Abstract
Full-length intercellular adhesion molecule 1 (ICAM-1) is an immunoglobulin-like expressed in various tissues, including the endothelium. The binding of ICAM-1 on an endothelial cell to lymphocyte function-associated molecule 1 (LFA-1) and macrophage-1 antigen (Mac-1) on neutrophils is important for neutrophil recruitment. ICAM-1 has five extracellular immunoglobulin domains and most investigations have studied neutrophil recruitment with full-length ICAM-1. Binding between LFA-1: ICAM-1 and Mac-1: ICAM-1 differ slightly in function and binding site; LFA-1 binds domain 1 of ICAM-1 while Mac-1 binds domain 3. Six alternatively-spliced ICAM-1 isoforms were previously discovered in ICAM-1-exon-deletion mice: full-length ICAM-1, ICAM-1(2-4), ICAM-1(2-6), ICAM-1(3-6), ICAM-1(4-6), and ICAM-1(2-5), named for the spliced exons. To better understand the role of ICAM-1 isoforms in Mac-1: ICAM-1-dependent neutrophil recruitment to the liver, we studied the relative binding of five ICAM-1 isoforms to Mac-1. These isoforms were transiently expressed in HEK293T cells using customized plasmids. A V-bottomed adhesion assay assessed the binding of these cells to Mac-1. The ICAM-1(2-4) isoform demonstrated the highest relative binding percentage to Mac-1 (relative binding percentage of 42.3% in the presence of Mg2+/EGTA), while ICAM-1(2-6) demonstrated the lowest (relative binding percentage of 9.9% in the presence of Mg2+/EGTA). We also began assessment of the existence of ICAM-1 isoforms in human blood, liver, and lung RNA samples using reverse transcription polymerase chain reaction (RT-PCR). Additional experiments are required to evaluate the binding of the six ICAM-1 isoforms to Mac-1 and LFA-1 using the V-bottomed adhesion assay, and will confirm the presence of ICAM-1 isoforms in humans.