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Biomedical subjects

V B Morhenn

Publications and source records attributed to V B Morhenn.

At least 55 records · Page 3Linked to original sources
PubMed1985

Induction of the synthesis of triton-soluble proteins in human keratinocytes by gamma interferon.

Recombinant human gamma interferon (r-IFN-gamma) induces the synthesis and expression of HLA-DR antigen on cultured, normal, human keratinocytes depleted of Langerhans cells. After removal of r-IFN-gamma from the culture medium of keratinocytes that are expressing HLA-DR antigen, the cells continue to express this antigen for at least 2 days. r-IFN-gamma induces, in a dose dependent fashion, the synthesis of several triton-soluble proteins with the most prominent having an apparent molecular weight of 53,000. Whereas normal keratinocytes do not express HLA-DR antigen in vivo, they do express HLA-DR in a variety of skin diseases such as lichen planus, graft-versus-host disease, and mycosis fungoides. We propose that an understanding of lymphocyte-keratinocyte interactions in vivo may be achieved by further studies of the mechanism of action of r-IFN-gamma on cultured keratinocytes and that the results may provide insight into the patho-physiology leading to a number of common inflammatory and neoplastic skin diseases.

Cells, Cultured↗
PubMed1985

A monoclonal antibody against basal cells of human epidermis. Potential use in the diagnosis of cervical neoplasia.

A murine monoclonal antibody was generated against human skin cells obtained from psoriatic plaques. The antibody, called VM-2, recognizes an epitope expressed on the basal cell layer of human skin and other epithelia. VM-2 also binds to cultured cells from a variety of human carcinomas including HeLa cervical carcinoma, A-431 vulvar carcinoma, A-549 lung alveolar carcinoma, and SCL-1 skin squamous cell carcinoma cells. In several primary human cell lines, including fibroblasts, endothelial cells, and cells from the hematopoietic lineage, the antigenic site recognized by VM-2 could not be detected. The cellular antigen when immunoprecipitated by VM-2 from both normal and transformed cells appears to be proteins of approximately 100,000 and 120,000 mol weight. In frozen sections from human tumor-containing tissues, VM-2 labels skin, cervical, and lung squamous carcinoma cells, as well as skin basal carcinoma cells. Malignant cells present in exfoliative smears from epidermoid invasive neoplasias of the cervix are also selectively recognized by VM-2 in distinction to normal squamous cervical cells. VM-2 is thus directed against an antigen associated with neoplastic cells when applied in selected sites of exfoliative cytology. This monoclonal antibody represents a new reagent that should prove useful in the diagnosis of cervical neoplasia.

Antibodies, Monoclonal↗
PubMed1984

Bullae in a patient with toxic shock syndrome.

A case of toxic shock syndrome presenting with macular erythema and subepidermal bullae is described. The patient meets the Centers for Disease Control criteria for toxic shock syndrome. A biopsy of the bullae and macular erythema revealed, respectively, subepidermal bullae and superficial perivascular infiltration. This case demonstrates the potential difficulty in differentiating between toxic shock syndrome and toxic epidermal necrolysis.

Adult↗
PubMed1984

T cell subpopulations in alopecia areata.

Alopecia areata remains a disease of unknown cause and unsatisfactory treatment. Histologically it is characterized by a lymphocytic infiltrate that surrounds the lower half of the hair follicle. Our findings show that 64% to 92% of this infiltrate is composed of T lymphocytes bearing the Leu 3a (helper-inducer) phenotype. The remaining lymphocytes demonstrate the Leu 2a (cytotoxic-suppressor) phenotype. This helper T cell predominance is present in all patients, irrespective of clinical activity or duration of disease. The Leu 3a/Leu 2a ratio ranged from 2.6 to 19.8, the higher ratios being seen in patients with clinically active disease. The cutaneous infiltrate also is Ia-positive, indicating the activated nature of these lymphocytes.

Adult↗
PubMed1984

Langerhans cells react with pan-leukocyte monoclonal antibody: ultrastructural documentation using a live cell suspension immunoperoxidase technique.

Langerhans cells are generally regarded as members of an Ia+ dendritic cell system capable of potent accessory cell function in immune responses. While it has been shown that murine Langerhans cells are bone marrow-derived, the ontogenic relationships among human Langerhans cells, other dendritic cells, macrophages, and leukocytes in general have yet to be fully clarified. Recently, several pan-leukocyte monoclonal antibodies have been produced which react with the human leukocyte common antigen. This antigen resembles the murine T200 antigen and is expressed by all leukocyte subtypes but not by nonhematopoietic cells. Using an immunoperoxidase technique for staining suspensions of live skin cells, we have documented Langerhans cell reactivity with pan-leukocyte monoclonal antibody L3B12 at the ultrastructural level. Reactivity with this highly sensitive and specific pan-leukocyte marker supports the concept of the human Langerhans cell as a specialized form of bone marrow-derived mononuclear leukocyte and defines an immunologic feature common to dendritic cells, macrophages, and leukocytes that is not shared by other cell types. This finding is discussed in the context of other recent data concerning the immunologic phenotype of Langerhans cells. Since the immunoultrastructural method employed does not require cell fixation of any kind prior to immunologic staining, it should prove particularly useful for studying cell surface antigens that are adversely affected by fixation.

Animals↗
PubMed1984

Langerhans cell production of interleukin-1.

Epidermal Langerhans cells (LC) are related to certain cells of the monocyte-macrophage lineage and to other dendritic cells. While epidermal LC and macrophages bear receptors for the Fc portion of IgG, other dendritic cells do not. However, unlike human dendritic cells from peripheral blood, LC bear the antigen against which the OKT6 antibody is directed. Within the skin this antibody binds only to LC or indeterminate cells. Functionally LC and dendritic cells can present antigen to sensitized T cells and are capable of stimulating allogenic T cells. Since lymphokines are thought to play an important role in T-cell activation and since LC are potent stimulators of antigen-specific T-cell proliferation, we investigated whether LC could produce interleukin-1 (IL-1). Our initial studies revealed that whole epidermal cell suspensions comprised of LC, keratinocytes, and melanocytes produce a factor that is similar to macrophage-derived IL-1. We termed this factor epidermal cell-derived thymocyte-activating factor ( ETAF ). Using a panning technique we obtained a highly enriched (up to 97%) population of LC which constitutively produced significant IL-1-like activity. Using an antibody against the monocyte-derived leukocytic pyrogen (LP) which has been shown to inhibit IL-1 activity, we were able to inhibit IL-1 activity from LC-enriched cultures. The results of this study indicate that within the epidermis there are at least two cell populations, keratinocytes and LC, that can constitutively secrete potentially important soluble immunostimulatory factors.

Fluorescent Antibody Technique↗
PubMed1984

Recombinant gamma interferon induces HLA-DR expression on cultured human keratinocytes.

In normal human epidermis, expression of HLA-DR antigen is restricted to Langerhans cells (LC) and acrosyringial epithelium. However, in diseases such as lichen planus and graft-vs.-host, HLA-DR antigen appears to be expressed by keratinocytes, although the exact source of the HLA-DR is unclear. Two possibilities are that (1) the HLA-DR is shed by neighboring immunocompetent cells, or (2) that the keratinocytes are synthesizing the antigen themselves. Recently, gamma interferon has been shown to induce HLA-DR biosynthesis and expression on human malignant melanoma cells lines and on normal vascular endothelium. We report here that pure recombinant human gamma interferon (100 units/ml) induces HLA-DR expression on 60-70% of cultured human adult keratinocytes depleted of LC within 2-4 days of culture as determined by fluorescence-activated cell sorter (FACS) analysis using monoclonal antibodies. No residual LC or lymphocytes could be detected in these cultures. This is the first demonstration of HLA-DR expression by cultured human keratinocytes. This expression may be of functional significance in antigen presentation and cell-mediated cytotoxicity involving the epidermis.

Antibodies, Monoclonal↗
PubMed1984

Antiproliferative effects of recombinant alpha- and gamma-interferons on cultured human keratinocytes.

To extend our initial observation that recombinant gamma-interferon induced expression of class II major histocompatibility (HLA-DR) antigen on normal cultured human keratinocytes, we studied the antiproliferative effects of recombinant alpha- and gamma-interferons. Both interferons reduced the number of attached cells (dose range 10-10(3) units/ml; 7.1 X 10(-11) to 7.1 X 10(-9) M) and gamma-interferon was 100 times more potent than alpha. This effect did not require the presence of Langerhans cells. gamma-Interferon reduced total cell production during the first week without increasing the percentage of cells shed. During the second week, gamma-interferon also increased the percentage of cell shedding. Although 10(2) units/ml of gamma-interferon was maximal for HLA-DR expression by cultured keratinocytes, there was increasing reduction of attached cells between 10(2) to 10(3) units/ml. The demonstration that recombinant gamma-interferon induces HLA-DR expression and also inhibits keratinocyte proliferation in vitro at nanomolar concentration expands the growing list of normal cells whose biologic function may be influenced by this lymphokine in vivo.

Cell Count↗
PubMed1983

Use of a murine monoclonal antibody which binds to malignant keratinocytes to detect tumor cells in microscopically controlled surgery.

A new murine monoclonal antibody, termed VM-1, stains basal cell carcinoma cells and at least some squamous cell carcinoma cells. This antibody was used to visualize the tumor margins during microscopically controlled surgery using immunofluorescence (IF) and immunoperoxidase (IP) technics. Our experiments demonstrate that VM-1 is helpful in detecting small islands of tumor cells in the tissue at the margins of skin cancers. Thus, tissue staining with this antibody should improve the accuracy of microscopically controlled excision, and hopefully will decrease the rate of tumor recurrence after surgery.

Animals↗
PubMed1983

Inhibition of a Langerhans cell-mediated immune response by treatment modalities useful in psoriasis.

Neither the pathogenesis of psoriasis nor the mechanism whereby seemingly diverse therapies alter the disease is understood. In this study, several antipsoriatic agents were tested for their effects on the skin cell lymphocyte reaction (SLR), an immunologic assay in which HLA-DR antigens on Langerhans cells (LC) stimulate proliferation of allogeneic lymphocytes. Every agent tested (cortisol, methotrexate, hyperthermia, anthralin) inhibited the SLR at therapeutic dose levels. By contrast, a variety of antibiotics, an anti-inflammatory agent, and lithium carbonate and propranolol, two drugs known to be ineffective in psoriasis, failed to inhibit the SLR. Finally, we have shown that hyperthermia and anthralin treatments are toxic for LC whereas they have little or no effect on keratinocyte viability. These results suggest that antipsoriatic agents may act in psoriasis by alteration or killing of LC.

Anthralin↗
PubMed1983

Selective enrichment of human epidermal cell subpopulations using monoclonal antibodies.

In studying the mechanisms that regulate the growth and differentiation of the human epidermis, it would be helpful to obtain relatively pure populations of the different epidermal cell types. We have used a solid-phase immunoabsorption method termed "panning" to positively select two types of epidermal cells: Langerhans cells and the keratinocytes found in the basal cell layer (basal cells). To attach basal cells to a goat anti-mouse IgG-coated plastic surface, we used murine monoclonal antibodies (VM-1 or VM-2), which were recently produced in our laboratory and bind specifically to antigens on human basal cells. Using antibodies VM-1 or VM-2, we panned for basal cells and obtained a yield of about 40 percent (an enrichment of about 2.5-fold). The cells enriched for basal cells demonstrated much better growth and DNA synthesis than did the cell fraction depleted of basal cells. For positive selection of Langerhans cells, we used OKT6, a murine monoclonal antibody that binds specifically to Langerhans cells in the epidermis. We determined that of those cells preincubated with OKT6 and adherent to an antibody-coated petri dish surface, about 70 percent demonstrated OKT6 binding by fluorescence microscopy. This represents a 15- to 20-fold enrichment for Langerhans cells. The nonadherent cell fraction contained less than 1 percent OKT6-positive cells. Ultrastructural studies showed that the cells thus separated were Langerhans cells. The OKT6-positive but not the OKT6-negative cells were capable of stimulating allogeneic lymphocytes in the skin-cell-lymphocyte reaction. Thus the panning technique is an effective method for obtaining greatly enriched subpopulations of viable epidermal cells.

Animals↗
PubMed1982

Cultured human epidermal cells do not synthesize HLA-DR.

All nucleated cells express HLA-A, B, and C antigens. However, only a few cells, including epidermal cells, demonstrate HLA-DR antigens which are potent transplantation immunogens in man. The current study was undertaken to determine if epidermal cell continue to synthesize and/or express HLA-DR antigens after prolonged in vitro culture. Epidermal cells cultured for 7 days or more no longer stimulated allogeneic lymphocytes in the epidermal cell-lymphocyte reaction. Indirect immunofluorescence light microscopy of cultured cells using mouse monoclonal antibody to HLA-DR antigen confirmed that these cells do not express HLA-DR antigens whereas they retain beta 2-microglobulin. Detergent extracts of 12-day cultured epidermal cells biosynthetically labeled with 35 S-methionine were immunoprecipitated with monoclonal anti-DR antibody and analyzed by the method of two-dimensional polyacrylamide gel electrophoresis. No radiolabeled proteins were found on these gels in the regions where HLA-DR molecules are known to migrate. These data indicate that HLA-DR antigen is absent from cultured epidermal cells. Finally, we describe a technique for growing epidermal cells on a gelatin membrane which allows subsequent removal of intact cell monolayers from the culture dish. Such monolayers may be useful for purposes of transplantation.

Cells, Cultured↗
PubMed1982

Use of the fluorescence-activated cell sorter to quantitate and enrich for subpopulations of human skin cells.

A variety of immunologic staining techniques were compared in a quantitative study of antigen expression by human epidermal cells. Virtually all nucleated epidermal cells express beta 2-microglobulin, which is associated with HLA-A, -B, and -C antigens, whereas only about 4% expressed T6, an antigen expressed by Langerhans cells but not other cells in the skin. With the fluorescence-activated cell sorter (FACS), epidermal cell suspensions were selectively enriched 10- to 15-fold for T6-positive Langerhans cells. An average of 6.5% of cells were specifically stained by anti-HLA-DR antibody. When dispersed cells stained with anti-DR plus peroxidase were examined with the technique of immunoelectron microscopy, only mononuclear leukocytes (probably Langerhans cells) were stained. After separating HLA-DR positive skin cells with the FACS, the DR-positive population but not the DR-negative population stimulated proliferation of allogeneic responder lymphocytes, indicating that sorted cells are metabolically active. We conclude that HLA-DR antigen is not expressed by keratinocytes in normal human skin cell suspensions and that the FACS can be used to selectively enrich or deplete skin cell suspensions of antigenically distinct subpopulations such as Langerhans cells.

Antibodies, Monoclonal↗
PubMed1982

Human skin cells synthesize HLA-DR molecules of the same charge and molecular weight as those synthesized by autologous lymphocytes.

Between 5 and 14% of human skin cells, separated by trypsinization, express HLA-DR antigen as detected by indirect immunofluorescence with monoclonal anti-DR antibody. To determine the source and structure of these DR molecules, skin cell suspensions were biosynthetically labeled with 35S-methionine and the radiolabeled DR molecules were analyzed by the method of two-dimensional polyacrylamide gel electrophoresis. The results indicated the presence of heavy, intermediate, and light chains indistinguishable from those of B lymphocyte DR. Gels in which DR immunoprecipitates from skin cells and autologous lymphocytes were run simultaneously confirmed this finding. Thus, skin cells and lymphocytes synthesize HLA-DR antigen of identical charge and molecular weight.

B-Lymphocytes↗
PubMed1982

Expression of HLA-DR antigen in skin from patients with psoriasis.

Using murine monoclonal antibodies against human HLA-DR antigen and against human T cells, we investigated the indirect immunofluorescence staining pattern of involved and uninvolved skin from patients with psoriasis. The staining pattern of involved psoriatic epidermis is different from the pattern seen in uninvolved skin from the same patient and consists of scattered, single HLA-DR positive cells alternating with groups of HLA-DR positive cells. These HLA-DR positive cell clusters can be seen at any level of the epidermis. In some patients, the dermis of involved skin shows prominent accumulations of T cells which are HLA-DR positive and thus represent activated T cells.

Fluorescent Antibody Technique↗