The effect of cytokines on the expression of MHC antigens and ICAM-1 by normal and transformed synoviocytes.

We report the expression on synovial cells of cell surface molecules known to be involved in T cell activation by antigen presenting cells. Normal human synovial fibroblasts and a human synovial cell line transformed with the SV40 large T antigen were used for in vitro stimulation studies with recombinant cytokines. We demonstrate an increase in MHC-A, B, C expression in normal synovial cells in response to recombinant interferon gamma (r gamma IFN), tumour necrosis factor alpha and beta (rTNF alpha and beta) and interleukin-1 (rIL-1 alpha). Intercellular adhesion molecular-1 (ICAM-1) expression was increased in parallel with MHC Class I. The combination of r gamma IFN and rTNF alpha was additive in its effect on ICAM-1 expression. Northern blot analysis suggests that ICAM-1 expression in synovial cells is controlled at the level of transcription. In contrast, MHC Class II (HLA-DR) was only significantly induced by r gamma IFN. Other stimuli including interleukin-4 (IL-4), interleukin 6 (IL-6), granulocyte macrophage colony stimulating factor (GM-CSF) and prostaglandin E2 (PGE2) did not affect the expression of ICAM-1 or MHC Class I and II. Leucocyte function antigen 3 (LFA-3) expression was not affected by any of the stimuli tested. Immunoperoxidase staining of rheumatoid synovial tissue confirmed enhanced in vivo expression of ICAM-1 in rheumatoid arthritis. These changes are discussed in the context of T cell activation in inflammatory arthritis.

Analysis of adhesion molecules in the immunopathogenesis of giant cell arteritis.

To explore the role of adhesion molecules in mediating mononuclear cell localisation, development of the granulomatous reaction, and cell mediated damage to the arterial wall in giant cell arteritis, 17 temporal artery biopsy specimens were examined. Eleven showed the histological features of giant cell arteritis and six showed no evidence of arteritis. All were examined for the expression of LFA-3, ICAM-1 and its receptor LFA-1, and HLA-DR. Temporal arteries with early features of arteritis, as well as histologically unaffected skip areas, showed a regional induction of ICAM-1 expression, but not HLA-DR, on smooth muscle cells of the media. ICAM-1 expression was detected in areas where a clinically important mononuclear cell infiltrate had not yet developed. In more florid cases of giant cell arteritis there was an additional widespread induction of ICAM-1 expression on intimal myofibroblasts. Strong expression of ICAM-1, HLA-DR, and LFA-3 was found on macrophages, epithelioid cells, and giant cells comprising the granulomatous lesion. The pattern of expression of these adhesion molecules suggests that they have a role in leucocyte traffic into the vascular lesion as well as in mediating the intercellular interactions which constitute the granulomatous response.

Isolation and characterization of the promoter region of the human intercellular adhesion molecule-1 gene.

We have isolated and restriction enzyme-mapped a human genomic DNA clone encompassing the first two exons and the 5' flanking sequence of the human intercellular adhesion molecule-1 (ICAM-1) gene. The transcription initiation site was identified using primer extension analysis, and 1.7 kb of DNA upstream of the transcription initiation site was sequenced. The 5' region and first exon of the ICAM-1 gene was found to be a CpG island as it was (i) (G + C)-rich with a high frequency of the dinucleotide CpG and (ii) hypomethylated irrespective of the level of ICAM-1 expression in the tissues examined. These features of the ICAM-1 promoter are similar to the promoters of many 'housekeeping' genes. However, consensus sequences for several potential regulatory elements were found in the 5'-flanking sequence, an observation in keeping with the pattern of strongly regulated ICAM-1 expression. Examination of the chromatin structure upstream of the ICAM-1 gene revealed the presence of a constitutive DNase I-hypersensitive site 1.5 kb upstream of the transcription initiation site. Direct evidence that the upstream region constitutes a promoter element was demonstrated in transient transfection assays. A series of chloramphenicol acetyltransferase gene (CAT) constructs containing 5' fragments ranging in size from 1054 to 310 bp had equivalent levels of promoter activity when transfected into HeLa cells. Using a CAT construct containing a 447 bp ICAM-1 promoter fragment, we demonstrate an increase in transcription in response to interferon gamma (IFN-gamma), suggesting that this proximal region of the promoter is responsible, at least in part, for IFN-gamma induction of ICAM-1 expression.

Functional studies of stable L cell transfectants expressing intercellular adhesion molecule 1 [ICAM-1].

High molecular weight DNA isolated from human tonsil was transfected into mouse L cells to produce transfectants expressing the human intercellular adhesion molecule [ICAM-1]. The transfected ICAM-1 molecule was highly expressed on the cell membrane and our studies show that the transfected ICAM-1 molecule was a fully functional adhesion protein. All leucocyte subtypes showed specific binding to the ICAM-1 transfectants, their adhesion being inhibited by Fab1 fragments of W-CAM-1 antibody and LFA-1 MoAb (leucocyte function antigen). B cell lines (RAJI, Nalm1) showed the highest degree of ICAM-1 mediated adhesion. Normal lymphoblasts showed comparable levels of binding whilst normal neutrophils (both resting and activated by (N-formyl-methionyl-leucyl-phenylalanine) fMLP) showed the least ICAM-1-mediated adhesion. Despite a significant level of adhesion to ICAM-1 transfectants shown by T lymphoblasts generated in a two way MLR there was no evidence of cytolysis of ICAM-1 transfectants. These studies demonstrate the potential of ICAM-1 transfectants as tools for analysis of the role of ICAM-1 in lymphoid adhesion.

Regulation of expression of a human intercellular adhesion molecule (ICAM-1) during lymphohematopoietic differentiation.

The intercellular adhesion molecule (ICAM-1) is a cell-surface molecule which binds to leukocyte function antigen-1 (LFA-1) and regulates both leukocyte adhesion to endothelial cells and immune functions requiring cell-cell contact. Membrane expression of ICAM-1 is highly regulated on all hematopoietic lineages. Cell membrane antigen is significantly expressed on a small subset of bone marrow (BM) progenitors but is weak or absent on all cell lineages once they enter the circulation. However, strong expression on tissue macrophages and germinal center B cells suggested that activated cells may show upregulated expression. When B cells, T cells, macrophages, or granulocytes were activated in vitro by suitable mitogens, ICAM-1 expression was induced in all cases. Parallel studies of hematopoietic tumors demonstrated a heterogeneity of expression which correlated with expression on their normal cellular counterparts. In particular, a striking correlation between expression on B-cell tumors and corresponding stages of B-cell differentiation was noted. The widely varying expression of ICAM-1 contrasts with LFA-1 which, while variable, is nevertheless significantly positive at all stages of differentiation. This suggests that the major regulation of homotypic adhesion mediated by the LFA-1/ICAM-1 linkage occurs through control of ICAM-1 expression. In keeping with this notion, ICAM-1 expression was also correlated with the "adhesiveness" of B-lymphoid tumors. Large solitary lymphoma masses showed intense expression of ICAM-1. Conversely, chronic lymphocytic leukemia (CLL) cells and lymphoma cells from tumors exhibiting diffuse, widespread lymph node disease showed weak expression. These observations are discussed in relation to the role of ICAM-1 in regulation of lymphoid recirculation and the biology of lymphoid tumors.

Intercellular adhesion molecule 1 (ICAM-1) has a central role in cell-cell contact-mediated immune mechanisms.

The role of intercellular adhesion molecule 1 (ICAM-1) in immune function was probed by using the Wehi-CAM-1 (W-CAM-1) monoclonal antibody. This antibody blocks aggregation of cell lines mediated by the ICAM-1 molecule and is here shown to block homotypic binding of purified populations of activated T and B lymphocytes (blasts) and also aggregation of mixed T- and B-cell blasts. We also demonstrate that W-CAM-1 inhibited T-cell adhesion to normal human endothelial cells, the first step in lymphocyte egress into the tissues. In tests of immune function, W-CAM-1 had a modest inhibitory effect on T- and B-cell activation by potent mitogens and no effect on the response of activated lymphocytes to lymphokines. By contrast, activation induced by cell-cell contact (mixed lymphocyte reaction, T-cell-mediated B-cell activation) was significantly inhibited. Moreover, the antibody was shown to block elements of both effector arms of the immune system (cytotoxic cell function and immunoglobulin production). These findings show that the ICAM-1 molecule is a central component of the mechanism of lymphocyte-endothelial cell adhesion. The studies of lymphoid function demonstrate a pivotal role for this molecule in both the T-cell/T-cell and T-cell/B-cell interactions, which underpin the regulation of the immune response, and in the mechanism of cell-mediated cytotoxicity.