Parasite accessory cell interactions in murine leishmaniasis. I. Evasion and stimulus-dependent suppression of the macrophage interleukin 1 response by Leishmania donovani

Interleukin 1 (IL 1) is a principal mediator of the host immune response to microbial challenge. Accessory cells of the monocyte-macrophage series are a major source of this cytokine and are also chronically parasitized by protozoa of the genus Leishmania. This suggests that characterization of the macrophage IL 1 response to Leishmania would increase our understanding of the regulation of host immunity to these organisms. In the present study, the macrophage IL 1 response to Leishmania donovani was examined because infections with this organism have findings consistent with parasite-specific T cell unresponsiveness. Cytokine activity was measured either by direct stimulation or by co-stimulation of thymocytes. Conditioned media from BALB/c resident peritoneal macrophages infected with amastigotes of L. donovani contained no more IL 1 than did supernatant fluids of control cells. In contrast, supernatants from cells stimulated with lipopolysaccharide or heat-killed Listeria monocytogenes had significantly increased cytokine content. Resident cells infected with L. donovani for 4 hr before being stimulated with Listeria demonstrated a suppressed IL 1 response (approximately 40% of Listeria alone) to this secondary particulate stimulus. In contrast, the secondary response of leishmania-preinfected cells to lipopolysaccharide was not affected. To examine whether accessory cell nonresponsiveness to L. donovani (with respect to IL 1) was related to the state of macrophage activation, elicited peritoneal macrophages obtained by injection of proteose peptone were also studied. These cells responded to stimulation with lipopolysaccharide and fixed Staphylococcus aureus with increases in intracellular, membrane, and secreted cytokine activities. In contrast, L. donovani failed to induce any of these activities. This was found to be the case irrespective of whether amastigotes were alive or killed or opsonized with specific antibodies. Elicited cells preinfected with Leishmania responded normally to secondary stimulation with lipopolysaccharide, but not S. aureus (64% of Staphylococcus alone). In addition, attachment and penetration of L. donovani promastigotes and their subsequent conversion to amastigotes within macrophages failed to induce IL 1 synthesis. The findings of this study indicate that L. donovani has the ability to both evade and suppress the macrophage IL 1 response. Because this monokine provides an obligatory signal during macrophage-dependent T cell activation, evasion of signal transduction for IL 1 synthesis may be related to defects in cell-mediated immunity which occur during infections with this organism.     

Gabaergic Pharmacological Activity of Propofol Related Compounds as Possible Enhancers of General Anesthetics and Interaction with Membranes

Phenol compounds, such as propofol and thymol, have been shown to act on the GABA_A receptor through interaction with specific sites of this receptor. In addition, considering the high lipophilicity of phenols, it is possible that their pharmacological activity may also be the result of the interaction of phenol molecules with the surrounding lipid molecules, modulating the supramolecular organization of the receptor environment. Thus, in the present study, we study the pharmacological activity of some propofol- and thymol-related phenols on the native GABA_A receptor using primary cultures of cortical neurons and investigate the effects of these compounds on the micro viscosity of artificial membranes by means of fluorescence anisotropy. The phenol compounds analyzed in this article are carvacrol, chlorothymol, and eugenol. All compounds were able to enhance the binding of [³H]flunitrazepam with EC₅₀ values in the micromolar range and to increase the GABA-evoked Cl^? influx in a concentration-dependent manner, both effects being inhibited by the competitive GABA_A antagonist bicuculline. These results strongly suggest that the phenols studied are positive allosteric modulators of this receptor. Chlorothymol showed a bell-type effect, reducing its positive effect at concentrations >100 μM. The concentrations necessary to induce positive allosteric modulation of GABA_A receptor were not cytotoxic. Although all compounds were able to decrease the micro viscosity of artificial membranes, chlorothymol displayed a larger effect which could explain its effects on [³H]flunitrazepam binding and on cell viability at high concentrations. Finally, it is suggested that these compounds may exert depressant activity on the central nervous system and potentiate the effects of general anesthetics.     

11 beta-chloromethyl-[3H]estradiol-17 beta: a very high affinity, reversible ligand for the estrogen receptor

It has been suggested that binding of 11 beta-chloromethyl estradiol (11 beta-CME2) to the estrogen receptor is irreversible, since its complex with receptor fails to undergo exchange with estradiol (E2). To investigate this behavior directly, 11 beta-CME2 was prepared in high specific activity, tritium-labeled form: The binding of [3H]11 beta-CME2 to the estrogen receptor from lamb and rat uterus and MCF-7 human breast cancer cells was shown to be fully reversible; the 11 beta-CME2 complex with receptor, as well as that of a structural analog 11 beta-ethyl estradiol, however, do not dissociate or exchange with [3H]E2 over a 22 h period at 25 degrees C. By competitive or direct binding assays, the affinity of 11 beta-CME2 for the estrogen receptor can be estimated to be as much as 10- to 30-fold higher than that of E2. The complexes of estrogen receptor from MCF-7 cells with [3H]11 beta-CME2 and [3H]E2 show identical velocity sedimentation profiles on sucrose gradients, under conditions when the receptor is either a monomer of a dimer. Because of its very high affinity and unusual dissociation kinetics, [3H]11 beta-CME2 should be a very useful ligand for studies of estrogen receptor dynamics and in the assay of estrogen receptor concentrations in tumors and tissues.