IgG subclass distribution in intravenous immunoglobulin preparations.
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Enzyme-Linked Immunosorbent Assay↗
Biomedical subjects
H Leibl
Publications and source records attributed to H Leibl.
Comparable modulation of human monocyte functions by commercial factor VIII concentrates of varying purity.
Our previous observation on immune modulation induced by a given factor VIII (F VIII) concentrate preparation was extended by showing that the immune-modulating capacity is a more general feature of F VIII products and is independent of product purity. Interaction of human monocytes with therapeutic concentrations of various F VIII concentrates (0.2 to 2 IU F VIII/mL, six different F VIII concentrates from four manufacturers) led to a significant reduction in the expression of IgG Fc receptors in the membrane of these cells (F VIII concentrate-induced downmodulation of the receptor). This Fc receptor downmodulation was achieved by a short (1-hour) incubation of human monocytes with F VIII concentrates 16 hours prior to the Fc receptor assay and did not correlate with the respective product's IgG content. Although the IgG concentrations of the different products varied greatly (from 1.0 to 177.3 mg/1,000 IU F VIII), all products behaved comparably with respect to Fc receptor downmodulation (F VIII-treated monocytes: 34% +/- 7% to 44% +/- 4% rosette-forming cells; controls in the absence of F VIII: 83% +/- 5%). Furthermore, we also were able to demonstrate that heat treatment of F VIII, now used by virtually every manufacturer to eliminate contaminating viruses, had no effect on the respective products' Fc receptor-modulating capacity. The immune-modulating component was characterized as being a high-molecular-range compound containing IgG, IgM, F VIII, and blood group substances (most likely a combination of immune complexes and immunoglobulin aggregates). This compound is present in comparable amounts in both high-purity and intermediate-purity products and apparently copurifies with F VIII during the manufacturing process.
Antigen-Antibody Complex↗
Effect of UVB radiation on the biosynthesis of HLA-DR antigens.
HLA-DR molecules on the surface of immunocompetent cells are thought to represent target structures for the immunomodulating effects of UV radiation during the induction of an immune response. We therefore investigated the effect of UVB radiation on the de novo synthesis of HLA-DR-gamma-chains in the cytoplasm and the expression of alpha- and beta-chains on the surface of the human lymphoblastoid B-cell line Raji. Raji cells were UVB irradiated before biochemical experiments were performed. Cells were then metabolically labeled or radioiodinated and detergent lysates immunoprecipitated using antibodies directed against the gamma- or the alpha- and beta-chain of the HLA-DR molecule. Over a wide dose range, UVB-irradiated Raji cells were shown to still express HLA-DR determinants on their surface and, even more importantly, to be capable of synthesizing HLA-DR-alpha, beta- and gamma-chains in a normal fashion. Despite this, the functional capacity of Raji cells was impaired in a dose-dependent manner. UV radiation thus seems to exert its immunomodulating effects primarily at a different level than the incriminated immune-response-associated antigens, which are expressed as recognition structures on the surface of immunocompetent cells.
Antigen-Presenting Cells↗
IgG subclasses in chimpanzees.
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Animals↗
Human epidermal cells synthesize HLA-DR alloantigens in vitro upon stimulation with gamma-interferon.
Under certain pathologic conditions, human keratinocytes synthesize and express HLA-DR antigens. Assuming that soluble mediators might be responsible for this phenomenon, differentiating, primarily DR-keratinocytes were grown in the presence or absence of mixed leukocyte culture supernatants and tested for Ia antigen expression. After 6 days of culture, keratinocytes displayed surface-bound HLA-DR alpha/beta complexes when exposed to mixed leukocyte culture supernatants but not when cultured in media alone. These HLA-DR moieties on keratinocytes result from active biosynthesis as evidenced by the demonstration of the intracytoplasmic HLA-DR gamma (invariant) chain within these cells. In view of reports that interferon-gamma promotes Ia-production in a variety of cell types, we reasoned that this lymphokine might be responsible for the Ia-inducing property of mixed lymphocyte culture supernatants. Indeed, recombinant interferon-gamma, but not interferon-alpha or interleukin-2, proved to be a potent stimulator of Ia expression by keratinocytes. The further finding that this event can be prevented by the addition of a monoclonal anti-interferon-gamma antibody strongly suggests that this cytokine is directly responsible for HLA-DR production by keratinocytes. The interferon-gamma-induced acquisition of HLA-DR antigens by primarily DR-keratinocytes may provide a useful tool to study the role of these alloantigens in T-cell activation and may also add to our understanding of mechanisms operative in epidermal cell-T-cell interactions.
Antibodies, Monoclonal↗
Expression of the Ly-5 alloantigenic system on epidermal cells.
The expression of Ly-5 alloantigens is confined to hemopoietic cell types and is therefore considered a valuable indicator for the bone marrow derivation of a given cell. The further finding that different hemopoietic cell lineages express different molecular forms of the Ly-5 alloantigens prompted us to investigate (1) whether murine epidermal cells or subpopulations thereof express Ly-5 specificities and if so, (2) whether the expression of particular molecular configurations of Ly-5 antigens would allow us to gain a clue about the derivation of certain epidermal cell populations. When epidermal sheets from BALB/c, C57Bl/6, and C3H/He mice, were exposed to monoclonal anti Ly-5.1 antibody in an indirect immunofluorescence technique, a system of evenly distributed, dendritic cells was visualized. Allelic exclusion of the Ly-5 system was demonstrated by replacing anti-Ly-5.1 antibody by anti-Ly-5.2 reagent and by using epidermal sheets from SJL/J mice. Studies on epidermal cell (EC) suspensions revealed that about 1.6-5.2% of C3H/He EC were Ly-5-reactive and that approximately equal numbers of Ly-5-positive cells bore either Thy-1 or Ia antigens. Electron microscopic studies disclosed two morphologically different Ly-5-positive cell populations, i.e., cells of the Langerhans cell lineage and a recently defined cell system, whose most prominent feature is the expression of the Thy-1 antigen. We have termed these cells dendritic Thy-1+EC (dTHY-1+EC). In order to define the molecular configurations of the Ly-5 alloantigens, EC and spleen cells were internally labeled and--after immunoprecipitation of cell-membrane detergent extracts with anti-Ly-5.1--were analyzed on sodium dodecyl sulfate-polyacrylamide gels. Spleen cells yielded 3 bands with a molecular weight of 180,000, 195,000, and 215,000, respectively, as is characteristic for T lymphocytes, non-T/non-B cells, and B lymphocytes. In contrast, a single 195,000-200,000 dalton band was found in precipitates of both untreated and Langerhans cell-depleted (anti-Ia+C) EC. These data demonstrate the existence and active biosynthesis of the Ly-5 alloantigenic system on certain EC populations, i.e., Langerhans cells and dThy-1+EC, and therefore imply that both cell types originate from a bone marrow-derived precursor. The expression of the same molecular configuration of Ly-5 alloantigens on both LC and dThy-1+EC suggest that these two cell populations do not belong either to the T-cell or to the B-cell lineage and imply an ontogenetic relationship between dThy-1+EC and Ia-positive EC.
Animals↗
Inhibition of DNA synthesis of melanoma cells by azelaic acid.
Azelaic acid was successfully used in the clinical treatment of 7 cases of lentigo maligna in that remission of the lesions was observed in all our patients. In order to elucidate mechanism(s) of the beneficial clinical effects, we studied the effect of azelaic acid on cultured melanoma cells. Cell numbers recovered from melanoma cell cultures grown for several days in the presence of 10 mM azelaic acid were 50-70% less than those recovered from control cultures or from cultures containing 10 mM adipic acid. This reduction of cell numbers was not due to a simple cytotoxic or cytolytic effect of azelaic acid but rather due to a dose-dependent inhibition of DNA synthesis. Interestingly, nontoxic concentrations of azelaic acid, which significantly reduced DNA synthesis of cultured melanoma cells, had no overt effect on the protein synthesis of these cells. It is conceivable that inhibition of DNA synthesis is one of the mechanisms by which azelaic acid prevents growth and proliferation of abnormal melanocytes.
Administration, Topical↗
Expression of Thy-1 antigen by murine epidermal cells.
We report on the occurrence of a cell population within the murine epidermis which, by both morphologic and surface property criteria, is distinct from all other epidermal cell types known so far. These previously unrecognized cells are evenly distributed within the epidermis, display a primarily dendritic shape, exhibit a lobulated nucleus, contain large amounts of vimentin type intermediate-sized filaments, but lack desmosomes, melanosomes, Merkel cell granules, and Birbeck granules. As opposed to melanocytes, these cells fail to display tyrosinase activity. Surface marker analysis reveals these cells to uniformly express the Thy-1 antigen and to lack I-A and I-E/C antigen specificities. A major portion of these Thy-1-bearing cells are reactive with a monoclonal antibody to the Ly-5 determinant whereas attempts to demonstrate Lyt-1,2,3 antigens consistently yield negative results. These findings strongly suggest that Thy-1+ epidermal cells originate from the bone marrow; however, their precise relationship to distinct members of the hemopoietic differentiation pathway remains to be established.
Animals↗
Differential stimulation by 17 beta-estradiol and synthetic estrogens of progesterone-receptor and of translocation of estrogen-receptor in rat pituitary and uterus.
Central and peripheral actions of physiological and synthetic estrogens with elevated biological potencies were investigated. The synthetic estrogens were 1,3-diacetoxy-17 alpha-ethinyl-7 alpha-methyl-1,3,5(10)-estratrien-17 beta-ol (DM-EE2) and 1,3-dibenzoyloxy-17 alpha-ethinyl-7 alpha-methyl-1,3,5(10)-estratrien-17 beta-ol (DB-EE2). More prolonged nuclear retention of the estrogen-receptor was found after application of DM-EE2 and DB-EE2, than in the pituitary and uterus after injection of the same dose of 17 beta-estradiol (E2). Longer nuclear retention of the estrogen-receptor was observed in the pituitary after DB-EE2 application and in the uterus after DM-EE2 than in E2-treated rats. Pituitary progesterone-receptor was raised to 706% of controls by treatment with E2 and 1187% and 2308% after injection of DM-EE2 and DB-EE2, respectively. On the other hand, only 208% and 225% stimulation, respectively, of progesterone-receptor was observed in the uterus after treatment with the synthetic estrogens DM-EE2 and DB-EE2. In vitro cytosol receptor binding assays revealed low relative binding affinities of the synthetic estrogens compared to E2. The results may be explained by the hypothesis that differences in biological activity of estrogens may be caused by different metabolic and/or cellular events localized in distinct target organs or by altered affinity of the nuclear estrogen receptor complex to chromatin receptor sites due to a change in receptor conformation.
Animals↗
Inhibition by tamoxifen of estrogen-stimulated accumulation of preprolactin messenger ribonucleic acid.
Estrogens are involved in the stimulation of prolactin synthesis in the rat anterior pituitary. After 5 days of treatment with 17beta-estradiol, strong enhancement of [3H]leucine incorporation into prolactin an stimulation of translatable preprolactin mRNA, respectively, were noted. Increase in prolactin synthesis was found following daily application with estriol and the synthetic estrogen 1,3-dibenzoyloxy-17 beta-methyl-1,3,5(10)-estratrien-17 beta-ol (DB-EE2). The antiestrogen tamoxifen (trans-1-(p-beta-dimethylaminoethoxyphenyl)-1,2-diphenylbut-1-ene) was demonstrated to inhibit estradiol-stimulated prolactin syntheses. Antagonistic effects of tamoxifen were dose dependent. Low loses of the antiestrogen were already sufficient to suppress 17 beta-estradiol-enhanced levels of prolactin mRNA. On the other hand, estriol potentiated 17 beta-estradiol-stimulated levels of serum prolactin and prolactin synthesis. Our results add further information about the transcriptional control by estrogens and demonstrate inhibitory actions of antiestrogens on distinct regulatory levels of protein synthesis in the rat pituitary.
Animals↗
Effects of 17 beta-estradiol, synthetic estrogens, and antiestrogens on prolactin secretion, nuclear translocation of pituitary estrogen receptor and on progesterone receptor levels in rat pituitary, hypothalamus and uterus.
Estrogen-receptor and progesterone-receptor values were studied following repeated application of estrogens and/or the antiestrogen tamoxifen. Nuclear estrogen-receptor contents of the pituitary as well as serum prolactin levels and uterus weight were increased by treatment with 17 beta-estradiol and synthetic estrogens DM-EE2 (1,3 diacetoxy-17 alpha-ethinyl-7 alpha-methyl-1,3,5(10)-estratrien-17 beta-ol) and DB-EE2 (1,3 dibenzoyloxy-17 alpha-ethinyl-7 alpha-methyl-1,3,5(10)-estratrien-17 beta-ol). Tamoxifen also increased pituitary estrogen receptor values, but no stimulation of serum prolactin levels was found. Tamoxifen decreased estradiol stimulated uterus weight, but had no repressor effects on serum prolactin levels. Progesterone receptor levels were stimulated in the pituitary by repeated injection of 17 beta-estradiol, DM-EE2 and DB-EE2 and to a smaller degree by tamoxifen. The stimulation of progesterone receptor levels by estrogen was greater in the pituitary than in the hypothalamus and uterus. The present data combine to suggest differences in metabolic and/or cellular events following stimulation with estrogens of different biological activities.
Animals↗
Estrogen interaction with the anterior pituitary of female rats: Differential cytosol binding, nuclear translocation and stimulation of RNA synthesis by 17 beta-estradiol and tamoxifen.
The interaction of tamoxifen (trans-1-(rho-beta-dimethylaminoethoxyphenyl)-1,2-diphenylbut-1-ene) with the cytosol estrogen receptor of the anterior pituitary of female rats was studied. No differences were recorded between incubations of cytosol samples with 17 beta-[3H]estradiol performed in the presence of absence of unlabeled 17 beta-estradiol and tamoxifen, respectively, thus suggesting that these interactions were at common receptor sites and excluding possible cooperative interactions. Competition experiments and Scatchard plot analysis of saturation experiments add further evidence for common receptor sites. A dissociation constant for tamoxifen of Kd - 2 nM was recorded. Tamoxifen was found to be bound to a moiety sedimenting the 4-5 S region, on a 6-24% linear sucrose density gradient at low salt concentrations, whereas 17 beta-estradiol sedimented in the 8-9 S area. These data suggest possible conformational changes of the receptor in the presence of tamoxifen. Furthermore, nuclear estrogen receptor levels remained elevated for at least 80 h after the application of tamoxifen alone or in a combination with 17 beta-estradiol, and a concomitant inhibition of cytosol receptor replenishment was noted. Tamoxifen and 17 beta-estradiol, respectively, were found to stimulate progesterone receptor levels when applied through 5 days. Tamoxifen plus 17 beta-estradiol administration elevated progesterone receptor contents above those found for each of the two compounds alone. On the other hand, tamoxifen enchanced the 17 beta-estradiol-induced prolactin serum levels, but did not stimulate prolactin serum levels by itself. These data combine to suggest that tamoxifen interacts with common estrogen receptor sites at the rat anterior pituitary.
Animals↗
Nuclear translocation of estrogen-receptor complex and stimulation of RNA synthesis by estrogens of different biological potencies in the female rat pituitary.
Stimulation of RNA synthesis and of nuclear translocation of estrogen-receptor complexes was investigated in isolated nuclei of anterior pituitaries of castrated female rats after injection with estrogens of different biological potencies. The assay system for the estimation of total RNA synthesis was validated and data suggest that incorporation of [3H]UMP into acid-precipitable material is consistent with RNA synthesis. An increase in RNA synthesis was seen 30 min after application of either 17 beta-estradiol, estriol or 1,3-diacetyl-17 alpha-ethinyl-7 alpha-methyl-1,3,5,(10)estratriene-17,3-ol (DMEE). RNA synthesis was maximal 90 min after estrogen application. Thereafter, RNA synthesis decreased slowly and reached pretreatment levels 3, 8 and 30 h after application of estriol, 17 beta-estradiol and the diacetyl derivative of ethinyl-estradiol, respectively. All estrogens were found to stimulate rapidly nuclear translocation of estrogen-receptor complexes. Peak levels of nuclear receptor contents were reached 30 min after administration of estrogens. A concomitant depletion of cytosol receptor levels was noted. Nuclear retention of estrogen-receptor complexes paralelled duration of enhanced RNA synthesis and correlated with biological potencies of the steroids. Data of present experiments combine to suggest that long-term nuclear retention is a requisite for expression of biological activity of estrogens at the anterior pituitary. Furthermore, the degree of biological activity seems to be associated with duration of stimulation of RNA synthesis, amount of estrogen-receptor complexes translocated to the nucleus, and duration of nuclear retention.
Animals↗