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1.
Adenosine 3',5' cyclic monophosphate in Euglena gracilis   总被引:2,自引:0,他引:2  
Euglena gracilis contains in high concentration the enzymes for the synthesis and degradation of cyclic AMP. The synthetic enzyme, adenyl cyclase is mainly associated with a particulate fraction which sediments at 7,000–30,000xg whereas the degradative enzyme, 3′5′ nucleotide phosphodiesterase, is soluble (does not sediment at 78,000xg). The adenyl cyclase activity is stimulated somewhat by prostaglandins and by catecholamines, agents which markedly stimulate cyclase in appropriate mammalian tissues. There is no detectable activity of guanyl cyclase, the enzyme which synthesizes cyclic GMP. Euglena also contains a cyclic AMP stimulated protein kinase which is associated with a particulate fraction sedimenting at 30,000xg.  相似文献   

2.
Cyclic nucleotide metabolism and reactive oxygen production by macrophages   总被引:2,自引:0,他引:2  
The production of reactive oxygen species by elicited rat peritoneal macrophages was assessed by in vitro measurement of chemiluminescence in the presence of luminol. The divalent ion ionophore A23187 stimulated the production of reactive oxygen species. This action was inhibited by monobutyryl and dibutyryl derivatives of cyclic AMP but was not affected by derivates of cyclic GMP. Cyclic AMP and cyclic GMP concentrations increased rapidly in macrophages exposed to A23187 or zymosan. Indomethacin (20 μmol/1) inhibited the increase in cyclic AMP concentration but not the increase in cyclic GMP concentration. Neither A23187 nor zymosan stimulated adenylate cyclase activity in broken cell preparations of macrophages. The observations are consistent with the hypothesis that PGE produced by macrophages after phagocytotic stimuli may inhibit certain macrophage functions and perform a regulatory role in these cells. This action of PGE may be mediated by cyclic AMP.  相似文献   

3.
Lipopolysaccharide from E. Coli (LPS) and BCG cell walls (BCGcw) are recognized immunoadjuvants that directly stimulate some macrophage functions. The macrophage cell line J774.1 and peritoneal exudate cells (PEC) from mice can be stimulated by LPS or other adjuvants in vitro to synthesize and release protein factor(s) that activate thymus-derived lymphocytes. We have utilized J774.1 cells and PEC to demonstrate that an increase in ornithine decarboxylase (ODC) activity is a marker of early biochemical changes in adjuvant-stimulated macrophages. BCGcw and LPS increased ODC within 2 hours in J774.1 cells as well as murine peritoneal exudate macrophages. Maximal increases in ODC were detected 4 hours after the addition of adjuvants to J774.1 cells. The marked increases (12–23 fold) in ODC observed with BCGcw (20 μg/ml) did not appear to involve an effect on cell proliferation which was suppressed by this adjuvant. Cycloheximide inhibited the induction of ODC by LPS and BCGcw in the macrophage cell line. Evidence that the induction of ODC may be promoted by an increase in cyclic AMP was provided by experiments demonstrating that prostaglandin E1 (PGE1) and 8-bromo-adenosine-3′:5′-monophosphate (8Br-cyclic AMP) can mimic the effects of LPS and BCGcw in J774.1 cells. These observations indicate that one of the early biochemical changes in macrophages promoted by adjuvants is an induction of ODC.  相似文献   

4.
The injection of β-ecdysone into chilled Hyalophora gloveri pupae resulted in the stimulation of adenyl cyclase activity in the wing epidermis as measured by the incorporation of label into cyclic AMP from a prelabeled endogenous pool. Stimulation was also obtained in pupal wings in vitro and in wing epidermal homogenates. Although the sequence of responses to β-ecdysone in vitro depended on the composition of the incubation medium, the stimulation of cyclic AMP synthesis always preceded increases in the rates of RNA and protein synthesis. The increase in adenyl cyclase activity is the earliest metabolic event thus far discerned as a result of β-ecdysone action. It is suggested that β-ecdysone stimulates adenyl cyclase (and guanyl cyclase) but that the hormone also exerts effects on target cells independent of the cyclic AMP system.  相似文献   

5.
Peritoneal macrophages from Schistosoma mansoni-infected mice are activated both for nonspecific tumor cytotoxicity and for killing of skin-stage schistosomula in vitro. In the current study, mechanisms for induction of macrophage tumoricidal and schistosomulacidal activity have been compared. Examination of macrophages activated in vivo by BCG infection or C. parvum treatment, or in vitro by exposure to lymphokine prepared from antigen-stimulated BCG-immune spleen cells, showed that these effector functions were closely linked. Indeed, fractionation of lymphokine-rich supernatant fluids by Sephadex G-100 gel filtraction showed that activities responsible for induction of schistomula killing by inflammatory macrophages and for induction of tumoricidal activity cochromatographed as a single peak in the 50,000 MW region. Thus, development of macrophage-mediated cytotoxicity against these two extracellular (tumor cell or helminth) targets was coincident in several cell populations activated in vivo or in vitro. However, activation for tumoricidal and schistosomulacidal capacity appeared to be quantitatively dissociated in macrophages from mice with chronic schistosomiasis; those cells demonstrated low, yet significant, levels of larval killing (13 to 15 those of BCG or lymphokine-activated cells) but maximal levels of tumor cell cytotoxicity. Furthermore, cytotoxicity by peritoneal cells from S. mansoni-infected mice was not increased in vitro by exposure to lymphokine. Identification of this functional alteration in S. mansoni-activated cells may help to clarify the role of macrophages in the partial immunity against challenge infection which is demonstrated by mice with chronic primary S. mansoni infection.  相似文献   

6.
To determine whether macrophages can discriminate in an immunologically specific manner between the intracellular pathogens which they inhibit or kill, unelicited peritoneal macrophages from mice infected with either of two related but antigenically dissimilar protozoa were challenged with these protozoa in vitro. Experimental conditions were varied in an attempt to establish a state in vivo in which macrophage specificity might be demonstrated. No differences could be discerned between the ability of macrophages from three different strains of mice infected with the protozoa to kill Besnoitia and Toxoplasma. The effect of macrophages on Toxoplasma as compared with Besnoitia did not evolve or vary during development, expression, or decline of an immune response, i.e., with varying times after infection of mice as well as with varying times after treatment of mice with irradiated Toxoplasma. The route of infection could not be shown to confer specificity on macrophages, as subcutaneous and intraperitoneal inoculation of Toxoplasma did not lead to differential ability of macrophages to inhibit or kill the protozoa. The different strains of protozoa used for infection of mice did not affect the ability of peritoneal macrophages from Besnoitia- and Toxoplasma-infected mice to inhibit multiplication of or kill Besnoitia and Toxoplasma comparably in vitro. Peritoneal macrophages of mice treated with Corynebacterium parvum kill both organisms efficiently. These macrophages were employed to determine whether stimulation of macrophages by treatment of mice with a substance unrelated to the protozoa would produce activated macrophages. Uninfected mice and mice infected with either Besnoitia or Toxoplasma were challenged with varying doses of the protozoa in parallel with examination of macrophages from the same groups of mice in vitro to determine whether the presence of stimulated macrophages in the peritoneal cavity was necessary for protection against Toxoplasma and Besnoitia, and if so if their presence was sufficient for protection. Only mice with activated peritoneal macrophages were protected. However, protection was greater when the primary infection was with the same organisms used for challenge at a time when macrophages inhibited or killed both protozoa efficiently in vitro. The possible role of other effector cells, subpopulations of macrophages of different functional abilities in various sites, and antibody or other lymphocyte products acting in concert with macrophages as factors which may explain the differences observed between in vivo protection and in vitro capacity to inhibit or kill the protozoa are discussed.  相似文献   

7.
Guanylate cyclase has been purified 60-fold from cell extracts of the bacterium Caulobacter crescentus. It has a molecular weight of approximately 140,000 and is dependent upon Mn2+ for activity. Enzymic activity is unaffected by cyclic AMP, cyclic GMP or N6,O2′-dibutyryl cyclic AMP but is stimulated by N2,O2′-dibutyryl cyclic GMP. The partially purified preparation of guanylate cyclase does not contain detectable adenylate cyclase activity.  相似文献   

8.
The natural affinity of various bacterial glycopeptides and lipopolysaccharides for mammalian cell membranes was estimated quantitatively by comparison with the adsorption of lipopolysaccharide from Escherichia coli NCTC 8623 to erythrocytes, thymocytes, bone marrow cells, spleen cells, peritoneal lymphocytes and macrophages. Immunopotentiating activity was estimated by measuring the ability of the bacterial fractions to stimulate a humoral response to ovalbumin in HAM/1CR mice. When the affinity for mammalian cell membranes was compared with the stimulation of the antibody response, it was found that a negative correlation for peritoneal macrophages (rs=?0.94, P<0.0005) and a positive correlation for peritoneal lymphocytes (rs=+0.97,P<0.0005) and spleen cells (rs=+0.76,P<0.005) existed.  相似文献   

9.
The adenyl cyclase of the oxyntic, or acid-secreting, cells of bullfrog gastric mucosa has been found to be a membrane-bound enzyme. A method has been developed to isolate the adenyl cyclase rich membrane fractions in a hypotonic medium containing dithiothreitol, which has been found to protect the hormonal resposivenes of the adenyl cyclase.Highest specific activity of adenyl cyclase was localized in a light membrane fraction which also had abundant K+-stimulated ATPase and K+-stimulated p-nitrophenyl phophatase and very low cytochrome c oxidase activty. The three gastric secretagogues tested, namely histamine, pentagastrin and methylcholine, significantly stimulated the adenyl cyclase activity of the light membrane fraction.After treatment with 10 mM Mg+ further subfractionation of the light membrane fraction on a sucrose density gradient yielded light membrane subfraction 1, light membrane subfraction 2 and light membrane subfraction 3 in order of increasing densities. The three subfractions had different enzymatic and chemical properties. Adenyl cyclase activity has been found to be distributed in all three subfractions. However, the hormonal responsiveness of the three fractions was quite different. Light membrane subfraction 2 could be stimulated by all three secretagogues, light membrane subfraction 1 by histamine and methylcholine, while light membrane subfraction 3 was refractory to all three secretagogues. On the basis of the cholesterol to phospholipid molar ratio, RNA content, glycoprotein content and the enzymatic data it is suggested that light membrane subfraction 1 and light membrane subfraction 2 are of general plasma-membrane type, while light membrane subfraction 3 is largely of cytoplasmic origin.  相似文献   

10.
CYCLIC adenosine 3′,5′-monophosphate (cyclic AMP) has been suggested to be the receptor site for neurotransmitters as well as a second messenger which mediates the action of a variety of hormones and neurohormones in animals and human tissues. Cyclic AMP is derived from adenosine triphosphate (ATP) by the activity of adenyl cyclase and it is metabolized into adenosine monophosphate (AMP) by phosphodiesterase. Beta receptor and adenyl cyclase may be the same in the peripheral tissues1. In vitro activation of adenyl cyclase activity by various putative neurotransmitters such as noradrenaline (NA), 5-hydroxytryptamine (5-HT), acetylcholine (ACh) and histamine in preparations from brain homogenates and slices is well established2. The in vivo effect, however, of these neurohormones on adenyl cyclase has not been studied.  相似文献   

11.
New Zealand Black (NZB) and NZB by New Zealand White (NZW) F1 hybrid (BW) mice develop clinical signs of autoimmune disease between 6 and 10 months of age but spleen cells from these strains have a greatly reduced in vitro response to sheep erythrocytes (SRBC) as early as 5–6 weeks of age. This hyporesponsiveness can be only partially restored with 2-mercaptoethanol, allogeneic macrophages or spleen cells, or allogeneic factor. The response of NZB and BW spleen cells to the thymic independent antigen DNP-Ficoll is nearly normal. The reduced in vitro SRBC response was found to be attributable to splenic T and B cells rather than macrophages. Macrophages from NZB mice were found to function normally. The in vitro behavior of NZB lymphocytes is very similar to non-autoimmune mice infected with common murine viral pathogens. NZB and BW mice may be making an active immune response as early as 5 weeks of age.  相似文献   

12.
Homogenates of adult Schistosoma mansoni (blood flukes), isolated from the porto-mesenteric veins of infected mice, contain substantial activities of adenylyl cyclase, cyclic AMP phosphodiesterase, and a cyclic AMP stimulated protein kinase. The adenylyl cyclase, which is largely sedimentable at 10,000xg, is stimulated 20-fold by 10mM sodium fluoride and 1.4 to 2-fold by serotonin, glucagon, prostaglandins E1, E2 or B1. The phosphodiesterase, which is largely sedimentable at 10,000xg, is inhibited by both aminophylline and papaverine but is not influenced by 10mM sodium fluoride. The protein kinase, which is present in the 10,000xg supernatant is stimulated 4 to 8-fold by either cyclic AMP or cyclic GMP. There is a preference for cyclic AMP (K12 = 1.1×10?7M) over cyclic GMP (K12 = 4.5×10?6M). If intact worms are incubated in a glucose free medium there is a mobilization of glycogen stores which is preceded by a rise in cyclic AMP concentration. In a medium with 5mM glucose there is neither a rise in cyclic AMP nor mobilization of glycogen.  相似文献   

13.
Treatment of Rana catesbeiana tail fin tissue in vitro with 0.1 mM or 0.5 mM cyclic AMP or with triiodothyronine induces an increase in the specific activity of hexosaminidase, a lysosomal marker enzyme, and a decrease in tissue area. Lithium chloride (8 mM), an inhibitor of adenylate cyclase, inhibits these changes when initiated by triiodothyronine but not when initiated by cyclic AMP. The levels of cyclic AMP, determined by radioimmunoassay techniques, increased 110 ± 10% over matched discs in culture after only one day's exposure to triiodothyronine. These results indicate the effect of triiodothyronine on fin resorption may be mediated by cyclic AMP.  相似文献   

14.
Butaclamol, a new neuroleptic agent, and its (+)- enantiomer caused a pronounced dose-related elevation of rat striatal homovanillic acid concentration invivo. In addition, each blocked the dopamine-induced increase in adenyl cyclase activity of homogenates of the olfactory tubercle, a limbic area in the brain. The (-)-enantiomer of butaclamol did not exhibit these activities indicating a stereochemical specificity for dopamine receptor-blockade activity. The (+)-enantiomer was 2–3 times more potent than butaclamol, exhibiting activities similar to those of fluphenazine. The present findings are consistent with the existence of relationships between changes in dopamine turnover in the striatum and the production of extrapyramidal side effects and between changes in adenyl cyclase activity of olfactory tubercle and antipsychotic activity.  相似文献   

15.
A cyclic AMP dependent protein kinase in Dictyostelium discoideum   总被引:4,自引:0,他引:4  
A cyclic AMP-dependent protein kinase was found to appear during the time course of development of Dictyosteliumdiscoideum. No cyclic AMP dependency was observed at any stage of development in crude 110,000 X G soluble extracts. After partial purification, however, extracts from post-aggregation stages contained enzyme that was activated up to 6-fold by cyclic AMP, whereas protein kinase from earlier stages was not affected by cyclic AMP. Likewise, cyclic AMP binding activity increased from the aggregation to the slug stage of development. Approximately one-half of the total cyclic AMP binding activity co-purified with the cyclic AMP dependent protein kinase. The enzyme from Dictyostelium showed similarities to mammalian protein kinases with respect to its kinetic properties but differed in its behavior on ion-exchange chromatography.  相似文献   

16.
Cyclic AMP was not found in vegetative cells or sporulating cells or dormant spores of Bacillusmegaterium using an assay which would have detected an invivo concentration of 1 – 2 × 10?9 M. Adenyl cyclase and cyclic AMP phosphodiesterase were also not detected in sonicates of vegetative or sporulating B.megaterium cells.  相似文献   

17.
The effect of morphine sulfate (MS) on adenylate cyclase (AC) and phosphodiesterase (PDE) activities in the rat striatum was investigated. MS produced a dose-dependent increase in basal AC activity and did not alter sodium fluoride-induced stimulation both invivo (7.5–30 mg/kg, 1 hr pretreatment, i.p.) and invitro (1–100μM). invitro, when submaximal effective concentrations of dopamine and MS were combined, there was an additive effect. However, administration of MS invivo did not alter dopamine-induced stimulation of AC activity. MS, invitro and invivo inhibited PDE activity in a dose-dependent manner only with the high substrate concentration (3.3 × 10−3M cyclic AMP). Preliminary results from this study indicate that morphine affects the cyclic AMP system.  相似文献   

18.
Adenylate cyclase activity and 3′, 5′ cyclic adenosinemonophosphate (cAMP) have been followed through the heat-synchronized cell cycle of Tetrahymena pyriformis. While the specific activity of adenylate cyclase remained essentially constant throughout the cycle, cAMP oscillated (between 10 and 50 pmoles/mg protein) through two cycles. Minima were observed at each division (DS border) and maxima at each SG2 border. Each heat shock caused slight temporary reduction in cyclase activity. Further observations suggest to us that adenylate cyclase shows conformational changes in response to temperature-induced alterations and to changes in lipid composition of membranes.  相似文献   

19.
The effect of dopamine on the cyclic AMP concentration in the rat spleen lymphocytes has been investigated invitro. It has been shown that dopamine in concentration above 10?6M induces a significant increase of cyclic AMP level. The maximal stimulatory effect was observed after 10 minutes of the lymphocytes incubation with dopamine. These data suggest that the dopamine receptor in lymphocyte belongs to D-1 category.  相似文献   

20.
The effects of a monoclonal antibody directed against immune response gene products on mouse NK activity were examined. In vivo administration of an anti-I-Ak antibody to C3H/He (H-2k) mice modulated their peritoneal cell (PC) and spleen cell (SC) natural killer (NK) activity against YAC-1 lymphoma target cells in vitro. No such effect was observed when BALB/c (H-2d) mice were treated with this antibody. Administration of anti-I-Ak antibody to mice before and after infection with Toxoplasma or treatment with poly(I:C) leads to suppression of NK activity in comparison to NK activity of mice infected with Toxoplasma or injected with poly(I:C) alone. A similar treatment regimen with M5/114 antibody which reacts with I-Ab, I-Ad, I-Ed, and I-Ek molecules resulted in decreased NK activity in B10.D2 (H-2d) but not in B10.BR (H-2k) mice. Serum and cell culture supernatant interferon (IFN) concentrations were not altered as a result of anti-I-Ak treatment. Removal of adherent cells did not restore NK activity of anti-I-Ak-treated Toxoplasma-infected mice to levels obtained with mice infected with Toxoplasma. In contrast, depletion of Ly 2.1+ cells from nylon-wool nonadherent SC of mice treated with anti-I-Ak antibody, before and after infection with Toxoplasma, resulted in restoration of NK activity to the same level as that observed in Toxoptasma-infected mice.  相似文献   

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