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Biomedical subjects

M Haftek

Publications and source records attributed to M Haftek.

At least 73 records · Page 4Linked to original sources
PubMed1986

Ultrastructural quantitation of desmosome and differentiation-related keratinocyte membrane antigen.

The keratinocyte membrane antigen KM 48 was defined by a new monoclonal antibody obtained after mouse immunization with normal human epidermal cell suspension. Specific reactivity of the antibody with desmosomal regions of keratinocyte cell membrane was demonstrated by immunoelectronmicroscopy. Langerhans cells, melanocytes, and indeterminate cells did not express the KM 48 antigen. Immunogold labelling permitted ultrastructural quantitation of KM 48 antibody binding on keratinocytes from various epidermal layers. A gradual increase in desmosome-related KM 48 antigen expression accompanied differentiation of keratinocytes during their transit from basal to granular layer. Distribution of the antigen on individual cells was uneven. The upper pole of a keratinocyte facing overlying more differentiated cells was always found to be laden with an immunogold marker about twice that of the opposite, lower surface of the cell. The results support the previous reports on gradual development of desmosomes during epidermal cell maturation and open up new possibilities for keratinocyte differentiation studies. They also underline the virtues of the immunogold-labelling method used for cell-surface antigen tracing and quantitation.

Adolescent↗
PubMed1986

Quantitative analysis of T6-positive Langerhans cells in human skin cancers.

Epidermal Langerhans cells (LC) are required for antigen-presentation and for stimulating antigen-specific T cell activation. Similar functions may be important in the immune response to malignant skin tumours. Monoclonal anti-T6 antibody was used to examine LC population in basal and squamous cell carcinomas. Positive control labeling was performed with monoclonal anti-HLA-DR antibody. The number of T6-positive LC per mm2 of section was significantly decreased (p less than 0.01) in the tumour group in comparison with a sex and age-matched control group. The number of sun-exposed and covered regions was taken into consideration in each respective group. Within the tumours, LC were found more frequently in the tumour periphery and in most differentiated tumour areas (horn pearls) than in the rest of the tumour mass. T6-positive LC were rarely found in the dermis. Moreover, LC exhibited morphological changes in specimens from tumours. Staining with anti-HLA-DR antibody revealed less numerous positive cells within tumour nests than labeling with OKT6. A relationship between T6-positive LC quantities and extent of HLA-DR-positive infiltrates around tumours could not be established. These results suggest that immunological surveillance of neoantigen-bearing tumour cells may be impaired in skin cancer. A reason for the reduced LC number may be an altered microenvironment in tumour tissue.

Aged↗
PubMed1986

Flow cytometry for separation of keratinocyte subpopulations from the viable epidermis.

Human epidermal cell suspensions were analyzed and sorted with flow cytometry. The desmosome and differentiation-related KM48 monoclonal antibody was used for indirect immunofluorescence and permitted staining of keratinocytes at various stages of the cell maturation. Intensity of the staining correlated with the degree of differentiation. Three sorting gates were chosen to obtain subpopulations which varied distinctly in KM48 expression. The flow cytometry-sorted cells were characterized by their ultrastructural appearance and by the bullous pemphigoid antigen expression. According to the ultrastructure criteria, about 50% of the cells obtained from the "IF negative" gate were basal layer keratinocytes (45.5% expressed bullous pemphigoid antigen); 90% of the "intermediate" gate cells were spinal layer keratinocytes, and over 80% of the cells sorted through the "strongly IF positive" gate were of the granular layer type. The method of keratinocyte separation proposed allows samples to be obtained for further biochemical and functional studies on keratinocyte subpopulations in normal and pathologic skin.

Antibodies, Monoclonal↗
PubMed1986

Immunogold labeling of keratin filaments in normal human epidermal cells with two anti-keratin monoclonal antibodies.

We report on application of the highly sensitive and specific immunogold labeling method for ultrastructural investigation of keratin intermediate filament antigens in human epidermal cell suspensions. Triton X-100 pretreated cells proved accessible to the colloidal gold conjugate, thus enabling keratin filament bundles to be labeled. Anti-keratin KL1 and KL2 monoclonal antibodies were raised in mice after immunization with either human stratum corneum-isolated keratins or keratins extracted from human epidermal cells suspensions, respectively. Immunoelectron microscopy confirmed immunofluorescence and immunoperoxidase results of epidermal keratinocyte staining, and revealed two different antibody reactivity patterns: KL2 reacted with keratin filaments in keratinocytes of all epidermal layers, whereas antigen to KL1 was detected only on keratin of the suprabasal layers, not on the basal keratinocyte tonofilaments. The monoclonal antibody-recognized epitopes were specific for the keratin filaments. Vimentin-rich cells (melanocytes) were not stained in the same epidermal cell suspensions. Additionally, two distinct ultrastructural patterns of keratin filament epitope labeling were observed. KL1 and KL2 monoclonal antibodies react with two different antigenic determinants, depending on the stage of keratinocyte differentiation, and may therefore be used for immunohistochemical studies of various keratin-containing cells in normal and pathologic conditions.

Antibodies, Monoclonal↗
PubMed1986

[Morphologic and functional study of a human insulin-secreting cell line].

Monolayer cell cultures were obtained from a human insulinoma (HIN) after collagenase digestion. HIN cells were initially plated on extracellular matrix (ECM) secreted by bovine corneal endothelial cells. Capsular integrity from cell clusters quickly interrupted and cell began to migrate as adhesive sheets onto ECM. After 2 months on ECM cell attachment and proliferation occurred on plastic allowing cloning of cells by limiting dilution. 9 clones were successfully cultured for 7 months with 20 subsequent passages. Immunoreactivity for insulin by indirect immunofluorescence typical secretory granules by electron microscopy and stable amounts of immunoreactive insulin in culture media suggest that HIN cells are beta cell related. One clone HIN D8 when challenged for half an hour with either 30 mM glucose, 1 mM isobutyl Methylxanthine 4 mM Tolbutamide, 10(-6) M glucagon responded respectively with a 1.5, 2, 3 and 1.5 fold increase in insulin output. Population doubling time of HIN D8 was 42 hrs. Establishment of such insulin secreting cell lines provides a valuable tool for diabetes research.

Adenoma, Islet Cell↗
PubMed1985

Specific cell-mediated immunity in patients with epidermodysplasia verruciformis and plane warts.

The characterization of different types of human papilloma viruses made it possible to study the specific immune responses to purified viral antigens in patients with warts. The specific cell-mediated immunity was investigated by means of a leukocyte migration inhibition factor test in 9 patients with epidermodysplasia verruciformis and 4 patients with regressing plane warts. The results show a significant increase in specific cell-mediated reactivity concomitant with the regression of the warts.

Adult↗
PubMed1985

A monoclonal antibody labelling the keratinocyte membrane: a marker of epidermal differentiation.

A murine hybridoma secreting an IgM monoclonal antibody (KL3) was produced by cell fusion of mouse myeloma cells with spleen cells from mice immunized with human epidermal keratins. On normal human epidermis KL3 stained the intercellular spaces from the stratum germinatum to the stratum granulosum with a fluorescence intensity increasing from the basal layer to the upper layers. Basal cells were not stained on the side facing the basement membrane. About 90% of free keratinocytes isolated after trypsinization were labelled by KL3 in a punctate staining. Immunoelectron microscopy allowed us to show that the antigen recognized by KL3 was exclusively localized on the keratinocyte membrane especially in the desmosomal plaques. KL3 reactivity was not modified by preincubation of skin sections with lectins showing a selective intercellular labelling of upper layers of epidermis or pemphigus antisera, nor by adsorption of the antibody on NP40 soluble proteins of the epidermis. Though KL3 reactivity was completely abolished after adsorption of purified keratins, no immunological reactivity of KL3 was detected with epidermal keratin polypeptides blotted on nitrocellulose paper. In psoriatic epidemis and epidermal tumors KL3 reactivity was drastically modified. These results suggest that KL3 recognized a keratinocyte membrane antigen implied in the epidermal differentiation process.

Animals↗
PubMed1983

Langerhans cells in skin from patients with psoriasis: quantitative and qualitative study of T6 and HLA-DR antigen-expressing cells and changes with aromatic retinoid administration.

Using a monoclonal antibody against human HLA-DR antigens and OKT6, we investigated by indirect immunofluorescence the distribution of Langerhans cells in normal human skin and involved and uninvolved skin from patients with psoriasis before, during, and after systemic aromatic retinoid administration. In parallel, enumeration of HLA-DR and of OKT6+ cells was also performed. In involved psoriatic epidermis the distribution of positive cells was disturbed; OKT6+ cells were reduced in number, as were HLA-DR+ cells which were seen in clusters. In control skin sections, a regular pattern of fluorescent dendritic epidermal cells was noted. In normal-appearing human skin, in nonlesional psoriatic skin, but not in diseased psoriatic skin, the number of OKT6+ cells per epidermal section surface unit was higher than that of HLA-DR expressing cells. Changes in the number and distribution of OKT6 and HLA-DR+ cells in psoriatic involved epidermis were corrected by oral retinoid treatment.

Adult↗
PubMed1981

Normal and psoriatic human skin grafts on "nude" mice: morphological and immunochemical studies.

124 normal and psoriatic human skin grafts were performed on congenitally athymic nude mice. 5, 10, 15, 20, and 30 days after grafting, histological and immunochemical studies (using bullous pemphigoid, pemphigus vulgaris and anti 67 K keratin polypeptide immune sera) were performed. The preservation of certain immunological identities of grafted skin, at least with regard to intercellular substance and basement membrane, was observed and the keratinization pattern of human epidermis also remained unaltered throughout the post-grafting period. The typical clubbing of rate ridges/acanthosis pattern in psoriatic skin was preserved in 40% of involved psoriatic skin grafts. The possible significance of these findings is discussed.

Animals↗
PubMed1980

Comparative studies on cell-mediated immunity in patients with different warts.

The distribution of peripheral blood T and B lymphocytes, the in vitro lymphocyte response to PHA, and in vivo experimental DNCB sensitization were studied in patients with different clinical forms of warts (common, 84; flat, 88; plantar, 22; genital, 14) and in 15 cases of epidermodysplasia verruciformis (EV). The percentage of T lymphocytes forming E rosettes was significantly decreased in patients with common (54.8%), flat (47.5%) and plantar (58.3%) warts, and those with EV (47.6%) in comparison with normal controls (68.4%). The DNCB sensitivity developed less frequently and it was less intensive in patients with common and flat warts than in the normal population. 60% of EV cases were anergic to challenging doses of DNCB. The lymphocyte response to PHA was reduced in all groups of patients studied as compared to normals. T cell function was found to be most defective in patients with EV and those with flat warts. Only a slight but statistically significant defect was demonstrated in the common wart group. CMI in patients with both plantar and genital warts was shown to be almost normal; except minor alterations of PHA-induced lymphocyte transformation and E rosetting T lymphocyte counts. These data have shown the divergency of CMI defect in the patients with different clinical forms of warts caused by various HPV types. This could indicate that distinct HPV types varied in their infectiveness and host cell-mediated resistance is a fundamental factor preventing viral infection.

Humans↗
PubMed1978

Defective function of T lymphocytes in psoriasis.

The distribution of thymus-derived (T) and bone marrow-derived (B) lymphocytes in 100 patients with psoriasis were studied by the rosetting techniques. Depression of the number of T lymphocytes forming spontaneous rosettes with sheep erythrocytes (E rosettes) occurred in 66% of patients, whereas no difference in B lymphocytes bearing C3 receptor (EAC rosettes) was observed between psoriatics and normals. The decrease in E rosettes was associated with the active phase of the disease. This disappeared 4-6 wk after onset of remission, which suggested that the abnormality in T-cell marker distribution is transitional. Lymphocytes forming neither E nor EAC rosettes, which were found to be significantly increased in active psoriasis, were identified as T lymphocytes since they reacquired normal E rosette function during short-term preincubation with concanavalin A (Con A). A serum factor was also demonstrated which inhibited E rosette formation by normal peripheral blood lymphocytes. Its activity increased linearly within 2 mo from the onset of skin lesions. The data suggest that in active psoriasis serum factors may be coated on the lymphocyte surface membrane which may be responsible for blocking of specific receptor for sheep erythrocytes and/or interfere with T lymphocyte function.

Animals↗
PubMed1978

Cell-mediated immunity (CMI) in psoriasis.

Patients with psoriasis were found to have less intensive experimental DNCB sensitization and decreased lymphocyte response to nonspecific mitogens, PHA, Con A, and PWM. E rosette formation was defective only in active psoriasis, in contrast to normal T and B cell counts. A reduction in DNCB hypersensitivity development and the percentage of E rosette forming lymphocytes were related to disease activity, but not to extention of skin lesions. The defect of E rosette function appeared to be transitional and completely disappeared in the remission. Abnormalities in CMI in psoriasis were found to be related at least partially to the existence in patients sera of a factor inhibiting normal T lymphocyte function. The study provides no evidence for the presence of primary CMI defect in psoriasis.

B-Lymphocytes↗
PubMed1978

Immunologic abnormalities in psoriasis: the inhibition of leucocyte migration by stratum corneum antigens.

The migration inhibition of peripheral blood leucocytes by stratum corneum (SC) antigens extracted from psoriatic scales and normal skin was studied in 25 patients with psoriasis and 12 normal controls. In 44% of patients, the radius of the area of spontaneous migration in the absence of antigen was significantly reduced. 9 out of 25 psoriatic cases showed inhibition of leucocyte migration after the addition of both SC antigens. Migration inhibition with SC antigens was observed only in patients in whom the radius of spontaneous migration area appeared to be normal. The data did not confirm the evidence of delayed hypersensitivity to epidermal SC antigens in patients with psoriasis. The decrease of spontaneous leucocyte migration in psoriasis might be dependent on the influence of antibodies against stratum corneum and/or immune complexes coated on lymphocyte or leucocyte surface membrane.

Adolescent↗