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S MacNeil

Publications and source records attributed to S MacNeil.

At least 73 records · Page 4Linked to original sources
PubMed1988

Calmodulin antagonists of improved potency and specificity for use in the study of calmodulin biochemistry.

Syntheses are described for a range of N-(omega-aminoalkyl)-5-iodo- and -5-cyanonaphthalene-1-sulphonamides. The selective activity of these compounds as inhibitors for calmodulin-dependent phosphodiesterase (EC 3.1.4.17) is compared with their activity for the calmodulin-independent but calcium-dependent enzymes protein kinase C and transglutaminase (EC 2.3.2.13). The results show a drastic improvement in the selectivity of effect for the 5-iodo-compounds compared with the widely-used drug, W7, N-(6-aminohexyl)-5-chloronaphthalene-1-sulphonamide.

Calmodulin↗
PubMed1988

Calcium/calmodulin regulation of the proliferation of human epidermal keratinocytes, dermal fibroblasts and mouse B16 melanoma cells in culture.

We have investigated the relationship between extracellular calcium, intracellular calmodulin and proliferation in normal keratinocytes. Keratinocyte proliferation and its sensitivity to calmodulin antagonists was compared with that of normal human dermal fibroblasts and neoplastic mouse B16 melanoma cells. Keratinocytes were similar to fibroblasts in showing reduced proliferation in low (0.15 mM) calcium medium and unlike B16 cells which continued to proliferate until calcium was reduced to submicromolar levels. Intracellular calmodulin was significantly higher in rapidly dividing keratinocytes in normal (1.15 mM) calcium medium than in slower dividing cells in low (0.15 mM) calcium. Fibroblasts and B16 cells maintained similar calmodulin levels in both low and normal calcium media. Calmodulin antagonists inhibited proliferation of all three cell types equally. Thus, keratinocyte calmodulin seems related to the proliferative state of the cell (unlike fibroblast calmodulin) and calmodulin antagonists may be of use in controlling the hyperproliferation of the psoriatic epidermis.

Animals↗
PubMed1988

Extracellular calmodulin and its association with epidermal growth factor in normal human body fluids.

In this study we describe the occurrence of a calmodulin-like protein in normal human biological fluids. Extraction of the calmodulin-like protein from breast milk, saliva, serum and urine provided an extract with enhanced calmodulin immunoreactivity which, in the case of milk and saliva, showed a protein band comigrating with authentic calmodulin (Mr 17,000) on sodium dodecylsulphate-polyacrylamide gel electrophoresis. However, in milk, saliva and serum a major protein band of Mr 14,000-15,000 was always observed, which we speculate may be related to calmodulin, possibly as a partially degraded form. Estimates of biologically active calmodulin in most normal extracellular fluids were of the order which we have found will stimulate cell division when added to the extracellular medium of cells in culture. Levels ranged from 0.03 nmol/l in urine to 18.6 nmol/l in breast milk, and exhibited a quantitative relationship (r = 0.79, P less than 0.01) to epidermal growth factor (EGF) levels in fluids. Where EGF concentrations varied from normal (increased in saliva 24 h after oral surgery and reduced in the urine of patients with renal failure) calmodulin concentrations were similarly affected. The presence of calmodulin in serum may in part be attributable to its release from platelets which are particularly rich in calmodulin. Release of calmodulin from the platelet was associated with that of EGF and other platelet products.

Blood Platelets↗
PubMed1987

Evidence for the involvement of calmodulin in natural cytotoxicity using a range of calmodulin antagonists of varying potency and improved specificity.

The calmodulin antagonist W7 and 4 of its analogues were examined for their ability to inhibit human NK cell mediated cytotoxicity. With the exception of one of these compounds, which is extremely hydrophobic, there was a good correlation between the ability of drugs to inhibit human NK antitumour cytotoxicity and calmodulin-dependent phosphodiesterase activity in vitro. The most potent of the compounds, 5-iodo-1-C8, an analogue of W7, has an IC50 of 3 microM upon biological and biochemical assay. This particular compound is both more potent and specific than the parent compound W7, is non-toxic to cells over the range used and is also capable of inhibiting the biological activity of NK cells upon pre-treatment of the effector cells, inferring the mechanism of NK cytotoxicity to be calmodulin dependent.

Calmodulin↗
PubMed1987

Activation of calmodulin by the essential trace element chromium.

Chromium at very low concentrations is an essential trace element--at higher concentrations it is associated with contact dermatitis and other toxicity problems. Its ionic radius is just outside that of other metal cations which have been found to activate calmodulin in vitro. We found that chromium was able to activate calmodulin at two different concentration ranges--over the micromolar range (which would probably never be achieved in man) a small degree of activation was found--but a much greater activation (76% of the maximum possible) was also found at nanomolar concentrations of chromium. In welders, who work with stainless steel and who were not reporting any physical symptoms of chromium toxicity, red cell chromium levels were 28.2 +/- 3.3 nM (n = 22) compared to 7.5 +/- 0.7 nM (n = 11) for normal controls. Thus, the concentration of chromium experienced within the cell can be of the order which will activate calmodulin in vitro. The possibility exists, therefore, that inappropriate activation of calmodulin could be relevant to chromium biology possibly contributing to the symptoms of chromium toxicity.

Animals↗
PubMed1987

A comparative study of the anti-proliferative effects of calmodulin antagonists in cultured cells--W7 derivatives of improved cytostatic potential.

We have compared the effects of the calmodulin antagonist N-(6-aminohexyl)-5-chloro-1-naphthalene-sulphonamide (W7) and relatives of the parent compound, modified by substituting the 5-chloro- for an iodo-residue and increasing stepwise the length of the carbon chain from 6 to 12, for their ability to inhibit cell proliferation in tissue culture. These species showed improved specificity and potency, as determined against a calcium calmodulin-dependent beef heart phosphodiesterase in vitro, exhibited time courses of inhibition similar to the parent compound W7, but were more potent at inhibiting DNA synthesis in K562 human leukaemic cells cultured in serum-free medium. The elevation observed in the percentage of cells in G1 of the cell cycle following drug exposure indicated that these drugs, like W7, arrested growth by inhibiting entry of cells into and through S phase. Higher doses of all drugs were irreversibly cytotoxic as determined by the inability of the cells to recover DNA synthetic capacity and to form colonies in soft agar or to recover their normal cell cycle distribution. We also discuss the possible implications of extracellular calmodulin and antagonism thereof on cell proliferation.

Animals↗
PubMed1986

Psoriasis.

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Calmodulin↗
PubMed1986

An investigation of the ability of antipsoriatic drugs to inhibit calmodulin activity: a possible mode of action of dithranol (anthralin).

Epidermal calmodulin (CaM) has been reported to be elevated in psoriasis and to decrease following clearance of psoriasis with treatment. We set out to investigate whether any of the principle drugs used in the treatment of psoriasis had inherent CaM antagonist activity. Utilizing a CaM-activated phosphodiesterase we have demonstrated that even at very high concentrations, the systemic drugs etretinate, methotrexate, and 8-methoxypsoralen, and the topical agents hydrocortisone and crude coal tar showed minimal CaM inhibitory activity. Dithranol (anthralin), however, whether freshly prepared or oxidized, produced substantial inhibition of CaM activity and was demonstrated to be a potent competitive antagonist of CaM, suggesting another possible therapeutic mode of action of dithranol in psoriasis.

Animals↗
PubMed1986

Calmodulin levels in psoriasis: the effect of treatment.

Epidermal calmodulin levels were measured in lesional and non-lesional skin in 20 psoriatics before and after treatment by a variety of established topical and systemic regimes. Pretreatment, 10 out of 20 patients showed elevated calmodulin levels in lesional epidermis. Clearance of psoriatic plaques was accompanied by a significant overall reduction (p less than 0.05) in epidermal calmodulin, irrespective of the treatment regime used. For the non-lesional epidermis, only 5 out of 17 patients initially showed elevated calmodulin levels and there was no significant reduction in those levels following treatment.

Adult↗
PubMed1985

Calmodulin regulation of adenylate cyclase activity.

Calmodulin-dependent stimulation of adenylate cyclase was initially thought to be a unique feature of neural tissues. In recent years evidence to the contrary has accumulated, calmodulin-dependent stimulation of adenylate cyclase now being demonstrated in a wide range of structurally unrelated tissues and species. Demonstration of the existence of calmodulin-dependent adenylate cyclase has in nearly all instances required the removal of endogenous calmodulin. It is not yet clear whether calmodulin-dependent and calmodulin-independent forms of the enzyme exist and whether some tissues (such as heart) lack a calmodulin-dependent adenylate cyclase. The presence of calmodulin appears largely responsible for the ability of the adenylate cyclase enzyme to be stimulated by submicromolar concentrations of calcium; it may not be relevant to the inhibition of the enzyme which occurs at higher concentrations of calcium. The physical relationship of calmodulin to the plasma membrane bound enzyme (or to the soluble forms of the enzyme) is not known nor is the mechanism of adenylate cyclase activation by calmodulin clear; current data suggest some involvement with both the N and C units of the enzyme. Finally, it is possible that in vivo calcium contributes to the duration of the hormone stimulated cyclic AMP signal. Thus current in vitro data suggest that optimal hormonal activation of calmodulin-dependent adenylate cyclase occurs at very low intracellular calcium concentrations, comparable to those found in the resting cell; conversely the enzyme is inhibited as intracellular calcium increases, following for example agonist stimulation of the cell. These higher calcium concentrations would then activate calmodulin-dependent phosphodiesterase. Such differential effects of calcium on adenylate cyclase and phosphodiesterase would ultimately restrict the duration of the hormone-induced cyclic AMP signal.

Adenylyl Cyclases↗
PubMed1984

Isolation, purification and cell-free synthesis of calmodulin from the pig anterior pituitary gland.

Calmodulin was extracted and purified from pig anterior pituitary gland. The protein was characterized by its migration on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis in the presence of Ca2+ or EGTA, its U.V. spectrum between 240 and 290 nm and the activation of calmodulin-deficient cyclic AMP phosphodiesterase. The yield was 370 mg/kg wet wt. mRNA was also extracted from the same tissue and translated in a wheat-germ cell-free translation system. Translated calmodulin was identified by its heat-stability, its co-migration with authentic anterior-pituitary calmodulin on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, its acidic isoelectric point (4.15) on flat-bed isoelectric focusing, its Ca2+-dependent binding to fluphenazine-Sepharose 6B, and its co-elution from this gel with authentic unlabelled calmodulin with EGTA buffer. Calmodulin was not translated as a precursor form. In this tissue it was calculated that calmodulin accounted for 0.5-1% of the total translated protein.

Animals↗
PubMed1984

Biologically active calmodulin levels are elevated in both involved and uninvolved epidermis in psoriasis.

The aim of this study was to determine whether levels of biologically active calmodulin are elevated in both lesional and uninvolved epidermis in psoriasis. Epidermal shave biopsies were obtained from normal controls and from both psoriatic plaques and nonlesional psoriatic skin. Following determination of the protein content, the calmodulin activity of the homogenized samples was then measured using a calmodulin-sensitive phosphodiesterase enzyme bioassay. In normal skin, calmodulin activity was 1.29 +/- 0.35 micrograms calmodulin mg-1 epidermal protein (mean +/- SEM, n = 12 volunteers) compared to 7.88 +/- 1.59 micrograms calmodulin mg-1 epidermal protein for plaque (n = 16 patients) and 10.19 +/- 2.35 micrograms calmodulin mg-1 epidermal protein for the uninvolved skin of 12 of these patients. The levels of biologically active calmodulin were therefore elevated in both plaque and uninvolved epidermis of patients with psoriasis compared to epidermis from normal volunteers. These results suggest that an abnormality in the regulation of calmodulin activity may be involved in the pathogenesis of psoriasis.

Biopsy↗
PubMed1984

Calmodulin.

The two best-defined intracellular signalling systems are cAMP and calcium. It is clear that the actions of cAMP depend on its interaction with an enzyme--cAMP-dependent protein kinase. One of the major developments in recent years has been the recognition that many of the intracellular actions of calcium occur through its binding to and activation of calmodulin.

Adenylyl Cyclases↗
PubMed1983

Calcium, calmodulin, and the production of prostacyclin by cultured vascular endothelial cells.

The production of prostacyclin (PGI2) by cultured porcine aortic endothelial cells, in response to serum and the calcium ionophore A23187, was inhibited by TMB-8, an antagonist of intracellular calcium mobilization. The calcium-channel blocker methoxyverapamil (D600) inhibited serum-induced PGI2 production in but had little effect on A23187-induced PGI2 production. Calmodulin activity was detected in endothelial-cell lysates and was inhibited by the calmodulin antagonist W7, which also inhibited PGI2 production in response to both agonists. Calcium and calmodulin appear to play an important role in mediating PGI2 production by the vascular endothelium.

Animals↗
PubMed1983

The relationship between prolactin secretion and calmodulin activity.

The possible role of calmodulin in the control of prolactin secretion was examined. The effects of a wide variety of agents on inhibition of prolactin secretion from isolated rat anterior pituitary cells and inhibition of calmodulin activity in an in-vitro system (calmodulin-activated cyclic GMP phosphodiesterase) were compared. A number of phenothiazines showed a close correlation of potencies in the two systems, as did the more specific calmodulin antagonist, N-(6-aminohexyl)-5-chloro-1-naphthalene sulphonamide (W7), suggesting that calmodulin may be involved in the control of prolactin secretion. Some other drugs also inhibited both prolactin secretion and calmodulin activity in addition to their other well-characterized biochemical effects. However, many of these drugs (including two phenothiazines) were more potent inhibitors of prolactin secretion than of calmodulin activity, suggesting that other intracellular systems in addition to calmodulin may be involved in the control of the secretory process.

Animals↗
PubMed1983

A possible role for calmodulin in human thyroid cell metabolism.

A protein which shared several characteristics with authentic calmodulin was extracted from human thyroid homogenates. The protein bound to fluphenazine--Sepharose and could be specifically eluted using EGTA. The eluted protein had a u.v. spectrum characteristic of calmodulin and migrated like authentic calmodulin with a calcium-dependent shift on sodium dodecyl sulphate polyacrylamide-gel electrophoresis. Calmodulin in thyroid cell extracts was shown to be biologically active, measured by its ability to activate a calmodulin-deficient cyclic GMP phosphodiesterase; this activation could be inhibited by trifluoperazine. A possible role for calmodulin in the action of TSH on the thyroid was demonstrated by studying the effects of phenothiazines and the naphthalene sulphonamide, W7, a more specific calmodulin inhibitor, on TSH-stimulated cyclic AMP levels in cultured thyroid cells. The phenothiazines and W7 were found to inhibit the accumulation of cyclic AMP in response to TSH in a concentration-dependent manner although low concentrations of W7 enhanced TSH-stimulated cyclic AMP accumulation.

3',5'-Cyclic-GMP Phosphodiesterases↗