Biomedical subjects

M A Talle

Publications and source records attributed to M A Talle.

At least 19 recordsLinked to original sources

PubMed1998

Detergent-salt resistance of LAP2alpha in interphase nuclei and phosphorylation-dependent association with chromosomes early in nuclear assembly implies functions in nuclear structure dynamics.

Lamina-associated polypeptide (LAP) 2 of the inner nuclear membrane (now LAP2beta) and LAP2alpha are related proteins produced by alternative splicing, and contain a common 187 amino acid N-terminal domain. We show here that, unlike LAP2beta, LAP2alpha behaved like a nuclear non-membrane protein in subcellular fractionation studies and was localized throughout the nuclear interior in interphase cells. It co-fractionated with LAP2beta in nuclear lamina/matrix-enriched fractions upon extraction of nuclei with detergent, salt and nucleases. During metaphase LAP2alpha dissociated from chromosomes and became concentrated around the spindle poles. Furthermore, LAP2alpha was mitotically phosphorylated, and phosphorylation correlated with increased LAP2alpha solubility upon extraction of cells in physiological buffers. LAP2alpha relocated to distinct sites around chromosomes at early stages of nuclear reassembly and intermediarily co-localized with peripheral lamin B and intranuclear lamin A structures at telophase. During in vitro nuclear assembly LAP2alpha was dephosphorylated and assembled into insoluble chromatin-associated structures, and recombinant LAP2alpha was found to interact with chromosomes in vitro. Some LAP2alpha may also associate with membranes prior to chromatin attachment. Altogether the data suggest a role of LAP2alpha in post-mitotic nuclear assembly and in the dynamic structural organization of the nucleus.

Animals↗

PubMed1995

Whole blood culture for measuring mitogen induced T cell proliferation provides superior correlations with disease state and T cell phenotype in asymptomatic HIV-infected subjects.

Proliferative responses to a panel of mitogens were compared in parallel for two sources of cells, whole blood (WB) and conventionally prepared peripheral blood mononuclear cells (PBMC), obtained from asymptomatic HIV seropositive and control subjects. Weak but statistically significant correlations of the proliferative responses were observed. Use of either lymphocyte source produced significant differences in the proliferative responses between the HIV seropositive and control subjects, but the use of WB was more powerful, with a smaller sample size being required to discriminate between the proliferative responses of the two study groups. Furthermore, proliferative responses using WB gave strong and highly significant correlations with a number of important changes in the surface marker phenotype of the lymphocyte populations in the HIV seropositive subjects including CD4, CD8, CD4:CD8 ratio and certain CD8 subsets, whereas strong correlations were not observed with the PBMC. The response of WB lymphocytes to staphylococcal enterotoxin B (SEB) was highly reproducible and provided the best discrimination between HIV-infected and control subjects. We conclude that the use of WB for measuring lymphoproliferation is easy, rapid, accurate, and discriminative for assessing and following the changes in immune function which occur in HIV seropositive subjects, applicable in the clinical as well as in the research setting.

Adult↗

PubMed1993

Immunosuppression by glucocorticoids: inhibition of production of multiple lymphokines by in vivo administration of dexamethasone.

Glucocorticoids are extremely potent immunosuppressive agents, capable of directly affecting the function of lymphocytes. We studied the effect of in vivo dexamethasone (DEX) administration on anti-CD3-induced lymphocyte proliferation and lymphokine production in mice. To characterize the kinetics and dose responsiveness of lymphocytes to DEX, splenocytes from BALB/c mice that had received a single dose of DEX in vivo were cultured in vitro with suboptimal and optimal concentrations of anti-CD3 monoclonal antibody. Cell proliferation in response to suboptimal concentrations of anti-CD3 was decreased by DEX doses of > or = 30 mg/kg; much higher doses (> or = 200 mg/kg) were required to inhibit cell proliferation in response to optimal anti-CD3 stimulation. Inhibition of suboptimal anti-CD3-stimulated proliferation was evident within 4 hr after DEX administration, was maintained for at least 24 hr, and was no longer evident at 7 and 14 days. Lymphokine secretion induced by optimal doses of anti-CD3 in vitro was differentially affected by in vivo DEX treatment. IL-1 alpha, IL-4, IL-6, IL-10, and IFN-gamma levels were decreased by treatment with low doses of DEX (30 mg/kg), whereas higher doses were required to inhibit production of IL-2, IL-3, and TNF. GM-CSF (granulocyte-macrophage-colony stimulating factor) was least susceptible to DEX inhibition. Low-dose (30 mg/kg) DEX treatment significantly reduced anti-CD3-stimulated production of most lymphokines tested at 4 and 12 hr; by 24 hr the levels of most lymphokines had begun to return to control values. Hence, our data indicate that administration of a single dose of DEX (30 mg/kg for 4 hr) results in significant suppression of lymphokine production and cell proliferation that precedes any significant cell loss and can be used as a reversible model of immunosuppression.

Animals↗

PubMed1992

Identification of anti-CD3 antibodies that do not modulate antigen but induce mitogenesis and block the mixed lymphocyte reaction.

Using a panel of anti-CD3-TCR monoclonal antibodies (OKT3 A-E), it appears possible to separate the ability to cause surface antigen modulation from inhibition of MLR or induction of mitosis. OKT3D, an antibody that recognizes the CD3 antigen at a site that can be differentiated from the epitopes recognized by other members of this panel by competition binding, does not cause antigen modulation when incubated with human T cells for up to 3 days. Despite this, OKT3D is mitogenic and is capable of blocking MLR. Two different isotypes were produced from the OKT3D clone, IgG1 and IgG2b. The IgG2b isotype of OKT3D blocked MLR even in individuals unable to respond mitogenically to this antibody. Use of members of this panel may now permit dissection of the types of signals delivered by the CD3-TCR complex inducing mitosis, receptor modulation, and other T-cell responses.

Animals↗

PubMed1991

OKT3E, an anti-CD3 antibody that does not elicit side effects or antiidiotype responses in chimpanzees.

Chimpanzees were injected with OKT3 and two other anti-CD3 antibodies, OKT3D and OKT3E. Both of the new antibodies were of the mouse IgG2b isotype. Administration of the antibodies was identical to the clinical regimen used for OKT3 in humans: 5 mg i.v., daily for 10 consecutive days. All animals were monitored for fever during administration of the antibodies, and blood samples were taken throughout the treatment period for monitoring the effects of the antibodies on peripheral lymphocyte subsets and the appearance of circulating cytokines. OKT3 produced similar clinical effects to those observed in humans; fever (2/3), as well as elevations in cytokines were observed. As in humans, peripheral T cells were cleared with the first dose and remained absent or modulated of their T cell receptor molecules throughout treatment. OKT3D, IgG2b also produced fevers (2/3) and elevations of cytokines. Although it also cleared circulating T cells with the first dose and T cell counts remained low throughout treatment, remaining circulating T cells were coated with administered antibody and were able to reexpress the CD3 antigen. OKT3E, IgG2b produced no temperature elevations and no elevations in cytokines. Although it cleared the circulation of T cells with the first does, cells reappeared during treatment, modulated of their CD3 antigens or coated with the administered antibody. All three antibodies raised antimouse antibodies, and OKT3 and OKT3D also produced blocking antiidiotype antibodies. OKT3E treatment did not result in anti-OKT3E blocking antibodies.

Animals↗

PubMed1991

Use of monoclonal antibodies to identify thymopoietin in cultured human thymic epithelial cells.

Murine monoclonal antibodies (mAbs) were developed to discriminate thymopoietin, a human thymic hormone, and thysplenin, a closely related molecule found in spleen. Three of these recognized both native and synthetic thymopoietin as well as thysplenin. Together they define two non-overlapping epitopes which withstand sodium dodecyl sulfate denaturation and can be detected by western blotting. We used these three mAbs to demonstrate the production of thymopoietin by cultured thymic epithelial cells for up to several weeks. Three additional mAbs were selective for thysplenin. Highly specific mAbs will be useful for characterizing further these physiologically distinct polypeptides.

Antibodies, Monoclonal↗

PubMed1987

Flow cytometric resonance energy transfer measurements support the association of a 95-kDa peptide termed T27 with the 55-kDa Tac peptide.

Two monoclonal antibodies (OKT27 and OKT27b) have been produced that react with distinct epitopes of a 95-kDa peptide. The T27 antigen is widely distributed, being expressed on B lymphocytes, monocytes, and adult T-leukemic cells but not on polymorphonuclear leukocytes or platelets. There was a low level of T27 expression on resting T cells that increased on T-cell activation. In preliminary studies, the OKT27b antibody coprecipitated a 55-kDa peptide, as well as the 95-kDa peptide, from the radiolabeled cells of the HuT 102B2 cell line. Preclearance with anti-Tac, a monoclonal antibody to the 55-kDa peptide of the multichain interleukin 2 receptor, removed the 55-kDa but not the 95-kDa peptide from subsequent OKT27b immunoprecipitates of HuT 102B2 extracts, suggesting the possibility that the T27 peptide was associated with the Tac peptide. However, the precipitation of the p55 Tac peptide by OKT27b was quite inconsistent. Thus, additional information was sought using a flow cytometric energy transfer technique to provide a physical estimation of the proximity between the Tac and the T27 peptides. The flow cytometric version of the fluorescence resonance energy transfer technique permits the determination of inter- and intramolecular distances at 2- to 10-nm levels on a cell-by-cell basis. Using this approach, there was a mean energy transfer of 7.3% with HuT 102B2 cells when fluorescein isothiocyanate anti-Tac served as the donor and tetramethylrhodamine isothiocyanate OKT27 served as the acceptor. In contrast, there was no energy transfer in comparable studies observed when fluorescein anti-Tac and rhodamine anti-transferrin receptor antibodies were used. These observations support the conclusion that there is a close nonrandom proximity in HuT 102B2 cells between the 95-kDa peptide identified by the OKT27 monoclonal antibody and the p55 Tac peptide of the multichain interleukin 2 receptor.

Antibodies, Monoclonal↗

PubMed1985

Null cell identification and characterization with OKT16: an anti-p40 monoclonal antibody.

A murine monoclonal antibody, OKT16, specific for human lymphocytes of T lineage, was isolated by standard immunization and hybridization techniques. The distribution of the antigen defined by OKT16 was similar to the antigen reactive with monoclonal antibodies 3A1 and WT1. This identity of antigen targets was confirmed in an enzyme-linked immunosorbent assay system and by sequential immunoprecipitation. Under reducing conditions, OKT16 reacted with an antigen of 40K daltons; however, under nonreducing conditions this antigen appeared as an 84K-dalton molecule, which suggests that the p40 antigen exists as a disulfide-linked dimer. By indirect immunofluorescence, OKT16 reacted with a greater fraction of nonrosetting, non-B (null) lymphocytes than with antibodies to other T cell-specific proteins. Two-color immunofluorescence demonstrated the coexpression of the T16 antigen and the C3bi receptor on most null cells. The T10 antigen (found on cortical thymocytes and activated peripheral T cells) was restricted to most T16-bearing null cells and expression of the Fc receptor for aggregated IgG (defined by monoclonal antibody 73.1) was restricted to a major subset of T16-bearing null cells. The T cell-specific markers defined by OKT8, OKT11, and OKT17, as well as the monocyte marker defined by OKM5, were expressed by smaller subsets of OKT16-reactive null cells. These studies support by phenotypic analysis the functional heterogeneity ascribed to null cells. The 40K-dalton T16 antigen has the most extensive null cell representation of all the T lineage markers described to date.

Antibodies, Monoclonal↗

PubMed1984

Distinct classes of human T-cell activation antigens.

The characterization of three groups of antigens expressed by activated human T lymphocytes and detected by monoclonal antibodies is reported. Antigens defined by OKT19, OKT21, and OKT22 do not appear on in vitro activated T cells until increases in DNA synthesis become apparent and are not detected on most Interleukin 2 (IL-2)-independent cell lines and normal peripheral blood lymphocytes, monocytes, and granulocytes. Cell surface molecules reactive with the monoclonal antibodies OKT23 and OKT24 are displayed prior to any notable increase in DNA synthesis and are present on IL-2 independent cell lines, irrespective of lineage. T23 and T24 do not appear on peripheral blood cells and their distribution more closely resembles that of the T9 antigen (the receptor for transferrin) than antigens of the other groups. The third group of antigens, T14 and T20, have been classified as "early" antigens relative to DNA synthesis. They are expressed by distinct populations of normal lymphoid cells as well as by some IL-2-independent cell lines. Display of each group of activation antigens on T lymphocytes can be induced by either phytohemagglutinin, purified protein derivative from tuberculin, or allogeneic non-T cells, is not restricted to the OKT4+ or OKT8+ subsets, and is predominant on cells exhibiting the light-scattering properties of blast cells. The relative lack of expression of these antigens among normal peripheral blood cells make them attractive candidates for identifying changes in the status of immune activation.

Antibodies, Monoclonal↗

PubMed1984

Possible involvement of the T4 molecule in T cell recognition of class II HLA antigens. Evidence from studies of CTL-target cell binding.

The present study examines the potential role of the T4 molecule in functional cell-cell interactions between target cells and human cytotoxic T lymphocyte (CTL) clones that are specific for HLA class II alloantigens encoded by the SB locus. There were marked differences (greater than 30-fold) between the seven SB-specific clones studied with respect to their susceptibility to inhibition by anti-T4 as well as anti-T3 antibodies. We wished to test the hypothesis that such variation among the clones would be due to differences in clonal "affinity" for antigen. To quantitate differences among the CTL clones in the tightness with which they bind target cells, the clones were analyzed using a previously published assay of susceptibility of CTL-target cell conjugates to dissociation in the presence of unlabeled targets. The results revealed that the clones that were most susceptible to inhibition by anti-T4 and anti-T3 were the weakest target cell binders, and vice versa. Anti-T4 antibody could partially induce dissociation of functional CTL-target cell conjugates in the absence of any added cold targets. For the "highest affinity" clone such anti-T4 antibody-induced dissociation could be observed at 4 degrees C but not 23 degrees C. These results indicate that the T4 molecule is functionally involved in target cell binding by CTL, and raise the possibility that although it is easiest to demonstrate the function of the T4 molecule in "low affinity" clones, that function may also be operative in the "high affinity" clones.

Adult↗

PubMed1984

Immunoassay for bovine serum thymopoietin: discrimination from splenin by monoclonal antibodies.

A polyclonal rabbit anti-bovine thymopoietin antiserum was used to develop a radioimmunoassay and sandwich enzyme-linked immunosorbent (ELISA) assay for the thymic hormone thymopoietin. Both assays showed slightly less sensitivity for the closely related splenic hormone splenin (SP) than thymopoietin (TP) and markedly less sensitivity for the human as compared with the bovine polypeptides. A number of murine monoclonal antibodies specific for bovine thymopoietin were generated; they were unreactive with bovine splenin and were also unreactive with human thymopoietin and splenin. A sandwich ELISA using these monoclonal anti-TP antibodies together with polyclonal rabbit anti-TP was specific for bovine thymopoietin and measured 300-500 ng/ml thymopoietin in bovine serum. Similar approaches are being pursued to develop an immunoassay for thymopoietin in human serum.

Animals↗

PubMed1984

Thymopoietin radioreceptor assay utilizing lectin-purified glycoprotein from a biologically responsive T cell line.

Radiolabeled thymopoietin that was biologically active and of high specific activity was prepared by a novel technique involving protection of free amino groups, selective excision of the protected N-terminal prolyl group with post-proline cleaving enzyme, reaction of the newly exposed alpha-amino group with a highly radioiodinated compound, and deprotection and purification of the polypeptide. Binding of this radiolabeled thymopoietin was not demonstrable by conventional techniques with cells, cell membranes, or solubilized cell membranes, apparently due to the presence of active proteases in these preparations. A glycoprotein with thymopoietin binding properties was prepared by lectin purification from the detergent-solubilized membranes of CEM cells, a human T cell line that responds to thymopoietin in vitro with increases in intracellular cyclic GMP. Presumably this procedure separated the thymopoietin binding protein from membrane proteases, thus permitting the development of a radioreceptor assay. Evidence is presented that the thymopoietin binding protein represents a thymopoietin receptor that is probably related to the mediation of immunoregulatory actions of thymopoietin on a subset of peripheral T cells.

Binding, Competitive↗

PubMed1984

Distinct epitopes on the T8 molecule are differentially involved in cytotoxic T cell function.

The present report attempts to determine if there are distinct epitopes on the T8 molecule that are involved in class I-restricted cytotoxic T lymphocyte (CTL) function. A panel of 9 monoclonal antibodies (OKT8A,B,C,E,F,G,H,I, and OKT5) was produced and all antibodies were shown to bind to the T8 molecule. This panel of antibodies was employed to characterize the distribution of distinct epitopes on the T8 molecule and to block the activity of class I-specific influenza virus-immune and allo-immune CTL effector function. Significant differences in the ability of the anti-T8 antibodies to block CTL function were observed: OKT8C and T8F blocked best (49 and 55% respectively); OKT8A,E,G,H,I, and OKT5 blocked less well (24-31%); and OKT8B blocked marginally (11%). There was no correlation between the capacity of the antibodies to block CTL function and their heavy chain isotype. Competitive binding of the different OKT8 antibodies to the cell surface and differential trypsin sensitivity of the epitopes recognized by the antibodies indicated that OKT8C and T8F were located on topographically distinct regions of the T8 molecule. These results indicate that specific epitopes on the T8 molecule are involved in CTL function, and that there could be more than one functional site on the molecule.

Animals↗

PubMed1984

The T4 surface antigen is involved in the induction of helper function.

The OKT4 monoclonal antibody reacts with a 62K m.w. glycoprotein present on a subset of human T cells with the capacity to help or induce B cell differentiation. The present studies were undertaken to determine whether the T4 molecule itself plays any role in the helper function mediated by T4+ cells. Using a series of monoclonal antibodies (OKT4, OKT4A-E), which react with distinct noncompeting epitopes of the T4 molecule, we found that OKT4A and OKT4E, but not OKT4, antibodies inhibited the induction of B cell differentiation by T4+ cells. This inhibition was apparent when using nonirradiated or irradiated T4+ cells, and was noted over a wide range of concentrations. Importantly, inhibition occurred only if the antibody was present during the first 24 hr of cell culture, and the presence of antibody did not alter the kinetics of induction of B cell differentiation. Thus, these data suggest that the T4 molecule plays an early, critical role in cellular interactions required for helper cell function.

Antibodies, Monoclonal↗

PubMed1984

Distribution of antigens defined by OKB monoclonal antibodies on benign and malignant lymphoid cells and on nonlymphoid tissues.

Monoclonal antibodies OKB1, OKB2, OKB4 and OKB7 have been previously shown to detect distinctive antigens displayed on B, but not on T, lymphocytes. Benign and malignant lymphoid cells were investigated for their reactivity with these antibodies in cell suspension by indirect immunofluorescence and in cryostat tissue sections by the avidin-biotin complex immunoperoxidase technique. Fetal liver pre-B cells and pre-B and common type acute lymphoblastic leukemia cells isolated from 15 patients were OKB1-OKB2+OKB4-OKB7-. All mature lymphoid tissue B cells and the neoplastic cell surface immunoglobulin-positive (SIg+) B cells isolated from each of 47 B cell neoplasms were OKB2+. OKB1 and OKB7 were expressed by interfollicular, follicular center, and many, but not all, mantle zone B cells. OKB4 was expressed by follicular center cells, but not by mantle zone or interfollicular B cells. The neoplastic SIg+ B cells isolated from 45 of 47 B cell malignancies were OKB1+OKB4+, and those isolated from 45 of 46 B cell malignancies were OKB7+. The neoplastic B cells of one mantle zone lymphoma were OKB1-, of one small lymphocytic cell lymphoma were OKB7-, of one large cell lymphoma were OKB4-, and of one small lymphocytic cell lymphoma with a monoclonal gammopathy were OKB1-OKB4-. Normal and myeloma plasma cells were OKB-. The malignant T cells isolated from 12 T cell neoplasms were OKB2-OKB4-, but were OKB1+ and/or OKB7+ in 3 cases. Thus, the OKB antibodies appear to detect distinctive antigens that may be expressed at different stages of B cell differentiation. In addition, OKB4 reacted with selected renal and respiratory epithelium, and OKB2 reacted with a wide range of epithelial tissues. The OKB antibodies should prove useful in the investigation of B cell differentiation and may aid in the identification and characterization of lymphoproliferative malignancies with significant therapeutic and prognostic differences not identifiable by conventional histopathologic and immunologic methods.

Animals↗

PubMed1983

Cell membrane perturbation of resting T cells and thymocytes causes display of activation antigens.

Three human lymphocyte antigens recognized by monoclonal antibodies OKIa1, OKT9, and OKT10 were found to be minimally represented on resting peripheral T cells (all three) and thymocytes (OKIa1 and OKT9). These antigens, which are present on "activated" T cells, were promptly displayed on "resting" T cells or thymocytes following cross-linking of surface-bound monoclonal antibody by horse alpha-mouse IgG. These experiments suggested that membrane perturbations may induce the expression of certain antigens that are normally present in an unexpressed form in resting cells.

Antibodies, Monoclonal↗

PubMed1983

Five epitopes of a differentiation antigen on human inducer T cells distinguished by monoclonal antibodies.

A series of mouse monoclonal antibodies reacting with human T cells of the helper/inducer subclass, OKT4, 4A, 4B, 4C, and 4D, have been reported. Using double-fluorescent staining and complement-mediated depletion, it was shown that the antigen(s) recognized by OKT4, 4A, 4B, 4C, and 4D antibodies are present on the same cell. Using FITC-labeled OKT4, 4A, 4B, 4C and 4D, a lack of competition between the antibodies for their epitopes was shown. Immunoprecipitation of the antigen recognized by each antibody yielded a molecule of approximately 60,000-62,000 Da. Sequential precipitation with several antibodies resulted in a minimum of additional precipitated antigen following removal of the first antigen. Capping of cell surface antigen with OKT4, followed by staining with OKT4, 4A, 4B, 4C, or 4D, indicated that the epitopes for all five antibodies co-cap. A sandwich ELISA assay using OKT4 and the other antibodies showed that molecules binding to OKT4A, 4B, 4C, and 4D also bound OKT4. It can therefore be concluded that monoclonal antibodies OKT4, 4A, 4B, 4C, and 4D recognize distinct epitopes present on a molecule of approximately 60-62,000 Da on human helper/inducer T cells.

Animals↗

PubMed1983

Patterns of antigenic expression on human monocytes as defined by monoclonal antibodies.

A series of seven monoclonal antibodies directed at determinants on human peripheral blood monocytes were produced and characterized. The antibodies were separated into three groups based on cell distribution and percentages of monocytes bearing antigen. Hybridoma antibodies, termed OKM1, OKM9, and OKM10, recognized antigen(s) expressed on the majority of adherent monocytes, null cells, and granulocytes. The second group, comprising OKM5 and OKM8, reacted with most adherent monocytes and platelets. OKM3 and OKM6, comprising a third group of antibodies, reacted with a subpopulation of adherent monocytes and platelets. OKM antibodies were not expressed on lymphocytes, thymocytes, and lymphoblastoid cells, with the exception of OKM3 which reacted with three B-cell lines. SDS gels of immunoprecipitates formed with OKM antibodies yielded the following tentative molecular weight results: OKM1 and OKM9 antigens appeared to be 160,000 (nonreduced) and 170,000 (reduced); OKM10 precipitated two polypeptides of 170,000 and 115,000 (reduced); OKM5 and OKM8 precipitated a single polypeptide of 88,000 (reduced, nonreduced); OKM6 antigen appeared to be 116,000 (nonreduced) and 130,000 (reduced).

Animals↗