Recombinant Gamma Interferon Increases the Binding of Peripheral Blood Mononuclear Leukocytes and a Leu-3+ T Lymphocyte Clone to Cultured Keratinocytes and to a Malignant Cutaneous Squamous Carcinoma Cell Line That Is Blocked by Antibody Against the LFA-1 Molecule

Department of Pathology, Stanford University School of Medicine, California.
Journal of Investigative Dermatology (Impact Factor: 7.22). 02/1988; 90(1):17-22. DOI: 10.1111/1523-1747.ep12462420
Source: PubMed


Because keratinocytes (KCs) express HLA-DR in a wide variety of skin diseases in which mononuclear leukocytes are observed in close apposition to KCs (i.e., graft-versus-host disease), and since gamma interferon (IFN-gamma) induces HLA-DR expression on KCs, we asked whether IFN-gamma treatment of KCs would influence the adherence of mononuclear leukocytes. When allogeneic peripheral blood mononuclear leukocytes (PBML) and a Leu-3+ T cell clone were coincubated with IFN-gamma-treated KCs (300 U/ml, 3 days), there was a marked increase in binding compared with nontreated KCs. Similar binding results were obtained using a cutaneous squamous carcinoma cell line (SCL-1) after IFN-gamma treatment. The IFN effect was relatively specific for IFN-gamma, as neither IFN-alpha nor -beta had any effect. Tumor necrosis factor exposure (500 U/ml, 3 days) increased the binding of the Leu-3+ T cell clone to both KCs and SCL-1 cells. Neutrophils displayed a less marked (but statistically significant) increase in binding to IFN-gamma-treated KCs. Using the Leu-3+ cell clone and SCL-1 cells, detailed kinetic analysis of the effect of IFN-gamma on binding was performed. The increased adherence between the cells began to appear after only 7 hours of treatment with r-IFN-gamma (300 U/ml) and reached a plateau at 48 hours, with significantly enhanced binding continuing for at least 48 hours after removal of IFN-gamma. The mechanism of binding was explored by preincubation of the PBML/Leu-3+ T cells with anti-LFA-1 (lymphocyte function-associated antigen) antibody (0.6-6.0 micrograms/ml), which totally inhibited the binding with no effect by anti-LFA-2 or -3 or class I or II antibodies despite documented binding of these antibodies to the cells. These results suggest that, after exposure to IFN-gamma, the ability of KCs to bind mononuclear leukocytes is strongly enhanced, and this adherence may be important in leukocyte trafficking in the skin as well as contributing to altered KC-leukocyte interaction, which may be of fundamental importance in a variety of skin disease.

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Available from: Alan M Krensky, Apr 23, 2014
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    ABSTRACT: Recombinant gamma interferon (r-IFN-gamma) increases the adherence of peripheral blood mononuclear leukocytes (PBMLs) to cultured keratinocytes and cutaneous microvascular endothelial cells (MECs). To determine which specific type of PBMLs bound to these r-IFN-gamma treated cells, we performed immunophenotyping on the adherent PBMLs. The adherent PBMLs were detached from the r-IFN-gamma treated keratinocytes and MECs by adding EDTA, and collected by cytocentrifugation, followed by immunocytochemical staining using a panel of monoclonal antibodies. Our results reveal that the relative adherent population of PBMLs was composed of approximately 60%-70% monocytes and 18%-24% Leu 2+ T lymphocytes (T-cytotoxic/suppressor) which preferentially bound to r-IFN-gamma treated keratinocytes and MECs. There was some lesser binding by Leu 3 + lymphocytes (T-helper/inducer); approximately 8%, and no binding of B lymphocytes. Since r-IFN-gamma also induced HLA-DR expression in keratinocytes and MECs, these in vitro data suggest that r-IFN-gamma may play an important role in the immunobiology of diverse skin diseases such as graft vs host disease, lichen planus, and other inflammatory dermatoses, because the keratinocytes express HLA-DR and the predominant T-cell subset in the epidermis is Leu 2 + (over the Leu 3 + T cell) in all of these conditions. These results represent a direct attempt to explain in situ immunophenotypic mononuclear leukocyte subset distribution patterns by using r-IFN-gamma and purified cultured cells such as keratinocytes and MECs. We propose that IFN-gamma, by both increasing the adherence of PBMLs, and promoting selective binding of monocytes and Leu 2 + T lymphocytes to both keratinocytes and MECs, may be important in regulating PBML localization and recirculation in the skin.
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    ABSTRACT: To extend our previous observation in which the binding of resting allogeneic peripheral blood mononuclear leukocytes (PBML) to recombinant gamma-interferon (IFN-gamma)-treated keratinocytes was characterized, we examined the influence of phorbol ester activation of the PBML to both autologous and allogeneic IFN-gamma-treated keratinocytes. The activation of PBML by phorbol esters (5 to 100 ng/ml) for brief periods of time (5 min to 1 h) at 37 degrees C led to an increase in the relative percentage of adherence to IFN-gamma-treated keratinocytes from 15% for non-activated PBML to 30% for phorbol ester-treated PBML. A biologically inert phorbol ester derivative did not enhance the binding reaction. No significant binding of phorbol ester-activated PBML was observed to non-IFN-gamma-treated keratinocytes. Both reduction in temperature to 4 degrees C and preincubation of the phorbol ester-treated PBML with anti-LFA-1 monoclonal antibody, led to complete inhibition of this adherence reaction indicating a role for the LFA-1 molecule in phorbol ester-activated PBML/IFN-gamma-treated keratinocyte reactions. Immunophenotypic analysis of the adherent cell population of the phorbol ester-activated PBML to the IFN-gamma-treated keratinocytes revealed that the predominant adherent cell type was the CD8+ T-cell subset (44%) versus the CD4+ T-cell subset (33%) with 23% monocytes and no binding of B lymphocytes. These results suggest that phorbol ester-activated PBML binds twice greater than resting PBML to IFN-gamma-treated keratinocytes, and this increased adherence may further contribute to homing of activated lymphocytes to the epidermis and mononuclear cell trafficking in the skin of inflammatory dermatoses.
    Journal of Investigative Dermatology 06/1988; 90(5):684-9. DOI:10.1111/1523-1747.ep12560903 · 7.22 Impact Factor
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    ABSTRACT: We investigated the in vivo effect of recombinant interferon-gamma (IFN-gamma) and tumour necrosis factor alpha (TNF-alpha) treatment of mice on the development of the delayed-type hypersensitivity (DTH) reaction and lichenoid tissue reaction (LTR) following the local injection of cloned autoreactive T cells. Both the DTH reaction and the LTR were significantly enhanced by pretreatment with IFN-gamma, but not with TNF-alpha. Induction of class II MHC antigens on keratinocytes was not essential for the enhancement by IFN-gamma. Administration of anti-IFN-gamma antibody reduced the DTH reaction and LTR, although complete inhibition was not observed with our treatment regimen. The ability of IFN-gamma to increase the number of the cloned T cells invading the epidermis in vivo, is in keeping with our previous observation that IFN-gamma treatment of cultured keratinocytes markedly increased the adherence reaction between T cells and keratinocytes in vitro.
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