Potentiation of adenylyl cyclase in human peripheral blood mononuclear cells by cell-activating stimuli.

The isoprenaline-induced production of cAMP in human peripheral blood mononuclear cells (PBMC) was potentiated significantly by incubating PBMC with isoprenaline in the presence of phytohaemagglutinin (PHA), Concanavalin A (Con A) or A23187. This potentiation, that proved to be dependent on the concentration of PHA, Con A or A23187, increased the maximal response but did not cause a change in the potency of isoprenaline. Potentiation could not be induced by the phorbol ester phorbol-myristate acetate, suggesting that protein kinase C-dependent pathways are not likely to be involved in potentiation of adenylyl cyclase. Potentiation could be inhibited by chelating extracellular Ca2+ with EGTA and also by N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamine, an inhibitor of calmodulin. Potentiation could not be induced by preincubation of PBMC with PHA, suggesting that transient biochemical changes are involved. It was concluded from these results that potentiation in PBMC probably involves the activation of Ca2+/calmodulin-dependent adenylyl cyclase subtypes. Potentiation of the adenylyl cyclase activity could be an important physiological mechanism in vivo preventing cells from becoming "over stimulated".

Differences in innervation and secretory behaviour of two nasal glandular parts in the rat.

The submucosal glands in the rat nose are characterized by the presence of both neutral and acid glycoproteins, which are important constituents of nasal and tracheobronchial secretions. This study is an attempt to gain a better insight into the secretion of these two types of glycoproteins and its neural regulation. Radiobiochemical experiments show a higher sensitivity to methacholine of the nasal glandular region producing acid glycoproteins than the area secreting neutral glycoproteins. Radioligand receptor binding suggests that the binding parameters of the muscarinic receptors in these two areas are mutually different. Furthermore, rat nasal glandular muscarinic receptors appear to be different from those in smooth muscle of rat ileum.

Desensitization of the adenylyl cyclase system in peripheral blood mononuclear cells from patients with asthma three hours after allergen challenge.

BACKGROUND: Bronchial hyperreactivity is a common characteristic of patients with asthma and is often associated with enhanced activities of peripheral blood cells. Signal transduction systems are important in regulating cellular activities and can be modified by allergen challenge. METHODS: Patients with allergic asthma (n = 15) were challenged with house dust mite allergen, resulting in an asthmatic response. Adenylyl cyclase activity was measured in membranes from peripheral blood mononuclear cells before, 3 hours after, and 24 hours after challenge. RESULTS: Allergen challenge proved to have opposite effects in two distinct subgroups of patients. In 10 patients (group I) a heterologous desensitization of the adenylyl cyclase system was observed after challenge, whereas in five patients (group II) an increase in adenylyl cyclase activity was found. Adenylyl cyclase activity before allergen challenge in group II was significantly lower than in group I and comparable to cyclase activity found in group I after allergen challenge. This suggests that in these five patients the adenylyl cyclase system was already desensitized before the start of the study, possibly as a result of natural allergen exposure. Heterologous desensitization in group I was found within 3 hours after allergen challenge, that is before the onset of the late bronchoconstrictive reaction. CONCLUSIONS: Because adenylyl cyclase is important in the regulation of cytokine production by mononuclear cells, alteration of cytokine production induced by desensitization of adenylyl cyclase could therefore play a role in the development of the late bronchoconstrictive reaction.

Defect in potentiation of adenylyl cyclase correlates with bronchial hyperreactivity.

BACKGROUND: Adenylyl cyclase is a transmembrane signaling system involved in the inhibition of cellular responses. Recently, we showed that the activity of adenylyl cyclase may be potentiated by stimuli that induce an increase of cellular responses but that do not activate adenylyl cyclase. This is probably an important physiologic feedback mechanism that prevents cells from becoming "overstimulated." OBJECTIVE: Because increased cellular activities are frequently observed in persons with asthma, we hypothesized that a defect in potentiation of adenylyl cyclase might be involved. METHODS: Potentiation of isoprenaline-induced adenosine cyclic monophosphate (cAMP) production with the mitogen phytohemagglutin (PHA; 45 micrograms/ml) or the calcium ionophore A23187 (1 mumol/L) was studied in peripheral blood mononuclear cells taken from patients with asthma (n = 8) and healthy control subjects (n = 11). RESULTS: Isoprenaline-induced cAMP production was potentiated significantly in the healthy control subjects (PHA, 110% +/- 15%; A23187, 92% +/- 25%). In contrast, potentiation was not seen with PHA or A23187 in the total group of patients with asthma. However, some patients showed weak potentiation, whereas in others PHA decreased isoprenaline-induced cAMP production. Moreover, the effect of PHA on isoprenaline-induced cAMP production correlated significantly with the degree of bronchial hyperreactivity in patients with asthma (r = 0.96; p = 0.0001). CONCLUSIONS: The observed defect in signal transduction could play an important part in bronchial hyperresponsiveness.

Production of diacylglycerol and arachidonic acid in peripheral blood mononuclear cells from patients with asthma and healthy controls.

BACKGROUND: Enhanced activities of peripheral blood cells are a common characteristic of patients with asthma. OBJECTIVE: Here we tested whether this could be due to a dysfunction in one or more signal transduction systems. METHODS: The production of 1,2-diacylglycerol (1,2-DAG) and arachidonic acid was compared in mononuclear blood cells from patients with asthma (n = 10) and healthy controls (n = 12). RESULTS: Using three different stimuli (concanavalin A, aluminium fluoride or the calcium ionophore A23187) no difference in the production of both 1,2-DAG and arachidonic acid could be found between patients and controls before allergen challenge. Concanavalin A-induced 1,2-DAG production could be inhibited completely in the presence of isoprenaline; concanavalin A-induced arachidonic acid production, partially. The inhibitory effect of adenylate cyclase activation on the production of 1,2-DAG and arachidonic acid was identical in patients and controls. Following allergen challenge, there was a tendency to an increased production of 1,2-DAG and arachidonic acid in controls, whereas in patients there was a tendency to a decreased production. CONCLUSIONS: Enhanced cellular activities found in patients with asthma are not caused by an intrinsic dysfunction in production of 1,2-DAG and arachidonic acid.