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M C Udey

Publications and source records attributed to M C Udey.

49 records · Page 3Linked to original sources
PubMed1992

Differential sensitivity of freshly isolated and cultured murine Langerhans cells to ultraviolet B radiation and chemical fixation.

Previous studies have demonstrated that low doses of ultraviolet B (UVB) radiation (100 J/m2) abrogate the accessory function of freshly isolated murine epidermal Langerhans cells (fLC) and cause a parallel decrease in the ability of LC to express increased amounts of ICAM-1 (CD54) in vitro. We have subsequently observed that the accessory cell function of cultured LC (cLC), as assessed by their ability to support anti-CD3 monoclonal antibody (mAb)-induced T cell mitogenesis, was not inhibited by levels of UVB exposure (100 J/m2) that completely inhibited the function of fLC, although exposure of cLC to UVB radiation (100 J/m2) resulted in a decrease in the level of ICAM-1 expression on most cLC and a concomitant decrease in cLC survival during a subsequent 24-h incubation. Time course studies revealed that T cells stimulated with anti-CD3 mAb in the presence of cLC became committed to proliferate 4-8 h after culture initiation, while 24-30 h of co-culture was required for irreversible T cell activation when fLC were utilized as accessory cells. In addition, paraformaldehyde (PFA)-fixed (non-viable) cLC supported anti-CD3 mAb-induced T cell proliferation, whereas PFA-fixed fLC were ineffective. We propose that cLC are functionally resistant to low doses of UVB radiation and chemical fixation because cLC express sufficient levels of the adhesion or co-stimulatory molecules [including ICAM-1 and Mac-1 (CD11b/CD18)] required to induce T cell activation. Conversely, fLC are sensitive to the effects of UVB radiation and chemical fixation because these physicochemical agents prevent acquisition of critically important surface molecules in culture.

Animals↗
PubMed1992

Effects of ultraviolet radiation on murine epidermal Langerhans cells: doses of ultraviolet radiation that modulate ICAM-1 (CD54) expression and inhibit Langerhans cell function cause delayed cytotoxicity in vitro.

Low doses (100 J/m2) of ultraviolet B (UVB) radiation from sunlamp fluorescent FS20 tubes inhibit the ability of freshly isolated murine epidermal Langerhans cells (LC) to support anti-CD3 MoAb-induced T-cell mitogenesis and selectively inhibit the upregulation of ICAM-1 expression by LC without causing appreciable cytotoxicity in short-term (less than or equal to 24 h) incubations (J Immunol 146:3347-3355, 1991). In the present study, epidermal cells (EC) were exposed to UVB radiation or were sham-irradiated and cultured for 24, 48, or 72 h when LC were recovered, enumerated, and assayed for simultaneous expression of I-A antigens and ICAM-1 by flow cytometry. UVB-irradiated LC that had been cultured for 24 h exhibited levels of I-A antigens comparable to those on unirradiated LC but expressed substantially less ICAM-1. After 48 and 72 h, cultured UVB-irradiated LC expressed somewhat lower levels of I-A antigens and markedly less ICAM-1 than unirradiated controls. Although similar numbers of LC were recovered from cultures initiated with UVB-irradiated and unirradiated epidermal cells after 24 h, far fewer identifiable LC were recovered from cultures seeded with irradiated cells at 48 and 72 h (approximately 50 and approximately 10% of control, respectively). The effect of UVB radiation on the survival of LC in vitro was not reversible with exogenous TNF alpha (125 U/ml) alone or granulocyte/macrophage colony-stimulating factor (5 ng/ml) and IL-1 (50 U/ml) in combination, although these cytokines had modest effects on the expression of I-A antigens and ICAM-1 by cultured UVB-irradiated LC. Results of survival studies performed with enriched LC preparations demonstrated that UVB radiation was clearly cytotoxic for LC and did not merely downregulate surface expression of I-A antigens or alter LC buoyant density. Exposure of LC to radiation from blacklight fluorescent (UVA) tubes (0.25 J/cm2) in the presence of 8-methoxypsoralen (1 micrograms/ml; PUVA) or monochromatic UVC radiation (20 J/m2) also inhibited LC accessory cell function. Results of survival studies performed with EC that had been exposed to PUVA or UVC radiation before culture were similar to those of studies performed with UVB-irradiated cells, although PUVA- and UVC-induced LC cytotoxicity was much more pronounced 48 h after culture initiation than UVB-induced cytotoxicity. UVA radiation alone augmented LC recovery at 24 and 48 h, but did not influence I-A antigen or ICAM-1 expression.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1992

Doses of ultraviolet radiation that modulate accessory cell activity and ICAM-1 expression are ultimately cytotoxic for murine epidermal Langerhans cells.

Mechanisms that underlie immunomodulatory properties of ultraviolet (UV) radiation remain incompletely characterized. Recently, we have studied effects of UV on the functional activity of epidermal Langerhans cells (LC) and have attempted to relate inhibitory effects of UV on LC function to modulatory effects of UV on adhesion molecule expression by LC. Exposure of LC in vitro to amounts of UVB, UVC, or psoralen+UVA (PUVA) radiation that inhibited LC function also prevented increased expression of intercellular adhesion molecule-1 by LC in vitro. Subsequent studies revealed that amounts of UV radiation that inhibited LC function and modulated ICAM-1 expression also decreased LC survival in vitro, although UV-induced LC cytotoxicity did not become apparent until 48-72 h after UV exposure. Our results are consistent with those of previous studies that suggested that low doses of UV radiation were cytotoxic for LC in situ. The potential cytotoxicity of UV radiation for LC should be considered when studies of effects of UV radiation on immune responses in skin are interpreted.

Animals↗
PubMed1991

Inhibition of epidermal Langerhans cell function by low dose ultraviolet B radiation. Ultraviolet B radiation selectively modulates ICAM-1 (CD54) expression by murine Langerhans cells.

Immunosuppressive effects of low levels of ultraviolet B (UVB) radiation on cutaneous immune responses have been attributed to deleterious effects of UVB radiation on epidermal Langerhans cells (LC). To determine how UVB radiation modulates LC function we examined the effect of in vitro UVB exposure on LC accessory cell activity and surface phenotype. Exposure of BALB/c murine epidermal cells to low dose (less than or equal to 200 J/m2) UVB radiation in vitro inhibited their ability to support the mitogenic response of unstimulated, accessory cell-depleted splenic T cells to anti-CD3 mAb. LC accessory cell activity was also inhibited when LC were exposed to UVB radiation in situ, although several-fold higher doses of UVB radiation were required to achieve complete inhibition of LC function. This dose-dependent inhibition was mediated through a direct effect on LC that could not be reversed by IL-1 or IL-6 alone or in combination, or granulocyte-macrophage-CSF. TNF-alpha did not inhibit LC accessory cell function in this assay and anti-TNF-alpha neutralizing antibodies did not reverse the inhibitory effects of UVB radiation. UVB irradiated LC failed to participate in the anti-CD3-dependent clustering that normally occurs between T cells and LC during the proliferative response of murine T cells to anti-CD3 mAb, suggesting that UV radiation may interfere with accessory cell function by preventing intercellular adhesion. Two-color flow cytometric studies revealed low levels of the ICAM-1 on freshly isolated LC and some keratinocytes. ICAM-1 expression on LC increased 15 to 20-fold within the first 24 h in vitro and continued to increase during a 72-h culture period. The integrin LFA-1 was not identified on freshly isolated or cultured LC but was detected on responding T cells. Prior exposure of LC to UVB radiation (50 or 100 J/m2) inhibited the increase in ICAM-1 expression that normally occurs in vitro by up to 70% whereas surface levels of class II MHC Ag, CD45 and Fc-gamma receptors were not affected. Blocking studies revealed that anti-CD3 induced T cell proliferation and T cell-LC cluster formation was inhibited by both anti-LFA-1 and anti-ICAM-1 mAb suggesting that ICAM-1 expressed on LC must bind to LFA-1 on T cells to facilitate proliferative responses of T cells to anti-CD3 mAb. We conclude that the in vitro inhibitory effects of low dose UVB radiation on LC accessory function may result because UVB radiation prevents upregulation of ICAM-1 expression by LC in culture.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Antigen-presenting cells in essential fatty acid-deficient murine epidermis: keratinocytes bearing class II (Ia) antigens may potentiate the accessory cell function of Langerhans cells.

Essential fatty acid deficiency (EFAD) is a useful model for studying the role of (n-6) fatty acid metabolism in normal physiology. Because cutaneous manifestations are among the earliest signs of EFAD and because abnormalities in the distribution and function of tissue macrophages have been documented in EFAD rodents, we studied the distribution and function of Class II MHC (Ia) antigen-bearing cells in EFAD C57B1/6 mouse epidermis. Immunofluorescence studies revealed 1.9-9.6 (mean +/- SEM = 5.2 +/- 2.6) times more class II MHC (Ia) antigen-bearing epidermal cells in suspensions prepared from EFAD as compared to normal skin. Analysis of epidermal sheets demonstrated similar numbers of dendritic Ia+ and NLDC145+ cells in EFAD and normal epidermis, however. This discrepancy occurred because some keratinocytes in EFAD epidermal sheets expressed class II MHC (Ia) antigens, whereas keratinocytes in normal mouse epidermis did not. Two-color flow cytometry confirmed that all Ia+ cells in normal epidermis are Langerhans (Ia+ NLDC145+) cells, whereas Ia+ cells in EFAD epidermis are comprised of Langerhans cells and a subpopulation of keratinocytes (Ia+ NLDC145-). Similar levels of Ia antigens were expressed on EFAD and normal Langerhans cells. EFAD and normal epidermal cells were also compared in several in vitro assays of accessory cell function. Epidermal cells prepared from EFAD C57B1/6 mice present the protein antigen DNP-Ova to primed helper T cells more effectively than epidermal cells prepared from normal animals. EFAD epidermal cells are also more potent stimulators of T cells in primary and secondary allogeneic mixed lymphocyte-epidermal cell reactions than normal epidermal cells. The functional differences between EFAD and normal epidermal cells do not appear to result from increased cytokine release or decreased prostaglandin production by EFAD epidermal cells. In view of these findings and the observation that the antigen-presenting cell activity of EFAD epidermal cells correlates with the number of Ia+ keratinocytes in epidermal cell preparations, Ia+ keratinocytes (in the presence of Langerhans cells) may potentiate cutaneous immune responses in vitro and perhaps in vivo as well. These results also suggest that (n-6) fatty acids or metabolites of (n-6) fatty acids are involved in regulating the expression of class II MHC (Ia) antigens by keratinocytes in vivo.

Animals↗
PubMed1982

Effect of inhibitors of arachidonic acid metabolism on alpha-aminoisobutyric acid transport in human lymphocytes.

The role of arachidonic acid metabolism (or metabolites) in the modulation of alpha-aminoisobutyric acid transport in resting and concanavalin A-stimulated human peripheral blood lymphocytes was evaluated using previously characterized inhibitors of arachidonic acid metabolism. Nordihydroguairetic acid (a nonselective antioxidant), 5,8,11,14-eicosatetraynoic acid (an inhibitor of lipoxygenase and cyclooxygenase activities), indomethacin and acetylsalicylic acid (selective cyclooxygenase inhibitors), and 1-benzylimidazole, Ro-22-3581 and Ro-22-3582 (thromboxane synthetase inhibitors) proved to be potent inhibitors of amino acid transport activity in normal resting and lectin-activated lymphocytes at concentrations known to decrease thromboxane A2 production. The rank order of effectiveness of these various inhibitors compared favorably with their relative potencies as inhibitors of thromboxane B2 synthesis under the same conditions, as determined by radioimmunoassay. Inhibitory effects noted were not due to overt cytotoxicity and seemed to involve changes primarily in the Vmax and not the Km of the transport process. Drug-induced alterations in the magnitude of concanavalin A binding were not observed. These results suggest that the activity of amino acid transport systems can be influenced by certain arachidonic acid metabolites, probably thromboxanes, in both stimulated and unstimulated lymphocytes. In addition, these findings may provide a partial explanation for the observation that inhibitors of thromboxane formation prevent lymphocyte mitogenesis.

5,8,11,14-Eicosatetraynoic Acid↗
PubMed1982

Properties of a surface antigen expressed on activated human thymus-derived lymphocytes.

We have identified previously a quantitatively minor membrane protein (p28) with an apparent reduced m.w. of 28,000, which is biosynthetically labeled in activated human lymphocytes. Rabbit antisera with activity directed against p28 (alpha-ATC) were prepared and p28 was identified by immunoprecipitation in NP-40 extracts of activated, extrinsically labeled lymphocytes. p28 was not expressed in appreciable amounts by unstimulated T cells, stimulated or unstimulated B cells, null cells, or adherent cells. Protein p28 was only minimally represented on resting thymocytes but was easily detected on 4-hr activated thymocytes and the T lymphoblastoid cell lines HSB2 and MOLT-4. Absorption and immunoprecipitation studies with alpha-ATC indicated that p28 was not present on erythrocytes, platelets, neutrophils, six B cell lines, six null cell lines, and seven other T lymphoblastoid cell lines. Protein p28 from HSB2 cells was absorbed by lentil lectin, concanavalin A, and wheat germ agglutinin affinity columns and was eluted with the appropriate sugars. Gel filtration column chromatography of unreduced p28 in the presence of 0.5% NP-40 or 0.1% deoxycholate gave elution characteristics consistent with a m.w. of approximately 60,000 to 100,000. In preparative isoelectric focusing (IEF) studies the isoelectric point (pI (p28) = 5.2 to 6.1) was similar or identical to that described for the reduced and denatured protein in two-dimensional polyacrylamide gels (pI = 5.5 to 6.2). Protein p28 was eluted from DEAE-cellulose (Whatman DE-52) ion exchange columns at 0.05 to 0.15 M NaCl. Experiments with monoclonal antibodies or heteroantisera specific for other T cell and B cell antigens and various lymphoblastoid cell lines and normal peripheral blood cells indicated that p28 is distinct from the human Ia-like antigens, from T3, T4, T5, T8, and from several other reported human T cell antigens that appear to correspond to Thy-1, the sheep erythrocyte receptor, and a human thymus-leukemia antigen.

Absorption↗
PubMed1981

Membrane protein synthesis in mitogen-stimulated human T lymphocytes.

Proteins were biosynthetically labeled with [35S]-methionine in resting and mitogen-stimulated human peripheral blood lymphocytes. Lymphocytes were disrupted by sonication, the membrane fraction was isolated by differential centrifugation, and radiolabeled membrane proteins were identified after 2-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and autoradiography. The T cell mitogen concanavalin A (Con A) produced selective increases in the incorporation of [35S]-methionine into a membrane protein (or family of 4 structurally related membrane proteins) with an apparent m.w. of 28,000 and approximate isoelectric points (pI) of 5.5, 5.7, 6.0, and 6.2 (p28). Quantitative microdensitometry was used to evaluate p28 synthesis in a large number of different lymphocyte donors. Consistent and highly significant differences between resting and stimulated cells were demonstrable whether the 4 p28 spots were considered individually (p less than 0.0025) or in sum (p less than 0.0005). The p28 response to Con A was shown to be contained within the T lymphocyte fraction of mononuclear cells, was dose dependent over the mitogenic concentration range for Con A, and was blocked by alpha-methyl-D-mannoside, a sugar inhibitor of Con A binding. Differences between stimulated and resting cells were readily apparent within 2 hr of activation and became progressively larger with time. Protein p28 was prominently represented in plasma membranes isolated on discontinuous sucrose density gradients and was identified as a cell surface glycoprotein based on studies in which intact stimulated cells were exposed to degradative enzymes. This protein was lost from cells treated with pronase, whereas neuraminidase digestion changed the pI of p28 from a range of 5.5 to 6.2 to a range of 6.1 to 6.4 without changing the apparent m.w.

Cell Separation↗
PubMed1981

Inhibition of lectin-induced lymphocyte activation by 2-cyclohexene-1-one: decreased intracellular glutathione inhibits an early event in the activation sequence.

The importance of normal intracellular glutathione (GSH) levels in the activation of human peripheral blood lymphocytes (PBL) by mitogenic lectins was explored using 2-cyclohexene-1-one (2-CHX-1), an agent that selectively decreases the levels of intracellular GSH. PBL incubated with mitogenic lectins and graded doses of 2-CHX-1 showed a dose-dependent inhibition of activation assayed by [3H]-thymidine uptake and percentage of blast transformation. 2.5 x 10(-5) M 2-CHX-1, a concentration that decreases GSH levels to less than 20% of control, caused an 80 to 90% suppression of both [3H]-thymidine uptake and blast transformation; lower concentrations were less effective. Time course studies showed that 2-CHX-1 was maximally effective only if added during the first 4 hr of culture. At the concentrations used, 2-CHX-1 was not cytotoxic to lectin-stimulated or unstimulated PBL and did not interfere with mitogen-lymphocyte interaction. These results suggest that adequate levels of glutathione are necessary for lymphocyte activation and that the glutathione requirement is exerted in the early activation sequence.

Cyclohexanes↗
PubMedDate not supplied

Bacterial DNA and CpG-containing oligodeoxynucleotides activate cutaneous dendritic cells and induce IL-12 production: implications for the augmentation of Th1 responses.

BACKGROUND: Unmethylated CpG sequences in bacterial DNA act as adjuvants selectively inducing Th1 predominant immune responses during genetic vaccination or when used in conjunction with protein Ag. The precise mechanism of this adjuvant effect is unknown. Because dendritic cells (DC) are thought to be crucially involved in T cell priming and Th1/Th2 education during vaccination via skin, we characterized the effects of bacterial DNA and CpG-containing oligodeoxynucleotides (CpG ODN) on cutaneous DC. METHODS AND RESULTS: Stimulation with CpG ODN 1826 (6 micrograms/ml) induced activation of immature Langerhans cell (LC)-like DC as determined by an increased expression of MHC class II and costimulatory molecules, loss of E-cadherin-mediated adhesion and increased ability to stimulate allogeneic T cells. Composition-matched control ODN 1911 lacking CpG sequences at equal concentrations was without effect. In comparison to LPS and ODN 1911, CpG ODN 1826 selectively stimulated DC to release large amounts of IL-12 (p40) and little IL-6 or TNF-alpha within 18 h and detectable levels of IL-12 p70 within 72 h. Stimulation with Escherichia coli DNA, but not calf thymus DNA, similarly induced DC maturation and IL-12 p40 production. Injection of CpG ODN into murine dermis induced enhanced expression of MHC class II and CD86 by LC in the overlying epidermis and intracytoplasmic IL-12 p40 accumulation in a subpopulation of activated LC. CONCLUSION: Bacterial DNA and CpG ODN stimulate DC in vitro and in vivo and may preferentially elicit Th1-predominant immune responses because they can activate and mobilize DC, inducing them to produce IL-12.

Animals↗
PubMedDate not supplied

Studies on the stimulation of cAMP metabolism by heparin solutions containing benzyl alcohol.

Heparin solutions containing benzyl alcohol as a preservative increase intracellular cAMP concentrations in human lymphocytes by as much as 5 fold. Similar changes but of a lesser magnitude were also observed in human polymorphonuclear leukocytes and platelets and rabbit alveolar macrophages. Further analysis demonstrated that benzyl alcohol used as a preservative was responsible for this finding and that heparin per se had no effect on cAMP metabolism. The cAMP response to benzyl alcohol was a time- and dose-dependent process. In human peripheral blood lymphocytes, cAMP levels increased in a linear fashion over a benzyl alcohol concentration range from 0.015-0.30% (v/v). The increased cAMP concentrations in lymphocytes as measured by radioimmunoassay were not due to formation of a cross-reacting adenosine-alcohol adduct since the immunoreactive material produced was destroyed by phosphodiesterase and co-migrated with 3H-cAMP in thin layer chromatographic studies. Concentrations of benzyl alcohol which affect cAMP metabolism are commonly present in biologic studies employing heparin solutions. In view of these findings certain previously described effects of heparin may need to be reinterpreted and in the future heparin solutions containing benzyl alcohol should not be used when studying biologic processes known to be influenced by alterations in cAMP metabolism.

Animals↗