Structural Requirements for Cub Domain Containing Protein 1 (CDCP1) and Src Dependent Cell Transformation

Cub domain containing protein 1 (CDCP1) is strongly expressed in tumors derived from lung, colon, ovary, or kidney. It is a membrane protein that is phosphorylated and then bound by Src family kinases. Although expression and phosphorylation of CDCP1 have been investigated in many tumor cell lines, the CDCP1 features responsible for transformation have not been fully evaluated. This is in part due to the lack of an experimental system in which cellular transformation depends on expression of exogenous CDCP1 and Src. Here we use retrovirus mediated co-overexpression of c-Src and CDCP1 to induce focus formation of NIH3T3 cells. Employing different mutants of CDCP1 we show that for a full transformation capacity, the intact amino- and carboxy-termini of CDCP1 are essential. Mutation of any of the core intracellular tyrosine residues (Y734, Y743, or Y762) abolished transformation, and mutation of a palmitoylation motif (C689,690G) strongly reduced it. Src kinase binding to CDCP1 was not required since Src with a defective SH2 domain generated even more CDCP1 dependent foci whereas Src myristoylation was necessary. Taken together, the focus formation assay allowed us to define structural requirements of CDCP1/Src dependent transformation and to characterize the interaction of CDCP1 and Src.     

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.     

Harn-steroidprofile hirsuter frauen

Urine steroid profiles of hirsute womenSteroid profiles of women suffering from idiopathic hirsutism show in more than 50% of the cases a 10–100 fold increase in the excretion of dehydroepiandrosterone (DHEA) compared with normal values.The excretion of DHEA was reduced much more than that of other 17-ketosteroids if the adrenals (NNR) were suppressed by dexamethasone (DXM). Within one week they reached values at the compound noise level of the gas chromatograms. If the ovaries were stimulated with human chorionic gonadotropin during continued suppression of the NNR with DXM no increase of DHEA could be detected.     

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.