Eosinophils and T lymphocytes are thought to be involved in allergic airway inflammation. Both cells express the alpha 4 beta 1-integrin, very late antigen-4 (VLA-4, CD49d/CD29); alpha 4-integrins can promote cellular adhesion and activation. Therefore, we examined the in vivo effects of a blocking anti-alpha 4 monoclonal antibody, HP 1/2, on antigen-induced early and late bronchial responses, airway hyperresponsiveness, inflammatory cell influx, and peripheral leukocyte counts in allergic sheep. Sheep blood lymphocytes, monocytes, and eosinophils expressed alpha 4 and bound HP 1/2. In control sheep, Ascaris antigen challenge produced early and late increases in specific lung resistance of 380 +/- 42% and 175 +/- 16% over baseline immediately and 7 h after challenge, respectively, as well as airway hyperresponsiveness continuing for 14 d after antigen challenge. Treatment with HP 1/2 (1 mg/kg, i.v.) 30 min before antigen challenge did not affect the early increase in specific lung resistance but inhibited the late-phase increase at 5-8 h by 75% (P < 0.05) and inhibited the post-antigen-induced airway hyperresponsiveness at 1, 2, 7, and 14 d (P < 0.05, for each time). Intravenous HP 1/2 given 2 h after antigen challenge likewise blocked late-phase airway changes and postchallenge airway hyperresponsiveness. Airway administration of HP 1/2 (16-mg dose) was also effective in blocking these antigen-induced changes. Response to HP 1/2 was specific since an isotypic monoclonal antibody, 1E6, was ineffective by intravenous and aerosol administration. Inhibition of leukocyte recruitment did not totally account for the activity of anti-alpha 4 antibody since HP 1/2 neither diminished the eosinopenia or lymphopenia that followed antigen challenge nor consistently altered the composition of leukocytes recovered by bronchoalveolar lavage. Because airway administration of HP 1/2 was also active, HP 1/2 may have inhibited cell activation. Reduction of platelet-activating factor-induced eosinophil peroxidase release from HP 1/2-treated eosinophils supports such a mechanism. These findings indicate a role for alpha 4-integrins in processes that lead to airway late phase responses and persisting airway hyperresponsiveness after antigen challenge.

Endothelial attachment is the initial step in leukocyte recruitment into developing atherosclerotic lesions. To determine whether vascular cell adhesion molecule-1 (VCAM-1) expression may play a role in inflammatory cell recruitment into human atherosclerotic lesions, immunohistochemistry was performed with a polyclonal rabbit antisera, raised against recombinant human VCAM-1, on 24 atherosclerotic coronary plaques and 11 control coronary segments with nonatherosclerotic diffuse intimal thickening from 10 patients. Immunophenotyping was performed on adjacent sections to identify smooth muscle cells, macrophages, and endothelial cells. To confirm VCAM-1-expressing cell types, double immunostaining with VCAM-1 antisera and each of the cell-specific markers and in situ hybridization were performed. All atherosclerotic plaques contained some VCAM-1, compared to 45% of control segments. VCAM-1 was found infrequently on endothelial cells at the arterial lumen din both plaques (21%) and in control segments (27%), but was prevalent in areas of neovascularization and inflammatory infiltrate in the base of plaques. Double immunostaining and in situ hybridization confirmed that most VCAM-1 was expressed by subsets of plaque smooth muscle cells and macrophages. The results document the presence of VCAM-1 in human atherosclerosis, demonstrate VCAM-1 expression by human smooth muscle cells in vivo, and suggest that intimal neovasculature may be an important site of inflammatory cell recruitment into advanced coronary lesions.

VCAM-1, a leukocyte adhesion molecule expressed by cytokine-activated endothelial cells in culture, may mediate mononuclear leukocyte infiltration in vessels and interstitium in solid organ allograft rejection. Using the avidin-biotin immunoperoxidase technique and an affinity-purified rabbit polyclonal antisera to recombinant human VCAM (rVCAM Ab) which works in methyl Carnoy's fixed tissues, we studied the expression of this molecule in biopsies of transplanted kidneys (N = 34) with and without features of rejection and allograft nephrectomies (N = 17) as well as nontransplanted control tissues (N = 26). The rVCAM Ab showed a population of reactive endothelial cells limited to sites of prominent subendothelial leukocytic cell infiltration in arteries and veins, and occasional peritubular capillaries (PTC) in rejecting allografts. Endothelial expression of VCAM was rarely identified in biopsies showing interstitial rejection only or cyclosporine toxicity, usually in PTC, and was only rarely encountered in nontransplanted control tissues. Apparent de novo expression of VCAM-1 by arterial smooth muscle cells and mesangial cells was present in cases of severe rejection. In addition, a population of cells (DC) with dendritic morphology was identified by rVCAM Ab within sites of lymphoid cell aggregation in rejecting allografts. Further evidence that these cells represent true DC was obtained by identification of VCAM-1 positive, morphologically similar cells in both germinal centers and interfollicular areas of all seven reactive lymph nodes tested; and by similar staining of these cells in the allografts and lymph nodes by antibodies to nerve growth factor receptor and the complement receptor CR1, previously shown to recognize DC. DCs were generally not seen in uninflamed normal control organs or portions of allografts uninvolved by lymphoid aggregates. Enhanced tubular epithelial cell expression of VCAM-1 was also present in rejecting allografts. All staining could be abolished by absorption of the antisera with VCAM-1 transfected, but not ICAM-1 or ELAM-1 transfected, CHO cells. In situ hybridization studies utilizing a cDNA probe to human VCAM-1 demonstrated mRNA production by glomerular, tubular and vascular cells corresponding to sites where the protein was immunohistochemically localized.(ABSTRACT TRUNCATED AT 400 WORDS)

B lymphocytes express several adhesion molecules that are involved in cell-cell and cell-extracellular matrix interactions. The alpha4beta1 integrin VLA-4, expressed on pre-B and mature/activated B cells, mediates adhesion of these cells to its two ligands, VCAM-1 and fibronectin. Recent evidence suggests that VLA-4 is involved in T lymphocyte activation; however, relatively little is known of the role of VLA-4 in B cell differentiation. To begin to assess the potential involvement of VLA-4 in B cell activation, we have examined the effect of ligation of VLA-4 on protein tyrosine phosphorylation in B cells. We found that cross-linking of VLA-4 by either mAb or natural ligands (i.e., VCAM-1 and the FN-40 cleavage fragment of fibronectin) induced the tyrosine phosphorylation of a 110-kDa protein in a human pre-B cell line (Nalm-6), an EBV-transformed B cell line (SB), and normal tonsillar B cells. These findings suggest that VLA-4 can activate a tyrosine kinase in B cells and B cell lines. These signals may be involved in the subsequent differentiation of pre-B and mature B cells within specific microenvironments where VLA-4 mediated adhesion is operational.