Molecular mechanisms underlying lymphocyte recirculation II. Differential regulation of LFA‐1 in the interaction between lymphocytes and high endothelial cells

Abstract

Although it has been suggested that LFA-1 is one of the important molecules mediating interaction between lymphocytes and high endothelial (HE) cells, the implication was based on the observation that lymphocyte binding to high endothelial venules in frozen lymph node sections was partially inhibited by anti-LFA-1 monoclonal antibody at 4 degrees C. However, it has previously been unequivocally demonstrated that LFA-1 molecule is unable to function at this low temperature. To assess the actual involvement of LFA-1 in lymphocyte-HE cell interaction at body temperature, we examined effects of newly developed anti-rat LFA-1 and anti-rat ICAM-1 monoclonal antibody on binding of resting or activated lymphocytes to a rat HE cell line at 37 degrees C. We found that (a) LFA-1/ICAM-1-independent pathway was predominant in the interaction between resting lymphocytes and HE cells, indicating that LFA-1 functions little, if any, in the interaction and (b) lymphocyte triggering through CD3 significantly increased binding inducing a LFA-1/ICAM-1-dependent pathway, whereas phorbol 12-myristate 13-acetate stimulation also enhanced lymphocyte binding but inducing a LFA-1-dependent/ICAM-1-independent pathway. In both cases no apparent alteration in LFA-1 expression was observed, suggesting that the increase in lymphocyte adhesion was not due to quantitative but a qualitative change induced in LFA-1 molecule upon lymphocyte stimulation. These findings suggest that LFA-1 is normally inert but can be "switched" upon lymphocyte stimulation to participate in lymphocyte-HE cell interaction, and that the LFA-1's ligand specificity can be differentially regulated at the lymphocyte-HE venule interface.

Impairment of selectin-mediated leukocyte adhesion to venular endothelium in spontaneously hypertensive rats.

“…The mesenteric microcirculation was visualized through an intravital microscope (x 55 water immersion objective; E Leitz Wetzlar GmbH, Wetzlar, Germany) via a CCD television camera (model 2362; Javelin Electronics, Tokyo, Japan) to investigate venular leukocyte behavior while monitoring arteriolar and venular erythrocyte velocity ( 11 ). Single unbranched venules (25)(26)(27)(28)(29)(30)(31)(32)(33)(34)(35)(36)(37)(38)(39)(40) prm in diameter) were selected for study. Centerline erythrocyte velocity (VR) was measured by the cross-correlation velocimeter (IPM, Inc., La Mesa, CA) and was calibrated against a rotating glass disk coated with erythrocytes rotating with known velocity.…”

“…Neutrophils in the marginating pool were isolated from WKY and SHR by a hetastarch exchange transfusion (26). The cell suspension (1 x 106 cell/ml) was incubated with 10IM 2', 7-bis (carboxyethyl)-5(6) carboxyfluorescein tetra-acetoxymethyl ester (BCECF-AM; Dojindo Laboratories, Kumamoto, Japan) for 1 h at 370C, washed and resuspended in the modified Eagle's medium (MEM), and served as a sample for cell binding assay (27). Unstimulated neutrophils (1 x 106 cells/ ml) or neutrophils stimulated with phorbol myristate acetate (50 pg/ ml, for 10 min) were inoculated onto the wells containing CHO cells in a final volume of 100 Ml and incubated for 30 min at 37TC.…”

“…The well plates were then inverted for 15 min at room temperature to remove nonadherent neutrophils. After the adherent cells were lysed with 50 Al of 0.1% NP-40, the number of these cells was calculated on the basis of a fluorescence readout at 530 nm under the excitation at 490 nm using an automated microplate fluorometer (MTP; Corona Electric Co., Ibaraki, Japan) (27). Effects of pretreatment with mAbs against CD18 (WT-3, 10 pg/ml at final concentration) or against ICAM-1 (1A29, F(ab')2 fragments, 16 pg/ml at final concentration) were evaluated in selected experiments.…”

Measurement of Human Intercellular Adhesion Molecule 1 in the Blood Is Useful for Predicting the Occurrence of Hepatocellular Carcinomas from Chronic Hepatitis C and Liver Cirrhosis

“…Of these, 6 patients with CH (7.5%) and 1 with LC ceased visiting the hospital within 2 years, and 1 patient with LC (5.3%) died of liver failure within 1 year. The remaining 74 patients with CH [mean age 52.9 years, proportion of males 60.8%, mean alanine aminotranferase (ALT) value 58.2 U/l, mean platelet count 187,000/mm 3 , mean observation period 6.94 years] and the remaining 17 patients with LC (mean age 63.1 years, proportion of males 55.6%, mean ALT value 57.4 U/l, mean platelet count 88,720/mm 3 , mean observation period 4.81 years) were included in this study. All of the patients were positive for serum HCV RNA and HCV antibody (second-generation enzyme-linked immunosorbent assay, EIA; Abbott, Tokyo, Japan) and were observed for more than 2 years.…”

Functional and Phenotypical Analysis of Subsets of Rat CD4+ T Cells

“…Murine mAb 1A29 and WT. 1 which detect rat antigen equivalent to human CD54 (intercellular adhesion molecule-1; ICAM-1) and CDlla (LFA-1), respectively, were kindly supplied by Dr. Miyasaka, Department of Immunology, Tokyo Metropolitan Institute of Medical Science (32,33). MRC OX-22 and OX-1 that recognize high molecular isoforms (200 and 220 kDa) of the rat CD45 and all isoforms (180,190,200, and 220 kDa) of the rat CD45, respectively (29), were purchased from Cedarlane Lab.…”