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Biomedical subjects

N H Hunt

Publications and source records attributed to N H Hunt.

At least 91 records · Page 5Linked to original sources
PubMed1985

Reactive-oxygen formation and its relationship to prostaglandin and cyclic AMP production by zymosan-treated rat peritoneal macrophages.

Addition of zymosan (20 particles/cell) to suspensions of resident rat peritoneal macrophages caused an increase in the concns of prostaglandins and cyclic AMP. Preincubation of the cells with inhibitors of arachidonate metabolism led to inhibition of prostaglandin, but not of cyclic AMP, formation, which suggested that the two processes may occur independently of each other in phagocytosing cells. The luminol-dependent chemiluminescence associated with the addition of zymosan to the cells consisted of a minor, Ca2+-dependent, glucose-independent component and a major, glucose-dependent, Ca2+-independent component. Only the minor, Ca2+-dependent component appeared to be related to the lipoxygenation of arachidonic acid. Close examination of the production of prostaglandins and cyclic AMP and of chemiluminescence after zymosan addition, indicated that the expanded pool of endogenous cyclic AMP was probably not a negative modulator of the other two processes, although they remained susceptible to inhibition by exogenously-added cyclic AMP analogues or PGE2. The events induced by zymosan may be relevant to the physiological roles of prostaglandins during the inflammatory response.

Animals↗
PubMed1985

Oxidative stress and protective mechanisms in erythrocytes in relation to Plasmodium vinckei load.

The protection of mouse erythrocytes (RBC) parasitized with Plasmodium vinckei vinckei against activated oxygen species was examined in relation to the intraerythrocytic parasite load. RBC from highly infected animals were separated by density gradient centrifugation into six bands with increasing parasite content and with parasitemias ranging from 17% to 100%. Increase in parasite load was accompanied by a decrease in the activities of the enzymes superoxide dismutase (EC 1.15.1.1), catalase (EC 1.11.1.6), glutathione peroxidase (EC 1.11.1.9), glutathione reductase [NAD(P)H] (EC 1.6.4.2), and NADH-methemoglobin reductase (EC 1.6.2.2; NADH:ferricytochrome b5 oxidoreductase) in the RBC lysates. In contrast, the total amount of reduced glutathione increased in the highly parasitized bands. Furthermore, the vitamin E content of all RBC bands, including the one that contained mainly nonparasitized erythrocytes, was 3- to 5-fold higher than that of control noninfected RBC. Increasing parasite load was accompanied by an increase in the production of malonyldialdehyde, indicating enhanced lipid peroxidation. Our results indicate that oxidative stress is experienced by all RBC during a malarial infection and is accompanied by a variety of changes in the antioxidant defense mechanisms of the host and the parasite. Furthermore, it appears that the plasma membrane of the host cell is better protected against oxidative injury than are the membranes surrounding the parasite.

Animals↗
PubMed1984

Activation of cyclic AMP-dependent protein kinase in macrophages.

The activity of cyclic AMP-dependent protein kinase (cyclic AMP-PK) was significantly higher (P less than 0.001) in thioglycollate-elicited than in resident rat peritoneal macrophages. The activity ratio of the enzyme (its activity in the absence of added cyclic AMP divided by that in the presence of 5 microM cyclic AMP) was similar in the two cell types. The divalent ion ionophore A23187 induced a rapid increase in the activity ratio of cyclic AMP-PK in both macrophage types. This effect was blocked by pretreating the cells with indomethacin or aspirin (inhibitors of cyclo-oxygenase) and bromo-phenacyl bromide (an inhibitor of phospholipase A2), implicating the synthesis of a prostanoid as an intermediary step. Prostaglandin (PG) E2, 8-bromo cyclic AMP and cholera toxin, all of which inhibit chemiluminescence and/or PG formation in macrophages, increased the activity ratio of cyclic AMP-PK in these cells. We propose that the activation of cyclic AMP-PK plays a central role in the response of macrophages to both endogenously-generated and exogenously added PGE.

Acetophenones↗
PubMed1984

Changes in hormone responsiveness and cyclic AMP metabolism in rat hepatocytes during primary culture and effects of supplementing the medium with insulin and dexamethasone.

Primary monolayer cultures of rat hepatocytes were used for studies of long-term and acute effects of hormones on the cyclic AMP system. When hepatocyte lysates were assayed at various times after plating of the cells three major changes in the metabolism of cyclic AMP and its regulation were observed: Glucagon-sensitive adenylate cyclase activity gradually declined in culture. In contrast, catecholamine-sensitive activity, being very low in normal adult male rat liver and freshly isolated hepatocytes, showed a strong and rapid increase after seeding of the cells. Concomitantly, there was an early elevation (peak approximately equal to 6 h) and a subsequent decrease in activity of both high-Km and low-Km cyclic AMP phosphodiesterase. These enzymic changes probably explained the finding that in intact cultured cells the cyclic AMP response to glucagon was diminished for 2-24 h after seeding, followed by an increase in the responsiveness to glucagon as well as to adrenergic agents up to 48 h of culture. Supplementation of the culture media with dexamethasone and/or insulin influenced the formation and breakdown of cyclic AMP in the hepatocytes. Insulin added at the time of plating moderately increased the adenylate cyclase activity assayed at 48 h, while dexamethasone had no significant effect. In the presence of dexamethasone, insulin exerted a stronger, and dose-dependent (1 pM - 1 microM), elevation of the adenylate cyclase activity in the lysates, particularly of the glucagon responsiveness. Thus, insulin plus dexamethasone counteracted the loss of glucagon-sensitive adenylate cyclase activity occurring in vitro. Kinetic plots of the cyclic AMP phosphodiesterase activity showed three affinity regions for the substrate. Of these, the two with high and intermediate substrate affinity (Km approximately equal to 1 and approximately equal to 10 microM) were decreased in the dexamethasone-treated cells. Insulin partly prevented this effect of dexamethasone. Accumulation of cyclic AMP in intact cells in response to glucagon or beta-adrenergic agents was strongly increased in cultures pretreated with dexamethasone. The results suggest that insulin and glucocorticoids modulate the effects of glucagon and epinephrine on hepatocytes by exerting long-term influences on the cyclic AMP system.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed1984

Activity of divicine in Plasmodium vinckei-infected mice has implications for treatment of favism and epidemiology of G-6-PD deficiency.

Intravenous injection of divicine into mice infected with Plasmodium vinckei rapidly killed the parasites and caused haemolysis. Degenerating parasites were observed frequently inside intact circulating erythrocytes, implying that parasite death was not a passive consequence of haemolysis. Both parasite death and haemolysis were prevented by the iron chelator desferrioxamine. In vitro, divicine caused the accumulation of malonyldialdehyde and the depletion of reduced glutathione in normal mouse erythrocytes. Desferrioxamine inhibited the former event, but not the latter. These observations support the hypothesis advanced by Huheey & Martin (Experientia, 31, 1145, 1975) to explain the patchy geographical distribution of glucose-6-phosphate dehydrogenase deficiency in historic malarial areas and also suggest that desferrioxamine, a drug already in clinical use, is a potential treatment for favism and other examples of oxidative haemolysis.

Animals↗
PubMed1984

Production of luminol-reactive oxygen radicals during Plasmodium vinckei infection.

We tested the ability of whole blood and enriched fractions of peripheral blood polymorphonuclear leukocytes obtained from mice during the course of infection with Plasmodium vinckei to produce luminol-mediated chemiluminescence in response to phagocytic and nonphagocytic stimuli. The chemiluminescence response of whole blood to all stimuli increased dramatically and nonlinearly as the infection progressed, and there was a concomitant increase (80%) and decrease (70%) in the total numbers of leukocytes and erythrocytes, respectively. The proportion of polymorphonuclear leukocytes in the total leukocyte population increased threefold. On a per cell basis and at a constant hematocrit, the chemiluminescence response of peripheral leukocytes from infected animals to phorbol myristate acetate or opsonized zymosan was only slightly greater than that of cells from uninfected animals. Polymorphonuclear leukocytes isolated from the blood of infected animals also showed no large increase per cell in chemiluminescence responsiveness. Thus, although leukocyte numbers increase during a murine malarial infection, there appears to be no major change in the capacity of individual peripheral blood leukocytes to produce activated species of oxygen. However, the physiological reduction in the total concentration of hemoglobin at high parasitemia, due to hemolysis and hemoglobin digestion by the parasites, increases the possibility of oxygen radical-mediated damage to tissues and intraerythrocytic parasites as a result of decreased antioxidant protection.

Animals↗
PubMed1984

Radical-mediated damage to parasites and erythrocytes in Plasmodium vinckei infected mice after injection of t-butyl hydroperoxide.

Intravenous injection of t-butyl hydroperoxide rapidly killed Plasmodium vinckei in mice, and caused haemolysis. The same dose seemed harmless to unparasitized mice. Many parasites disintegrated inside circulating erythrocytes, so parasite death was not simply a passive consequence of haemolysis. Injection of desferrioxamine, which removes the traces of free iron that promote the dissociation of t-butyl hydroperoxide into radical species, prevented both parasite death and haemolysis. Lipid peroxidation, as measured by accumulation of malonyldialdehyde over 2 h in vitro, occurred in erythrocytes exposed to t-butyl hydroperoxide, and was particularly marked in erythrocytes from parasitized mice. These erythrocytes accumulated appreciable malonyldialdehyde even without exposure to t-butyl hydroperoxide. Desferrioxamine inhibited the accumulation of malonyldialdehyde, but did not prevent depletion of reduced glutathione by t-butyl hydroperoxide. This suggests that t-butyl hydroperoxide damaged parasites and erythrocytes by dissociating into radical species, rather than by decreasing intraerythrocyte anti-oxidant capacity. In earlier experiments we suggested that intraerythrocytic parasite death and haemolysis caused by alloxan were mediated by radical species, and these experiments with t-butyl hydroperoxide add weight to this interpretation. We regard both of these systems as models for macrophage-induced parasite death and host pathology in acute malaria.

Animals↗
PubMed1983

Reactive oxygen production, arachidonate metabolism and cyclic AMP in macrophages.

Chemiluminescence was used as an indicator of the production of reactive oxygen species by thioglycollate-elicited rat peritoneal macrophages stimulated by A23187. This action of the ionophore was inhibited by bromophenacyl bromide and nordihydroguaiaretic acid, inhibitors of the phospholipase A2 and lipoxygenase enzymes, respectively. The cyclo-oxygenase inhibitors, indomethacin and aspirin, did not diminish the light output. Preincubation of the cells with the 8-bromo- or dibutyryl analogues of cyclic AMP or with the cyclic AMP-phosphodiesterase inhibitors theophylline and RO-20-1724, or with PGE2, inhibited the A23187-evoked chemiluminescence. The results suggest that he lipoxygenase pathway of arachidonic acid metabolism may make a significant contribution to reactive oxygen production. This process may be modulated, and its duration limited, by cyclic AMP-mediated actions of prostaglandins, which are products of the cyclo-oxygenation of arachidonate.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1983

Cyclic AMP and the regulation of prostaglandin production by macrophages.

The ionophore A23187 (6.7 microM) increased the rates of formation of prostaglandins and cyclic AMP in suspensions of thioglycollate-elicited rat peritoneal macrophages. Both effects were inhibited by the calmodulin blocker trifluoperazine (50 microM) and the calcium channel blocker verapamil (500 microM). Inhibitors of phospholipase A2 and cyclo-oxygenase also blocked both actions of A23187. The stimulated prostaglandin formation was markedly reduced when the cells were preincubated with 8-bromo-cyclic AMP (1mM), dibutyryl cyclic AMP (1mM) or cholera toxin (500ng/ml). Addition of exogenous arachidonic acid (30 microM) alleviated this inhibition. We propose that the effect of A23187 on macrophages includes a 'self-limiting' mechanism whereby newly-synthesized prostaglandins can inhibit, via cyclic AMP, a step(s) prior to the transformation of arachidonic acid and thus modulate their own production.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1983

Glycogen synthesis in resident and thioglycollate-elicited rat peritoneal exudate cells.

The values of the A0.5 for glucose-6-P, apparent Km for UDPglucose and -/+glucose-6-P activity ratio are similar for glycogen synthase derived from rat resident and thioglycollate-elicited peritoneal macrophages; the specific activity is 7-fold higher for the enzyme from thioglycollate-elicited macrophages. The rate of incorporation of [14C]glucose into macrophage glycogen is 7-fold greater in the elicited population that that in the resident one; the values of the S0.5 for glucose are similar. The in vitro activation of glycogen synthase proceeds at a greater rate and extent for the enzyme from elicited macrophages; thus, phosphatase activity may be reduced in resident macrophages relative to that in thioglycollate-elicited ones.

Animals↗
PubMed1983

Evidence for reactive oxygen intermediates causing hemolysis and parasite death in malaria.

A rapid reduction in parasitemia associated with damage to intraerythrocytic parasites was observed in Plasmodium vinckei-infected mice after they had received a single intravenous injection of alloxan. This was not prevented by prior injection of glucose, but was prevented by desferrioxamine or diethyldithiocarbamate. Prior injection of propanol partially blocked the phenomenon. A transient hemolysis was observed in malaria-infected mice, but not in controls, after injection of alloxan. This was also blocked by desferrioxamine, but not by glucose. Both the fall in parasitemia and hemolysis occurred, but less dramatically, when phenylhydrazine or hydrogen peroxide was injected into parasitized mice. Again, the hemolysis was blocked by desferrioxamine. These observations are consistent with the parasite death and hemolysis being mediated by reactive oxygen species, possibly hydroxyl radicals, and have implications for our understanding of hemolysis, endothelial damage, and parasite suppression in acute malaria. Our evidence that malaria parasites are susceptible to free oxygen radicals supports the view that high intraerythrocytic oxidative stress may contribute to the high frequencies in malarial areas of genes for certain erythrocyte-related traits and suggests that some antimalarial drugs may suppress parasites partly through oxidative damage.

Alloxan↗
PubMed1980

RMI 12330A, an inhibitor of cyclic nucleotide phosphodiesterases and adenylate cyclase in kidney preparations.

N-(cis-2-phenylcyclopentyl)azacyclotridecan-2-imine hydrochloride (RMI 12330A) inhibited cyclic AMP and cyclic GMP phosphodiesterase activities in kidney preparations from rat and mouse. The drug was effective in the concentration range 0.1-1 mM. The agent was much less effective in inhibiting chick kidney cyclic nucleotide phosphodiesterases. The onset of inhibition of rat particulate cyclic AMP phosphodiesterase activities was rapid (less than 30 s) and irreversible. The inhibition of the low Km forms of cyclic AMP phosphodiesterase in mouse kidney homogenates was of the non-competitive type. RMI 12330A inhibited cyclic AMP phosphodiesterase activities in intact rat renal tubules. Adenylate cyclase activity, both basal and stimulated, was inhibited in all three species by the drug. Since RMI 12330A affects cyclic GMP metabolism as well as cyclic AMP metabolism, caution must be exercised in interpreting its effects upon cellular processes in terms of its actions upon the adenylate cyclase-cyclic AMP pathway alone.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed1980

Cyclic nucleotide excretion in human malignancies.

1. Urinary cyclic nucleotide excretion was studied in 51 patients with malignant tumours and in 24 control subjects. 2. Adenosine 3':5'-cyclic monophosphate excretion was normal in patients with tumours. 3. Urinary excretion of guanosine 3':5'-cyclic monophosphate was significantly greater than normal in patients with lymphoid tumours (both sexes) and in male patients with myeloid tumours or malignant melanoma. 4. Guanosine 3':5'-cyclic monophosphate excretion was normal in patients with breast carcinoma or colonic adenocarcinoma.

Adolescent↗