Showing papers by "Charles A. Dinarello published in 1982"

The biology of saa: identification of the inducer, in vitro synthesis, and heterogeneity demonstrated with monoclonal antibodies

Abstract: Continued studies of the macrophage-derived mediator of SAA synthesis (SAA Stimulating Factor) confirm our previous observations that SAASF copurified with leukocytic pyrogen (LP) and lymphocyte activating factor (LAF). Moreover, new data demonstrate three separate isoelectric points for human LP-LAF-SAASF each of which possess the three biological activities. During the purification of 15,000 MW LP from crude stimulated mononuclear cell supernatants, only those fractions with pyrogenic activity in rabbits caused augmented stimulation of lymphocytes (LAF) and induced SAA synthesis in mice. Purified human LP stimulated isolated mouse hepatocytes in vitro to synthesize SAA in a dose-responsive manner. Colchicine treatment of hepatocytes led to decreased secretion of SAA into the medium and to an intracellular accumulation of SAA. Messenger RNA was isolated from the livers of endotoxin-stimulated mice and translated in a wheat-germ cell-free system. A major product was identified at 13-14,000 MW. Immunoprecipitation with anti-mouse AA identified several bands on autoradiography of polyacrylamide gels. These larger SAA precursors may account for the previously noted heterogeneity of human SAA, comprising at least 6 SAA isomers, of similar molecular weight but different solubility and electrophoretic charge characteristics. Two monoclonal antibodies (IgM-K and IgG1-K) have been prepared using standard cell hybridization techniques. They are directed at the variable COOH terminal region of SAA since they detect differences between the 6 human SAAs but do not react with human, monkey, dog or mouse AA proteins, human AP, C-reactive protein, IgG nor albumin. These antibodies will be useful in examining the origin, structure and function of SAA.

Ability of human leukocytic pyrogen to stimulate brain prostaglandin synthesis in vitro.

Abstract: Fever is thought to be mediated by leukocytic pyrogen (LP), a polypeptide synthesized by phagocytic leukocytes and which is responsible for the upwards resetting of the hypothalamic thermostat. In an attempt to study the effects of LP directly on brain tissue, purified human LP was incubated with rabbit brain slices in vitro. Because of the well-documented role of prostaglandin (PG) synthesis in both the production of fever and antipyresis, PGE levels were measured on the supernates of brain slices incubated 30 min with LP. Levels of PGE increased 3- to 4-fold in rabbit anterior and posterior hypothalami. In addition, PGE levels were similarly increased in temporal cortex slices when exposed to LP. In another set of experiments, PGE levels increased 4- to 5-fold when brain tissue was incubated with a highly purified preparation of bacterial endotoxin (ET). The ability of ET to increase brain PGE levels was not affected by moderate heating (56 degrees C, 30 min), whereas this temperature destroyed the PGE-inducing properties of LP. The antipyretic ibuprofen markedly reduced the amount of PGE measured in the brain slice supernates after stimulation with LP, suggesting that LP brings about synthesis of PGE and not the release of preformed PG. The results demonstrate that LP is a potent inducer of PGE synthesis in rabbit brain and that receptors for LP are not restricted to the thermoregulatory center, but rather may be distributed throughout the brain.

Human monocyte-derived soluble product(s) has an accessory function in the generation of histamine- and concanavalin A-induced suppressor T cells.

Abstract: We have analyzed the cellular interactions required for the generation of histamine- and concanavalin A (Con A)-induced suppressor T cells by employing a co-culture assay and techniques for fractionation of human blood mononuclear cells (PBMC). PBMC cultured in the presence of histamine (0.1 mM-1 mM) or Con A (20 micrograms/ml) for 24 h, mitomycin treated and subsequently combined with autologous mitogen-stimulated mononuclear cells, significantly suppressed a subsequent blastogenic response. PBMC fractionated over nylon wool columns and depleted of adherent cells and enriched for T cells (NWNA-T) were unable to generate suppressor activity. However, suppressor cell function by NWNA-T cells was reconstituted by the addition of autologous monocytes. In both the histamine and ConA suppressor systems, the requirement for monocytes in the activation process was enhanced by suspending the NWNA-T population in supernatants derived from allogeneic monocytes stimulated with heat-killed Staphylococcus albus. These crude supernatants contained leukocytic pyrogen (LP) and lymphocyte activating factor (LAF). Sequential purification and separation of the crude supernatants using gel-filtration, immunoadsorption, and isoelectric focusing demonstrated that only those fractions containing LP and LAF were capable to reconstituting NWNA-T cell histamine and Con A-induced suppressor activity. Thus, these studies suggest that the accessory role of supernatants derived from activated monocytes in the generation of suppressor cells may be mediated by LP/LAF. Further studies are in progress to explore the mechanism by which soluble factors stimulate suppressor T cells.

Studies on the active site of human leukocytic pyrogen

Abstract: Leukocytic pyrogen, a polypeptide produced by phagocytic mononuclear cells, is thought to be the endogenous mediator of fever. In addition to its effects on thermoregulation, leukocytic pyrogen has been shown to induce synthesis of acute-phase proteins, increase lymphocyte blastogenesis to mitogens, and cause release of neutrophil-specific granule contents. Despite its important role in biologic responses, little is known concerning the structure-function relationship of the molecule. In the present studies several protein-modifying conditions were used in order to examine specific amino acid participation at the active site. Because the state of purity of leukocytic pyrogen may be critical during certain reaction conditions, highly purified preparations were used. Experiments suggest that the active site requires the gamma-carboxyl group of glutamic acid and that blocking arginine reduces both the pyrogenic and neutrophil releasing properties of the molecule. Other studies demonstrate that the pyrogenicity of human leukocytic pyrogen is not due to serine esterase or carboxypeptidase B activity and that the 15,000-dalton molecule may be a glycoprotein. These experiments provide further evidence that the lymphocyte-activating and neutrophil-granule-releasing properties of human leukocytic pyrogen require the same active site which produces fever.

Production of fever and its effects on the host.

Abstract: Leukocytic pyrogen (LP) is a polypeptide that is released from phagocytic leukocytes and mediates fever, by direct action on the thermoregulatory centers in the hypothalamus. During the mediation a complex series of biochemical events occurs in the hypothalamus, including increases in prostaglandin synthesis. The production of LP by phagocytic leukocytes (predominantly monocyte/macrophage) requires synthesis of protein and new messenger-RNA. Recently, it has been discovered that LP has other direct effects on the host. Amongst these effects are: increases in acute phase reactants, including serum amyloid A protein (SAA); release of specific granule contents from neutrophils in vitro; a similarity (if not identity) to human lymphocyte activating factor (LAF) which is a monokine. Thus, LP may have a much broader role to play in the inflammatory response than just mediating fever.

Theories of Fever From Antiquity to the Enlightenment

Abstract: Despite the fact that the availability, acceptance, and use of clinical thermometers in medical practice is relatively recent medical history (Wunderlich's classic paper on clinical thermometry and fever was published in 1842), there is a wealth of treatises, books, pamphlets, and illustrations on the subject of fever as a general sign of disease as well as "fevers" as a diagnostic pool of the prevailing infectious diseases in the world. Medical historians considering various theories on fever during these 2,000 years also encounter a history of scientific philosophy. Theories on Fever From Antiquity to the Enlightenment is a compilation of seven essays that were delivered at a meeting of the Wellcome Institute for the History of Medicine in June 1980. The seven articles are each expertly researched and clearly presented in the context of the specific theories of fevers and, in addition, the social, religious, and political effects such theories had