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T K Smith

Publications and source records attributed to T K Smith.

At least 91 records · Page 5Linked to original sources
PubMed1995

On the cytotoxicity and genotoxicity of allyl and phenethyl isothiocyanates and their parent glucosinolates sinigrin and gluconasturtiin.

Four compounds commonly found in the human diet, allyl isothiocyanate (AITC), phenethyl isothiocyanate (PEITC) and their parent glucosinolates sinigrin and gluconasturtiin, were tested for cytotoxic and genotoxic effects in a Chinese hamster ovary cell line (CHO). The isothiocyanates were found to be more than one thousand times more cytotoxic than the glucosinolates, showing significant cytotoxic activity at concentrations below 1.0 microgram/ml. AITC was unable to induce either chromosome aberrations or sister chromatid exchanges (SCEs) even at highly cytotoxic doses. In contrast, PEITC was found to induce both aberrations and SCE at concentrations of 0.9-1.2 micrograms/ml whilst sinigrin and gluconasturtiin induced aberrations at concentrations above 2 mg/ml.

Animals↗
PubMed1995

Selective toxicity of compounds naturally present in food toward the transformed phenotype of human colorectal cell line HT29.

It has previously been observed that allyl isothiocyanate, a compound naturally present in the diet, is more cytotoxic toward the human colorectal adenocarcinoma cell line HT29 in its control transformed state than after exposure to sodium butyrate or to dimethylformamide, which slow growth and induce differentiation (detransformation). In the present study, a range of other dietary compounds were assayed for such selective toxicity. These compounds were chosen as constituents of foodstuffs that have been identified from epidemiologic studies as being potentially antitumorigenic and also as having anticarcinogenic activity in experimental models. Benzyl and phenethyl isothiocyanate, benzyl thiocyanate, and quercetin showed decreased toxicity towards HT29 after detransformation of the cells by one or both treatments, whereas no change was observed in the sensitivity to diallyl sulfide or diallyl disulfide. It is proposed that the presence of such selectively toxic compounds in the diet may inhibit the development of tumors by interfering with the growth of preneoplastic lesions while having little effect on normal cells. The cumulative effects of these inhibitions may contribute to the chemopreventive properties of the parent foodstuffs observed in epidemiologic studies.

Adenocarcinoma↗
PubMed1995

Toxicity and growth-promoting potential of spermine when fed to chicks.

Previous studies have shown that the feeding of putrescine, a biogenic amine and the precursor of the mammalian polyamines, can promote whole-body growth of chicks. The current study was undertaken to determine the effect of spermine, also a biogenic amine and the most cationic of the polyamines, under similar conditions. In Exp. 1, 120 week-old chicks were fed purified crystalline amino acid-based diets containing 0, .2, .4, .6, .8, or 1.0% spermine for 14 d. Spermine proved highly toxic and growth rates were reduced compared with controls when even .2% was fed. In Exp. 2, chicks were fed 0, .0375, .0750, or .1000% spermine. These concentrations proved less toxic than those used in Exp. 1. Supplemental dietary cysteine was then provided at 0, .3, .6, and .9% together with 0, .025, .050, or .400% spermine (Exp. 3) because depletion of cellular glutathione has been suggested as contributing to spermine's toxicity. Even high levels of cysteine supplementation did not overcome spermine's toxicity. Subsequent dietary provision of L-2-oxothiazolidine-4-carboxylic acid (OTC, Exp. 4), a cysteine prodrug, showed that depletion of cellular glutathione was not likely a cause of spermine toxicosis. A trend toward increased weight gain and feed efficiency was observed when low concentrations of spermine were fed. It was concluded, however, that dietary spermine was more toxic to chicks than was previously seen for putrescine, that any growth-promoting effects of dietary spermine are small, and that supplements of dietary cysteine or OTC are unlikely to increase these effects by overcoming spermine toxicosis.

Adenosylmethionine Decarboxylase↗
PubMed1995

Altered tissue amino acid metabolism in acute T-2 toxicosis.

T-2 toxin is a Fusarium trichothecene mycotoxin that has been shown to alter brain neurochemistry and eating behavior in animals eating contaminated diets. Experiments were conducted to determine the role of altered tissue amino acid metabolism in the etiology of acute T-2 toxicosis. Fasted weanling rats were orally dosed with 0 or 2.0 mg T-2 toxin/kg body weight. Blood, brain, liver, and muscle tissue were excised 4 and 8 hr after dosing, and amino acid concentrations were determined. Hepatic enlargement coupled with reduced liver concentrations of free small neutral, large neutral, and basic amino acids were seen 4 hr after dosing. Brain and muscle amino acid concentrations were largely refractory to treatment, while the plasma concentrations of tyrosine and lysine, and the sum of the basic amino acids fell. Hepatic amino acid concentrations returned to control levels 8 hr after dosing at which time aminoacidemia was seen. This was due partially to an increase in plasma concentrations of large neutral amino acids including particularly the branched-chain amino acids. A subsequent experiment was conducted to determine the effect of T-2 toxin on 14C-leucine uptake and incorporation into protein in liver slices 4 hr after dosing. Exposure to T-2 toxin reduced total (free + protein-bound) uptake of leucine due primarily to reduced incorporation of leucine into newly-synthesized hepatic protein. It was concluded that reduced amino acid uptake by liver preceded aminoacidemia in acute T-2 toxicosis, although it is not clear how this might influence subsequent changes in brain neurochemistry and behavior.

Amino Acids↗
PubMed1994

Interaction of gamma-glutamyl transpeptidase with acivicin.

Inactivation of gamma-glutamyl transpeptidase by acivicin (L-(alpha S,5S)-alpha-amino-3-chloro-4,5-dihydro-5-isoxazole acetic acid) is rapid, thought to be irreversible, and associated with binding of close to 1 mol of inhibitor/mol of enzyme. Previous studies with [3-14C]acivicin indicated binding (prevented by substrate) to a specific hydroxyl group (threonine 523) of the rat kidney enzyme. In the present work, we found that such inactivation can be reversed by treating the inhibited enzyme with hydroxylamine. Reactivation (more than 85% complete) is associated with release from the inactivated enzyme of compounds that exhibit the properties of threo-beta-hydroxy-L-gamma-glutamyl hydroxamate and 3-hydroxypyrrolidone-2-carboxylate. We found that the enzyme acts very slowly on acivicin, at a rate that is about 10(-9) that of its normal catalytic rate with glutathione, to form threo-beta-hydroxy-L-glutamate and hydroxylamine. The findings indicate that inhibition by acivicin involves its transformation on the enzyme to an inhibitory species which is attached, apparently by ester linkage, to a specific hydroxyl group of the enzyme. The very slow rate of release of this intermediate appears to account for the observed inhibition.

Animals↗
PubMed1994

Active deglycosylated mammalian gamma-glutamyl transpeptidase.

gamma-Glutamyl transpeptidase, a highly glycosylated heterodimeric enzyme that is usually attached to the external surface of cell membranes, is of major importance in the metabolism of glutathione. The enzyme, which has been isolated from many animal sources, contains a large amount of carbohydrate, which is linked to both protein subunits. Previous work has not shown whether such carbohydrate is needed for enzyme activity nor indicated its functional role. Notably, gamma-glutamyl transpeptidase isolated from Escherichia coli, which exhibits about 80% amino acid sequence homology with the rat enzyme, has only about 0.1% of its specific enzymatic activity and is not glycosylated. Here we treated the highly glycosylated gamma-glutamyl transpeptidases isolated from rat and pig kidneys with a mixture of glycosidases and then separated two completely active gamma-glutamyl transpeptidase fractions from each species. One fraction was completely devoid of carbohydrate and was fully active as compared with the respective isolated enzymes, but differed in solubility and stability. The other fraction, which contained 10-20% of the initially bound carbohydrate, exhibited a marked increase in susceptibility to proteases. The oligosaccharide chains of gamma-glutamyl transpeptidase may protect against protease action (including self-destruction by the inherent protease activity of the light subunit) during synthesis of the active enzyme from its single chain precursor, as well as after enzyme synthesis.

Animals↗
PubMed1994

Cytokines decrease apolipoprotein accumulation in medium from Hep G2 cells.

Cytokines, important biochemical mediators of inflammation, cause a rapid fall in the plasma concentration of cholesterol in vivo. One mechanism by which cytokines may cause acquired hypocholesterolemia is by decreasing the hepatic synthesis and secretion of apolipoproteins. To test this hypothesis, we incubated Hep G2 cells with human recombinant tumor necrosis factor-alpha, interleukin-1 beta, and interleukin-6. Each of the cytokines resulted in a dose-related reduction in the concentrations of apolipoprotein (apo) A-I, apoB, and lecithin:cholesterol acyltransferase (LCAT) activity in the medium after 24 hours of incubation. The effect of cytokines on apolipoprotein accumulation was not affected by preincubation of Hep G2 cells with fatty acids. Cytokines decreased the concentration of cellular apoA-I mRNA in a dose-related fashion but did not affect cellular concentrations of apoB mRNA. The concentrations of triglyceride and cholesterol were also reduced in the medium of cells incubated with cytokines. Total cell sterol synthesis rates were calculated by [14C]acetate incorporation. Cells incubated with interleukin-6 had a 31% increase in sterol synthesis rate but a 41% decrease in sterol secretion. These data suggest that these cytokines can decrease the hepatic synthesis and/or secretion of apolipoproteins and that this may explain, in part, the acquired hypocholesterolemia seen during acute and chronic inflammation.

Apolipoprotein A-I↗
PubMed1994

Mechanism of cyclic AMP-induced hyperpolarization in canine colon.

The mechanism of forskolin (FSK)-induced hyperpolarization was investigated in strips of canine colonic circular muscle. FSK responses were compared to those of the K+ channel opener lemakalim (LEM). Both FSK (10 microM) and LEM (10 microM) hyperpolarized cells near the myenteric border by 10 to 20 mV. Responses to both agents were abolished by 35 mM external K+, indicating a probable mediation by K+ channels. FSK increased the open probability of Ca(++)-activated K+ channels in isolated colonic myocytes. However, in muscle strips charybdotoxin (100 nM) and tetraethylammonium (10 mM) failed to reduce FSK- and LEM-induced hyperpolarizations whereas tetrapentylammonium (50 microM) and 4-aminopyridine (10 mM) blocked both responses. Phencyclidine (100 microM), Ba++ (1 mM) and the antagonist of ATP-sensitive K+ currents glybenclamide (10 microM) blocked LEM- but not FSK-induced hyperpolarizations. Delayed rectifier current in isolated myocytes was activated near -20 mV and was blocked by (order of potency): nifedipine > tetrapentylammonium > phencyclidine > 4-aminopyridine > tetraethylammonium. Charybdotoxin (100 nM), Ba++ (1 mM) and glybenclamide (10 microM) were without effect. Ca(++)-activated K+ current was activated near +30 mV and was blocked by: charybdotoxin > tetraethylammonium > tetrapentylammonium >> phencyclidine = 4-amino-pyridine. These data suggest that LEM induces membrane hyperpolarization by activation of a K+ current with a pharmacology similar to ATP-sensitive K+ current whereas cyclic AMP-induced hyperpolarization appears to involve activation of a current other than delayed rectifier current, Ca(++)-activated K+ current or ATP-sensitive K+ current.

Animals↗
PubMed1993

Methionine toxicity in the rat in relation to hepatic accumulation of S-adenosylmethionine: prevention by dietary stimulation of the hepatic transsulfuration pathway.

Rats were fed toxic levels of methionine with or without simultaneous dietary supplements of glycine and serine. Feed intake, growth rate, and metabolite concentrations in intestine, plasma, liver, skeletal muscle, and kidneys were monitored. Both toxic amounts of methionine and supplemental glycine and serine affected the tissue distribution of several amino acids resulting in similar, opposite, and diet-specific effects on the parameters studied. These changes were considered to be normal responses of amino acid metabolism to diet and to reflect metabolite flows between tissues. The feeding of toxic levels of methionine resulted in the accumulation of methionine, taurine, and glutathione in all tissues measured, but caused marked accumulation of S-adenosylmethionine and its catabolites only in liver. Hepatic accumulation of S-adenosylmethionine was accompanied by 40% stimulation of methionine adenosyltransferase and 40% repression of spermine synthase over a 2-week period. Simultaneous dietary supplements of glycine and serine combined with toxic levels of methionine markedly stimulated hepatic methionine catabolism. As a result, tissue distribution of methionine and glutathione returned close to normal in all tissues measured and accumulation of hepatic S-adenosylmethionine and its catabolites was prevented. Concentrations of taurine in liver, blood, and kidneys were further elevated, suggesting increased conversion of methionine to taurine followed by urinary excretion. These changes were accompanied by normalization of the above enzyme activities and the absence of symptoms of methionine toxicity. It was concluded that methionine toxicity is likely to be linked to hepatic accumulation of S-adenosylmethionine, resulting in liver dysfunction probably due to nonenzymatic methylation of liver macromolecules. Accumulation of tissue glutathione may also contribute to toxicity.

Amino Acids↗
PubMed1993

Beta-adrenergic inhibition of electrical and mechanical activity in canine colon: role of cAMP.

The effects of beta-adrenergic receptor stimulation on the electrical and mechanical activity of canine colonic circular muscles were compared with forskolin (Fsk), a known stimulator of adenylate cyclase. The actions of isoproterenol (Iso) were mediated by beta 2-receptors. Iso and Fsk increased intracellular adenosine 3',5'-cyclic monophosphate (cAMP) levels in both the presence and absence of acetylcholine (ACh), whereas ACh (0.3 microM) alone reduced cAMP levels. These agents caused inhibition of spontaneous and ACh-induced contractions. Inhibition was associated with a reduction in the amplitude and duration of electrical slow waves recorded near the submucosal border. Near the myenteric border, Iso and Fsk hyperpolarized the membrane by up to 30 mV and changed the pattern of electrical rhythmicity. These effects were mimicked by 8-bromo-cAMP (1-3 mM). Contractile inhibition with Fsk and Iso was associated with a decrease in the amplitude and duration of Ca2+ transients measured with fura-2 fluorescence. cAMP (10-300 microM) reduced the Ca2+ sensitivity of the contractile apparatus in muscles permeabilized with staphylococcal alpha-toxin. The actions of Iso appear linked to cAMP. We hypothesize that cAMP produces relaxation both by modulation of membrane ionic channels with a consequent decline in the entry of Ca2+ as well as through a decrease in the sensitivity of the contractile apparatus to Ca2+.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1992

Effect of interleukin-1 alpha on lipoprotein lipids in cynomolgus monkeys: comparison to tumor necrosis factor.

Acute inflammation is associated with changes in lipoprotein metabolism. Cytokines are thought to mediate the metabolic effects of the inflammatory process. This study was undertaken to compare the effects of interleukin-1 alpha (IL-1 alpha) to tumor necrosis factor (TNF) on lipoprotein metabolism in non-human primates. Recombinant human IL-1 alpha (100 micrograms/kg), TNF alpha (20 micrograms/kg) and lipopolysaccharide (20 micrograms/kg) were injected into cynomolgus monkeys. Lipoprotein concentrations, plasma activities of post-heparin lipase (PHLA) and lecithin:cholesterol acyltransferase (LCAT) were measured prior to and 24 and 48 h after, injection. All three injections caused afebrile response in the animals. Interleukin-1 alpha had no effect on plasma lipoprotein concentrations, composition of lipoproteins or enzyme activity. In contrast, injection of TNF caused significant changes in lipoprotein concentrations. There was a 38% increase in plasma triacylglycerol and 30% decrease in plasma cholesterol at 48 h after injection. Concentrations of apolipoproteins A-I and B were decreased 20% and 44%, respectively, at 48 h. Compositional analyses of lipoprotein particles after TNF injection showed that both the LDL and HDL particles had decreased content of cholesterol ester and increased triacylglycerol after injection, and plasma activities of PHLA and LCAT were decreased. These changes were qualitatively similar to those seen after LPS injection. These data suggest that, unlike TNF, IL-1 alpha is not an important mediator of the inflammatory process on lipoprotein metabolism in cynomolgus monkeys.

Animals↗
PubMed1992

Dexamethasone increases apolipoprotein A-I concentrations in medium and apolipoprotein A-I mRNA abundance from Hep G2 cells.

Glucocorticoid hormones increase high-density lipoprotein (HDL) levels in vivo. However, there is little known about the mechanism by which glucocorticoids alter HDL metabolism. Hep G2 cells were incubated with dexamethasone to determine the effect of glucocorticoids on apolipoprotein (apo) A-I secretion. Dexamethasone increased apo A-I concentration in a dose-dependent fashion. After 24 hours, 5.5 x 10(-5) mol/L dexamethasone increased apo A-I accumulation in culture medium by 54%. Detectable increases in apo A-I concentration were noted in medium by 5 hours of incubation and persisted up to 48 hours. Cellular apo A-I mRNA concentration increased by 28% after incubation with dexamethasone for 24 hours. The increase in apo A-I mRNA concentration was detectable within 3 hours after incubation with dexamethasone. In contrast, incubation with dexamethasone decreased apo B concentration by 43% in culture medium, but it had no effect on cellular apo B mRNA concentrations. Dexamethasone had little effect on cholesterol and triglyceride accumulation in the medium. Incubation with albumin alone did not affect apo A-I concentration, but it decreased apo B concentration by 30% in the medium. Incubation with albumin and dexamethasone had no effect on apo A-I concentration in medium and had no additive effect on apo B concentration. These data suggest dexamethasone increases secretion of apo A-I by Hep G2 cells by increasing mRNA levels.

Albumins↗
PubMed1992

Recent advances in the understanding of Fusarium trichothecene mycotoxicoses.

Recent concepts in the etiology of Fusarium trichothecene mycotoxicoses have been reviewed. The effect of orally administered trichothecenes on tissue metabolism has been traced from the gastrointestinal tract to the liver and subsequently to blood. It is proposed that the hyperaminoacidemia resulting from trichothecene toxicoses contributes to the behavioral changes observed, including loss of appetite and vomiting. Studies with several species and several trichothecenes have shown that elevated brain tryptophan arising from trichothecene-induced aminoacidemia can subsequently alter regional brain serotonin concentrations. This may produce behaviors such as loss of appetite and muscle incoordination characteristic of the firing of serotonergic neurons. Support is also presented for the concept that other Fusarium metabolites such as fusaric acid may act synergistically with trichothecenes to produce these effects.

Animals↗
PubMed1992

Effect of dietary cysteine supplements on canola meal toxicity and altered hepatic glutathione metabolism in the rat.

Experiments were conducted to determine the effects of feeding canola meal (Brassica campestris and Brassica napus) on the rat hepatic glutathione detoxification system and whether dietary cysteine supplements might modify such effects. Rats were fed test diets for 14 d. Body weight change, feed consumption, hepatic glutathione concentration, and hepatic glutathione-S-transferase (GSH-S-T) activities were determined. Weight gain was decreased when canola meal was fed, whereas hepatic glutathione concentrations increased, as did hepatic GSH-S-T activity. All effects correlated with total glucosinolate concentration in the canola meal. Dietary cysteine supplements, however, did not influence the growth reduction and increased hepatic glutathione concentrations caused by feeding canola meal. Supplemental cysteine prevented the elevation in hepatic GSH-S-T activity. The elevation in hepatic glutathione concentration caused by canola meals was not an overcompensation caused by an initial depletion and therefore reflected a general hepatotoxicity. Feeding supplemental cysteine increased hepatic glutathione levels at early time intervals and delayed the induction of GSH-S-T caused by canola meal toxicity. There was no beneficial effect of supplemental dietary cysteine in overcoming the toxicity of high levels of canola meal, but supplemental cysteine did modify the canola meal-induced changes in hepatic glutathione metabolism.

Animal Feed↗
PubMed1992

Convergence of reflex pathways excited by distension and mechanical stimulation of the mucosa onto the same myenteric neurons of the guinea pig small intestine.

The effects on morphologically and electrophysiologically characterized myenteric neurons of activation of intestinal reflex pathways were examined in vitro. Opened segments of guinea pig small intestine were pinned serosa down in an organ bath that had two balloons set into its base. A 5-10-mm-wide strip of myenteric plexus between the balloons was exposed from the mucosal side, and neurons were impaled with microelectrodes. Reflex pathways were stimulated by inflation of the balloons to distend the intestinal wall, and by deforming the exposed mucosal villi with a brush. Impaled neurons were classified electrophysiologically as AH-neurons or S-neurons (Hirst et al., 1974) and were injected with biocytin to determine their shapes and projections. None of the 58 AH-neurons responded to distension. In contrast, 63 of 131 S-neurons responded to distension with a burst of fast EPSPs; about one-third of the responding S-neurons received input from ascending reflex pathways, one-third received input from descending reflex pathways, and one-third received input from both ascending and descending pathways. Most neurons in this last group supplied extensive varicose branches to the tertiary plexus and were probably longitudinal muscle motor neurons. Neurons receiving input from only one pathway usually projected in the direction of that pathway; many of these were circular muscle motor neurons. Almost all neurons responding to distension were also excited by deforming the villi. Responses evoked by distension or deforming the mucosa declined when stimuli were repeated at intervals less than 10 sec. This was seen in ascending and descending pathways but was more prominent in the former. Deforming the mucosa evoked a normal response even when the response to repeated distensions had disappeared. It is concluded that distension and deforming the mucosa excite separate populations of sensory neurons to activate reflex pathways that converge onto common motor neurons and probably onto common interneurons.

Animals↗
PubMed1991

Active transport of amino acids by gamma-glutamyl transpeptidase through Caco-2 cell monolayers.

The possible role of the gamma-glutamyl cycle in the transport of amino acids, using the Caco-2 cell monolayer as an in vitro model of the small intestine, has been investigated. The transport of [2-3H]glycine and [2-3H]glycylglycine through the Caco-2 monolayer has been shown to occur by two modes of action. Active transport is unidirectional from apical to basolateral region and is a carrier mediated system. The enzyme gamma-glutamyl transpeptidase seems to be involved in this process, since when the enzyme is inhibited, the active transport is also inhibited. However transport still takes place, and this occurs by a slower non-active process, which is bidirectional and is mediated by passive diffusion. The rate of transport of [2-3H]glycylglycine and [2-3H]glycine were 585 (+/- 24) and 287 (+/- 16) pmolcm-2min-1 respectively, while the non-active transport takes place at 87 (+/- 6) pmolcm-2min-1. Thus, amino acid translocation in Caco-2 cells is shown to occur by two methods, one of which involves the gamma-glutamyl cycle.

Amino Acids↗