Interferon-gamma (IFN-γ) down-regulates the rhinovirus-induced expression of intercellular adhesion molecule-1 (ICAM-1) on human airway epithelial cells

Human rhinoviruses (HRV) are a major cause of upper respiratory tract infections in man, and can exacerbate existing pulmonary disease. The major group of HRV attach to ICAM-1, which is expressed on nasal and bronchial epithelial cells. To study the influence of biological mediators on ICAM-1 expression, and consequently HRV attachment and infection, we compared the effects of various cytokines, alone and in combination, on ICAM-1 expression by an uninfected and HRV-infected bronchial epithelial cell line H292. Cytokines known to be released soon after viral infection, such as tumour necrosis factor-alpha (TNF-α), IL-1β and the chemokine IL-8 increase ICAM-1 expression on uninfected cells. Epithelial cells infected with live HRV-14 displayed marked up-regulation of ICAM-1 compared with baseline. TNF-α further enhanced the HRV-induced increase in ICAM-1 expression on epithelial cells, peaking at day 4 after infection, whilst IL-8 exhibited a steady increase in ICAM-1 expression over 14 days. In contrast, IFN-γ, a known Th1 antiviral lymphokine, whilst increasing the level of ICAM-1 on uninfected cells, induced a significant persistent down-regulation of ICAM-1 expression on HRV-infected epithelial cells. With combinations of TNF-α and IFN-γ, ICAM-1 expression on HRV-infected cells was reduced to basal levels. The effects of IFN-γ were paralleled by a reduction in viral titres. Our in vitro model has provided useful insights into the early pathogenic events of HRV infection at the level of the host cell–v irus interaction. Our data confirm that biological mediators play a crucial role in the pathogenesis as well as the course of HRV infection which is modulated by the types, and time kinetics of inflammatory cytokines in the immediate microenvironment.     

Intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) expression and function on cultured human glomerular epithelial cells.

Glomerular epithelial cells are involved in extracapillary inflammation (crescents) but the mechanisms of this extracapillary accumulation of macrophages, epithelial cells and occasional lymphocytes are unknown. Human glomerular parietal epithelial cells express ICAM-1 and VCAM-1 on immunohistological stains of renal biopsies. We studied the expression of these cell adhesion molecules on cultured human glomerular epithelial cells (HGEC), their regulation by pro-inflammatory cytokines, and their role in mediating the adhesion of concanavalin A (Con A)-activated peripheral blood mononuclear cells. Human glomerular epithelial cells in culture constitutively express ICAM-1 and VCAM-1. The expression of ICAM-1 was not significantly altered by tumour necrosis factor-alpha (TNF-alpha) (P = 0.32), IL-1 beta (P = 0.24), interferon-gamma (IFN-gamma) (P = 0.66) or IL-4 (P = 0.85). VCAM-1 expression was increased by all four cytokines, but only significantly so by IL-4 (P = 0.0001). Con A-stimulated, monocyte-depleted peripheral blood lymphocytes bound to human glomerular epithelial cells, median 28.9% (range 14.5-37.9%). This adherence was significantly inhibited by anti-ICAM-1 (P = 0.03) and anti-LFA-1 (P = 0.02), but not by anti-VCAM-1 (P = 0.13) or by antibody to von Willebrand factor (P = NS). The interaction between ICAM-1 on HGEC and LFA-1 on mononuclear cells may be important in the pathogenesis of extracapillary inflammation in glomerulonephritis.     

Podocyte expression of MHC class I and II and intercellular adhesion molecule-1 (ICAM-1) in experimental pauci-immune crescentic glomerulonephritis

We examined immunopathological changes of podocytes in vivo which, based on in vitro studies, are thought to be relevant for the pathogenesis of renal diseases. We investigated the alterations of podocytes in local inflammation in a recently developed model of pauci-immune necrotizing crescentic glomerulonephritis (NCGN) in the rat. Frozen and plastic embedded kidney sections at different time points of the disease were incubated with antibodies directed to MHC class I, MHC class II, ICAM-1 and to relevant cytokines. Strong glomerular expression of MHC class I. II and ICAM-1 was found within 4 days, and plastic embedded sections clearly demonstrated increased cell membrane staining of podocytes. Increased glomerular interferon-gamma (IFN-γ) was detected within 24 h of induction of NCGN. and IL-IβT and tumour necrosis factor-alpha (TNF-α) were found from day 4. The potency of these cytokines to induce adhesion molecules on podocytes was investigated on rat glomerular epithelial cells in vitro. By using IACS analysis and electron microscopical techniques, we found that the in vivo expression of MHC class I, II and ICAM-I by podocytes could in vivo be simulated by IFN-γ IFN-α weakly induced MHC class I. while IL-Iβ and TNF-α were ineffective. We hypothesize that podocytes in this in vivo model are important to maintain the local inflammatory process in the glomerulus by expression of relevant adhesion molecules and MHC molecules upon stimulation with specific cytokines.     

Expression of interferon-gamma receptors and interferon-gamma-induced up-regulation of intercellular adhesion molecule-1 in basal cell carcinoma; decreased expression of IFN-γR and shedding of ICAM-1 as a means to escape immune surveillance

The peritumoural inflammatory infiltrate in basal cell carcinoma (BCC) of the skin consists mainly of T lymphocytes which hardly invade the tumour nests. The absence of intercellular adhesion molecule-1 (ICAM-1) on BCC cells may explain the lack of tumour-infiltrating cells and the lack of an active cell-mediated immune response in this tumour. In this study, the induction of ICAM-1 was investigated in BCC biopsies using recombinant human interferon-gamma (rHuIFN-γ). The expression of interferon-gamma receptors (IFN-γR) in the biopsies was also investigated. The results showed that BCC cells expressed ICAM-1 after incubation with rHuIFN-γ, but to a lesser degree than normal epidermal cells. The levels of shed ICAM-1 were significantly increased in the culture supernatants of tumour biopsies compared with those from normal skin biopsies, after culturing in the presence of rHuIFN-γ. The expression of IFN-γR was significantly decreased on the tumour cells compared with the overlying epidermis. The decreased expression of IFN-γR on the tumour cells and the shedding of ICAM-1 into the peritumoural stroma may be a plausible mechanism by which the tumour cells are protected against an active cell-mediated immune response. © 1998 John Wiley & Sons, Ltd.     

Adhesion molecules (E-selectin, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1)) in sera from patients with Staphylococcus aureus bacteraemia with or without endocarditis

The aim of this prospective study was to evaluate if patients with endocarditis display a more extensive endothelial activation than those with bacteraemia but without endocarditis. Sixty-five patients with blood culture-verified Staphylococcus aureus bacteraemia were included and serum samples collected on admission were analysed by enzyme immunoassays. Elevated serum concentrations of adhesion molecules were found in most of the patients with S. aureus bacteraemia. Patients with endocarditis (n = 15) showed significantly higher serum E-selectin (median 156 ng/ml) and VCAM-1 (median 1745 ng/ml) concentrations compared with those with S. aureus bacteraemia but without endocarditis (80 ng/ml and 1172 ng/ml, respectively; P = 0.01 and P = 0.003). No significant difference was found between the groups concerning ICAM-1 (median 451 ng/ml versus 522 ng/ml). In addition, serum tumour necrosis factor-alpha (TNF-alpha) concentrations were significantly correlated (P < 0.002) to serum levels of E-selectin, ICAM-1 and VCAM-1.     

Circulating soluble intercellular adhesion molecule-1 (sICAM-1) in patients with sarcoidosis

sICAM-1 has been elevated in sera of specific inflammatory diseases, and circulating sICAM-1 concentrations reflect disease activity in these diseases. We measured circulating sICAM-1 concentrations and serum angiotensin-converting enzyme (SACE) activity in patients with sarcoidosis. Patients with sarcoidosis had significantly increased circulating sICAM-1 concentrations (62.8±33.5U/ml) and SACE activity (23.7±7.4U/l) compared with controls (circulating sICAM-1 50.9±12.1U/ml, and SACE 13.5±3.8U/l). Successive measurements showed that circulating sICAM-1 values changed in parallel with disease activity in sarcoidosis. In the progressive disease group (progressed or without change for 2 years or more), circulating sICAM-1 values (102.2±35.3U/ml) at the time of diagnosis were significantly increased compared with those in the regressive disease group (disappeared or regressed within 2 years) (46.4±12.6U/ml). However, there was no significant difference in SACE activity of the regressive and progressive disease groups. Fifteen patients with a high value of circulating sICAM-1 (> 75U/ml, mean of controls+2s.d.) all had progressive disease, while only 15 of 44 patients with a high value of SACE had progressive disease. Circulating sICAM-1 will be a useful blood marker to predict outcome and to monitor disease activity in sarcoidosis.     

Cellular mRNA expression of interferon-gamma (IFN-γ), IL-4 and transforming growth factor-beta (TGF-β) in rats nasally tolerized against experimental autoimmune myasthenia gravis (EAMG)

Nasal administration of nicotinic acetylcholine receptor (AChR) to Lewis rats prior to myasthenogenic immunization with AChR plus Freund's complete adjuvant (FCA) resulted in prevention or marked decrease of the severity of EAMG, suppression of AChR-specific B cell responses and of AChR-reactive T cell functions. To examine the involvement of immunoregulatory cytokines and the underlying mechanisms involved in tolerance induction, in situ hybridization with radiolabelled synthetic oligonucleotide probes was adopted to enumerate mononuclear cells (MNC) expressing mRNA for the proinflammatory cytokine IFN-γ, the B cell stimulating IL-4 and the immune response-down-regulating TGF-β. Popliteal and inguinal lymph nodes from EAMG rats contained elevated numbers of AChR-reactive IFN-γ, IL-4 and TGF-β mRNA-expressing cells compared with control rats receiving PBS nasally and injected with FCA only. Nasal tolerance to EAMG was accompanied by decreased numbers of AChR-reactive IFN-γ and IL-4 mRNA-expressing cells, and strong up-regulation of TGF-β mRNA-positive cells in lymphoid organs compared with non-tolerized EAMG control rats. The relative affinity of anti-AChR antibodies was lower, but muscle AChR amounts were higher in nasally tolerized rats compared with non-tolerized EAMG control rats. The results suggest that IFN-γ and IL-4 are central effector molecules in the development of EAMG, and that TGF-β plays an important role in tolerance induction to EAMG.     

Blood levels of transforming growth factor-beta 1 (TGF-β1) are elevated in both relapsing remitting and chronic progressive multiple sclerosis (MS) patients and are further augmented by treatment with interferon-beta 1b (IFN-β1b)

The serum levels of TGF-β1, measured by solid-phase ELISA, were determined to be significantly augmented in patients with both relapsing remitting (RR) and secondary chronic progressive (CP) MS compared with sex- and age-matched healthy controls. Moreover, in RR MS patients, the blood levels of the cytokine were further augmented either during relapses or, in a rapid but reversible fashion, by s.c. injection with 8 million International Units (MIU) IFN-β1b. Because TGF-β1 possesses multiple anti-inflammatory activities, we hypothesize that the increase in its circulating levels in RR and CP MS patients might represent an endogenous anti-inflammatory mechanism aimed at counteracting ongoing immunoinflammatory events, and that IFN-β may further potentiate this natural defensive apparatus.     

Transforming growth factor-beta (TGF-β) expression and interaction with proteinase 3 (PR3) in anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis

TGF-β is a multifunctional cytokine modulating the onset and course of autoimmune diseases as shown in experimental models. The aim of this study was to investigate TGF-β expression in ANCA-associated vasculitis (AAV), and the possible interactions of this cytokine with lysosomal enzymes identified as ANCA autoantigens (e.g. PR3). This included TGF-β effects on the translocation of the lysosomal enzymes to the cell surface of polymorphonuclear neutrophils (PMN), and the presumed activation of non-bioactive, latent TGF-β by these enzymes. Patients with various types of systemic vasculitis (SV) were studied, including three different types of AAV (Wegener's granulomatosis (WG), Churg–Strauß syndrome (CSS) and microscopic polyangiitis (MPA)). Regardless of the type of assay applied, the TGF-β1 isoform was found to be overexpressed in SV, including AAV, and to correlate with disease activity as shown for WG. Mean TGF-β1 plasma levels in AAV patients ranged from 8.9 ng/ml (WG) to 13.3 ng/ml (CSS) (control 4.2 ng/ml; P< 0.01), while TGF-β2 levels were not elevated. Flow cytometry analysis showed TGF-β1 to be a potent translocation factor for PR3 comparable to other neutrophil-activating factors such as IL-8. PR3 membrane expression on primed PMN increased by up to 51% after incubation with TGF-β1. PR3 itself was revealed as a potent activator of latent TGF-β, thus mediating bioeffects of this cytokine. These findings, together with other features of TGF-β such as induction of angiogenesis and its strong chemotactic capacity, indicate that TGF-β might serve as a proinflammatory factor in SV, especially in AAV.     

Circulating intercellular adhesion molecule-1 (ICAM-1) as an early and sensitive marker for virus-induced T cell activation

The effect of systemic virus infection on the level of circulating ICAM-1 (cICAM-1) in serum, and the role of virus-activated T cells in this context, were studied using the murine lymphocytic choriomeningitis virus infection as primary model system. A marked virus-induced elevation in cICAM-1 in serum was revealed, the presence of which coincided with the phase of virus-induced T cell activation. However, high levels of cICAM-1 in serum were observed well before maximal T cell activation could be demonstrated. No increase in cICAM-1 was observed in the serum of infected T cell-deficient nude mice, clearly demonstrating that T cells were mandatory. Analysis of MHC class I and MHC class II-deficient mice revealed that either CD4⁺ or CDS⁺ T cells alone are sufficient, despite a markedly reduced inflammatory exudate in the former animals. These results indicate that virus-activated T cells induce shedding of ICAM-1 into the circulation, and this parameter may be used as an early and sensitive marker for immune activation.     

Pirfenidone induces intercellular adhesion molecule-1 (ICAM-1) down-regulation on cultured human synovial fibroblasts

Pirfenidone has been shown to modify some cytokine regulatory actions and inhibit fibroblast biochemical reactions resulting in inhibition of proliferation and collagen matrix synthesis by fibroblast. We have investigated the effect of pirfenidone on the expression of cell adhesion molecules. The synovial fibroblasts were treated with IL-1α in the presence or absence of pirfenidone (range 0–1000 μM ), and assayed for the expression of adhesion molecules such as ICAM-1 and endothelial-leucocyte adhesion molecule-1 (E-selectin) by cell ELISA. Pirfenidone significantly down-regulated the expression of ICAM-1 on cultured synovial fibroblasts in a dose-dependent manner. In contrast, expression of E-selectin was not affected. Furthermore, we examined whether pirfenidone affects the cellular binding between cultured lymphocytes and IL-1α-stimulated synovial fibroblasts by in vitro binding assay and found their mutual binding was significantly suppressed in a dose-dependent manner by pirfenidone. It is speculated that down-regulation of ICAM-1 might be one of the novel mechanisms of action of pirfenidone. These data indicate a novel mechanism of action for pirfenidone to reduce the activation of synovial fibroblasts.     

Increased expression of intercellular adhesion molecule-1 (ICAM-1) in a murine model of pulmonary eosinophilia and high IgE level.

T lymphocytes and eosinophils are probably involved in the pathogenesis of allergic bronchopulmonary aspergillosis (ABPA), a disease characterized by pulmonary eosinophilia and high serum and lavage IgE levels. We recently developed a murine model of ABPA. To investigate the mechanisms of T lymphocyte and eosinophil recruitment to the lung in this disease, we examined the expression of ICAM-1 in the lung tissue of mouse challenged with Aspergillus fumigatus (Af) antigen. C57B1/6 mice were intranasally exposed to Af (Af group) or saline (control group) three times a week for 1, 2 or 3 weeks. On days 4, 7, 14 and 21, mice were killed and lung tissue was fixed in acetone and embedded in glycol methacrylate. Serial 2-microns sections were stained with chromotrope 2R and MoAbs against ICAM-1, CD11a/CD18 (LFA-1) and CD3. Af-challenged mice presented significant increases in eosinophil, T lymphocyte and LFA-1-positive cell count and up-regulated expression of ICAM-1 in the lung tissue at all the time points examined. ICAM-1 expression intensity correlated with the number of T lymphocytes (r = 0.59, P < 0.01), LFA-1-positive cells (r = 0.68, P < 0.001), but not of eosinophils (r = -0.24, P > 0.05). These findings suggest that up-regulation of ICAM-1 expression is involved in the inflammatory process of this murine model of ABPA, and that this up-regulation may be more relevant to the the T lymphocyte accumulation in the lung.     

Immunocytochemical localization of inducible nitric oxide synthase and transforming growth factor-beta (TGF-β) in leprosy lesions

Inducible nitric oxide synthase (iNOS) and TGF-β were localized by immunocytochemistry in skin lesions from patients across the leprosy spectrum, and from patients undergoing reversal reaction. iNOS expression was highest at the tuberculoid pole of the spectrum, and increased during reversal reaction. TGF-β was observed throughout the leprosy spectrum, but was highest at the lepromatous pole. Levels of TGF-β decreased during reversal reaction. Reduced levels of TGF-β may contribute to unregulated inflammatory responses during reactional episodes.     

Release of transforming growth factor-beta (TGF-β) and fibronectin by alveolar macrophages in airway diseases

Asthma and chronic bronchitis are associated with airway remodelling, and airway macrophages are present in bronchial inflammation. TGF-β and fibronectin released by alveolar macrophages possess a fibrogenic potency. The potential role of alveolar macrophages in airway remodelling was studied in asthma and chronic bronchitis by the release of TGF-β and fibronectin. Alveolar macrophages were isolated by bronchoalveolar lavage in 14 control subjects, 14 asthmatics and 14 chronic bronchitics. The spontaneous and lipopolysaccharide (LPS)- or concanavalin A (Con A)-induced release of TGF-β and fibronectin was measured by ELISA. Alveolar macrophages from chronic bronchitics spontaneously release greater amounts of TGF-β and fibronectin than those from asthmatic and control subjects. Alveolar macrophages from asthmatics release greater amounts of TGF-β and fibronectin than those from control subjects. The spontaneous release of TGF-β is significantly correlated with that of fibronectin. Fibronectin release was significantly reduced after LPS stimulation, and TGF-β release was significantly increased after LPS stimulation, except in chronic bronchitis patients. Con A increased the release of TGF-β in cells from normal subjects. This study suggests that activated macrophages play a role in airway remodelling in chronic bronchitis and to a lesser extent in asthma.     

Roles of tumour necrosis factor-alpha (TNF-alpha), transforming growth factor-beta (TGF-beta), and IL-10 in the modulation of intercellular adhesion molecule-1 (ICAM-1) expression by macrophages during mycobacterial infection

Profiles of ICAM-1 expression on cultured murine peritoneal macrophages infected with Mycobacterium avium complex (MAC) were examined, with special reference to modulating roles of TNF-alpha, TGF-beta, and IL-10. When macrophages were infected with MAC, ICAM-1 expression, measured by microscopic counting of ICAM-1+ macrophages stained with anti-ICAM-1 antibody, ELISA, and flow cytometric analysis, was rapidly increased, peaking at day 3 (early-phase up-regulation) due to endogenous TNF-alpha, and thereafter gradually declined to the normal level within 1 week or more (late-phase down-regulation). The late-phase ICAM-1 down-regulation was also seen in macrophages phagocytosing heat-killed MAC and those stimulated with lipopolysaccharide but not in macrophages phagocytosing latex beads. ICAM-1 mRNA expression was augmented markedly at day 1 after MAC infection and thereafter decreased. While TNF-alpha and IL-10 production by MAC-infected macrophages was observed during the first 3 days, TGF-beta production was initiated from day 3 and continued until day 14. Exogenously added TGF-beta strongly inhibited the early-phase increase in ICAM-1 expression by infected macrophages, and the blockade of endogenous TGF-beta with anti-TGF-beta antibody markedly inhibited late-phase ICAM-1 down-regulation. Moderate blocking effect was also observed for anti-IL-10 antibody. On the other hand, late-phase ICAM-1 down-regulation was not prevented by the addition of exogenous TNF-alpha. Therefore, TGF-beta and IL-10, especially the former, appear to play active roles in the late-phase down-regulation of ICAM-1 in MAC-infected macrophages during long-term cultivation.     

Dynamic expression of transforming growth factor-betas (TGF-β) and their type I and type II receptors in the synovial tissue of arthritic rats

A well characterized animal model that shares many characteristic features with rheumatoid arthritis (RA) is collagen-induced arthritis (CIA) in DA rats. Recent studies have demonstrated that TGF-β, a multifunctional cytokine, is an important modulator of the immune response in CIA, and possibly also in RA. In this study we have investigated the expression of the precursor forms of TGF-β1, TGF-β2, TGF-β3, as well as TGF-β type I receptor (TGF-βRI) and TGF-β type II receptor (TGF-βRII) in the synovial tissue of arthritic rats during the course of the disease. By using immunohistochemical techniques, an abundant expression of all three TGF-β isoforms and their receptors was observed in the arthritic synovia, an expression that increased with time after onset of disease. Antibodies to TGF-β1, TGF-β2, TGF-βRI and TGF-βRII stained blood vessels intensively, already at the early onset of inflammation, whereas the synovial lining layer and chondrocytes expressed strong immunoreactivity later on in the inflammatory process. The most intense staining with these antibodies was detected in fibroblasts within fibrotic tissue, in particular at the cartilage–pannus junction. Interestingly, TGF-β3 only stained macrophage-like cells and chondrocytes in the synovia. The data suggest that the abundant expression of TGF-β1, TGF-β2, TGF-β3 as well as TGF-βRI and TGF-βRII in the synovia, is of pathogenic importance in the development of CIA, although the question of how the different TGF-β isoforms may enhance or counteract different arthritogenic events remains open.     

Hypoxia augments cytokine (transforming growth factor-beta (TGF-β) and IL-1)-induced vascular endothelial growth factor secretion by human synovial fibroblasts

Vascular endothelial growth factor (VEGF) is abundant in synovium and synovial fluids, where it probably contributes to vascular permeability and angiogenesis in arthritic joints. To investigate the probable sources of VEGF in synovium, we compared the ability of several cytokines (TGF-β, platelet-derived growth factor (PDGF), IL-1, tumour necrosis factor (TNF), basic fibroblast growth factor (bFGF) that are associated with arthritis and angiogenesis, to stimulate secretion of VEGF protein by human synovial fibroblasts. TGF-β was the strongest inducer of VEGF secretion; six times more VEGF was secreted when cells were stimulated by TGF-β than when stimulated by PDGF or IL-1 for 24 h. TNF-α and bFGF did not stimulate any secretion of VEGF. The stimulatory effects of TGF-β and IL-1 on VEGF secretion were additive. Hypoxic culture alone also stimulated VEGF secretion, but more importantly, hypoxic culture conditions doubled the rate of VEGF secretion stimulated by the cytokines TGF-β and IL-1. When dermal and synovial fibroblasts were stimulated identically by hypoxia and cytokines (TGF-β and IL-1), synovial fibroblasts secreted four times more VEGF than did dermal fibroblasts. Thus in rheumatoid arthritis, the capacity of synovial fibroblasts in the hypoxic environment to secrete large amounts of VEGF in response to cytokines such as TGF-β probably contributes significantly to angiogenesis in the synovium.