Intercellular adhesion molecule-1 (ICAM-1) is a membrane protein, exists as a dimer on the cell surface, and interacts with leukocyte function associated antigen-1 (LFA-1), a member of beta2-integrin family. A soluble circulating form of ICAM-1 (sICAM-1) is also detected in human serum, and has been implicated as a regulator for LFA-1-dependent cell-cell interaction in vivo. However, previous reports demonstrated that sICAM-1 shows little inhibitory effect on LFA-1 binding to ICAM-l, indicating that sICAM-1 is unlikely to antagonize LFA-1/ICAM-1-mediated cellular events in vivo. Here, we investigated the property of the dimeric sICAM-1 as an inhibitor of LFA-1 interaction with ICAM-3, since the lower avidity of LFA-1 for ICAM-3 compared with ICAM-1 or ICAM-2 had been speculated. Using recently constructed heterodimeric sICAM-1 joined at the C terminus via an a-helical coiled coil (ACID-BASE) (Jun, CD. et al., 2001, Proc Natl Acad Sci 98, 6830-6835), we also tested whether the structural integrity in dimer could affect the inhibitory action of sICAM-1. Engineered sICAM-1 dimer that contained intact ectodomain (E34/E34) significantly blocked SKW3 cell (LFA-1+) binding to ICAM-3, but not to ICAM-1 and ICAM-2, indicating the lower avidity of LFA-1 to ICAM-3 than that of both ICAM-1 and ICAM-2. A one binding site knock out mutant (E34/K34) showed -2-fold reduction in efficiency compared with E34/E34 to inhibit cell binding. Interestingly, a one binding domain deletion mutant (E34/deltaD1-D2) showed significant reduction (~5-fold) compare with E34/K34, suggesting that structural integrity, which is precluded in E34/deltaD1-D2, is necessary for optimal binding of dimeric sICAM-1 to LFA-1, thereby inhibiting LFA-1/ICAM-3-dependent adhesion. Furthermore, BIAcore affinity measurements revealed that E34/deltaD1-D2 bound to immobilized soluble open LFA-1 I domain with an -3-fold reduced affinity compared with E34/K34. Overall, our results demonstrate that maintaining the structural integrity in dimer is necessary for optimal binding of sICAM-1 to LFA-1, and further suggest the therapeutic potential of dimeric sICAM-1 to antagonize LFA-1/ICAM-3-mediated cellular events in vivo.
Binding Sites ; Humans ; Intercellular Adhesion Molecule-1 ; Leukocytes* ; Lymphocyte Function-Associated Antigen-1* ; Membrane Proteins

Emerging evidence suggests that gap formation and opening of the endothelial junctions during leukocyte extravasation is actively controlled to maintain the integrity of the vascular barrier. While the role for endothelial cells to this process has been well defined, it is not clear whether leukocytes are also actively contributing to endothelial barrier function. We have recently showed that extravasating leukocytes deposit microparticles on the subendothelium during the late stages of extravasation, which is LFA-1 dependent. Using multiphotonintravital microscopy (MP-IVM) of mouse cremaster muscle vessels in the current work, we show that microparticle formation and deposition maintains the integrity of the microvascular barrier during leukocyte extravasation. Inhibition of neutrophil-derived microparticle formation resulted in dramatically increased vascular leakage. These findings suggest that deposition of microparticles during neutrophil extravasation is essential for maintaining endothelial barrier function and may result in temporal difference between neutrophil extravasation and an increase in vascular leakage.
Animals ; Endothelial Cells ; Leukocytes ; Lymphocyte Function-Associated Antigen-1 ; Mice ; Microscopy ; Monocytes ; Muscles ; Neutrophils

BACKGROUND: Psoriasis is a chronic relapsing disease characterized by epidermal hyperproliferation and epiderrnal and dermal inflammatory cell infiltration. The etiology of this disease is still unclear. Recently, there has been growing interest in the probable role of a T cell mediated immune response in the pathogetiesis of psoriasis. The infiltrating cells in psoriatic lesions have been iden- tified by monoclonal ant~ibodies and T cells were found to be the major infiltrating type. OBJECTIVE: This stud was done to investigate the difference of cellular infiltration and adhesion molecule exg!ressions between early and late skin lesions of psoriasis using immunohistochernical studies. Methpds : Patients with psoriatic lesions were divided into two groups. The early gr oup were defined as having skin lisions that had lasted for about 4 weeks, and late group were defined as having skin lesions that had lasted for more than 8 weeks. Then biopsy specirnens were stained using monoclonal antibidies for CD4, CD8, CD1, LFA-1, and ICAM l. RESULTS: 1. CD4 positive cells,vere tly increased in both the early and late groups compared with CD8 positive cells. Z. CD8 positive cells were significantly increased in the late g~roup compared with the early group. 3. CDl-posit,ive dendri!ic cells were more nurnerous in the late group than the early group. 4. There were no significant differences between the early and late group with regard to numbers of LFA-1 positive (ells. 5. ICAM-1 were more strongly expressed on epidermal keratinocytes in the late than the early group. CONCLUSION: CD4-positive cells are important in early and late psoriatic lesions and CD8 positive cells playi more important role in late than early lesions. ICAM-1 and LI'A-1 play a role in cell adhesion of infiltrating cells and lymphocytic rnigration to the epidermis.
Biopsy ; Cell Adhesion ; Epidermis ; Humans ; Intercellular Adhesion Molecule-1 ; Keratinocytes ; Lymphocyte Function-Associated Antigen-1 ; Psoriasis ; Skin ; T-Lymphocytes

BACKGROUND: Psoriasis is a chronic relapsing disease characterized by epidermal hyperproliferation and epiderrnal and dermal inflammatory cell infiltration. The etiology of this disease is still unclear. Recently, there has been growing interest in the probable role of a T cell mediated immune response in the pathogetiesis of psoriasis. The infiltrating cells in psoriatic lesions have been iden- tified by monoclonal ant~ibodies and T cells were found to be the major infiltrating type. OBJECTIVE: This stud was done to investigate the difference of cellular infiltration and adhesion molecule exg!ressions between early and late skin lesions of psoriasis using immunohistochernical studies. Methpds : Patients with psoriatic lesions were divided into two groups. The early gr oup were defined as having skin lisions that had lasted for about 4 weeks, and late group were defined as having skin lesions that had lasted for more than 8 weeks. Then biopsy specirnens were stained using monoclonal antibidies for CD4, CD8, CD1, LFA-1, and ICAM l. RESULTS: 1. CD4 positive cells,vere tly increased in both the early and late groups compared with CD8 positive cells. Z. CD8 positive cells were significantly increased in the late g~roup compared with the early group. 3. CDl-posit,ive dendri!ic cells were more nurnerous in the late group than the early group. 4. There were no significant differences between the early and late group with regard to numbers of LFA-1 positive (ells. 5. ICAM-1 were more strongly expressed on epidermal keratinocytes in the late than the early group. CONCLUSION: CD4-positive cells are important in early and late psoriatic lesions and CD8 positive cells playi more important role in late than early lesions. ICAM-1 and LI'A-1 play a role in cell adhesion of infiltrating cells and lymphocytic rnigration to the epidermis.
Biopsy ; Cell Adhesion ; Epidermis ; Humans ; Intercellular Adhesion Molecule-1 ; Keratinocytes ; Lymphocyte Function-Associated Antigen-1 ; Psoriasis ; Skin ; T-Lymphocytes

Endotoxin-induced uveitis model was produced in Lewis rats by footpad injection of Salmonella endotoxin(lipopolysaccharide, LPS). The clinical course and histological examination were observed at intervals of two hours. Immunohistochemical staining of intercellular adhesion molecule-1(ICAM-1) and lymphoyte function associated antigen-1(LFA-1) were also peformed. Initial intraocular inflammatory signs were observed at 8-10 hours after injection. Clinical and histological abnormalities peaked at 24-48 hours and were resolving after 72 hours. Histological changes were limited to anterior uvea. ICAM-1 expression was first noted on cells of iris and ciliary body and LFA-1 was expressed on infiltring inflammatory cells 8 hours after the injection and increased by 24 hours and disappeared after 72 hours. LPS induced uveitis in the rat provides a simple, reproducible model for anterior uveitis. ICAM-1 and LFA-1 expression in uveal tract are increased during early phase of uveitis and may enhance the adherence of inflammatory cells with the subsequent initiation of inflammation.
Animals ; Ciliary Body ; Inflammation ; Intercellular Adhesion Molecule-1 ; Iris ; Lymphocyte Function-Associated Antigen-1 ; Rats ; Salmonella ; Uvea ; Uveitis* ; Uveitis, Anterior

AIMTo investigate the mechanism of the vascular endothelial cell (VEC) injury caused by freezing/thawing.

METHODSThe frozen/thawed neutrophil (PMN) model was founded by freezing PMNs with a rate cooling instrument and then rewarming them in a water bath, the PMNs used here were separated from rat's peripheral blood using density gradients centrifugation techniques. The expression of LFA-1 on the surface of frozen/thawed PMNs was observed at 4 h,12 h and 24 h after freezing/thawing. After co-incubating untreated VECs with frozen/thawed PMNs, we detected the VEC injury and the changes in PMN-VEC adhesion.

RESULTS(1) The PMNs LFA-1 expression increased in a time-dependent manner within 24 h after the freezing/thawing of PMNs. (2) After co-incubating untreated VECs with frozen/thawed PMNs, the adhesion between frozen/thawed PMNs and VECs increased and VEC injury occurred. (3) Monoclonal antibody against LFA-1 could block the PMN-VEC adhesion and subsequently attenuated the VEC injury.

CONCLUSIONThe freezing/thawing of PMNs can elicited an increase in PMN LFA-1 expression and trigger the PMN-VEC adhesion and subsequently bring about the VEC injury.

Animals ; Cells, Cultured ; Endothelial Cells ; cytology ; Freezing ; Lymphocyte Function-Associated Antigen-1 ; metabolism ; Neutrophils ; cytology ; metabolism ; Rats ; Rats, Wistar

LFA-1/ICAM-1 costimulation plays an important role in immunologic reaction of many different T cell populations. After TCR/CD3 complex cross-linking MHC/peptide, LFA-1, expressed on T cell increases a higher affinity and avidity for ICAM-1 rapidly. LFA-1 is a key molecule in formation of the immune synapse. LFA-1/ICAM-1 costimulation can engage various signaling events of T cell by up-regulating the activities of PI 3-kinase, sphingomyelinase, and c-Jun NH2-terminal kinase. With the costimulation of LFA-1/ICAM-1, engagement of TCR molecules results in a significant increase of T cell activities, including higher Th1 cytokines production, strongly proliferative response and higher T cell cytotoxicity.
Animals ; Cytokines ; biosynthesis ; Cytotoxicity, Immunologic ; Humans ; Intercellular Adhesion Molecule-1 ; physiology ; Lymphocyte Activation ; Lymphocyte Function-Associated Antigen-1 ; physiology ; Signal Transduction ; T-Lymphocytes ; immunology

The basic route and mechanism for diapedesis has not yet to be fully defined. Here we present evidence that "cell-cell separation" between endothelial cells (ECs) may provide a route for leukocyte diapedesis. We unexpectedly found that extensive interaction between peripheral blood leukocytes and ECs that were activated by TNF-alpha induced the opening of EC contacts and, surprisingly, resulted in cell-cell separation. This event was specific to the intercellular adhesion molecules-1 (ICAM-1)/leukocyte function-associated antigen-1 interaction, as demonstrated by the following: (1) ICAM-1 expression correlated with increased EC contraction; and (2) the blocking of ICAM-1 selectively inhibited EC separation. Thus, we suggest that "cell-cell separation" could be a mechanism for diapedesis in situations that may require massive leukocyte infiltration.
Cell Movement ; Cells, Cultured ; Endothelial Cells/*cytology/metabolism ; Flow Cytometry ; Fluorescent Antibody Technique ; Humans ; Intercellular Adhesion Molecule-1/*metabolism ; Leukocytes/cytology/*immunology ; Lymphocyte Function-Associated Antigen-1/*metabolism

The basic route and mechanism for diapedesis has not yet to be fully defined. Here we present evidence that "cell-cell separation" between endothelial cells (ECs) may provide a route for leukocyte diapedesis. We unexpectedly found that extensive interaction between peripheral blood leukocytes and ECs that were activated by TNF-alpha induced the opening of EC contacts and, surprisingly, resulted in cell-cell separation. This event was specific to the intercellular adhesion molecules-1 (ICAM-1)/leukocyte function-associated antigen-1 interaction, as demonstrated by the following: (1) ICAM-1 expression correlated with increased EC contraction; and (2) the blocking of ICAM-1 selectively inhibited EC separation. Thus, we suggest that "cell-cell separation" could be a mechanism for diapedesis in situations that may require massive leukocyte infiltration.
Cell Movement ; Cells, Cultured ; Endothelial Cells/*cytology/metabolism ; Flow Cytometry ; Fluorescent Antibody Technique ; Humans ; Intercellular Adhesion Molecule-1/*metabolism ; Leukocytes/cytology/*immunology ; Lymphocyte Function-Associated Antigen-1/*metabolism

LFA-1 and ICAM-1 mediate a bi-directional signaling across the cell membrane which is essential for biological functions of lymphocyte, including exudation, activation, adhesion, immunosurveillance as well as immuno-logical synapse formation. The signal transducing is a dynamic process and dependent on the binding capacity between LFA-1 and ICAM-1. The affinity and the avidity of LFA-1 are two major regulation forms in this process. Phosphorylation of LFA-1 and cytoskeleton protein talin 1 play a critical role in signal transducing. In biology of lymphocyte, LFA-1 and ICAM-1 interaction forms the co-stimulatory signal to promote activation, proliferation and division. In this article the regulation of binding capacity between LFA-1 and ICAM-1, the regulation of LFA-1 subunit phosphorylation, the role of talin1 in signaling transduction of LFA-1 and ICAM-1, the synergic stimulatory signaling of LPA-1 and ICAM-1 were reviewed.
Humans ; Intercellular Adhesion Molecule-1 ; metabolism ; physiology ; Ligands ; Lymphocyte Function-Associated Antigen-1 ; metabolism ; physiology ; Lymphocytes ; cytology ; immunology ; metabolism ; Phosphorylation ; Signal Transduction ; physiology ; Talin ; metabolism