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Biomedical subjects

M W Spence

Publications and source records attributed to M W Spence.

At least 55 records · Page 3Linked to original sources
PubMed1987

Prevalence of major mental retardation and associated disabilities in the Canadian Maritime Provinces.

A cross-sectional survey of the prevalence of major mental retardation in 7- to 10-year-olds in the three Canadian Maritime provinces was carried out in 1980. Case ascertainment from multiple sources was considered to be virtually complete and resulted in a study group of 307 children, for whom 72% of the parents consented to personal data collection. The overall regional prevalence of major mental retardation was 3.65/1,000. Variation in county prevalence rate (based on parental residence at the time of birth) was associated, by regression analysis, with maternal age, areas of greater population concentrations, and number of physicians per capita. These associations were still present when children with Down syndrome were excluded from the analysis. Speech and behavior disorders and epilepsy were the major associated disabilities.

Canada↗
PubMed1986

Turnover of phospholipid fatty acyl chains in cultured neuroblastoma cells: involvement of deacylation-reacylation and de novo synthesis in plasma membranes.

Cultured neuroblastoma cells (NIE-115) rapidly incorporated the essential fatty acid, linoleic acid (18:2 (n = 6), into membrane phospholipids. Fatty acid label appeared rapidly (2-10 min) in plasma membrane phospholipids without evidence of an initial lag. Specific activity (nmol fatty acid/mumol phospholipid) was 1.5-2-fold higher in microsomes than in plasma membrane. In these membrane fractions phosphatidylcholine had at least 2-fold higher specific activity than other phospholipids. With 32P as radioactive precursor, the specific activity of phosphatidylinositol was 2-fold higher compared to other phospholipids in both plasma membrane and microsomes. Thus a differential turnover of fatty acyl and head group moieties of both phospholipids was suggested. This was confirmed in dual-label (3H fatty acid and 32P), pulse-chase studies that showed a relatively rapid loss of fatty acyl chains compared to the head group of phosphatidylcholine; the opposite occurred with phosphatidylinositol. A high loss of fatty acyl chain relative to phosphorus indicated involvement of deacylation-reacylation in fatty acyl chain turnover. The patterns of label loss in pulse-chase experiments at 37 and 10 degrees C indicated some independent synthesis and modification of plasma membrane phospholipids at the plasma membrane. Lysophosphatidylcholine acyltransferase and choline phosphotransferase activities were demonstrated in isolated plasma membrane in vitro. Thus, studies with intact cells and with isolated membrane fractions suggested that neuroblastoma plasma membranes possess enzyme activities capable of altering phospholipid fatty acyl chain composition by deacylation-reacylation and de novo synthesis at the plasma membrane itself.

1-Acylglycerophosphocholine O-Acyltransferase↗
PubMed1986

Causal origins of major mental handicap in the Canadian Maritime provinces.

From a retrospective epidemiological survey, the prevalence of major mental handicap among seven- to 10-year-old children in the Maritime region of Canada in 1980 was estimated to be 36.5 per 10,000 children. Based on information obtained for 221 of the 307 children ascertained by the survey, prenatal origins dominated (58 per cent), followed by perinatal (10 per cent) and postnatal (4 per cent). No specific cause could be determined for 27 per cent of the children, but 41 per cent of this group had epilepsy and/or cerebral palsy in addition to major mental handicap. Significant differences were found in the causal spectrum between the Maritime region of Canada and regions studied in other developed countries. The results of this survey have implications for planning prevention programs, and for epidemiological surveillance and monitoring of adverse reproductive outcomes.

Birth Weight↗
PubMed1986

Acid and alkaline ceramidases of rat tissues.

Ceramidase (N-acylsphingosine deacylase) was measured in rat tissue homogenates with N-[1-14C]oleoyl- and N-[9,10-3H]palmitoyl-sphingosine. Two activities were observed with pH optima of approximately 5 and 8. The activity at pH 8 exceeded that at pH 5 in all tissues and tissue areas examined, and both activities decreased in the following order: kidney greater than brain stem greater than cerebrum greater than cerebellum greater than liver greater than spleen greater than cardiac muscle greater than lung greater than psoas muscle. The distribution of activity is not as tissue specific as that of neutral Mg2+-stimulated sphingomyelinase which is enriched in cells of neural origin and thus suggests a less specialized role for the alkaline ceramidase.

Amidohydrolases↗
PubMed1985

Rapid isolation of neuroblastoma plasma membranes on Percoll gradients. Characterization and lipid composition.

A purified plasma membrane fraction was isolated from cultured neuroblastoma (N1E-115) cells on a discontinuous gradient of 5, 25 and 35% Percoll within 1 h of cell disruption by nitrogen cavitation. Yield of plasma membrane, banding in the 25% Percoll (d = 1.051), was high as judged by the recoveries of the marker enzymes, 5'-nucleotidase (58.0 +/- 5.4%, n = 5), alkaline phosphatase (46.0 +/- 3.0%, n = 4) and Mg2+-stimulated neutral sphingomyelinase (48.0 +/- 4.2%, n = 3); enrichment of specific activities of these enzymes relative to total cell homogenate (lysate) were 10.9 +/- 1.0-, 9.1 +/- 1.0- and 9.6 +/- 0.4-fold, respectively. Levels of marker enzymes for other organelles were less than 3% of total activity, except for microsomes (less than 9%). The plasma membrane fraction was further characterized by 2-, 5- and 6-fold higher content (nmol/mg protein) of total phospholipids, free cholesterol and sphingomyelin, respectively, compared to lysate. Ratios of free cholesterol to phospholipids and of sphingomyelin to phosphatidylcholine in the plasma membrane fraction were about 2-fold greater than that of lysate. The cholesterol ester content of plasma membrane (36 +/- 8 nmol/mg protein) was 2-3-fold higher than that of lysate. Sphingomyelin of the plasma membrane fraction had a higher concentration of long-chain fatty acids (more than 18 carbon atoms) relative to lysate or microsomes. Significant differences also were observed in the fatty acyl composition of diphosphatidylglycerol, cholesterol esters and triacylglycerol of plasma membrane. Thus, we have devised a rapid and reliable method for isolation of highly purified plasma membranes of cultured neuroblastoma cells that is suitable for comparison of metabolic relationships between the plasma membrane and other cellular organelles.

Animals↗
PubMed1985

Studies on the turnover and subcellular localization of membrane gangliosides in cultured neuroblastoma cells.

To compare the subcellular distribution of endogenously synthesized and exogenous gangliosides, cultured murine neuroblastoma cells (N1E-115) were incubated in suspension for 22 h in the presence of D-[1-3H]galactose or [3H]GM1 ganglioside, transferred to culture medium containing no radioisotope for periods of up to 72 hr, and then subjected to subcellular fractionation and analysis of lipid-sialic acid and radiolabeled ganglioside levels. The results indicated that GM2 and GM3 were the principal gangliosides in the cells with only traces of GM1 and small amounts of disialogangliosides present. About 50% of the endogenously synthesized radiolabelled ganglioside in the four major subcellular membrane fractions studied was recovered from plasma membrane and only 10-15% from the crude mitochondrial membrane fraction. In contrast, 45% of the exogenous [3H]GM1 taken up into the same subcellular membrane fractions was recovered from the crude mitochondrial fraction; less than 15% was localized in the plasma membrane fraction. The results are similar to those obtained from previously reported studies on membrane phospholipid turnover. They suggest that exogenous GM1 ganglioside, like exogenous phosphatidylcholine, does not intermix freely with any quantitatively major pool of endogenous membrane lipid.

Animals↗
PubMed1985

Catabolism of exogenous and endogenous sphingomyelin and phosphatidylcholine by homogenates and subcellular fractions of cultured neuroblastoma cells. Effects of anesthetics.

Cultured murine neuroblastoma cells contain a neutral, Mg2+-stimulated sphingomyelinase and an alkaline phosphatidylcholine-hydrolyzing activity that are enriched in the plasma membrane fraction. The reaction products of sphingomyelin catabolism are phosphocholine and ceramide and those of phosphatidylcholine, glycerophosphocholine and fatty acid. These reactions were studied with endogenous as well as exogenous liposomal substrates. With both exogenous and endogenous substrates, the sphingomyelinase activity was stimulated two- to threefold by Mg2+ and a further three- to fourfold by volatile anesthetic agents. Stimulation was concentration-dependent and corresponded to anesthetic potency: methoxyflurane greater than halothane greater than enflurane. Greater than 80% of the plasma membrane sphingomyelin was hydrolyzed within 2 h in the presence of Mg2+ and anesthetic. In contrast, the activity with exogenous and endogenous phosphatidylcholine was unaffected by Mg2+ or Ca2+ and was markedly inhibited (50-80%) by anesthetic agents. The degree of inhibition was concentration-dependent and corresponded to anesthetic potency. The quantitative importance of choline-containing lipids in cell membranes, the relatively exclusive localization of the neutral Mg2+-stimulated sphingomyelinase in cells of neural origin, the totally different type of hydrolytic attack on phosphatidylcholine, and the reciprocal effects of anesthetics on the hydrolysis of these two lipids strongly suggest important roles for these activities in cell membranes in general and in the neuron in particular.

Anesthetics↗
PubMed1985

Triacylglycerol as a precursor in phospholipid biosynthesis in cultured neuroblastoma cells: studies with labeled glucose, fatty acid, and triacylglycerol.

Neuroblastoma cells rapidly incorporate exogenous fatty acids into cellular triacylglycerol and relationships between triacylglycerol and phospholipid biosynthesis have been indicated by the relative time course of labeling of these lipids. To evaluate this further, neuroblastoma cells were labeled using potential precursors of phospholipid including radiolabeled triacyglycerol, glycerol, glucose, and fatty acid. With [2-3H]glycerol or a mixture of [2-3H]glycerol trioleate and glycerol tri[1-14C]oleate, phospholipids were labeled at very low levels (less than 0.1 and less than 0.5%, respectively). With [6-3H]glucose, labeling of lipids (0.5-3.5%) was greatest in medium containing 19 mM fructose, whereas labeling with [1-14C]18:2(n-6) was similar in media containing either 19 mM fructose or 25 mM glucose. Labeling of the glycerol moiety of triacylglycerol with [6-3H]glucose increased with 40-200 microM 18:2(n-6) present and occurred predominantly in 2 h. Some [6-3H]glucose label was in fatty acyl chains (chiefly 16:0) of triacylglycerol by 16 h, but was unaffected by exogenous 18:2(n-6). Triacylglycerol was the only lipid to increase in mass (threefold with 200 microM 18:2(n-6)). During the chase of cells pulsed with [6-3H]glucose, label in triacylglycerol declined within 0.5 h, whereas in phospholipid it increased transiently up to 2 h and then declined. Changes were inversely proportional to 18:2(n-6) levels in the chase medium and labeled acyl chains moved in parallel with the glycerol moiety. Thus, a major portion of acyl chain transfer from triacylglycerol was accompanied by glycerol.(ABSTRACT TRUNCATED AT 250 WORDS)

Cells, Cultured↗
PubMed1984

Evidence that neutral sphingomyelinase of cultured murine neuroblastoma cells is oriented externally on the plasma membrane.

The activity of the neutral, Mg2+-stimulated sphingomyelinase of cultured neuroblastoma cells (N1E-115) is enriched in the plasma membrane fraction and is reduced following treatment of intact or broken cells with trypsin, alpha-chymotrypsin, papain, and protease. Two protease-sensitive enzymes of the cell interior (lactate dehydrogenase and NADPH-cytochrome c reductase) are not affected by protease treatment of intact cells. These results indicate that the neutral, Mg2+-stimulated sphingomyelinase is oriented externally on the plasma membrane of the cultured neuroblastoma cell.

Animals↗
PubMed1983

Studies on the turnover of endogenous choline-containing phospholipids of cultured neuroblastoma cells.

Previous studies on neuroblastoma cells in culture showed that the presence of partially purified rat liver phospholipid-transfer protein had a marked differential effect on the uptake and apparent subcellular distribution of radioactively labeled sphingomyelin and phosphatidylcholine (PC) added to the medium as mixed phospholipid (PC/sphingomyelin) liposomes. To determine the effect of phospholipid-transfer protein and exogenous phospholipids on the turnover and subcellular distribution of endogenous phospholipids, neuroblastoma cells were preincubated for 48 h in the presence of [methyl-3H]choline and washed. Aliquots of prelabeled cells were reincubated immediately in medium containing phospholipid-transfer protein mixed phospholipid liposomes, cytochalasin B and 2-deoxyglucose for 45 min at 37 degrees C; additional aliquots were chased first for 2 or 18 h with unlabeled choline before reincubation. The extent of labeled phospholipid degradation and accumulation in the medium, and the subcellular distribution of cell-associated labeled choline-containing phospholipids were determined. During incubation with phospholipid-transfer protein and mixed phospholipid liposomes, 25-35% of the cell-associated radioactive label from prelabeled cells, chased or unchased, was lost to the medium in 45 min. Over 50% of the label appearing in the medium was in water-soluble phospholipid degradation products. The loss of cell-associated label into the medium from unchased cells was stimulated significantly by phospholipid-transfer protein; however, prelabeled cells which had been chased for 18 h with unlabeled choline were unaffected by the presence of transfer protein. Endogenously synthesized radioactively labeled PC and sphingomyelin were distributed throughout all subcellular membranes, but least of all in the crude mitochondrial membrane fraction. Analysis of the subcellular distribution of cell-associated label remaining in chased or unchased cells after 45 min incubation with PC/sphingomyelin liposomes showed proportionate losses from all membrane fractions, except the crude mitochondrial fraction, which showed relative retention of labeled phospholipid. Phospholipid-transfer protein had no effect. The results are in distinct contrast to observations on the turnover, metabolism and subcellular distribution of labeled exogenous phospholipids under the same conditions, indicating that exogenous phospholipids do not intermix freely with any quantitatively major pool of endogenous phospholipid.

Animals↗
PubMed1983

Pathways of sphingomyelin metabolism in cultured fibroblasts from normal and sphingomyelin lipidosis subjects.

The metabolism of endogenous sphingomyelin labeled with 32P or [methyl-3H]choline and of exogenous [choline-methyl-3H], [32P]-, or [N-acyl-1-14C]sphingomyelin was studied in normal and Niemann-Pick Type A (NP-A) cultured fibroblasts. Despite a greater than 96% decrease in lysosomal sphingomyelinase activity in the NP-A cells, they were able to degrade endogenously produced [32P]- or [methyl-3H]sphingomyelin at normal or near normal rates. Exogenous [methyl-3H]-, [methyl-3H, 32P]-, and [methyl-3H, N-acyl-1-14C] sphingomyelin was taken up intact by normal and NP-A cells, with NP-A cells accumulating 4-8 times more lipid. By 20 h, 50% of the control cell-associated 3H and 32P was recovered in lecithin, and the ratio of activities (3H/32P) indicated most of the phosphorylcholine derived from sphingomyelin had been transferred intact. By comparison in NP-A cells, after a 40-h incubation only 20% of the labeled phosphorylcholine derived from sphingomyelin was recovered in lecithin. With both cell lines, 20 to 50 times more sphingomyelin was hydrolyzed than was taken up by the cells; the reaction products in the medium were ceramide and a mixture of water-soluble compounds such as phosphorylcholine and choline. These results indicate that there are at least two metabolic pathways for sphingomyelin modification in cultured fibroblasts in addition to degradation by the lysosomal acid sphingomyelinase. One route is hydrolysis by a cellular sphingomyelinase. The second is the hydrolysis and/or transfer of phosphorylcholine from sphingomyelin and results in the synthesis of lecithin.

Carbon Radioisotopes↗
PubMed1983

Concerted stimulation and inhibition of desaturation, chain elongation, and esterification of essential fatty acids by cultured neuroblastoma cells.

Neuroblastoma (N1E-115) cells in culture rapidly incorporated exogenous fatty acyl chains suspended as albumin complexes in the medium. The essential fatty acids, linoleic (18:2(n - 6)) and linolenic (18:3(n - 3)) acids, were converted to polyunsaturated acids by delta 6 and delta 5 desaturation and chain elongation. The major end products (20:4 from 18:2(n - 6) and 20:5 and 22:5 from 18:3(n - 3)) were preferentially esterified to phospholipids, whereas intermediates were esterified primarily or equally to triacylglycerol. The effects of unlabeled exogenous fatty acids (eg. 40 microM 18:2(n - 6), 18:3(n - 6), 18:3(n - 3), 20:3(n - 6), 20:4(n - 6), trans-18:2(n - 6), 18:1(n - 9), trans-18:1(n - 9), or 16:0) on the conversion of 2 microM [1-(14)C]18:2(n - 6), 18:3(n - 3), 20:3(n - 6), 20:4(n - 6), or 16:0 and on accumulation of products and unaltered substrates in phospholipids and triacylglycerol were examined after incubations of 2-24 h. With [1-(14)C]18:2, formation and esterification of 20:4 to phospholipids was (i) stimulated 4-8-fold by 18:2(n - 6), 20:3(n - 6), or 20:4(n - 6), (ii) inhibited by 18:3(n - 3) or trans-18:2(n - 6), or (iii) unaffected by 18:3(n - 6), 18:1(n - 9), trans-18:1(n - 9), and 16:0. Specific but less marked effects were observed with the other 1-(14)C-substrate acids. Thus, various fatty acids influenced the metabolism of essential fatty acids both at the level of conversion by desaturation and elongation and at esterification to complex lipids by mechanisms specific to individual acids. Product inhibition was not a major feedback mechanism; however, the complement of available fatty acids evidently modulates the acyl chain composition of membrane phospholipids through processes in addition to acyltransferase selectivity. The data support a closely coordinated or concerted enzyme system for directed synthesis of esterified polyunsaturated acyl chains.

Animals↗
PubMed1983

Phospholipid transfer protein-mediated incorporation and subcellular distribution of exogenous phosphatidylcholine and sphingomyelin in cultured neuroblastoma cells.

Murine neuroblastoma cells (strain N1E-115) were incubated with 2-[fatty acyl-14 C]acylphosphatidylcholine/sphingomyelin or phosphatidylcholine/[choline-3H]sphingomyelin liposomes (1:1, mol/mol; 1.2 mumol total lipid/mg cell protein) in the presence of partially purified rat liver phospholipid transfer protein (2.5 mg/ml), cytochalasin B (50 microM) and 2-deoxyglucose (50 mM) for 10 min. Washed cells were chased for periods of up to 45 min at 37 degrees C with medium containing transfer protein and unlabeled liposomes. Total transfer protein-dependent incorporation of [14C]phosphatidylcholine ([14C] PC) and [3H]sphingomyelin was 136.7 +/- 26.5(n = 5) and 23.7 +/- 5.4(n = 6) nmol/mg protein per 10 min incubation, respectively, (mean +/- S.D.). Incorporation of [14C]PC into the mitochondrial membrane fraction was 128-fold greater (nmol/mg protein) than incorporation of [3H]sphingomyelin. In contrast, incorporation of [3H]sphingomyelin into a fraction enriched in plasma membrane and into microsomes was 1.4- and 2.6-fold greater, respectively, than incorporation of [14C]PC. During the chase periods, the specific activities of total cellular phospholipids decreased as intact [14C]PC and [3H]sphingomyelin accumulated in the culture medium. In the case of cells labeled with [14C]PC, the effect was due primarily to a decrease in the amount of labeled phospholipid in the mitochondrial fraction; in the case of cells labeled with [3H]sphingomyelin, the decrease in activity was greatest in microsomal and plasma membrane phospholipids. The rate and extent of non-endocytotic incorporation of exogenous phosphatidylcholine into the cell membrane of cultured neuroblastoma cells, and its subsequent subcellular disposition, is different from that of exogenous sphingomyelin. Whereas PC is evidently incorporated into and turned over most rapidly in fraction enriched in mitochondrial membranes, sphingomyelin appears to be preferentially incorporated into microsomal and plasma membrane.

Animals↗
PubMed1983

Stimulation of neutral, magnesium-stimulated sphingomyelinase activity in the neurohypophysis of the rat by hypertonic saline ingestion.

Neutral, magnesium-stimulated sphingomyelinase and acid sphingomyelinase activities in the neurohypophysis, adenohypophysis and cerebrum of rats given 2.25% NaCl drinking water for 7 or 14 days were compared with the same enzyme activities in the tissues of control animals allowed free access to distilled drinking water. Neutral sphingomyelinase activity in the neurohypophysis was significantly increased in both experimental groups (7-day, 85.2 +/- 8.4 units/mg protein; 14-day, 110.1 +/- 14.8 units/mg; control, 61.4 +/- 5.5 units/mg). Acid sphingomyelinase activity was significantly but only transiently increased in the tissue (7-day, 73.2 +/- 3.4 units/mg; 14-day, 66.3 +/- 7.5 units/mg; control, 54.3 +/- 2.8 units). Enzyme activities in adenohypophysis and cerebrum of the experimental animals were not significantly different from those of controls. The results suggest a specific role for neutral, Mg2+-stimulated sphinomyelinase in neurosecretion.

Animals↗
PubMed1983

Inability of rabbit peritoneal polymorphonuclear leukocytes to synthesize arachidonic acid from linoleic acid.

In PMN leukocytes isolated from rabbit peritoneal exudate the major phospholipids were choline phosphoglycerides (40%), ethanolamine phosphoglycerides (26%) and sphingomyelin (20%) with lesser amounts (3-6%) of serine and inositol phosphoglycerides. The essential fatty acid, linoleic acid, predominated (greater than 35%) in each phospholipid except in inositol phosphoglycerides where it was slightly less than arachidonate and in sphingomyelin where saturated acids predominated. However, on a total mass basis there was more arachidonate in ethanolamine and choline phosphoglycerides than in inositol phosphoglycerides. The uptake, incorporation and metabolism of [1-14C] fatty acids of varying chain length and degrees of unsaturation were examined. All fatty acids were taken up but incorporation of saturated acids varied inversely with chain length. Arachidic acid and trans-isomers of 18:1 and 18:2 were esterified primarily to triacylglycerol whereas phospholipids contained a large portion of the other acids. Icosatrienoic and arachidonic acids were esterified to ethanolamine, serine and inositol phosphoglycerides to a comparatively greater extent, reflecting the normal distribution of these fatty acids. PMN leukocytes had a low capacity for delta 9 desaturation and chain elongation and no delta 6 or delta 5 desaturation could be detected. Thus, PMN leukocytes lack the ability to form arachidonate from 18:2 precursor molecules available in the cellular neutral lipids and phospholipids and arachidonate per se is an essential fatty acid for these cells.

Animals↗
PubMed1983

Neutral and acid sphingomyelinases of rat brain: somatotopographical distribution and activity following experimental manipulation of the dopaminergic system in vivo.

The activities of neutral, magnesium-stimulated, and acid sphingomyelinases were measured in five regions of rat brain. Neutral enzyme activity was 2-3-fold higher in striatum than in parietal cortex and 13-fold higher than in cerebral white matter. Acid sphingomyelinase activity was more evenly distributed throughout these regions. Striatal neutral sphingomyelinase activity was not affected by treatment of rats with reserpine or haloperidol and was reduced (16%) by 6-hydroxydopamine. Striatal acid sphingomyelinase was unaffected by reserpine and 6-hydroxydopamine, and was increased (17%) by haloperidol. We conclude that neutral, magnesium-stimulated sphingomyelinase activity differs in various regions of rat brain and is particularly enriched in the corpus striatum. However, it appears to be a constitutive component of tissue rather than a readily modulated regulatory element of the catecholaminergic system.

Animals↗