Transport measurements across Caco-2 monolayers of different organic and inorganic selenium: influence of sulfur compounds.

The transport and uptake of the most common Se compounds, selenate (SeO42-), selenite (SeO3(2-)), selenomethionine, and selenocystine, were investigated using confluent monolayers of Caco-2 cells, a human carcinoma cell line. Comparative measurements were performed in the absorptive (apical to basolateral side) and exsorptive (basolateral to apical side) directions. Apparent permeability coefficients (Papp), calculated from transport experiments in the absorptive direction, showed increasing values in the following rank order: about 1 x 10(6) cm/s < mannitol < SeO3(2-) < or = selenocystine < selenomethionine < SeO4(2-) < or = about 16 x 10(4) cm/s. The ratios of the Papp measured in the absorptive versus exsorptive directions indicated that only the organic forms presented a net polarized transport (Papp ratio >> 1), suggesting the presence of a transcellular pathway. No significant excretion was observed. The transport of selenomethionine was inhibited by its sulfur analog, methionine, suggesting a common transport mechanism. In contrast, an inhibition of the transport of selenocystine by cysteine was not observed. From the two substrates tested, sulfate and thiosulfate, only thiosulfate inhibited the transport of SeO4(2-) . This effect was also observed for SeO32- (i.e., was unspecific), which questioned the assertion of a common transport for sulfate and SeO4(2-) and may confirm the paracellular pathway of SeO42- suggested by the Papp ratio of about 1. The addition of glutathione (GSH) in large excess had no consequence on the passage of SeO3(2-) but strongly increased the uptake (about fourfold). The liquid chromatography - mass spectrometry (LC-MS) data showed that, in the ionic condition of incubation medium, GSH promptly reduced SeO3(2-) (< or = 2 min) in its elemental form Se0, which cannot ascribe to selenodiglutathione a direct role in the effect of GSH.

1,25-dihydroxyvitamin D3 regulates the synthesis of gamma-glutamyl transpeptidase and glutathione levels in rat primary astrocytes.

Astrocytes play a pivotal role in CNS detoxification pathways, where glutathione (GSH) is involved in the elimination of oxygen and nitrogen reactive species such as nitric oxide. We have previously demonstrated that the specific activity of gamma-glutamyl transpeptidase (gamma-GT), an enzyme of central significance in GSH metabolism, is regulated in vivo in astrocytes by 1,25-dihydroxyvitamin D3 (1,25-D3). The aim of the present work was to investigate, in primary cultures of newborn rat astrocytes, the effects of this hormone on gamma-GT synthesis and on GSH and nitrite levels after lipopolysaccharide (LPS) treatment. This study demonstrates that both gamma-GT gene expression and specific activity, induced by LPS, are potentiated by 1,25-D3. In contrast, 1,25-D3 does not regulate the expression of other enzymes involved in astrocyte detoxification processes, such as superoxide dismutase or GSH peroxidase. In parallel, 1,25-D3 enhanced intracellular GSH pools and significantly reduced nitrite production induced by LPS. Taken together, these results suggest that gamma-GT, GSH, and 1,25-D3 play a fundamental role in astrocyte detoxification pathways.

Manganese transport by Caco-2 cells.

The uptake and transport kinetics of manganese (Mn) were investigated in the human intestinal Caco-2 cell line both from the absorption side (apical to basolateral) and from the exsorption side (basolateral to apical). With regard to the former, transport versus time revealed (as uptake) a biphasic pattern with an initial transient phase followed by steady-state conditions. Uptake versus Mn concentrations showed saturation-type kinetics with a 100% increase of Mn binding capacity when measurements were made from 0.5 to 2 h of incubation. The transport characteristics in steady-state conditions exhibited two components, saturable (Vmax = 3.70+/-0.07 nmol/cm2/h, K(m) = 32.2+/-3.4 microM) and nonsaturable (slope = [1.4+/-0.2] x 10(-6)cm(-2)/h) usually presumed to reflect transcellular (carrier mediated) and paracellular (diffusional) pathways, respectively. Mn fluxes were decreased by calcium and calcium antagonists, almost 100% inhibited at 4 degrees C, and affected by quinacrine and ouabain. The inhibition of ATP synthesis was apparently ineffective. From the exsorption side, the Mn fluxes, without a transient period, had an approx 20-fold smaller rate than in the absorptive direction and showed mainly a nonsaturable route (slope = [0.6+/-0.1] x 10(-6) cm(-2)/h). The mechanisms participating in the Mn movements through the monolayer are discussed and proposed to be in common, at least partly, with other divalent cations such as calcium, zinc, or iron.

Influence of thyroparathyroidectomy and thyroxine replacement on Cu and Zn cellular distribution and on the metallothionein level and induction in rats.

Thyroid hormones are involved in copper and zinc distribution in rat tissues. We examined the influence of thyroparathyroidectomy (TPTY) and of a replacement therapy by T4 on Cu and Zn organ distribution. MT levels were also measured both in basal conditions and after induction by cadmium. The results confirm that a lack of T4 modified Cu and Zn in serum and tissues. In serum, TPTY increased Cu (+15%) and ceruloplasmin (+18%), and decreased Zn (-18%). In tissues, Cu was altered in liver (+13%), kidney (-24%), heart (-16%) duodenum (-18%), and Zn in liver (+25%) and kidney (-10%). The soluble fractions (100,000 g supernatant) were mainly affected in liver and kidney, and the subcellular fractions in heart and duodenum. MT levels were modified in basal conditions only in liver (+57%) and kidney (-36%). T4 administration partially prevented the effect of TPTY on both elements and MT concentrations. Therefore, no evidence is provided for a direct role of T4 in the metabolism of MT in a way comparable to the effects of glucocorticoids. However, MT could mediate the consequences of TPTY on metal distribution in certain organs, such as liver and kidney.

Effects of thyroparathyroidectomy and of thyroxin and calcitonin on the tissue distribution of twelve elements in the rat.

The effects of thyroparathyroidectomy (TPTY) and of replacement therapy using thyroxin (T4) and calcitonin (CT) on the tissue distribution of elements were studied in the rat under semichronic conditions. The elements Na, K, Ca, Mg, Fe, S, P, Rb, Sr, Mn, Cu, and Zn were determined in whole blood, plasma, brain, liver, heart, kidney, skeletal muscle, and bone. TPTY modified concentrations of all elements tested but only small changes were observed for K, Mg, S, and P. The mineral bone composition was slightly modified, 28 d after TPTY, whereas plasma was the most altered. The consequences of TPTY were corrected fairly well by T4 for Na, Cu, Zn, Fe, and S, and by CT for K, P, Rb but with less efficiency for Ca. This study revealed that hormones of the thyroid gland, mainly T4, play an important role in the plasma and tissue balance of elements. It is suggested that T4 participates in tissue fixation of Cu, Zn, and Fe and that CT influences phosphoremia and cellular Ca binding.

The effects of captopril and enalapril on tissue levels of various elements in spontaneously hypertensive rats.

The effects of two inhibitors of angiotensin I converting enzyme, captopril and enalapril, on the concentrations of Na, K, Ca, Mg, Fe, S, P, Sr, Mn, Cu and Zn ions in blood, plasma, heart, skeletal muscle, liver and kidney of spontaneously hypertensive rats (SHR) were studied. Captopril and enalapril were given by the intraperitoneal route for 15 days, at 160 mumols/kg/d and 40 mumols/kg/d, respectively. Elements in tissues were determined by inductively coupled plasma emission spectrometry with a JY 48 instrument. The common changes produced with the two drugs were: a decrease of Na in muscle (-10%), a decrease of Ca in plasma and kidney (less than -10%) and a decrease of Mn in liver (-15%). The main effects observed with only one of the two drugs were: an increase of Cu in plasma (+26%) with captopril, and increases of Sr in heart (+56%), muscle (+79%) and liver (+74%) with enalapril. Zinc concentration in tissues was not modified, except for an increase in liver with captopril (+13%) and a decrease in heart with enalapril (-11%).

Effect of aluminum and deferoxamine on biliary iron elimination in the rat.

Iron (Fe) and aluminum (Al) eliminations in bile were studied in rats after intravenous administration of Fe, Al, deferoxamine mesylate (Desferal, Ciba) (DFA), feroxamine (FeA), and aluminoxamine (AlA) at the dose of 50 mumole/kg body weight. Bile was obtained from the bile duct of anesthetized rats, and the concentrations of Fe and Al in bile were measured by an inductively coupled plasma optical emission spectrometer. The results showed an increase of Fe elimination in bile, from 10 to more than 20 mumole/liter after Fe and also after Al administration; an increase to about 350 mumole/liter after DFA administration; to 250 mumole/liter after FeA administration; and to 100 mumole/liter after AlA administration. Aluminum elimination in bile was increased only after Al and particularly after AlA administration but not after Fe and FeA administration. In conclusion, Al and AlA were able to increase Fe elimination in bile. Thus Al overload observed in hemodialyzed patients may induce an excessive iron loss in bile and partly explain microcytic anemia.

Distribution of 14 elements in various rat tissues following hypophysectomy, thyroparathyroidectomy, adrenalectomy, and castration.

The tissue distribution of 14 elements was simultaneously determined in rats 28 d after hypophysectomy (HPY), thyroparathyroidectomy (TPTY), adrenalectomy (ADY), and castration (CTN). The elements Na, K, Ca, Mg, Fe, S, P, Rb, Sr, Mn, Cu, and Zn were investigated in whole blood, plasma, brain, liver, kidney, heart, skeletal muscle, and bone. Additionally Mo was determined in kidney and liver. The following results were obtained: 1) With regard to hormone deficiency: HPY induced the most noticeable variations on all the elements tested owing probably to the direct and indirect effects of adenohypophyseal hormones. ADY led to the expected modification of Na and K but also to a Sr accumulation and a Rb depletion. TPTY induced a sharp decrease in plasma and tissues Ca, an increase in plasma P, but did not disturb the two elements in bone. An increase of Rb in many tissues and of Fe in heart, kidney, and liver were also observed. CTN had little consequences except in bone whose Cu and Fe contents were increased; 2) With regard to element variations: K, Mg, and S underwent little change. Discriminations were revealed between elements such as K and Rb, Ca and Sr, Ca and Mg, and Cu and Zn. The changes of Rb and Sr were consistent with regulatory mechanisms. The accumulation of Fe and Cu in tissues such as liver after HPY, TPTY, and ADY, suggest that the hormonal deficiencies could worsen the hemochromatosis and Wilson's disease; 3) With regard to plasma and tissues: No correlation appeared in element levels between plasma and other tissues. Brain was the least affected and liver, kidney and bone the most.

Effects of adrenalectomy and of adrenal hormones on the tissue distribution of 14 elements in the rat.

The effects of adrenalectomy (ADY) and of replacement therapy using a mineralocorticoid, deoxycorticosterone (DOC) and a glucocorticoid, dexamethasone (DEX) on the tissue distribution of elements in the rat, were studied under semichronic conditions. The elements Na, K, Ca, Mg, Fe, S, P, Rb, Sr, Mn, Cu, and Zn were determined in whole blood, plasma, brain, liver, kidney, heart, skeletal muscle, spleen, thymus, and bone. Additionally, Mo was determined in kidney and liver and Ba in bone. ADY modified concentrations of all elements tested. Small changes were observed for K, Mg, Ca, S, and P, whereas much larger changes were noted for Na, Rb, and Sr. Cu, Zn, and Fe were mainly modified in liver and kidney, organs involved in storage and/or elimination. The consequences of ADY were corrected fairly well by DEX for Mg, Mn, Ca, Cu, and Mo; by DOC for Na and K, and by the two corticoids for Zn, Fe, Sr, and Rb. This study revealed that corticoids, mainly glucocorticoids, play an important role in the plasma and tissue balance of elements. It is suggested that these results may have a pathological and clinical significance.

Altered element concentrations in tissues of spontaneously hypertensive rats.

The involvement of elements in the pathological process of primary hypertension has been established. The tissue distribution of 12 elements was studied in spontaneously hypertensive rats (SHR) and normotensive homologous rats (WKY). A multi-element analytical technique allowed simultaneous determination of sodium, potassium, calcium, magnesium, strontium, rubidium, manganese, copper, zinc, iron, sulphur and phosphorus in blood, plasma, brain, liver, kidney, skeletal muscle, heart and bone. Most elements were modified in SHR, except Ca, Rb and S. In plasma, an increase in Cu (+22%) and a decrease in K (-8%), Mg (-15%) and P (-11%) were observed. These variations, qualitatively similar to those found in man, suggest that the results in animal tissues could be extrapolated to man. Modifications were observed in all the tissues tested. Among them significant variations were noted in Na (+18%), Mn (+12%) and Cu (+29%) in kidney, and in K (+5%), Mg (+9%), Sr (-29%) and Zn (+14%) in heart. The role of these plasma and tissue variations in hypertension is discussed, as well as the possible involvement of the hypertensive process and/or hormones.

Tissue distribution of some elements in rats.

The concentrations of 14 elements were determined in rat whole blood, plasma, brain, heart, skeletal muscle, liver, kidney, spleen, thymus, and bone, using multielement ICP optical emission spectrometry. The tissue to plasma concentration ratios were calculated in order to determine tissue distribution and its homogeneity. The results show the likely sequences of element concentrations in tissues: RB>P≥K>(Fe)>Zn≥Mg>Cu>S>(Sr≥Ca>Na); and of the homogeneity of the distribution: (Fe)

Modification of tissue distribution of trace elements by cyclosporin in rats.

Cyclosporin-induced hypomagnesaemia, as observed in patients, could also be induced in rats by intramuscular administration of the drug (20 mg/kg/d) for 12 d. moreover, cyclosporin administration induced modifications in the concentrations of other elements in tissues, particularly an increase in Mg, Cu, and Zn in the thymus and an increase in Mo in the spleen and Sr in the liver.

A thiol oxidation interpretation of the Cu2+ effects on rat liver mitochondria.

A comparative study of the Cu2+ effects, binding and reduction, has been performed on rat liver mitochondria. In the first minutes, Cu2+ (less than or equal to 50 micron) is massively bound and reduced to the extent of 70%-80% while a simultaneous activation of respiration takes place. Then the remaining 20% or so of Cu2+ are progressively bound and reduced while respiratory inhibition, Ca2+ and Mg2+ effluxes, and swelling are observed. EDTA, used as a copper chelator, prevents or reduces the copper effects and removes part of the bound copper, according to the time of introduction in the incubation medium after Cu2+. The results suggest that the two steps of the copper binding and the effects following involve mainly first the outer (cytosol side) proteins of the inner membrane and then those of the inner membrane. 100 microM dithiothreitol and 100 microM glutathione used as antioxidant thiol reagents prevent, as does EDTA, but do not reverse the 25 microM copper effects. They also decrease the copper binding; however, no relationship between binding and preventive action is observed. It is shown that glutathione and dithiothreitol have a specific potent ability to reduce Cu2+, which explains that in presence of these reagents copper may react with mitochondria partly or entirely in the form of Cu+. These findings suggest that Cu2+ in its Cu+ form has no mitochondrial effect. A mechanism of copper action involving oxidation of some membrane thiol groups is discussed.

Ca2+ uptake and energy supply of sheep heart mitochondria in presence of some calcium antagonists.

The action of the calcium antagonists nifedipine, bepridil, diltiazem and of an alpha adrenergic antagonist, prazosin, was studied on Ca2+ uptake and energy supply (i.e. oxidative phosphorylation) of sheep heart mitochondria. In order to appreciate an inhibition, uncoupler like, of the Ca2+ influx, a comparative study was made with DNP. The Ca2+ uptake was inhibited, at the concentrations tested, (less than or equal to 10(-3) M), only by bepridil, nifedipine and DNP: a 2.5.10(-4) M concentration of these drugs brought an inhibition of 95%, 65% and 100% respectively. The inhibition was accompanied by a decrease of the ADP:O ratio and the RCI except with nifedipine. It is suggested that bepridil could act more as an inhibitor-uncoupler and nifedipine as an inhibitor of the Ca2+ uptake uniporter; both leading to a decrease of the mitochondrial participation in the regulation of the cellular homeostasis. This phenomenon could contribute to the known clinical properties of bepridil and nifedipine.

Blood and brain aluminium concentrations in mice after intra-pertioneal injection of different aluminium compounds.

Aluminium chloride, nitrate, lactate and gluconate was injected by intraperitoneal route to mice at equimolecular doses, equivalent to 54 and 5.4 mg/kg of metal. The blood aluminium concentrations were increased to very high values with gluconate (99.5 mg/l) to high values with lactate (4.5 mg/l) and to low values with nitrate and chloride (0.3 mg/l). Brain aluminium concentration was increased only with glutconate at the dose of 54 mg/kg. The quantity of aluminium found in blood and brain after intraperitoneal administration at equimolecular doses is very dependent of the preparation of aluminium used.

[Anti-inflammatory effect of aluminum or aurin tricarboxylic acid]. / Mise en évidence de l'action anti-inflmmatoire de l'aluminon ou acide aurine tricarboxylique.

The anti-inflammatory effect of aurin tricarboxylic acid is studied in rats after parenteral injection of 20 and 50 mg/kg. Aurin tricarboxylic acid reduces: the carrageenan-induced oedemas in paws, the carragenan-induced pleural effusion, the carragen-induced granulation tissue growth and the Freund adjuvant-induced arthritis. As non steroïdal antiinflammatory drugs, the aurin tricarboxylic acid inhibits (ED50 = 0.34.10-2 M) the human erythrocyte delta aminolevulinic acid dehydratase, a zinc dependent enzyme. A possible interaction between zinc-enzymes and non steroïdal antiinflammatory compounds is suggested.