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Biomedical subjects

S I Katz

Publications and source records attributed to S I Katz.

At least 19 recordsLinked to original sources
PubMed1995

Induction of IL-10 gene expression in human keratinocytes by UVB exposure in vivo and in vitro.

Numerous studies have demonstrated that ultraviolet B (UVB) irradiation has profound effects on the skin and systemic immune systems. Because many of the effects of UVB result in suppression of contact sensitivity responses and because IL-10 induces a Th2 rather than a Th1 response, we sought to determine whether UVB irradiation induces IL-10 transcription and subsequent protein secretion by human epidermal cells. Skin of nine volunteers was exposed to UVB or sham irradiation, and epidermal cell suspensions were prepared from suction blister roofs 24 h thereafter. mRNA was extracted using oligo dT-coated magnetic beads, and IL-10 cDNA was amplified with a sensitive RT-PCR technique. We found that IL-10 was constitutively expressed by epidermal cells in 5 of 9 volunteers and that IL-10 message was up-regulated by UVB exposure in all experiments. Since epidermis consists of a heterogeneous cell population with distinct cytokine profiles, we determined whether UVB caused enhanced IL-10 transcription and protein secretion in human keratinocyte cultures. In these experiments, IL-10 was constitutively expressed by keratinocytes and UVB up-regulated IL-10 gene expression in a dose-dependent manner 24 h after in vitro irradiation, coinciding with IL-10 protein secretion into the culture supernatants. Taken together, the findings indicate that UVB irradiation induces IL-10 in human keratinocytes and suggest that keratinocyte-derived IL-10 may be an important component of the immunosuppression that results from UVB irradiation.

Adult↗
PubMed1995

Functional studies of epidermal Langerhans cells and blood monocytes in HIV-infected persons.

APC dysfunction may be important in immune dysregulation associated with HIV disease. Langerhans cells, epidermal APC, can be infected with HIV, although their function in HIV-infected persons has not been studied. Therefore, we studied the immunologic function of Langerhans cells, in parallel with blood APC (enriched for monocytes/macrophages (M phi) function, in 21 HIV-seropositive (HIV+) and 21 HIV-seronegative volunteers, including three monozygotic twin pairs discordant for HIV serology. Langerhans cells from HIV+ patients were quantitatively normal and expressed normal levels of HLA-DR. However, Langerhans cells from AIDS patients and M phi from both AIDS and HIV+ non-AIDS patients stimulated allogeneic T cells less well compared with control APC. In addition, decreased recall Ag- and mitogen-induced T cell responsiveness was observed in HIV+ patients using either autologous Langerhans cells or autologous M phi as APC/accessory cells. Interestingly, Langerhans cells and M phi isolated from HIV+ twins (CD4+ cell counts of 181, 271, and 562/mm3) were able to present recall Ag normally to HIV-uninfected syngeneic T cells. In summary, APC from HIV+ patients were impaired in their ability to generate a primary immune response (i.e., alloantigen-induced T cell stimulation), but they retained the ability to generate a secondary immune response (i.e., recall Ag-induced syngeneic T cell stimulation). Thus, these findings suggest that defects in secondary immune responses commonly observed in HIV disease are dependent on T cell dysfunction alone, whereas defective primary immune responses may be secondary to both T cell and APC dysfunction.

Adult↗
PubMed1995

Characterization of epitopes recognized by hapten-specific CD4+ T cells.

Although protein-derived nominal Ags have, in many instances, been precisely determined, the epitopes recognized by hapten-specific CD4+ T cells responsible for contact sensitization have not been defined. To better understand the nature of the precise epitopes generated after hapten interaction with Langerhans cells (LC), we assessed the ability of TNP-modified I-Ak- and I-Au-binding peptides to activate hapten-specific CD4+ T cells obtained respectively from TNCB-primed C3H (H-2k) and PL/j (H-2u) mice. Using LC as APC, I-Ak-restricted TNP-specific CD4+ T cells proliferated in the presence of the synthetic peptide hen egg lysozyme 52-61 derivatized with TNP at position 56, and less so when TNP was coupled at positions 53 or 59. Similarly, I-Au-restricted TNP-specific CD4+ T cells from PL/j mice were triggered by the synthetic I-Au-binding 13 mer poly(A)-Y5-R6 TNP-modified at position 4, and to a limited extent with TNP coupled in positions 7 or 10. Our results indicate that hapten-modified MHC class II binding nonautologous peptides are recognized by hapten-specific CD4+ T cells and that precise positioning of hapten molecules on peptides binding MHC class II molecules is required for optimal CD4+ T cell recognition. These findings provide insight into the manner in which haptens are recognized by T cells involved in contact sensitivity and should facilitate the study and design of specific therapies for the manipulation of hapten-specific CD4+ T cell responses.

Amino Acid Sequence↗
PubMed1995

Contact sensitivity as a model for T-cell activation in skin.

Contact sensitivity has served as a useful model for the primary activation of T cells in skin and skin-associated lymphoid tissue. We have been interested in the early signals necessary for the induction of an allergen-specific T-cell response, as well as the factors controlling the intensity and extent of such an immune reaction. Because cytokines qualified as possible candidate molecules involved in directing primary immune responses in skin, we studied the early changes in the cytokine pattern of the epidermis. Apart from defining a cytokine pattern specifically induced only after application of allergen, we also identified Langerhans-cell-derived interleukin (IL)-1 beta as the first cytokine to be induced by allergen, within 15 min of hapten application. Further experiments demonstrated that IL-1 beta also subserved an essential function for the induction of contact sensitivity reactions, as injection of IL-1 beta into the ears of mice mimicked the application of contact allergen on the morphologic, phenotypic, and functional level. In addition, injection of anti-IL-1 beta monoclonal antibody before application of allergen completely prevented sensitization. We also identified the production of IL-10 by murine keratinocytes. IL-10 may serve as a counterregulatory molecule in contact hypersensitivity, as it functionally suppressed Langerhans cell accessory cell function by preventing the expression of certain costimulatory molecules on the surface of these antigen-presenting cells. Indeed, IL-10 converts those cells from potent inducers of primary immune responses to tolerizing antigen-presenting cells. The tolerizing function of IL-10 has also been described in vivo. Intradermal injection of IL-10 before application of the allergen induces hapten-specific tolerance in vivo. In aggregate, our data indicate that epidermally derived cytokines are intimately involved in the modulation of immune reactions in skin, in some cases enhancing sensitization and in others inducing specific immunologic tolerance and anergy.

Animals↗
PubMed1995

The skin as target, vector, and effector organ in human immunodeficiency virus disease.

Langerhans cells are CD4+ antigen-presenting cells in the dendritic cell family that can initiate primary and secondary immune responses after emigration from skin and mucosa. Because of these properties, Langerhans cells have been proposed as potential targets for human immunodeficiency virus (HIV) infection and as potential vectors for the transmission of primary HIV infection to T cells after mucosal exposure. In support of this theory, previous investigative studies have demonstrated that Langerhans cells are targets for HIV infection both in vivo and in vitro and that HIV-pulsed Langerhans cells, as well as blood dendritic cells, induce a productive infection in co-cultured T cells in vitro. In addition, Langerhans cell dysfunction has been proposed as contributing to the pathogenesis of some of the cutaneous manifestations observed in HIV+ individuals. In a recent study, we detected Langerhans cell dysfunction in patients with acquired immune deficiency syndrome, but not in earlier stages of HIV disease. Here we review previous and current investigative studies on HIV and the skin, with an emphasis on Langerhans cells, and discuss possible future investigations in this field.

Animals↗
PubMed1995

Inhibition of interferon-gamma-induced intercellular adhesion molecule-1 expression on human keratinocytes by phosphorothioate antisense oligodeoxynucleotides is the consequence of antisense-specific and antisense-non-specific effects.

Expression of intercellular adhesion molecule-1 (ICAM-1) by keratinocytes is an important event in the pathogenesis of T-cell-mediated inflammatory skin diseases. To determine if ICAM-1 expression could be selectively modulated, two antisense phosphorothioate oligonucleotides (S-ODN) targeting the translation initiation and 3' untranslated regions of ICAM-1 mRNA were added as lipid complexes to cultures of keratinocytes. Interferon-gamma was added after 24 h to induce ICAM-1 expression, which was quantitated by flow cytometry after 48 h. The S-ODN targeting the translation initiation site did not inhibit ICAM-1 expression at 0.2-20.0 microM. In contrast, 0.2-1.0 microM of the S-ODN targeting a site in the 3' untranslated region abrogated ICAM-1 expression in up to 75% of the keratinocytes; this inhibition was reversible when complementary sense S-ODN was added. Phosphodiester ODN (PD-ODN) targeting the same sites did not inhibit ICAM-1 expression on keratinocytes, most likely as a consequence of rapid degradation. Inhibition of ICAM-1 by the antisense S-ODN was selective; expression of beta 2-microglobulin, alpha 3-integrin, and beta 1-integrin remained largely unaffected and interferon-gamma-induced HLA-DR expression was inhibited to a much lesser extent than ICAM-1. Antisense-non-specific inhibition was also noted in that two scrambled S-ODN with an identical nucleotide (14 of 20 cytosines) composition inhibited ICAM-1 expression in up to 44% of the keratinocytes, whereas a degenerate S-ODN did not. The data demonstrate the complex effects exerted by antisense S-ODN in that ICAM-1 expression was inhibited via antisense-non-specific mechanisms probably due to the intrinsic properties of the S-ODN as well as via the anticipated sequence-specific mechanisms.

Base Sequence↗
PubMed1995

Human Langerhans cells express E-cadherin.

Murine Langerhans cells (LC) synthesize and express E-cadherin, a Ca(++)-dependent homophilic cell adhesion molecule that mediates LC-keratinocyte (KC) binding in vitro. In vivo, E-cadherin expression by LC may promote localization and persistence of LC within the epidermis through LC-KC adhesion. In addition, changes in LC E-cadherin expression or affinity may be an important factor in the egress of LC from the epidermis after exposure to antigen. The aim of the present study was to determine if human LC also express E-cadherin. Suction blister roofs were obtained from normal volunteers and epidermal cell (EC) suspensions were prepared by limited trypsinization in the presence of 1 mM Ca++. EC were then incubated with antibodies to E-cadherin and CD1a or HLA-DR, and examined by two-color analytical flow cytometry or immunofluorescence microscopy. Most (82.9% +/- 7.4% [mean +/- SD], range 67-89%, n = 7) freshly prepared human LC expressed E-cadherin, as did the majority of KC. The amount of E-cadherin (as determined by mean fluorescence intensity) expressed by LC and KC was similar. Trypsin/EDTA treatment of freshly prepared EC abrogated expression of E-cadherin by LC and KC, whereas E-cadherin was not degraded by trypsin in the presence of Ca++. LC expressed lower levels of E-cadherin after 3 d in culture. Thus, human LC, like murine LC, express the homophilic adhesion molecule E-cadherin, which may be important in establishing and maintaining interactions between LC and KC in mammalian epidermis.

Animals↗
PubMed1994

Quantitative differences in cell surface expression of class I MHC antigens on murine epidermal Langerhans cells.

Epidermal Langerhans cells are derived from cells of bone marrow origin and, as the primary APC population in the skin, are responsible for initiation of many immune responses. Consequently, cell surface expression of MHC Ags by Langerhans cells is central to their function. Although murine Langerhans cells express class II MHC Ags at high levels, their level of expression of class I MHC has been controversial. In this study, cell surface expression of multiple individual class I MHC Ags on murine epidermal Langerhans cells was analyzed using quantitative immunofluorescence. It was found that Langerhans cells differentially express products of distinct class I genes. Langerhans cells expressed low cell surface amounts of H-2K and Qa-2, whereas expression of surface H-2D and H-2L by the same cells was high. Murine epidermal Langerhans cells therefore express low cell surface amounts of some but not all class I MHC Ags. Differential surface expression of products of distinct class I MHC genes by Langerhans cells may have a profound effect on cutaneous immune responses.

Animals↗
PubMed1994

Heat-stable antigen is an important costimulatory molecule on epidermal Langerhans' cells.

Heat-stable antigen (HSA), expressed by activated B cells, has been described as a costimulatory molecule for CD4+ T cells. Because epidermal Langerhans cells (LC) are known to express HSA, we determined whether LC HSA also served as a costimulator of Th cells. We have confirmed that HSA is expressed by freshly prepared (fresh) and, to a lesser extent, short-term cultured (cultured) LC and we demonstrate that costimulatory effects of HSA are prominent on fresh and 1-day cultured LC, whereas 2- to 4-day cultured LC exhibit less HSA-costimulatory activity. The anti-HSA mAb 20C9 almost completely blocked the proliferative response of Th1 or lymph node T cells induced by fresh or cultured LC. 20C9 also specifically inhibited LC-dependent IL-2 production by Th1 cells. The inhibitory effect of 20C9 was not observed when Th2 cells were substituted for Th1 cells or peripheral lymph node T cells. Furthermore, Th1 cells rescued from cocultures of T cells and 20C9-treated, Ag-pulsed LC, (but not from control-treated cocultures) were anergic to restimulation with untreated Ag-pulsed LC. These data suggest that LC HSA is an important costimulatory molecule in Th1 cell-dependent cutaneous immune reactions.

Animals↗
PubMed1994

Murine epidermal Langerhans cells and splenic dendritic cells present tumor-associated antigens to primed T cells.

We examined the ability of epidermal Langerhans cells and splenic dendritic cells to present tumor-associated antigens (Ag) to immune T cells. Methylcholanthrene (MCA)-induced subcutaneous fibrosarcomas derived from C57BL/6 mice were used as tumor models. Our data demonstrate that both murine Langerhans cells and splenic dendritic cells have the capacity to present tumor-associated Ag to primed T cells. We found that variously treated tumor preparations (irradiated viable tumor cells, irradiated frozen-stored tumor cells, mitomycin C-treated viable tumor cells, and snap freeze-thawed tumor cell lysates) can be utilized for tumor Ag-pulsing. Primed CD4+ T cells demonstrated in vitro specificity towards their respective tumors and did not cross-react to other syngeneic MCA-induced or non-MCA-induced tumors. The T cell proliferative response critically depended on the presence of immune CD4+ T cells. We discuss the implications of these findings for the adoptive immunotherapy of cancer using immune CD4+ T cells.

Animals↗
PubMed1994

Extraction and quantitation of cytokine mRNA from human epidermal blister roofs.

The demonstration of cytokine mRNA expression in epidermal cells by the polymerase chain reaction technique preceded by reverse transcription (RT-PCR) requires linear test conditions (i.e. that the product obtained after amplification reflects the relative amounts of starting material) and high reproducibility, specificity, and sensitivity. By combining well-defined techniques for mRNA extraction and concentration measurement with a sensitive and well-calibrated RT-PCR technique, we demonstrated the presence of IL-1 alpha, IL-1 beta, IL-6, and TNF alpha in epidermal cells obtained from suction blister roofs from normal volunteers. Messenger RNA was extracted with superparamagnetic oligo(dT)25 Dynabeads, and the amount of mRNA was measured by spectrophotometry using a Beckman 5-microliters Ultra-Microcell prior to RT-PCR. Linear PCR conditions were obtained by carefully titrating the amounts of mRNA and the number of cycles. Reproducibility was estimated at different steps of the procedure, and the specificity of the enhanced cDNA products was verified by liquid hybridization with end-labelled probes. We suggest that this combination of techniques might prove useful for the simultaneous assessment of the expression of various cytokines from small samples of fresh human epidermal cells.

Blister↗
PubMed1993

Inhibition of Langerhans cell antigen-presenting function by IL-10. A role for IL-10 in induction of tolerance.

IL-10 is a product of activated keratinocytes and is released during the induction phase of contact sensitivity. As IL-10 effects have been described as being mediated by APC, we investigated effects of IL-10 on epidermal Langerhans cells (LC), the resident APC in the epidermis. Initial studies failed to demonstrate effects of IL-10 on MHC class II Ag expression by LC or anti-CD3 mAb- or alloantigen-induced LC-dependent T cell proliferation. However, production of IFN-gamma and IL-2, (but not IL-6) was markedly reduced in these assays. When the soluble-protein Ag specific T cell clones AE7 (Th1) and D10.G4 (Th2) were substituted for unprimed T cells, differential effects of IL-10 on T-cell proliferation were observed. Whereas IL-10-pretreated and untreated LC supported Th2 cell proliferation equally well, IL-10-pretreated LC were essentially unable to induce Th1 cell proliferation in response to native protein or peptide Ag. The inhibitory influence of IL-10 on Th1 cells was observed when fresh or 1 day cultured LC were used; 2- or 3-day cultured LC were affected to a much lesser extent by IL-10 pretreatment. Further, coculture experiments using IL-10-pretreated or untreated LC of a different haplotype suggest that IL-10 negatively regulates a costimulatory signal required for induction of Th1 cell proliferation. To assess whether T cells incubated with Ag and IL-10-pretreated LC were responsive to further stimulation, T cells were rescued after 1 day of coculture with IL-10-pretreated LC and restimulated, either immediately or after 1 to 5 days of rest, with untreated LC in the presence of Ag. T cells incubated with IL-10-pretreated LC were found to be anergic, whereas T cells incubated with untreated LC proliferated normally after further stimulation. However, anergic T cells responded vigorously to IL-2. These data indicate that although IL-10-pretreated LC are effective APC for Th2 cells, they fail to induce Th1 cell proliferation and rather induce clonal anergy in these cells.

Animals↗
PubMed1992

Identification and induction of keratinocyte-derived IL-10.

It has previously been demonstrated that the epidermis is a rich source of proinflammatory cytokines and growth factors and that complex interactions between these factors may affect inflammatory responses in skin. To investigate whether IL-10 (cytokine synthesis inhibitory factor) is part of this complex process, RNA was extracted from normal epidermis at various times after application of various chemicals to murine skin and mRNA signals for IL-10 were sought using a quantitative reverse-transcriptase-polymerase chain reaction technique. IL-10 signal strength was normalized to that of beta-actin in each sample. IL-10 mRNA signals were occasionally identified in normal epidermis but were uniformly enhanced 4 h after hapten application, and were maximal after 12 h. Contact allergens induced IL-10 mRNA signals whereas vehicles and irritants did not. Depletion of Langerhans cells, Thy-1+ dendritic epidermal cells, and T lymphocytes demonstrated that keratinocytes were the main source of IL-10 mRNA. IL-10 signals were also detected in mRNA derived from PAM 212 (spontaneously transformed keratinocyte) cells. IL-10 protein could be detected by immunoprecipitation, with an IL-10 mAB, of supernatants obtained 16 h after cultured epidermal cells were coupled with hapten. This study demonstrates that murine keratinocytes are capable of producing IL-10 mRNA and protein, and that signal strength of IL-10 mRNA is enhanced by hapten application.

Allergens↗
PubMed1992

Early molecular events in the induction phase of contact sensitivity.

To assess changes in epidermis-derived cytokine mRNA levels early in the afferent phase of allergic contact sensitivity, total epidermal mRNA was analyzed at various times after painting skin with haptens. We used a sensitive reverse transcriptase-polymerase chain reaction technique to quantitatively compare the regulation patterns of the following mRNAs: class II major histocompatibility complex I-A alpha, tumor necrosis factor alpha (TNF-alpha), interleukin (IL) 1 alpha, IL-1 beta, interferon (IFN) gamma, granulocyte/macrophage colony-stimulating factor, IFN-induced protein 10, and macrophage inflammatory protein 2. Enhanced Langerhans cell-derived IL-1 beta mRNA signals were detected as early as 15 min after skin painting with allergens. TNF-alpha, IFN-gamma, and granulocyte/macrophage colony-stimulating factor mRNAs were found to be upregulated after application of allergens, irritant, and tolerogens, but class II major histocompatibility complex I-A alpha, IL-1 alpha, IL-1 beta, IFN-induced protein 10, and macrophage inflammatory protein 2 mRNAs were upregulated only after allergen painting. Depletion of specific cell populations demonstrated that Langerhans cells were the primary source of the IL-1 beta and class II major histocompatibility complex I-A alpha mRNAs, keratinocytes were the primary source of TNF-alpha, IL-1 alpha, IFN-induced protein 10, and macrophage inflammatory protein 2, and infiltrating T lymphocytes were the source of IFN-gamma. Relevance of the molecular findings was demonstrated by the identification of biologically active IL-1 alpha and immunoreactive TNF-alpha in culture supernatants. These studies demonstrate that Langerhans cell-derived and certain keratinocyte-derived cytokine mRNAs are selectively upregulated by allergens in the very early afferent phase of contact sensitivity.

Allergens↗
PubMed1992

Cultured human Langerhans cells process and present intact protein antigens.

Epidermal Langerhans cells (LC) undergo profound phenotypic and functional alterations when cultured for 2 to 3 d. To determine whether the in vitro culture of human LC modulates their capacity to process and present intact protein antigens, we compared the ability of freshly isolated LC (fLC) and cultured LC (cLC) to stimulate in vitro T-cell proliferative responses to recall antigens. We found that human fLC and cLC were able to process and present recall antigens to primed T cells, inducing significant proliferative responses. For tetanus toxoid and Candida albicans extract, T-cell proliferative responses at 6 d to antigen-pulsed fLC were slightly greater than responses to antigen-pulsed cLC. For live influenza A virus, the T-cell responses induced by antigen-pulsed cLC were comparable or slightly greater compared with fLC. Allogeneic T-cell proliferation for both LC preparations were also comparable. The exogenous pathway of antigen processing was demonstrated by chloroquine inhibition.

Antigens↗
PubMed1992

Early events in the induction phase of contact sensitivity.

After allergen application to skin, there is enhanced class II major histocompatibility complex antigen expression, as well as enhanced T-cell-stimulatory function by epidermal Langerhans cells (LC). In this study, we investigated the early changes in the epidermal cytokine profile using a sensitive reverse transcriptase PCR technique to determine whether cytokines may be related to LC activation. We found that, on the mRNA and protein level, changes in the epidermal cytokine pattern caused by allergens in the induction phase of contact sensitivity are distinct from those caused by irritants or tolerogens. The earliest of the changes is the LC-derived interleukin (IL) 1 beta mRNA signal strength that is increased within 15 min of allergen painting.

Cytokines↗