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G C Manara

Publications and source records attributed to G C Manara.

At least 37 records · Page 2Linked to original sources
PubMed1990

Human normal-resting epidermal Langerhans cells do express the type 3 complement receptor.

The expression of CR3 by murine-resting epidermal Langerhans cells (LC) is well established, but CR3 expression by human normal-resting epidermal LC has not yet been demonstrated. In this study, highly sensitive immunostaining techniques, such as immunogold labelling in transmission- and scanning-electron microscopy, were used on freshly isolated, LC-enriched, normal human epidermal cells. Human normal resting epidermal LC were found to be CR3+, since a low but significant number of gold granules labelled the plasma membrane of all the LC observed under transmission-electron microscopy, and all the epidermal cells showing LC morphology as observed by scanning-electron microscopy.

Cell Membrane↗
PubMed1990

Immunogold scanning electron microscopy applied to the study of Langerhans cells immunophenotype.

An immunogold technique in scanning electron microscopy was applied to the detection of Langerhans cell surface CD1a antigen in a heterogeneous human epidermal cell suspension. Scanning electron microscopy investigations performed in the secondary electron imaging mode and in the backscattered electron imaging mode allowed to define Langerhans cell surface morphology and the labeling degree, respectively. In particular, the backscattered electron imaging mode of the scanning electron microscope appeared very useful in identifying colloidal gold particles and, as a result, made it possible to make total counts of the labeled surface antigenic sites. Finally, the combination of secondary electron and backscattered electron signals generated mixed images with the Langerhans cell surface morphology well recognizable together with distinct gold particles. The immunogold labeling in scanning electron microscopy here described represents a highly sensitive method and yields a new improvement in the study of Langerhans cells immunophenotype.

Antigens, Surface↗
PubMed1990

Subsets of keratinocytes and Langerhans' cells express epitopes associated with suppressor-inducer capabilities in resting normal human epidermis.

In recent years two cell populations with down-regulatory immune capabilities have been identified in murine epidermis. The present report demonstrates that even in human epidermis at least two populations of cells expressing suppressor-inducer phenotypes (i.e. CD45R-positive) exist, namely small subsets of keratinocytes and Langerhans' cells, respectively. Highly specific and sensitive 5-nm colloidal gold-immunoelectronmicroscopic techniques were carried out using anti-CD45R monoclonal antibodies, on freshly isolated crude epidermal cell suspensions, and 4000 cells were scrutinized in the electron microscope. Over 2% of the total epidermal cell population was CD45R+. Subpopulation analysis revealed that approximately 2% of keratinocytes and about 5% of the total Langerhans' cell population showed strong gold-plasma membrane staining, whilst the remaining epidermal cells were absolutely negative. Heterogeneity of staining together with this somehow surprising distribution of CD45R positivity on non-lymphoid epidermal cells was confirmed by the negative controls. These CD45R+ Langerhans' cells and keratinocytes are clearly candidates for the cells which have been functionally demonstrated as being capable of inducing down-regulation responsiveness in the human epidermis. However, functional investigations are needed to clarify the roles of the CD45R+ keratinocyte and Langerhans' cell subsets in the modulation of cutaneous immune responses.

Antigens, CD↗
PubMed1989

A multiple-staining ultrastructural procedure simultaneously detecting three membrane antigens on suspended cells by monoclonal antibodies and pre-embedding immunogold labelling.

A triple ultrastructural immunogold staining method for the simultaneous demonstration of three surface antigens of peripheral blood mononuclear cells at the electron microscope level is described. A six-step pre-embedding immunoelectron microscopy procedure was developed, using commercially available reagents. The CD11b antigen was first detected, through a two-step (indirect) method with 40 nm-sized gold particles; after a blocking step, the HLA-DR surface antigen was subsequently detected, through a two-step (biotin-streptavidin) method with 20 nm-sized gold particles; the CD4 antigen was finally detected, through a one-step (direct) method, using 5 nm-sized gold particles. Electron microscopic examination revealed firstly the presence of a triple-labelled cell subpopulation, which showed gold granules of the three sizes simultaneously decorating the cell membrane. Thus, the cells of such a subset simultaneously expressed the three antigens investigated. In contrast, either gold particles of only one size or no gold particles were observed on the cell surface of other subpopulations. This technique is a model demonstrating the importance of varying the size of particles in pre-embedding gold immunoelectron microscopy for a better analysis of the expression of surface antigens in isolated cells.

Antibodies, Monoclonal↗
PubMed1989

Adhesion molecules on the plasma membrane of epidermal cells. I. Human resting Langerhans cells express two members of the adherence-promoting CD11/CD18 family, namely, H-Mac-1 (CD11b/CD18) and gp 150,95 (CD11c/CD18).

The CD11/CD18 family of leukocyte adhesion-promoting proteins is comprised of three members, each composed of a shared beta subunit (CD18) noncovalently associated with unique alpha subunits (CD11a, CD11b and CD11c respectively). Such three heterodimers, named LFA-1 (CD11a/CD18), H-Mac-1 (CD11b/CD18) and gp150,95 (CD11c/CD18), are involved in mediating leukocyte adhesion in virtually all phases of the immune responses. Since Langerhans cells are regarded as cutaneous leukocytes, we investigated the expression of the members of the CD11/CD18 family on Langerhans cells. A vast series of immunostaining procedures was carried out, using monoclonal antibodies anti-CD11a, -CD11b, -CD11c, and -CD18. Normal skin frozen sections and epidermal sheets were investigated by immunohistology and immunofluorescence; suspended freshly isolated epidermal cells were processed using immunogold techniques, performed in both transmission and scanning electron microscopy, including double labeling procedures and semiquantitative analysis of the labeled cells. The results demonstrated the expression on the membrane of Langerhans cells of the CD11b, CD11c and CD18 antigens, thus indicating that at least both the H-Mac-1 (CD11b/CD18) and the gp150,95 (CD11c/CD18) members of the CD11/CD18 family are detectable on the cell surface of human resting Langerhans cells. Since both such moieties serve as adhesion molecules in (a) cell-cell interactions and in (b) leukocyte migration and localization, the present results suggest that H-Mac-1 and gp150,95 might display a key role (a) in promoting interactions between Langerhans cells and other cells, and (b) in guiding the migration and localization of Langerhans cells.

Adolescent↗
PubMed1989

Further characterization of the "incipient lesion of chronic stationary type psoriasis vulgaris in exacerbation". The CD4-positive lymphocytes are the prominent cell population infiltrating the dermis.

In previous studies, others and we have demonstrated by immunohistological methods in light microscopy that the prominent cell population infiltrating the psoriatic lesional dermis is represented by CD4-positive cells, thus suggesting an implication of T4 lymphocytes in the development of psoriatic lesion. However, recent findings have indicated that not only T4 lymphocytes, but also macrophage-like cells, are CD4-positive. In the present study, therefore, a highly specific and sensitive 'in situ' gold-immunoelectronmicroscopy procedure developed by us was performed to identify ultrastructurally with precision the nature of the CD4-positive cells infiltrating the dermis of the incipient psoriatic lesion. Although a consistent proportion (28%) of slightly CD4-positive macrophage-like cells was detected, the strongly CD4-positive lymphocytes were the prominent cell subset (51%). The important role presumably played by such CD4-positive lymphocytes in the pathogenesis of the psoriatic lesion is discussed. Our gold-immunoelectronmicroscopy technique improved the study of psoriatic cell subpopulations in situ, since (a) in comparison with immunocytochemistry in light microscopy, it allowed us not only to recognize the ultrastructure of the labelled cells, but also to detect even small amounts of antigen on the cell surface; and (b) in comparison with the commonly used peroxidase immunoelectronmicroscopy techniques, it allowed us to quantitatively assess the expression of antigens (e.g. CD4) on different cell subpopulations (e.g. the low CD4 expression by macrophage-like cells versus the high CD4 expression by T4 lymphocytes).

CD4 Antigens↗
PubMed1989

[Treatment of dry skin by the use of a "physiologic" bath oil].

The results obtained by using a bath oil (Idrovel bath) for treating skin dryness, both primary and associated with pathological conditions, are reported. Twenty patients out of 40 considered for treatment were undergoing photochemotherapy. The patients were evaluated both subjectively and, in order to obtain an objective measuring of skin hydratation, by instrumental capacitance method. The results were very encouraging, characterized by a statistically significant increase of capacitance data after treatment. These results seem to be due to the bath oil formulation, i.e. vegetal polyunsaturated oil mixture with little liquid paraffin content. In conclusion, this formulation is very similar to normal epidermal lipids composition, and seems to represent a good compromise between the occlusive and integrative effects of topical emulsions.

Administration, Cutaneous↗
PubMed1988

Hairy cell leukemia cells express CD1a antigen.

Five patients with hairy cell leukemia (HCL) were studied. Peripheral blood leukocytes, rosette-forming cells (T) and non-T-cells were stained in immunofluorescence by a panel of monoclonal antibodies to investigate the phenotype of HCL cells (HCLC). In all patients HCLC showed B-lymphocyte phenotype, although they were not stained by antibodies reactive for monocytes, natural killer cells, or T-cells. However, in all instances the large majority of HCLC were unexpectedly stained with an antibody (anti-CD1a) usually detectable only in early thymocytes and on Langerhans cells. This finding was further confirmed by immunoelectron microscopy. This type of ambiguity in the lineage of HCL could imply that HCLC might arise from cells differentiated towards the B-cell lineage, still sustaining an early antigen of a different (T) lineage. These results, moreover, extend the range of the known distribution of the CD1a antigen, which could be useful in diagnosing HCL.

Antibodies, Monoclonal↗
PubMed1988

The S-100 beta protein in normal human peripheral blood is uniquely present within a discrete suppressor-T-cell compartment.

The S-100-positive T lymphocytes, and, particularly, the S-100 beta subunit, are restricted, as demonstrated by quantitative subset analysis and double-labeling (gold-peroxidase) immunoelectron microscopy of T-cell subpopulations, to an unique T8-positive cell subset which interestingly was 9.3-negative and CD11b-positive. Since both the T8-positive, 9.3-negative and the T8-positive, CD11b-positive subpopulations have been demonstrated to show suppressive activities, the S-100-positive T cells seem to be closely restricted to a small T-suppressor-cell compartment. Although functional studies on viable isolated S-100 beta-positive cells are impossible to achieve, due to the lack of this protein on the cell membrane, its presence in a discrete T-suppressor compartment might suggest a possible role for the S-100 beta-positive T cells in the regulation of the immune system.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1988

Immunoelectronmicroscopic demonstration of S-100 protein in hairy cell leukemia cells.

Hairy cell leukemia (HCL) is generally considered a neoplastic proliferation of B-cell lineage. Among immunocompetent cells, however, S-100 protein is also demonstrated in "dendritic" cells (i.e., interdigitating reticulum cells, Langerhans cells, follicular dendritic cells) and in a small percentage of T8-cells. This study reports the surprising detection of S-100 protein in peripheral blood HCL cells of B-cell type, using immunofluorescence and immunoelectronmicroscopy. However, negative results for S-100 protein staining were obtained in paraffin splenic sections of HCL cases. This report stresses the need for caution in the interpretation of immunocytochemical methods and results, and emphasizes the diagnostic ambiguity of the S-100 protein.

Blood Cells↗
PubMed1988

Simultaneous colloidal gold immunoelectronmicroscopy labeling of CD1a, HLA-DR, and CD4 surface antigens of human epidermal Langerhans cells.

The simultaneous demonstration of three surface antigens of Langerhans cells (LC) within LC-enriched fresh epidermal cell suspensions from normal human skin was achieved, by means of a triple immunogold (IG) staining, using commercially available monoclonal antibodies (moAb) and immunoreagents, in a simple pre-embedding immunoelectronmicroscopy (IEM) procedure. As a result, suspended LC were triple-stained as follows: gold particles of 40 nm revealed the CD1 a antigen; gold particles of 20 nm revealed the HLA-DR antigen; and gold particles of 5 nm revealed the CD4 antigen. All the observed epidermal Birbeck granule-bearing LC were triple IG stained, thus simultaneously expressing the three surface differentiation antigens, which are therefore different from but coexisting with each other. The present investigation assesses the constant simultaneous expression by Birbeck granules bearing LC of not only CD1a and HLA-DR antigens, but also CD4 antigen. The occurrence is therefore excluded of both CD1a-positive HLA-DR-negative LC subpopulation and CD4-negative LC subpopulation, presumably due to the different sensitivity of the various procedures performed. The hypothetical occurrence of CD4-positive, CD1a-, and/or HLA-DR-negative LC subpopulations is ruled out. This study reaffirms indeed the high specificity and sensitivity of the IG-IEM method for a precise detection of the cell surface antigens of LC, and states the suitability of the IG labeling even for accurate multiple IEM stainings of LC.

Adult↗
PubMed1988

Detection of cell surface antigens in tissue sections by means of pre-embedding immunogold staining.

The immunogold technique has been used in electron microscopy to detect cytoplasmic and extracellular antigens by postembedding techniques. It has also been used to detect plasma-membrane-associated molecules on suspended cells and, recently, to visualize cell surface antigens in ultrathin sections of Lowicryl embedded specimens. In the present study, cell surface antigens of rat kidney and human skin were identified in tissue sections by using pre-embedding immunogold labeling. Brush border microvillar antigens and dermal lymphocyte antigens both bound numerous gold particles. The immunogold staining described here has the advantage over immunoperoxidase procedures that is not subject to diffusion or reabsorption artifacts, and allows estimation of the antigen density on labeled cells. Furthermore, this pre-embedding immunogold technique is ideally suited to detecting cell surface-associated antigens since it preserves antigenicity, allows gold particle penetration and enhances cell membrane profiles.

Animals↗
PubMed1988

One half of the CD11b+ human peripheral blood T lymphocytes coexpresses the S-100 protein.

The expression of the CD11b antigen and the presence of the S-100 (and, specifically, its beta subunit) protein within the T4- subpopulation of normal human peripheral blood lymphocytes were investigated by panning techniques, immunofluorescence analysis and immunoelectronmicroscopy. Both antigens are known to be absent in the T4+ lymphocytes. However, CD11b+ T lymphocytes represented about 30% of the T4- population; a part of them (over 1/3) belonged to and completely filled up the T4- T8- subpopulation, whereas the remaining part (almost 2/3) shared the T8 positivity. Interestingly, S-100+ T lymphocytes, which always were CD11b+ too, represented about one half of the CD11b+ T cells, but were excluded from the T4- T8- CD11b+ subpopulation, whereas they represented up to 80% of the T4- T8+ CD11b+ subset. Such findings demonstrate that the S-100+ T lymphocytes are exclusively restricted to a discrete T cell compartment which shows the T8+ CD11b+ immunophenotype. Since such T8+ CD11b+ cells had been shown to possess suppressive capabilities, we herein propose that S-100+ lymphocytes might to some extent modulate the immune responses. However, the exact functional significance of the S-100 protein still remains unknown.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1987

S-100-positive T cells are largely restricted to a CD8-positive, 9.3-negative subset.

The present report concerns the demonstration of the exclusive detection among peripheral blood T-lymphocytes of the S-100 protein within the CD8-positive subpopulation which lacks the antigen recognized by the 9.3 monoclonal antibody. Highly purified human peripheral blood T-cell subsets, obtained by means of panning techniques, were first stained, by an immunofluorescence method, with purified anti-S-100 protein antibodies. The vast majority of S-100 protein- (and, specifically, its beta subunit) positive cells were detected in the CD8-positive, 9.3-negative subset. This subset had previously been shown to comprise all the alloantigen-specific and histamine-inducible suppressor T-cells. Other T-subsets, even those showing either CD8-positivity (but 9.3-positivity) or 9.3-negativity (but CD8-negativity), were, as a rule, S-100 negative. Immunoelectronmicroscopy confirmed that the S-100-positive cells, showing peroxidase activity within the cytoplasm, were found exclusively within the CD8-positive, 9.3-negative subset. This finding of S-100 protein in cells of a specific T8 suppressor subset extends the range of the known distribution of this protein and may have important implications concerning its role in the modulation of immune responses.

Antibodies, Monoclonal↗