beta (40%), TNF-alpha (approximately 30%), and IL-10 (approximately 40%) mRNA levels and reduced concentrations of IL-1 beta (52%), TNF-alpha (62%), IL-10 (43%), and neutrophil counts (72%) in the BAL. Activation of NF-kappaB p65 subunit and phosphorylation of I-kappaB alpha were almost suppressed in diabetic rats. Treatment of diabetic rats with insulin completely restored mRNA and protein levels of these cytokines and potentiated lung ICAM-1 mRNA levels (30%) and number of neutrophils (72%) in the BAL. Activation of NF-kappaB p65 subunit and phosphorylation of I-kappaB alpha were partially restored by insulin treatment. In conclusion, data presented suggest that insulin regulates transcription of proinflammatory (IL-1 beta, TNF-alpha) and anti-inflammatory (IL-10) cytokines, and expression of ICAM-1 via the NF-kappaB signaling pathway.

Effect of activation of nuclear factor-KappaB on modulation of secretion of intercellular adhesion molecule-1 by A549 cell

To investigate the effects of interleukin-1beta (IL-1beta) on intercellular adhesion molecule-1 (ICAM-1) expression on A549 cell and underlying signal transduction pathways. A549 cells were pre-incubated with SC-514 [nuclear factor-KappaB (NF-KappaB) inhibitor (IKappaB) kinase-2 (IKK-2) inhibitor] and/or pre-treated with 1 microg/L IL-1beta. The phosphorylated IKappaBalpha (pIKappaBalpha) and degradation of IKappaBalpha were determined by Western blotting with specific antibody at 5, 10, 30 and 60 minutes. Laser scanning confocal microscope (LSCM) was used to examine the nuclear translocation of p65 at 30 minutes after stimulation. The DNA binding activity of p65 in nuclear extracts was detected at 1 hour following IL-1beta treatment. Chromatin immunoprecipitation (ChIP) assays combined with polymerase chain reaction (PCR) were used to evaluate interaction between p65 and ICAM-1 promoter site DNA at 1 hour after stimulation. The expression of ICAM-1 mRNA was assessed by reverse transcription (RT)-PCR at 4 hours, and the ICAM-1 expression on A549 cell surface was measured by enzyme linked immunosorbent assay (ELISA) at 24 hours after IL-1beta was added. IL-1beta induced rapid pIKappaBalpha augmentation and its subsequent degradation. LSCM graphs showed that IL-1beta stimulated the translocation of p65 from the cytosol to the nucleus. IL-1beta significantly increased the DNA binding ability of p65 (P<0.01) in cell nuclear extracts. ChIP-PCR suggested that both acetylated histone 4 and p65 were recruited to ICAM-1 promoter. IL-1beta significantly augmented ICAM-1 mRNA level at 4 hours and expression of ICAM-1 on A549 cell surface at 24 hours (both P<0.01). The IKK-2 inhibitor, SC-514, inhibited IL-1beta induced IKappaBalpha protein activity, blocked p65 nuclear translocation, caused a significant reduction in IL-1beta induced DNA binding activity for p65 and ICAM-1 mRNA expression, and suppressed ICAM-1 expression on A549 cell surface (all P<0.01). These results suggest that the activation of NF-KappaB mediates IL-1beta induced ICAM-1 expression in A549 cells.

The gene expression of TNF-&amp;alpha; and ICAM-1 as novel markers for atherosclerosis in Egyptian hemodialysis patients

Atherosclerosis is an inflammatory vascular disorder frequently associated with renal disease and considered one of the major causes of death in patients with End Stage Renal Disease (ESRD). An early phase of atherosclerosis involves recruitment of inflammatory cells from circulation by transendothelial migration. This process is predominantly mediated by cellular adhesion molecules which are expressed on the vascular endothelium and circulating leukocytes in response to several inflammatory stimuli. The goal of this study is to evaluate the prognostic value of the Tumour Necrosis Factor-α (TNF-α) and Intercellular Adhesion Molecule-1 (ICAM-1) in Hemodialysis (HD) patients by means of the quantitative determination of ICAM-1 and TNF-α mRNA levels using Real-Time Polymerase Chain Reaction (PCR) (RT-PCR), where specific primers for each were used. Meanwhile, the levels of both ICAM-1 and TNF-α mRNA gene expressions in the peripheral blood of the studied groups were correlated with the other biochemical parameters of atherosclerosis in HD patients. Statistical analysis of the data revealed a highly significant increase in both ICAM-1 and TNF-a mRNA expressions in HD patients as compared to the healthy controls. Moreover, both TNF-α and ICAM-1 showed a positive correlation with triglycerides and Very Low Density Lipoprotein (VLDL). There was also a strong positive correlation between TNF-α and ICAM-1.

Impact of rapamycin on liver regeneration

The remarkable capacity of the liver to regenerate after injury and the prospects of organ self-renewal have attracted much interest in the understanding and modulation of the underlying molecular events. We investigated the effect of mammalian target of rapamycin (mTOR) inhibitor rapamycin (RAPA) on liver by correlating intravital microscopy, immunohistochemistry, and reverse transcriptase polymerase chain reaction in a rat model of 2/3 hepatectomy. RAPA significantly retarded proliferation of hepatocytes, endothelial cells, and hepatic stellate cells (HSCs) mostly between days 2 and 4 after hepatectomy and downregulated major cytokines and growth factors (tumor necrosis factor alpha, hepatocyte growth factor, platelet-derived growth factor, platelet-derived growth factor receptor, insulin-like growth factor-1, transforming growth factor beta 1) important for liver regeneration. These effects were almost absent at later time points. RAPA also had a transient, but broad effect on angiogenesis, and impaired sinusoidal density as well as mRNA levels of vascular endothelial growth factor, vascular endothelial growth factor receptor 1, vascular endothelial growth factor receptor 2, and angiopoietin-1. Activation of HSC was also transiently suppressed as observed by smooth muscle protein 1 alpha protein expression and intercellular adhesion molecule-1 mRNA levels. The rate of apoptosis in liver was significantly increased by RAPA between day 3 and day 7. The effect of RAPA on liver repair, angiogenesis, and HSC activation is confined to the phase of active cell proliferation. This transient effect might allow further exploration of mTOR inhibitors in clinical situations that involve liver regeneration, and seems to have implications beyond immunosuppression.

Dexamethasone Inhibits Leukocyte Accumulation and Vascular Permeability in Retina of Streptozotocin-Induced Diabetic Rats via Reducing Vascular Endothelial Growth Factor and Intercellular Adhesion Molecule-1 Expression

Intravitreal injection of corticosteroid has been used to treat diabetic macular edema, however, the exact mechanism remains unknown. In the present experiment, four weeks after streptozotocin administration, intravitreal injection of dexamethasone (50 microg/10 microl) was performed. After 2 d injection, we investigated the effect of dexamethasone on leukocyte accumulation, vascular permeability and the expression of vascular endothelial growth factor (VEGF) and intercellular adhesion molecule-1 (ICAM-1) in streptozotocin-diabetic rats. Accumulated leukocytes were counted in vivo by acridine orange leukocyte fluorography, the retinal vascular permeability was measured by the Evans blue assay. The mRNA and protein level of VEGF and ICAM-1 were analyzed with real-time quantitative polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA) respectively. Dexamethasone downregulated VEGF and ICAM-1 expression in diabetic rats which correlated with its effect on leukocytes accumulation and retinal vascular permeability. The present data revealed that dexamethasone may inhibit retinal accumulation and leukostasis accumulation and vascular permeability through its blockage on VEGF and ICAM-1 expression.

Peroxisome proliferator‐activated receptor‐γ agonists inhibit respiratory syncytial virus‐induced expression of intercellular adhesion molecule‐1 in human lung epithelial cells

Respiratory syncytial virus (RSV) is the major causative agent of severe lower respiratory tract disease and death in infants worldwide. The epithelial cells of the airways are the target cells for RSV infection and the site of the majority of the inflammation associated with the disease. However, despite five decades of intensive RSV research there exist neither an effective active vaccine nor a promising antiviral and anti-inflammatory therapy. Recently, peroxisome proliferator-activated receptor-gamma (PPAR-gamma), a member of the nuclear hormone receptor superfamily, has been shown to possess anti-inflammatory properties. Therefore, we hypothesized whether the detrimental increase of intercellular adhesion molecule-1 (ICAM-1) on RSV-infected lung epithelial cells (A549 and primary normal human bronchial epithelial cells (NHBE)) might be modulated by natural and synthetic PPAR-gamma agonists (15d-PGJ2, ciglitazone, troglitazone, Fmoc-Leu). Our data show that all PPAR-gamma agonists under study significantly down-regulated the RSV-induced expression of ICAM-1 on A549- and NHBE cells in a dose-dependent manner resulting in a reduced beta2 integrin-mediated adhesion of monocytic effector cells (U937) to RSV-infected A549 cell monolayers. In contrast, the PPAR-alpha agonist bezafibrate had no impact on the RSV-induced ICAM-1 expression. The reduced ICAM-1 expression was associated with a diminished ICAM-1 mRNA level and binding activity of nuclear factor-kappaB (p65/p50) in A549 cells. These findings suggest that PPARgamma agonists have beneficial effects in the suppression of the inflammatory response during RSV infection and therefore might have clinical efficacy in the course of severe RSV-infection.

Increased peritoneal and endometrial gene expression of biologically relevant cytokines and growth factors during the menstrual phase in women with endometriosis

To examine differential messenger RNA (mRNA) expression of relevant cytokines, metalloproteases, growth and adhesion factors in endometrium and peritoneum from women with endometriosis when compared with women without the disease during menstrual and luteal phases of the cycle. Patients with endometriosis were compared with control patients. University hospital. A total of 35 patients (20 patients during the luteal phase and 15 patients during the menstrual phase) were selected for this study on the basis of cycle phase and presence or absence of endometriosis. In this study, endometriosis was laparoscopically and histologically confirmed in 24 women with endometriosis of revised American Society for Reproductive Medicine (ASRM) stage I-II (n = 12) and revised ASRM stage III-IV (n = 12), and the presence of a normal pelvis was documented by laparoscopy in 11 control patients. The macroscopically normal peritoneum tissues were collected from lateral wall left or right, near the colon ascendens or descendens. The expression levels were determined as ratios between the target molecules and beta-actin as housekeeping gene. In women with endometriosis, peritoneal mRNA levels of matrix metalloproteinase (MMP)-3, transforming growth factor-beta, interleukin (IL)-6, and intercellular adhesion molecule-1 and endometrial mRNA levels of MMP-3, tumor necrosis factor (TNF)-alpha, and IL-8 were significantly higher during the menstrual phase when compared with luteal phase. During the menstrual phase of the cycle, both endometrial expression of TNF-alpha, IL-8, and MMP-3 mRNA levels and peritoneal expression of transforming growth factor-beta, IL-6, and intercellular adhesion molecule-1 mRNA levels were significantly higher in women with endometriosis when compared with controls. Immunohistochemical staining confirmed the presence of TNF-alpha in peritoneum and endometrium in both women with endometriosis and controls. Increased endometrial and peritoneal cytokine mRNA expression during menstruation may contribute to a pelvic inflammatory microenvironment favoring the development of endometriosis.

Time Course of Gene Expression of Inflammatory Mediators in Rat Lung after Diesel Exhaust Particle Exposure

Diesel exhaust particles (DEPs) at three concentrations (5, 35, and 50 mg/kg body weight) were instilled into rats intratracheally. We studied gene expression at 1, 7, and 30 days postexposure in cells obtained by bronchoalveolar lavage (BAL) and in lung tissue. Using real-time reverse transcriptase-polymerase chain reaction (RT-PCR), we measured the mRNA levels of eight genes [interleukin (IL)-1β, IL-6, IL-10, iNOS (inducible nitric oxide synthase), MCP-1 (monocyte chemoattractant protein-1), MIP-2 (macrophage inflammatory protein-2), TGF-β1 (transforming growth factor-β1), and TNF-α (tumor necrosis factor-α)] in BAL cells and four genes [IL-6, ICAM-1 (intercellular adhesion molecule-1), GM-CSF (granulocyte/macrophage-colony stimulating factor), and RANTES (regulated upon activation normal T cell expressed and secreted)] in lung tissue. In BAL cells on day 1, high-dose exposure induced a significant up-regulation of IL-1β, iNOS, MCP-1, and MIP-2 but no change in IL-6, IL-10, TGF-β1, and TNF-α mRNA levels. There was no change in the mRNA levels of IL-6, RANTES, ICAM-1, and GM-CSF in lung tissue. Nitric oxide production and levels of MCP-1 and MIP-2 were increased in the 24-hr culture media of alveolar macrophages (AMs) obtained on day 1. IL-6, MCP-1, and MIP-2 levels were also elevated in the BAL fluid. BAL fluid also showed increases in albumin and lactate dehydrogenase. The cellular content in BAL fluid increased at all doses and at all time periods, mainly due to an increase in polymorphonuclear leukocytes. In vitro studies in AMs and cultured lung fibroblasts showed that lung fibroblasts are a significant source of IL-6 and MCP-1 in the lung.

Activation of nuclear factor-kappa B and effects of pyrrolidine dithiocarbamate on TNBS-induced rat colitis

To explore the changes of nuclear factor-kappa B (NF-kappaB) DNA-binding activity, the expression of intercellular adhesion molecule-1 (ICAM-1) regulated by NF-kappaB at various times and to evaluate the effects of pyrrolidine dithiocarbamate (PDTC) on trinitrobenzene sulfonic acid (TNBS)-induced rat colitis. TNBS of 0.6 mL was mixed with ethanol of 0.3 mL solution and instilled into the lumen of the rat colon. The rat models were divided into 6 groups, which were killed at 24 h, 3, 7, 14, and 21 d after enema. Colonic inflammation and damage were assessed by macroscopical and histological criteria. Activity of NF-kappaB DNA-binding was analyzed by electrophoresis mobility shift assays (EMSA). Expression of ICAM-1 was detected by in situ hybridization (ISH) and immunohistochemistry (IH). Then various doses of PDTC were injected into rat abdomen 30 min before enema with TNBS/ethanol as pretreatment. The rats were killed 4 h after enema and the colonic inflammation, myeloperoxidase (MPO) activity, malondialdehyde (MDA) level, and DNA-binding activity of NF-kappaB were assessed. Finally, PDTC was injected intraperitoneally after colitis was induced. Changes of morphology were assayed. During the first week, hyperemia, hemorrhage, edema and ulceration of the colonic mucosa appeared with predominant infiltration of leukocytes. Neutrophils, macrophages, lymphocytes infiltrated in mucosa and submucosa 14 d later. Fibroblasts and granuloma-like structures were also obviously seen. The binding activity of NF-kappaB began to increase at 24 h time point and reached a peak at 14 d, then decreased but still was higher than control group at 21 d (P<0.01). Levels of ICAM-1 mRNA and protein significantly elevated at 24 h and the peak was at 21 d. Pretreatment with PDTC could attenuate the development of inflammation but not by reducing NF-kappaB activity. This attenuation of inflammation had a positive relationship with the dose of PDTC. PDTC at the dose of 100 mg/kg had no therapeutic effect after colitis was induced. NF-kappaB activation is an important event that may be involved in acute and chronic inflammation development and may contribute to self-protection against early inflammation damage. NF-kappaB also regulates ICAM-1 expression during colonic inflammation. Pretreatment of PDTC may attenuate the inflammation development. But PDTC has no therapeutic effect after the colitis is induced.

The inhibitory effect of alacepril, an angiotensin-converting enzyme inhibitor, on endothelial inflammatory response induced by oxysterol and TNF-α

The objectives were to determine the effects of alacepril, an angiotensin-converting enzyme inhibitor, on the expression of adhesion molecules and monocyte adherence to endothelial cells induced by 7-ketocholesterol (7-KC) and tumor necrosis factor (TNF)-α. We used human aortic endothelial cells (HAECs) and U937 monocytic cells. Surface expression and mRNA levels of intercellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule 1 (VCAM-1) were determined by EIA and RT-PCR. Adherence of U937 to HAECs was assessed by adhesion assay. Incubation of HAEC with 7-KC increased the surface expression of protein and mRNA levels of ICAM-1 and VCAM-1 on HAECs and the production of reactive oxygen species (ROS) in HAECs. Pretreatment with alacepril reduced the enhanced expression of these molecules in a dose-dependent manner. The inhibitory effect of alacepril against 7-KC or TNF-α-induced CAMs expression was stronger than that of captopril or enalapril. Alacepril inhibited the production of ROS in HAECs stimulated by 7-KC or TNF-α. These results suggest that alacepril works as anti-atherogenic agent through inhibiting endothelial-dependent adhesive interactions with monocytes induced by 7-KC and TNF-α.

A novel chimeric ribozyme vector produces potent inhibition of ICAM‐1 expression on ischemic vascular endothelium

Inhibition of intercellular adhesion molecule-1 (ICAM-1) expression can ameliorate the inflammation induced by ischemia-reperfusion injury (IRI) in animal models. However, current strategies to reduce ICAM-1 expression have been limited by the lack of stability, poor specificity, and the transient nature of synthesized regulatory molecules (antisense/ribozyme). A chimeric expression vector was generated by fusing a ribozyme targeting sequence against ICAM-1 to stabilizing stem-loop structures and nuclear localization signals that are components of endogenous U1 small nuclear RNA. Oligonucleotide scanning was used to predict accessible sites for targeting within the rat ICAM-1 transcript. Efficacy of the chimeric ribozyme vector was tested by transfection of rat aortic endothelial (RAE) cells (in vitro) and intraportal delivery in a rat hepatic IRI model (in vivo). Transfection of RAE cells with the chimeric ribozyme vector produced potent and specific inhibition of ICAM-1 mRNA and protein levels by >65%. This reduction in ICAM-1 expression was accompanied by a proportional decrease in neutrophil adhesion to RAE cells. In vivo intraportal delivery of the chimeric targeting vector to rats sustaining hepatic IRI produced a marked reduction in ICAM-1 expression on liver endothelium after reperfusion. A chimeric ribozyme vector effectively inhibited ICAM-1 expression in vascular endothelial cells and in rat liver following IRI, demonstrating a novel gene targeting technique that may be ideally suited to clinical applications aimed at ameliorating IRI.

Vitamin C blocks TNF-α-induced NF-kB activation and ICAM-1 expression in human neuroblastoma cells.

Interactions of the cell adhesion molecules are known to play important roles in mediating inflammation. The proinflammatory cytokine, tumor necrosis factor-α (TNF-α), activates the NF-kB signaling pathway, which induces the expression of various genes, such as intercellular adhesion molecule-1 (ICAM-1). In this study, the effect of vitamin C on the ICAM-1 expression induced by TNF-α in a human neuroblastoma cell line, SK-N-SH was investigated. Treatment with vitamin C resulted in the downregulation of the TNF-α-induced surface expression and ICAM-1 mRNA levels in a concentration-dependent manner. Moreover, a gel shift analysis indicated that vitamin C dose-dependently inhibited the NF-kB activation and lkBα degradation induced by TNF-α. Taken together, these results suggest that vitamin C downregulates TNF-α-induced ICAM-1 expression via the inhibition of NF-kB activation.

Periovulatory expression of intercellular adhesion molecule-1 in the rat ovary.

Leukocytes, especially neutrophils and macrophages, traditional cellular regulators of the immune system, reside within the tissue architecture of the rodent and human ovary and dramatically increase in number, in response to gonadotropin, in the theca of preovulatory follicles. Evidence strongly suggests a modulatory role for leukocytes in ovarian tissue remodeling events, such as ovulation, luteinization, and luteolysis. The present study investigates the ovarian localization and potential gonadotropin regulation of intercellular adhesion molecule-1 (ICAM-1), an important factor in neutrophil and monocyte attachment to endothelium. Reverse transcription-polymerase chain reaction and immunohistochemical detection and quantification of ICAM-1 mRNA and protein were carried out in ovaries of immature and eCG/hCG-primed rats during the periovulatory period (0, 6, 12, and 24 h post-hCG). While whole ovarian ICAM-1 mRNA levels did not vary significantly during the preovulatory period, ovarian follicles exhibited ICAM-1 mRNA and protein specifically within the thecal region, where mRNA expression increased 5-fold and protein expression increased 6-fold when comparing pre-hCG levels with those at the estimated time of ovulation (12 h post-hCG). Thecal ICAM-1 was most prevalent in highly vascularized regions as evidenced by serial staining with an endothelium-specific antibody. Granulosa layer ICAM-1 immunoactivity was acquired only during/after follicle rupture. These results show ICAM-1 is localized within the ovarian theca and its expression is associated with follicular development in periovulatory follicles, peaking in expression at the time of rupture. Additionally, ICAM-1 is expressed among granulosa-lutein cells of the ovulating follicle and developing corpus luteum. Taken together, these findings suggest rat ovarian ICAM-1 may be instrumental in the active recruitment of leukocytes into the preovulatory ovary and may have a role in corpus luteum formation.