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C Dezutter-Dambuyant

Publications and source records attributed to C Dezutter-Dambuyant.

At least 91 records · Page 5Linked to original sources
PubMed1988

Do circulating OKT6-reactive cells belong to Langerhans cell lineage?

A few studies have been made in characterizing the phenotype, ontogeny, ultrastructure, and cytochemistry of circulating cord blood cells. Such studies have been hampered by the difficulty in obtaining pure populations of these cells. We therefore sought to obtain T6-positive cell-enriched populations from human cord blood using panning method. In cord blood, dendritic OKT6-labeled cells were classified in two types: some were similar to lymphocyte-like cells and some were consistent with monocyte-like cells. Both types lacked intracytoplasmic Birbeck granules. Homogeneous labeling of HLA-DR antigens were observed in both types. Heterogeneous labeling of T6 antigen was observed in both types. The monocyte-like cells have developed Birbek granule-like structures after exposure to OKT6 and immuno-adherence. The results suggest several different putative lineages of these OKT6-reactive cells: early precursors of dendritic cells, immature neonatal lymphoid subsets, and immature macrophagic leukocytes.

Adult↗
PubMed1988

Culture of putative Langerhans cell bone marrow precursors: characterization of their phenotype.

We searched for the presence of human CD1-positive cells in bone marrow populations in order to characterize putative Langerhans cell precursors. Bone marrow progenitors were cultured in 0.8% methylcellulose supplemented with 10% granulocyte-macrophage (GM) colony-stimulating factor(s) GCT and HTB9. We compared the kinetics of these two factors and found that GCT was the more appropriate for our study. After 8 days of culture, colony-forming units of granulocyte-macrophages (CFU-GM) were tested for the presence of CD1-positive cells using the immunofluorescence technique. Positive cells were counted by cytofluorometric analysis: 9.4% CD1a (BL6), 13.4% CD1c (L161), 4.3% CD1b (NuT2), 4.6% CD2 (T11), and 25.5% CD33 (My9). Ultrastructural features and phenotype were then specified by the immunogold labeling technique using electron microscopy. A subpopulation of CD1-positive cells showed the ultrastructural morphology of bone marrow pro-monocyte/monocyte cells. By using well-characterized monoclonal antibodies, it was demonstrated that these cells expressed the following phenotype: CD14+, CD33+, CD4+, HLA-DR+, HLA-DP+, HLA-DQ-, OKT10-, CD2-. These data indicate that these bone marrow promonocyte/monocyte progenitors express a phenotype similar to that of epidermal Langerhans cells but the density of each antigen is much lower than that observed on mature skin dendritic cells.

Antibodies, Monoclonal↗
PubMed1987

Effects of trypsin on the in situ identification of epidermal cell membrane antigens.

In this work the role of trypsin in revealing epidermal cell surface antigens, with the use of immunological markers, was investigated. Two monoclonal antibodies (MCA) were used, the first: D47, belongs to the first cluster of differentiation and recognizes a membrane antigen of human thymocytes; the second HLA-ABC-m3, is an anti-HLA-B27 MCA. Preliminary treatment with various concentrations of trypsin was performed on frozen skin sections and followed by indirect immunofluorescence. D47 reacted with epidermal dendritic cells only after trypsin pretreatment of skin sections. In addition a mild preliminary trypsinization was shown to increase in situ immunoreactivity of MCA HLA-ABC-m3 with epidermal cells. Best results were obtained when trypsin concentrations ranging from 2.5 to 5 micrograms/ml were applied for 10 min at 37 degrees C. Preliminary trypsinization may be of interest for a better exposure of some surface antigens to immunohistochemical markers.

Antibodies, Monoclonal↗
PubMed1987

Antigenic thymus-epidermis relationships. Reactivity of a panel of anti-thymic cell monoclonal antibodies on human keratinocytes and Langerhans cells.

We have investigated on human skin the reactivity of a panel of 42 anti-thymus monoclonal antibodies (MCA) supplied by the Third International Workshop and Conference on Human Leucocyte Differentiation Antigens, Oxford, 1986. MCA of the first cluster of differentiation (CD1) define a group of surface molecules expressed by cortical thymocytes. Some of them (OKT6, M241 and Na1/34) have been shown to react in normal human skin with the epidermal Langerhans cells (LC). Twenty-two CD1 MCA were investigated in the present study. On normal human skin, 13 MCA reacted with LC in situ. This result suggests and confirms the heterogeneity of CD1 MCA. Recently, some of them were shown to recognize biochemically different molecules and/or epitopes of thymocytes. In addition, 20 anti-thymic epithelium MCA were tested on human skin. The MCA which only reacted with the thymic epithelial cell network (except Hassall's corpuscles) decorated only the epidermal basal cell layer. The MCA which reacted with all the thymic epithelial cells (including Hassall's corpuscles) decorated all the epidermal cell layers. These results confirm the heterogeneity of the thymic epithelial microenvironment and underline the antigenic similarities between the thymic epithelial structures and the different epidermal cell layers. The existence of bone-marrow-derived CD1-positive cells (thymocytes or LC) in an epithelial cell network (the thymus and the epidermis) focus the speculation around the immunological role of the epidermal basal cell layer in the T cell education and the exact lineage of the epidermal LC.

Antibodies, Monoclonal↗
PubMed1986

Loss of allogeneic T-cell activating ability and Langerhans cell markers in human epidermal cell cultures.

Human Langerhans cells are the only epidermal cells that express the T6 and HLA-DR antigens and are responsible for the in vitro allogeneic T-cell proliferative responses in the mixed skin cell lymphocyte reaction (MSLR). To investigate the presence of Langerhans cells in normal human epidermal cell cultures, epidermal cell suspensions obtained from normal human skin specimens and from the subsequent epidermal cell cultures were analyzed by indirect immunofluorescence for the presence of T6 and HLA-DR determinants. In parallel, MSLRs were conducted with suspensions of cultured epidermal cells as stimulatory cells. These studies present evidence that when human epidermal cells are grown in culture, they loose both the ability to stimulate the proliferation of allogeneic T lymphocytes in vitro and their expression of HLA-DR and T6 antigens. The T6 antigens were lost during the first 2 weeks of culture, while HLA-DR determinants were still expressed by a small number of cells and were progressively lost through duration of cultures. The loss of HLA-DR antigens closely paralleled the progressive inability of human epidermal cells in culture to stimulate allogenic T cells in MSLR.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1986

Comparative phenotypic and ultrastructural characteristics of OKT6-positive cells in normal peripheral blood (adult and infant), in cord blood and in epidermis.

Simple and rapid procedures have been developed to isolate T6-positive cells in epidermal cell suspensions, in peripheral blood of adult, and in cord blood. The isolated T6-positive cells were tested by double immunogold labelling at ultrastructural level to analyze their phenotype (T6/HLA-DR antigens) and their morphological features in order to understand their origin, their fate, and their relationship to the early precursors of dendritic cells, immature neonatal lymphoid subsets, and immature macrophagic leukocytes.

Antibodies, Monoclonal↗
PubMed1986

Subclustering of CD1 monoclonal antibodies based on the reactivity on human Langerhans cells.

Among the differentiation antigens expressed by lymphoid cells, CD1 monoclonal antibodies (MCA) distinguish a group of antigens expressed by cortical thymocytes. Some of them (OKT6 and BL6) have been shown to react with normal human skin Langerhans cells (LC). Using indirect immunofluorescence on skin sections and LC-enriched epidermal cell suspensions obtained by skin trypsinisation, we have studied the reactivity of eight new CD1 MCA. Our immunocytochemical observations indicate that four groups of CD1 MCA can be distinguished on the basis of their reactivity on LC. NA.1.84, SK9/Leu6, 10 D.12.2 and I19 identify LC in skin and in suspensions, but 4A.76 and NUT2 were negative in both. M.241 became negative after trypsinisation of epidermal cells and D47 reacted with an antigen expressed on LC only after enzymatic treatment. These results demonstrate the heterogeneity of the antigens recognized by CD1 MCA. Some of them do not react with normal human LC. Some antigens appear to be masked in the epidermis and are expressed only after trypsin treatment. Our data confirm the heterogeneity of CD1 molecules recently documented on the basis of biochemical analysis.

Antibodies, Monoclonal↗
PubMed1986

Reactivity pattern of anti-CD1 and anti-HLA class II monoclonal antibodies with human eccrine sweat glands.

The known cross-reactivity of monoclonal antibodies prepared against CD1 and HLA-DR antigens with skin components prompted us to study the reactivity pattern of human eccrine sweat glands with a panel of monoclonal antibodies directed against CD1 antigens (OKT6, BL6, D-47) and against HLA-class II antigens (anti-DR, BL2, LEU-10, IV-D12, MAJA-7). The labelling pattern of eccrine glands with the panel of monoclonal antibodies used in this study permits to establish three different antigenic compartments on eccrine glands: 1) acrosyringium and distal part of dermal duct anti-DR+, BL2+, LEU-10+, IV-D12+; 2) proximal part of dermal duct MAJA-7+; 3) secretory part D-47+. The immunological markers used in this work provide a useful tool for investigation of eccrine gland differentiation and human eccrine glandular pathology.

Antibodies, Monoclonal↗
PubMed1986

[Expression of class II HLA antigens (HLA-DR, DQW1, DQW3) in normal skin and cutaneous pathology].

We report the cutaneous expression of class II HLA antigens disclosed by immunohistochemical staining of normal and lesional skin with monoclonal antibodies (MoAbs) reacting with HLA-DR, DQW1 and DQW3 antigens. Briefly, we disclosed: on normal human skin: 1) The Langerhans cells (LC) HLA-DR+, DQW1+; 2) the acrosyringium HLA-DR+, DQW1+, DQW3+; 3) the dermal vessel endothelial cells HLA-DR+. On lesional skin: 1) The LC were found HLA-DQW3+ in the lesional skin of some cutaneous diseases; this expression was never shown on LC of normal human skin; 2) the epidermal keratinocytes disclosed an uniform membrane expression of HLA-DR antigens in some cutaneous diseases; this kind of expression was not found by immunostaining with MoAbs directed against HLA-DQ antigens; 3) in psoriatic lesions some keratinocytes disclosed an heterogeneous expression of HLA-DR, DQW1 and DQW3 antigens; 4) tumoral cells from cutaneous malignant melanomas were shown to be HLA-DR+, DQW1+, DQW3+. The HLA-DQW3 expression on the LC of lesional skin is in favour with a modulation of HLA-DQW3 expression by unknown factors present in pathological skin. The HLA-DR expression on epidermal keratinocytes suggests a functional collaboration of keratinocytes with LC in the genetic restriction of cutaneous immune reactions.

Antibodies, Monoclonal↗
PubMed1985

Flow cytometry sorting of unlabelled epidermal Langerhans cells using forward and orthogonal light scatter properties.

A variety of techniques based on the presence of specific markers has been proposed to enrich Langerhans cells from epidermal cell suspensions. Computer analysis of multiparameter flow cytometry records involving forward angle and orthogonal scattered light and immunofluorescence of epidermal cells allowed us to determine the scattering properties of Langerhans cells. Unlabelled cells sorted according to these properties were shown to be Langerhans cells by electron microscopy and/or subsequent labelling by anti-HLA-DR monoclonal antibody. The relevance of this method is discussed to sorting viable Langerhans cells which may be used in functional studies and for establishing long-term culture.

Female↗
PubMed1985

Immunogold technique applied to simultaneous identification of T6 and HLA-DR antigens on Langerhans cells by electron microscopy.

A double-labeling immunogold technique in electron microscopy and specific monoclonal antibodies to surface antigens of Langerhans cells (OKT6 and BL2) were applied to assess directly the coexpression of two cell surface antigens (T6 and HLA-DR antigens) in a heterogeneous epidermal cell suspension. Electron microscopic examination of double-labeled cells revealed that all Birbeck granule-containing Langerhans cells bound OKT6 and BL2. The preparation of markers with colloidal gold particles and the procedure for double labeling are described. Several problems related to the steric hindrance and current artifacts are illustrated by micrographs and also discussed.

Antibodies, Monoclonal↗
PubMed1985

Recent advances of ultrastructural immunocytochemistry of epidermal Langerhans cells.

Using electron microscopy, the immunological visualization of the membrane antigens of Langerhans cells (LC) can be performed by immunoperoxidase and immunogold techniques. The immunoperoxidase labelling permits the identification of only one antigen and the observation of qualitative variations of surface antigens. The immunogold method allows the identification of one antigen or simultaneously two or three surface antigens using gold particles of various sizes. This technique can be used to quantify the surface density of antigens on the cell membrane. The simultaneous identification of different surface antigens can be correlated with the ultrastructural characteristics of the cells. Using this technique we have recently demonstrated the existence of LC subsets in normal epidermis, and the presence of circulating T6-positive cells in normal subjects. In addition, a very low density of T4 antigenic sites on the LC membrane surface was observed. Several problems of a double-labelling immunogold technique related to steric hindrance and current artifacts are discussed.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1985

Langerhans cell induced cytotoxic T-cell responses against normal human epidermal cell targets: in vitro studies.

We investigated the role of class II MHC (HLA-DR Ia-like) antigen-bearing Langerhans cells in the in vitro generation of human alloreactive cytotoxic T lymphocytes (CTL) directed against epidermal cells (EC). Normal EC in suspensions (from reconstructive surgery specimens) and anti-DR monoclonal antibody plus complement (C')-treated EC were used to stimulate peripheral blood (PB) T cells (T cells allogeneic or autologous to EC) in a classical 6-day human mixed skin cell-lymphocyte culture reaction (MSLR). CTL responses were then tested in 51Cr release assays against cultured EC (EC grown on collagen-coated plates in parallel to MSLR). CTL responses to EC were observed only after allogeneic MSLR and when targets and EC used in MSLR were from the same donor. They were comparable in magnitude to those seen in parallel studies using PBL from the same donor as the stimulating EC as target cells. They were abolished when EC used in MSLR were depleted in class II MHC Langerhans cells (preincubation of EC suspensions with anti-DR + C'). These data show: (a) alloreactive CTL to human normal EC may be generated in MSLR, as previously shown in animals; (b) the MSLR is an in vitro model of primary immunization against EC; (c) Langerhans cells are necessary for the generation of cell-mediated cytotoxicity reactions occurring against epidermal cells.

Cytotoxicity, Immunologic↗
PubMed1985

Human epidermal cell-induced generation of alloreactive cytotoxic T-lymphocyte responses against epidermal cells. Requirement for DR-positive Langerhans cells.

Human epidermal cells act as stimulators in the mixed-skin cell lymphocyte culture reaction (MSLR). To analyse the generation in MSLR of alloreactive cytotoxic T lymphocytes (CTL) in cell-mediated cytolysis of human epidermal cells. a phenomenon suggested by various observations of skin inflammatory processes in vivo, 18-h 51Cr-release assays against epidermal cells cultivated on collagen-coated plates (epidermal cells autologous to the stimulator cells in MSLR) were conducted after allogenic human MSLR. To analyse the role of human Langerhans cells and related epidermal dendritic cells, which are the only cells expressing the DR-Ia-like (class II) antigens in normal epidermis and in suspensions of normal epidermal cells, MSLR and CTL assays were conducted with, as stimulator cells, suspensions of normal human epidermal cells as controls, and, in parallel, suspensions of epidermal cells after preincubation with anti-class II monoclonal antibody and complement. The generation of alloreactive CTL to epidermal cells occurred only after allogenic MSLR and when targets and stimulator cells were from the same donor; it was abolished when epidermal cell suspensions used in MSLR were depleted in HLA-DR-expressing cells. These findings demonstrate that an epidermal cell-induced generation of cell-mediated cytotoxicity to human epidermal cells may occur in vitro. Langerhans cells and other class II antigen-expressing epidermal cells (dendritic indeterminate cells) are necessary for an optimal in vitro sensitization in MSLR and the subsequent generation of alloreactive CTL towards epidermal cells in man.

Cytotoxicity, Immunologic↗
PubMed1984

Ultrastructural immunogold labelling of human Langerhans cells enriched epidermal cell suspension.

Colloidal gold particles are well suited as markers in electron microscopy. Indirect immunogold staining was used to identify cell membrane antigens defined by monoclonal antibodies OKT6 and BL6 on human Langerhans cells (LC) in suspensions. Isolated epidermal cells were obtained by skin trypsinization and enriched or depleted in OKT6 positive on BL6 positive LC using the panning method: incubation of OKT6 or BL6 preincubated cells on immunoglobulin coated dishes. Indirect immunogold staining was then performed after prefixation in 2% paraformaldehyde. In LC enriched suspensions, only LC exhibited a specific membrane labelling with OKT6 or BL6 recognized by the presence of small evently distributed gold granules. Neither Birbeck granules, nor other cytoplasmic organelles, were labelled. No other epidermal cells were found positive. In LC depleted suspensions, no labelling was observed. Immunogold labelling on LC enriched suspensions after panning is now in progress for the qualitative evaluation and the quantitative analysis of cell surface constituents and antigens expressed by human dendritic epidermal cells.

Antibodies, Monoclonal↗
PubMed1984

Role of HLA-DR bearing Langerhans and epidermal indeterminate cells in the in vitro generation of alloreactive cytotoxic T cells in man.

Human epidermal cells (EC) act as stimulator cells in the mixed-skin cell lymphocyte culture reaction (MSLR). To analyze the role of human epidermal Langerhans cells (LC) and indeterminate cells (IC), which are the only cells expressing the DR-Ia-like antigens in normal epidermis, in the generation of alloreactive cytotoxic T lymphocytes (CTL) in cell-mediated cytolysis, 18-hr 51Cr-release assays against PBL targets (targets autologous to stimulator EC) were conducted after allogeneic human MSLR. MSLR and CTL assays were conducted with, as stimulator cells, suspensions of normal human EC as controls, and EC after: (1) preincubation with anti-HLA-DR or OKT6 (specific for LC in EC suspension) monoclonal antibodies; (2) panning, a monolayer technique used to deplete EC suspensions in OKT6 or DR-positive cells. The generation of alloreactive CTL was found to occur only after allogeneic MSLR and when targets and stimulator cells were from the same donor; it was reduced by EC incubation: cytotoxic activity 26.66 +/- 3.84 (controls); 8.8 +/- 3.6 and 7.7 +/- 3.7 (EC incubated with OKT6 or anti-DR, respectively); it was reduced or abolished when the EC used in MSLR were depleted in OKT6 or DR-positive cells by panning. These findings demonstrate that human LC and IC are necessary for an optimal in vitro sensitization in MSLR and the subsequent in vitro generation of alloreactive CTL in man.

Antibodies, Monoclonal↗
PubMed1984

Simultaneous detection of T6 and HLA-DR antigens distinguishes three cell subpopulations in dispersed normal human epidermal cells.

Normal human dendritic epidermal cells (EC) show specific surface membrane markers (T6 and DR antigens). This study presents evidence that, in dispersed EC suspensions, these membrane antigens may distinguish three antigen-positive EC subsets by means of double immunofluorescence labelling: 97.3 +/- 1.2% of the labelled cells were DR(+)/T6(+) while 2.2 +/- 0.9% were DR(+)/T6(-) and 0.5 +/- 0.6% were DR(-)/T6(+). Immunoelectron microscopy with gold particles confirms the co-existence of T6-positive and T6-negative epidermal cells.

Antibodies, Monoclonal↗