Biomedical subjects

R D Trochelman

Publications and source records attributed to R D Trochelman.

4 recordsLinked to original sources

PubMed1990

A structural characterization of the Mo3 activation antigen expressed on the plasma membrane of human mononuclear phagocytes.

Mo3 is an activation Ag expressed by human monocytic cells after stimulation in vitro by PMA, LPS, certain cytokines, and muramyl dipeptide. The structural characterization of Mo3 has been made possible by the development of a mAb (anti-Mo3f) that immunoprecipitates Mo3 from Nonidet P-40 lysates of radiolabeled PMA-stimulated U-937 cells and LPS-activated monocytes. On SDS-PAGE (nonreducing conditions) of anti-Mo3f immunoprecipitates, U-937 Mo3 is a single broad band of 39 to 66 kDa, whereas monocyte Mo3 is smaller with an apparent molecular mass of 32 to 56 kDa. Under reducing conditions, there is an increase in the m.w. of both species of Mo3 suggesting the existence of internal disulfide bonds. Mo3 is a glycoprotein with carbohydrate of the N-linked complex type as evidence by a reduction in m.w. by 40 to 50% after treatment with endoglycosidase F or N-glycanase; neuraminidase treatment produces a 3-kDa reduction in m.w. Deglycosylated Mo3 isolated from U-937 and monocytes have similar m.w. suggesting that the molecular heterogeneity of the native Mo3 may be due to differences in glycosylation. Mo3 is sensitive to phosphatidylinositol-specific phospholipase C with the release of native Mo3 from the surface of PMA-stimulated U-937 cells. These results indicate that Mo3 is a member of the glycosylphosphatidylinositol-linked family of surface glycoproteins.

Antibodies, Monoclonal↗

PubMed1990

Human mononuclear phagocyte activation antigens.

Activation of mononuclear phagocytes causes changes in plasma membrane composition that include the expression of surface antigens and receptors. Monoclonal antibody technology has made it possible to identify and characterize newly expressed surface antigens. Among these "activation antigens" is a glycoprotein, Mo3, which (among hematopoietic cells) is selectively expressed by human mononuclear phagocytes that have been exposed to inflammatory factors in vitro and in vivo. Progress toward a functional and structural analysis of Mo3 is described.

Animals↗

PubMed1989

Identification of a human monocyte antigen (Mo3e) associated with cellular activation and lymphokine responsiveness.

A murine monoclonal IgM antibody recognizes an antigen (Mo3e) found on the surface of human peripheral blood monocytes stimulated with phorbol 12-myristate 13-acetate (PMA), lipopolysaccharide and muramyl dipeptide and blocks the monocyte response to migration inhibitory factor (MIF). We utilized Western blot and immunoprecipitation analyses of whole cell lysates to investigate the biochemical nature of this monocyte antigen. Two distinct bands (75 kDa and 50 kDa) were detected by anti-Mo3e after monocyte lysates were resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and transblotted onto nitrocellulose paper. Stimulation of monocytes with PMA resulted in a marked increase in the amount of the 50-kDa species. Immunoprecipitation of Mo3e from lysates of surface-iodinated cells demonstrated one broad band at 55-80 kDa which increased after PMA stimulation. The epitope identified by anti-Mo3e was resistant to 2-mercaptoethanol and heat treatment (100 degrees C/5 min) and was sensitive to trypsin or papain treatment. Two-dimensional SDS-PAGE analysis revealed that the 75-kDa species has an isoelectric point of 7.0 and the 50-kDa species is more acidic with an isoelectric point of 5.3. These results indicate that anti-Mo3e antibody defines unique monocyte proteins that may play a role as suggested by previous studies, in monocyte activation and responsiveness to MIF.

Antibodies, Monoclonal↗

PubMed1988

Expression of Mo3e antigen by cultured human umbilical vein endothelial cells (HUVEC) stimulated by phorbol myristate acetate (PMA) and related pharmacological inducers of protein kinase C.

Mo3e is a protease-sensitive membrane antigen (p75,50) selectively expressed by human monocytic cells (monocytes and U-937 cells) stimulated in vitro by exposure to a variety of activating factors, including phorbol diester compounds, bacterial lipopolysaccharide (LPS), and muramyl dipeptide (MDP)(R.F. Todd et al., J. Immunol. 135, 3869, 1985). Here we report that primary and multiply-passaged cultures of HUVEC also express the Mo3e determinant after stimulation by phorbol myristate acetate (PMA) and related inducers of protein kinase C. As measured in a radioimmunoassay of anti-Mo3e antibody binding to monolayer cultures of HUVEC, unstimulated cells bore little if any Mo3e. After culture for 4-120 hr in medium containing PMA, 4 beta-phorbol dibutyrate, 4 beta-phorbol didecanoate, or mezerein (each at a concentration of 81 nM), or 1-oleoyl-2-acetoyl-sn-3-glycerol (1 mM), HUVEC were found to selectively express the Mo3e determinant. The magnitude of expression was dependent upon the concentration of the stimulus, maximal by 24 hr, and inhibited by cycloheximide. The combination of PMA and the calcium ionophore, ionomycin, had an additive or synergistic effect on HUVEC Mo3e expression. The biologically inactive phorbol compounds 4 beta-phorbol and 4 alpha-phorbol didecanoate failed to stimulate Mo3e expression. Also inactive as inducers of HUVEC Mo3e expression were crude lymphokine and monokine supernatants, recombinant human lymphokines (interferon-gamma and interleukin-2), recombinant human monokines (interleukin-1 and tumor necrosis factor), bacterial cell wall products including LPS and MDP, pharmacologic agents that increase intracellular cyclic adenosine monophosphate (prostaglandin E2, cholera toxin, theophylline, isoproterenol and isobutylmethylxanthine), lectins (Con A and PHA), and heparin. These results indicate that Mo3e is an inducible plasma membrane antigen of not only mononuclear phagocytes but also cultured HUVEC.

Animals↗