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W Newman

Publications and source records attributed to W Newman.

At least 91 records · Page 5Linked to original sources
PubMed1988

Regulation of colony-stimulating factor 1-dependent macrophage precursor proliferation by type beta transforming growth factor.

Transforming growth factor (TGF) type beta, a potent growth modulator, has recently been shown to inhibit the proliferation and function of several types of immune cells. This report investigates the effect of human platelet purified TGF-beta on CSF-1-induced proliferation in liquid cultures. We used two cell types to study TGF-beta effects, bone marrow precursors and a c-myc partially transformed CSF-1-dependent macrophage cell line designated BMM-8. We found that CSF-1-dependent proliferation of both cell types was strongly inhibited by TGF-beta in a dose-dependent manner. Approximately 1.6 and 8 pM TGF-beta inhibited 50% of CSF-1 proliferation of the bone marrow precursors and BMM-8, respectively. Inhibition appeared to be reversible, as bone marrow and BMM-8 cells proliferated in response to CSF-1 after preincubation of the cells in TGF-beta. Interestingly, inhibition of hematopoietic cells was observed only after a lag period of 24 to 48 h after onset of cultures. TGF-beta inhibition was partially diminished when increasing amounts of CSF-1 were added to the cultures. TGF-beta inhibition did not involve secondary inhibitory factors such as IFN or PG, both of which have been previously shown to suppress CSF responsiveness. Finally, flow cytometric analysis of the cell cycle indicated that within 48 h, TGF-beta-treated BMM-8 cells were prevented from entering S phase. These results suggest that TGF-beta may play an important role in the negative regulation of macrophage production.

Animals↗
PubMed1986

Early events in lymphocyte activation as defined by three new monoclonal antibodies.

Three new lymphocyte activation antigens are described whose kinetics of appearance place them very early in the activation pathway. The 78,000 dalton early antigen (Ea) 1 is present at low levels on resting lymphocytes, and its expression is enhanced twofold to threefold within 3 hr of stimulation. Ea2, a nondisulfide-bonded 86,000 and 73,000 dalton heterodimer, is first detectable 3 hr after activation and peaks by 9 hr. Its presence on all but a few cell lines, plus the variable association with a lower m.w. (28,000) structure, suggest that it may serve as a receptor for a growth factor. Neither Ea1 nor Ea2 are restricted to lymphocytes. The 31,000 dalton Ea3 antigen is induced only by PHA but not by other means of activation, and may pre-exist within the cell. The Ea3 antibody blocks PHA-induced but not OKT3-induced mitogenesis, suggesting differences in the pathways of activation by these two stimuli. These reagents, and OKT3, were used to define the cyclosporine A (CSA)-sensitive stage of lymphocyte activation. CSA blocks at a point before the biosynthesis of Ea1 and after that of T3/T cell receptor loss from the cell surface, at a point close to Ea2 biosynthesis.

Animals↗
PubMed1986

K562 killing by K, IL 2-responsive NK, and T cells involves different effector cell post-binding trigger mechanisms.

The monoclonal antibody 13.3 specifically blocks the trigger process of the NK-K562 cytolytic sequence at a post-binding effector cell level. This antibody was used to define differences in the lytic trigger processes of NK and other mechanisms of K562 lysis. Monoclonal antibody 13.3 inhibited lysis of K562 target cells by freshly isolated peripheral blood lymphocytes (PBL) and purified large granular lymphocytes (LGL), but had no inhibitory effect on antibody-dependent cell-mediated cytotoxicity to K562 by these effectors. Lectin-dependent cellular cytotoxicity (LDCC) to this target cell was also unresponsive to 13.3. The 13.3-induced inhibition of NK-K562 lytic activity persisted when PBL were activated in culture with interleukin 2 (IL 2) for periods up to 48 hr. After 48 hr of culture, the degree of inhibition diminished progressively in medium containing fetal calf serum but not in medium containing autologous serum. This 13.3-unresponsive lytic activity in cultured PBL could be attributed to more than one cell type and was present in both the LGL and Fc gamma receptor-depleted T cell fraction. Thus, K562 lysis by freshly isolated human lymphocytes via NK, K, and LDCC mechanisms is characterized by heterogeneity of the post-binding effector cell trigger mechanism. K562 lysis by lymphocytes cultured with IL 2 is similarly heterogeneous.

Antibodies, Monoclonal↗
PubMed1985

Heterogeneity of human natural killer recognition demonstrated by cloned effector cells and differential blocking of cytotoxicity with monoclonal antibodies.

Monoclonal antibodies (MoAb) against cell surface determinants were employed to investigate the specificity of natural killer (NK)-like lysis by cloned human effector cells recognizing only K562, only HSB2, or both K562 and HSB2 target cells. MoAb W6/32.HL, TU39, YD1/48.HLK, and anti-Tac failed to inhibit lysis despite the expression of antigens bound by these MoAb on the effector cell surface. MoAb OKT3 moderately (less than or equal to 50%) blocked lysis of K562 and HSB2 targets, whereas MoAb 13.1, which binds T200 molecules, strongly (up to 95%) blocked lysis of K562, but not HSB2, targets. MoAb 13.1 inhibited lysis by clones which killed only K562, as well as lysis by those which killed both HSB2 and K562. In the latter case, however, only lysis of K562 was inhibited. Taken together, these results may suggest the existence of multiple receptor specificities on a single NK-active clone.

Antibodies, Monoclonal↗
PubMed1985

Activated human B cells display a functional IL 2 receptor.

These studies deal with the expression of a functional IL 2 receptor on activated primary human B cells. Antibody against the receptor (alpha-TAC) reacted with 25 to 65% activated B cells, inhibited B cell proliferation by 50% and inhibited B cell secretion of Ig by greater than 90%. These effects were shown to be independent of contaminating T lymphocytes. Anti-TAC immunoprecipitated a molecule of identical size (65,000 daltons) from T and B lymphocytes; B cells were also shown to actively synthesize the IL 2 receptor. The chymotryptic peptide chromatograms of TAC antigen from T and B cells show these molecules to be indistinguishable.

Antibodies, Monoclonal↗
PubMed1984

Immunobiological and immunochemical aspects of the T-200 family of glycoproteins.

We present some new structural and functional data on the family of high mol. wt glycoproteins of human hematopoietic cells which strongly suggests they play a role proximate to certain Ca2+-dependent events. Structural data on this complex are presented which describe elements of its tertiary structure and associations between the different glycoproteins in the plasma membrane. The mol. wt profile of the T-200 structures on highly enriched natural killer (NK) cells is used to support an argument for a T-cell vs a myeloid nature for NK cells. Furthermore, certain murine monoclonal antibodies directed to a particular epitope of T-200 on NK cells are shown to block cytolysis. The specificity of this blockade strongly suggests that T-200 may be an NK receptor. A mapping of this effect has allowed us to define a new stage of the lytic cycle, termed "triggering". This very rapid event occurs subsequent to conjugation between the NK and the target cell, and prior to the Ca2+-dependent second-phase programming for lysis. A close association between "triggering", as defined by antibodies to T-200, and Ca2+-dependent stimulus-secretion events support the concept of T-200 as a receptor structure.

Antibodies, Monoclonal↗
PubMed1983

The mechanism of augmentation of natural killer cell activity by syngeneic tumor cells: role of macrophage-derived factor in NK boosting.

The role of natural killer (NK) cells in controlling tumor growth was investigated using an NK-susceptible (c127v-IC2) and an NK-insusceptible (c127av) subline of the lymphoma L5178Y. Syngeneic DBA/2 mice inoculated intraperitoneally with c127v-IC2 tumor cells survived significantly longer than did c127av-bearing mice. Similarly, c127v-IC2, but not c127av tumor cells, were found to augment NK activity of spleen and peritoneal exudate cells in both DBA/2 and BALB/c nu/nu mice when inoculated into the peritoneal cavity. C127v-IC2 tumor cells incubated with either DBA/2 or BALB/c nu/nu spleen cells in vitro boosted NK activity and induced the production of gamma-type interferon (IFN), whereas incubation with c127av tumor cells induced neither NK activity nor IFN. Two kinds of cells cooperated in the production of IFN in response to c127v-IC2 tumor cells, namely, cells which were nonadherent, bore asialo-GM1, NK-1.2 and a low level of Thy-1.2 antigen and thus closely resembled NK cells, and those which were adherent and phagocytic and lacked both asialo-GM1 and NK-1.2 markers, presumably macrophages. Further analysis strongly suggested that c127-v-IC2 tumor cells stimulate macrophages to produce factor(s) which can induce the production of IFN by NK cells. The induced IFN was shown to be of the gamma type by its lability at pH 2.0 and insusceptibility to anti-IFN alpha, beta serum. This suggests a novel pathway for NK cell activation, and strongly supports the importance of macrophages and NK cells in natural resistance against certain tumors.

Animals↗
PubMed1983

Identification and functional characterization of two distinct epitopes on the human T cell surface protein Tp50.

The structural and functional domains of Tp50, the human T lymphocyte surface protein associated with the E rosette receptor, were probed with the use of two murine monoclonal antibodies. Lysostripping, immune precipitation, and competitive binding experiments demonstrated that antibodies 9.6 and 35.1 bind to Tp50 epitopes in close proximity. In functional studies, both antibodies caused a similar degree of antigenic modulation, inhibited T cell proliferative responses, and inhibited cytotoxic T lymphocyte function without affecting cells that mediate antibody-dependent cell-mediated cytotoxicity. The antibodies were strikingly different, however, in that antibody 9.6 inhibited E rosette formation and natural killer cell-mediated lysis, whereas antibody 35.1 did not. These results could not be ascribed to differences in antibody class or binding characteristics, because Scatchard analysis demonstrated that these two IgG2a antibodies have comparable avidity and that T cells bind each antibody in equivalent amounts. The differential binding of antibodies 9.6 and 35.1 to T cells from nonhuman primates further supports the interpretation that the differences between the antibodies in their effects on E rosette formation and natural killer function stem from the fact that they bind to distinct epitopes of Tp50. The implications of these findings for understanding the functions of Tp50 molecules are discussed.

Animals↗
PubMed1983

Blockade of NK cell lysis is a property of monoclonal antibodies that bind to distinct regions of T-200.

The previously described NK inhibitory monoclonal antibody 13.1 is shown to immunoprecipitate a series of high m.w. glycoproteins homologous with the murine T-200/Ly-5 molecules. Not all antibodies to the human T-200 molecule, however, have an inhibitory effect on NK cell function. A comparison is made between two noninhibitory anti-T-200 antibodies, 13.5 and 13.6, and two inhibitory anti-T-200 antibodies, 13.1 and 13.3. All antibodies are of the IgG1 subclass. Sequential immunoprecipitation experiments show that these antibodies react with the same set of molecules. The differences in NK-blocking activity could not be explained by the amount of antibody bound per cell in NK-enriched populations, nor by the avidity with which they bound. It is shown by competitive radiobinding assays that the 13.1 and 13.3 antibodies define a region, termed region A, distinct from that defined by the nonblocking antibodies 13.5 and 13.6, termed region B. Region B is shown to reside between the membrane and region A. These findings show that the inhibition of NK lysis by anti-T-200 antibodies is a function of the site on that molecule to which these antibodies bind. This may also explain the ability of antibodies to the A region of T-200 to block selectively the lysis of myeloid and erythroid tumor targets, with no effect on the lysis of T lymphoma targets.

Animals↗
PubMed1983

Interleukin 2 induces gamma-interferon production: participation of macrophages and NK-like cells.

Interleukin 2 (IL 2) has been shown to be a potent stimulator of natural killer (NK) cells. In the present studies, partially purified mouse and human IL 2 preparations were also found to induce interferon (IFN) from mouse spleen cells. By the criteria of sensitivity to treatment at pH 2 and failure to be neutralized by a potent anti-alpha, beta IFN serum, the species of IFN produced was of type gamma. Cooperation between two types of cell, a macrophage and an NK-like cell, was required for IFN production by murine spleen cells treated with IL 2. The requirement for macrophages could be replaced with supernatant obtained by incubating macrophages for 24 hr with lymphokine preparations containing IL 2. Interestingly, mature T cells apparently played no role in the process. Furthermore, the beige (bg/bg) mutation, which severely impairs NK cell lytic activity, had no effect on the ability of NK-like cells to participate in IFN production. Cell fractionation experiments revealed no dissociation between the requirements for augmentation of NK cytotoxic activity and for IFN production, and it is concluded that at least a portion of the NK boosting induced by IL 2-containing preparations is mediated through gamma-IFN.

Animals↗
PubMed1983

T cell nature and heterogeneity of recognition structures of human natural killer (NK) cells.

Very low doses of trypsin (5 micrograms/ml) are sufficient to ablate NK cell activity. This finding was used to make several observations, and we have attempted to relate these observations to specific cell surface macromolecules. First, trypsinized effector cells no longer lysed seven different NK-susceptible targets, but the lysis of three additional targets was unaffected. These results suggest a heterogeneity of recognition potential that is inconsistent with the notion that there is only one class of NK "receptors" and one class of "target structures." Trypsin does not affect the conjugation of effector and target cells. Secondly, we have tried to identify those cell surface molecules that are affected by this low dose of enzyme. The examination of the 125I-labeled glycoprotein fraction NK-enriched cells showed that at least four molecules are cleaved, one of which may be in the T200 family. The examination of the [3H]galactose-labeled cell surface glycoproteins suggested in particular that some high m.w. glycoproteins were affected at the dose of trypsin that ablates NK function. Analysis of those molecules that we previously implicated in NK function, defined by monoclonal antibodies that block NK lysis, allowed us to rule out a role for the Tp 50 and Lp95-150 structures, while providing additional evidence of a role for the T200 glycoproteins in the trypsin-sensitive stage of cytolysis. Finally, closer examination of the electrophoretic mobilities and trypsin sensitivity of the T200 structures on highly enriched NK cells showed these structures to be indistinguishable from the T cell form of T200, yet quite distinct from the monocyte form. These results are therefore consistent with the possibility that NK cells are of the T rather than the monocyte lineage, and furthermore support a role for the T200 structure in the post-binding trypsin-sensitive stage of the NK cytolytic process.

Antibodies, Monoclonal↗
PubMed1983

Definition of a "trigger" stage in the NK cytolytic reaction sequence by a monoclonal antibody to the glycoprotein T-200.

The monoclonal antibody 13.1 recognizes an epitope on the T-200 glycoprotein and blocks natural killer (NK) lysis of the erythroleukemia target K562, but not of the acute lymphoblastic leukemia T cell target Molt-4. The inhibitory effect is at the killer cell level and not the target cell level, which suggests that 13.1 may react with a receptor on NK cells. This hypothesis was tested in assays to delineate precisely where in the NK cytolytic reaction sequence 13.1 interferes with lysis; 13.1 did not block initial NK-target cell interaction as measured in a target binding cell assay. With the use of a Ca++ pulse technique, 13.1 did not block any events occurring during Ca++-dependent programming. If the antibody was added after conjugate formation but before the addition of CaCl2 to initiate programming, however, full inhibition of NK lysis occurred. Therefore, 13.1 antibody defines a distinct stage in the NK reaction sequence that links target binding to the initiation of calcium-dependent programming events. NK cell binding alone is not sufficient to trigger lytic events, and the presence of a second structure or a distinct portion on a single structure is required to trigger lysis. We show that 13.1 blocks the ability of K562 to inhibit NK killing of Molt-4 in a cold target inhibition assay. Therefore, despite the fact that 13.1 does not disrupt conjugate formation, NK specificity may exist at a post-binding site rather than at the initial NK-target binding interaction. Our data suggest that the T-200 glycoprotein on NK cells triggers the initiation of the lytic events.

Antibodies, Monoclonal↗