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A Felipe

Publications and source records attributed to A Felipe.

At least 55 records · Page 3Linked to original sources
PubMed1995

Phospholipid and fatty acid composition in stored sheep erythrocytes of different densities.

The mammalian erythrocyte is an ideal model for studies of membrane aging under conditions of storage. The present study describes the variations in the membrane lipid composition of three density groups (light, 1.110 < d < 1.125; intermediate, 1.125 < d < 1.130, and dense, 1.130 < d < 1.140) of sheep erythrocytes separated by centrifugation in a discontinuous Ficoll density gradient after storage at 4 degrees C in a nutrient medium for up to 6 days. The only changes in phospholipid composition took place in the erythrocyte light fraction where sphingomyelin (SM) and phosphatidic acid increased (p < 0.05), whereas phosphatidylethanolamine (PE) decreased (p < 0.05). Moreover, polyunsaturated fatty acids (20:4 and 22:6) decreased during storage, whereas lipid fluorescence increased (p < 0.01) after 24 hours of storage in all the fractions separated. These observations suggest a lipid peroxidation process in all three erythrocyte groups during storage.

Animals↗
PubMed1995

[In vitro sensitivity of Mycobacterium chelonae strains to various antimicrobial agents].

The in vitro susceptibility of 32 Mycobacterium chelonae strains to 10 antimicrobial agents was determined. The sources of the different strains were: clinical samples from patients treated at the Hospital Universitario de Canarias and Hospital del Tórax (General and Chest facilities) and from environmental sources (water supply, sewage, swimming pools and the sea). The susceptibility tests were performed by a broth microdilution method (Mueller-Hinton Broth). The results showed amikacin as the most effective antimicrobial agent against M. chelonae isolates, then ofloxacin and cefoxitin. However no statistical difference was detected among them. The least effective was imipenem, followed by ciprofloxacin and norfloxacin.

Amikacin↗
PubMed1994

Primary structure and differential expression during development and pregnancy of a novel voltage-gated sodium channel in the mouse.

Until recently, all cloned vertebrate voltage-dependent sodium channels exhibited high sequence homology to one another and appeared to comprise a single multigene subfamily. An exception is the human Nav2.1 channel proposed to represent a second Na+ channel (NaCh) gene subfamily since comparison with previously cloned voltage-gated NaChs revealed only 40-45% identity. We have now cloned a mouse NaCh (mNav2.3) from an atrial tumor cell line that shows high amino acid sequence identity to hNav2.1 in functionally relevant regions such as the pore-forming segments, S4 segments, and inactivation gate sequence. Overall sequence identity is 68%. mNav2.3 mRNA was most abundant in heart and uterus, and the transcript levels in heart, brain, and skeletal muscle were differentially regulated during development. Transcript levels in heart were greatest immediately after birth. mNav2.3 transcript levels in pregnant uterus increased 3-fold between day 15 of pregnancy and birth and then declined 15-fold during the 2 days following delivery. The mNav2.3 amino acid sequence indicates that the Nav2 NaCh gene subfamily is well conserved across species, and the tissue-specific and developmental regulation of mRNA expression suggests these channels play important physiological roles in cardiac and uterine muscle.

Aging↗
PubMed1994

Coordinate induction of Na(+)-dependent transport systems and Na+,K(+)-ATPase in the liver of obese Zucker rats.

Solute uptake into liver plasma membrane vesicles from either lean or obese Zucker rats was monitored. D-Glucose and L-leucine uptakes at physiological concentrations of the substrate were not different in lean and obese Zucker rats. In agreement with a previous report (Ruiz et al. (1991) Biochem. J. 280, 367-372) L-alanine uptake was significantly enhanced in those preparations from obese animals. Na(+)-coupled uridine transport was markedly enhanced also in obese rats. The effect was due to an increase in Vmax (5.5 +/- 0.6 vs. 2.1 +/- 0.2 pmol/mg protein per 3 s, P < 0.01) without any significant change in Km (11.0 +/- 2.8 vs. 9.0 +/- 2.7 microM for obese and lean rats, respectively). Na+,K(+)-ATPase activity was also higher in liver plasma membrane vesicles from rat liver and it correlated with a higher amount of alpha 1-subunit protein in both, plasma membrane vesicles and homogenates from obese rat livers. In summary, in the hypertrophic liver of obese Zucker rats a coordinate induction of several Na(+)-dependent transport systems occurs and, in order to sustain the metabolic pressure associated with this adaptation, a significant induction of the Na+,K(+)-ATPase expression is also found. These data also provide new evidence for regulation of the recently characterized Na(+)-dependent nucleoside transporter.

Animals↗
PubMed1994

Evidence for a regulatory protein involved in the increased activity of system A for neutral amino acid transport in osmotically stressed mammalian cells.

System A for neutral amino acid transport is increased by hypertonic shock in NBL-1 cells previously induced to express system A activity by amino acid starvation. The hypertonicity-mediated effect can be blocked by cycloheximide but is insensitive to tunicamycin. The activity induced may be inactivated irreversibly by the addition of system A substrates, by a rapid mechanism insensitive to cycloheximide. In CHO-K1 cells, hypertonicity increases system A activity, as has been shown in NBL-1 cells. This effect is additive to the activity produced by derepression of system A by amino acid starvation and is insensitive to tunicamycin. Furthermore, the alanine-resistant mutant CHO-K1 alar4, which bears a mutation affecting the regulatory gene R1, involved in the derepression of system A activity after amino acid starvation, is still able to respond to the hypertonic shock by increasing system A activity to a level similar to that described in hypertonicity-induced derepressed CHO-K1 (wild type) cells. These results suggest (i) that the hypertonicity-mediated increase of system A activity occurs through a mechanism other than that involved in system A derepression and (ii) that a regulatory protein coded by an osmotically sensitive gene is responsible for further activation of preexisting A carriers.

Amino Acids↗
PubMed1994

Up-regulation of liver system A for neutral amino acid transport in euglycemic hyperinsulinemic rats.

To determine the role of insulin on the in vivo modulation of liver system A activity, we used the euglycemic hyperinsulinemic clamp coupled to the measurement of solute uptakes into plasma membrane vesicles partially purified from livers of hyperinsulinemic rats and their saline-infused controls. The clamp was performed in chronically catheterized rats, either in the fasted state, 24 h after surgery (Group I), or after 3 days of recovery (Group II). System A activity, measured as the MeAIB-inhibitable L-alanine uptake, was selectively induced by hyperinsulinemia, although the effect was much greater in Group II than in Group I rats (137% vs. 24% over the basal values, respectively). This might be explained by the higher basal levels found in those liver plasma membrane vesicles from Group I fasted animals. Hyperinsulinemia also decreased blood amino acids but to a similar extent in both experimental groups. This suggests that amino acid depletion by itself may not cause up-regulation of system A. Other transport activities involved in neutral amino acid transport (Systems ASC, N and L) were not modified by the clamp. The induction of system A cannot be explained by changes in the dissipation rate of the Na+ transmembrane gradient, because the differences between insulin- and saline-infused rats remained even when the electrochemical Na+ gradient was disrupted in the presence of monensin. Thus, hyperinsulinemia might induce an increase in the number of transporters inserted into the plasma membrane.

Amino Acid Transport Systems↗
PubMed1994

Differential expression of Isk mRNAs in mouse tissue during development and pregnancy.

The molecular isoform of the cDNA clone Isk present in the AT-1 atrial tumor cell line was characterized by molecular cloning of Isk cDNA. Since Isk mRNA was found in mouse heart, kidney, and uterus, a complete study of its expression during development in the heart and kidney was performed, in addition to its expression in the uterus during pregnancy. In the heart, Isk showed a 4-fold upregulation during the perinatal period followed by a 20-fold decrease between birth and the adult state. Furthermore, the two 0.9- and 3.4-kb transcripts were differentially regulated after birth. In the kidney, Isk progressively increased 10-fold, reaching steady-state adult values at 21 days. Isk mRNA levels in the uterus increased threefold at late pregnancy and decreased sixfold rapidly after birth. The Isk gene is differentially expressed during development in kidney and cardiac tissue, and both Isk transcripts appeared to be differentially regulated. Furthermore, the drastic changes in transcript levels before delivery and after birth suggest that Isk plays a significant role in myometrium during late pregnancy and delivery.

Amino Acid Sequence↗
PubMed1994

K+ currents and K+ channel mRNA in cultured atrial cardiac myocytes (AT-1 cells).

Atrial tumor myocytes derived from transgenic mice (AT-1 cells) maintain a well-differentiated cardiac biochemical and histological phenotype. In addition, they beat spontaneously in culture and exhibit long action potentials whose repolarization resembles that observed in native mammalian myocytes. In this study, we identified the major depolarization-activated outward currents in AT-1 cells; also, the presence of mRNAs that encode outwardly conducting ion channels was determined by cloning from an AT-1 cDNA library or by Northern hybridization. Among K+ channel isoforms, Kv2.1, minK, and Kv1.4 were readily detected in tumors and at 1 day in culture. Their abundance remained relatively stable (twofold or less change) after 14 days. The major outward current in AT-1 cells is a delayed rectifier that displays prominent inward rectification, activates rapidly (eg, 182 +/- 27 milliseconds [mean +/- SEM] at + 20 mV, n = 12), exhibits biexponential deactivation kinetics, and is extremely sensitive to the methanesulfonanilide dofetilide (IC50, 12 nmol/L). These characteristics identify this current as IKr, a delayed rectifier observed only in cardiac cells. IKr in AT-1 cells displayed slow inactivation: dofetilide-sensitive deactivating tails were greater after 1-second than after 5-second pulses. When IKr was blocked by > or = 0.5 mumol/L dofetilide, time-independent current was usually recorded (50 of 65 experiments); rapidly inactivating (6 of 65) or slowly inactivating (9 of 65) outward currents were occasionally observed. We conclude that AT-1 cells express mRNAs encoding cardiac K+ channels and display a cardiac electrophysiological phenotype.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Contrast sensitivity of the visual system in speckle imagery.

The contrast sensitivity function (CSF) of the whole visual system is determined with the use of coherent diffuse illumination. This function provides supplementary data about the effect of speckle on the ability of the visual system to perceive the spatial information contained in an image. The results show that speckle not only prevents perception of the finest details (highest frequencies) but also reduces the visibility of lower frequencies (especially where contrast is low). The difference between the CSF's determined with and without speckle is quantitatively very important. And the ratio between the two CSF's is a measure of the retinal ability to perceive contrast in the presence of speckle noise. The influence of the pupil size and luminance level on the CSF with speckle is studied and compared with the influence of the same parameters on the classical CSF.

Contrast Sensitivity↗
PubMed1994

Epidermal growth factor increases protein and messenger RNA expression levels of Ras GTPase activating protein.

The Ras GTPase activating protein (RasGAP) is a key regulatory enzyme in the Ras signaling pathway, which is crucial for growth factor-induced mitogenesis. In this study, it is shown that epidermal growth factor (EGF) increases RasGAP protein expression by 100-150% in NIH 3T3 cells, which overexpress the human EGF receptor, and mouse keratinocytes but does not increase RasGAP protein expression in A431 cells, where EGF does not have a mitogenic effect. In contrast, EGF does not affect the expression of other signal transduction SH2-containing proteins, such as phospholipase C gamma 1 and the p85 subunit of phosphatidylinositide 3-kinase. The growth factor-induced increase of RasGAP protein parallels an increase in RasGAP activity. EGF stimulates RasGAP protein synthesis and does not affect its degradation rate. The mechanism for RasGAP protein induction by EGF involves an increase in rasGAP mRNA levels. The growth factor-stimulated up-regulation of rasGAP is a delayed response, since the increase in the mRNA levels and protein synthesis rate begin after 3 h and reach maximal values between 9 and 24 h of growth factor treatment. EGF fails to increase rasGAP mRNA levels in the presence of cycloheximide, suggesting that this effect is dependent on de novo protein synthesis. However, cycloheximide alone is able to increase by 6-fold rasGAP mRNA expression. Since EGF does not modify rasGAP mRNA stability, the increase in rasGAP mRNA expression is likely due to an increase in transcription of the rasGAP gene.

Animals↗
PubMed1993

Changes in the fatty acid composition of stored erythrocytes from sheep of different ages.

This study examined the profile in sheep erythrocyte fatty acids from animals of different ages during storage at 4 degrees C in a nutritive medium for up to 6 days. The changes found in the fatty acyl profile were a decrease (P < 0.01) in the percentage of arachidonic acid and an increase (P < 0.01) in the percentage of minor fatty acids (representing < 2% in each case; 20:2, 22:0 and 22:1) with respect to fresh erythrocytes in all age groups. However, the saturated/unsaturated ratios and unsaturated index started almost constant in all cases. The changes observed occurred after 24-48 h of storage, with significant increases (P < 0.01) in the fluorescence detected in the lipid extracts from stored erythrocytes during this period. The above findings suggest peroxidative damage and changes in the erythrocyte lipid membrane during storage.

Aging↗
PubMed1993

Up-regulation of system A activity in the regenerating rat liver.

System A activity for neutral amino acid transport, measured as the MeAIB-sensitive Na(+)-dependent L-alanine uptake, is induced 6 h after partial hepatectomy in plasma membrane vesicles from rat livers. Other Na(+)-dependent transporters, like system ASC (MeAIB-insensitive Na(+)-dependent L-alanine transport) and the nucleoside carrier show similar inductions. Up-regulation of system A is not explained by changes in the dissipation rate of the Na+ transmembrane gradient, as deduced from uptake measurements performed in the presence of monensin. To determine whether induced system A shared any similarity with the activity found in hepatoma cell lines, we analyzed the N-ethylmaleimide (NEM) sensitivity of system A in both regenerating and control rat liver plasma membrane vesicles. NEM treatment was equally effective in inhibiting system A in both experimental groups. Thus, during the prereplicative phase of liver growth, a transport activity similar to basal system A is up-regulated in liver parenchymal cells, by a stable mechanism that does not involve changes in the Na+ transmembrane gradient.

Alanine↗
PubMed1993

L-alanine uptake by rat liver parenchymal and haematopoietic cells during the perinatal period.

Alanine disposal by liver parenchymal and haematopoietic cells from 21-day fetuses, newborns and adult rats was studied. Preparations selectively enriched in either haematopoietic cells or hepatocytes were obtained by direct perfusion of fetal- and neonatal-rat livers. L-Alanine transport into liver parenchymal cells was best fitted to two Na(+)-dependent saturable systems. The high-affinity system showed a much higher activity (Vmax.) in hepatocytes from fetuses and newborns than in those from adult rats (2.4, 4.3 and 0.3 nmol/8 min per 10(6) cells for fetuses, newborns and adults respectively). Vmax. for the low-affinity component was slightly lower during the perinatal period than in the adult (about 30 nmol/8 min per 10(6) cells for hepatocytes from fetuses and newborns, versus 48 nmol/8 min per 10(6) cells for adult rat parenchymal cells). Haematopoietic cells from fetal-rat livers showed significant Na(+)-dependent L-alanine uptake which was completely abolished after birth. These results show that the transport systems involved in L-alanine uptake by liver parenchymal cells are fully developed before birth. This probably contributes to fulfilling the high requirement for neutral amino acids for protein synthesis during development. Haematopoietic cells may play an important role in liver amino acid metabolism during fetal life.

Alanine↗
PubMed1993

Hyperosmolarity leads to an increase in derepressed system A activity in the renal epithelial cell line NBL-1.

Hyperosmolarity induced an increase in Na(+)-dependent L-alanine uptake in confluent monolayers of the established renal epithelial cell line NBL-1. This induction was attributable to system A and was only seen when the cells had been previously deprived of amino acids in the culture medium to derepress system A activity. It was additive to the adaptive regulation induction, and both were inhibited by cycloheximide. However, the hyperosmolarity effect was inhibited by colcemid (an inhibitor of microtubular function), but adaptive regulation was not. Otherwise, when cell monolayers were incubated in a control medium, basal Na(+)-dependent L-alanine uptake mediated by system B0 decreased. The results of this study show that: (i) system A activity was not induced by cell shrinkage and subsequent swelling due to extracellular hyperosmolarity when cells were incubated in control medium; (ii) previous expression of system A activity induced by amino acid starvation seems to be a prerequisite for further induction due to hyperosmolarity; and (iii) the effects of adaptive regulation and hyperosmotic stress are mediated by different mechanisms.

Alanine↗
PubMed1993

Early induction of Na(+)-dependent uridine uptake in the regenerating rat liver.

Na(+)-dependent uridine transport into liver plasma membrane vesicles from partially hepatectomized and sham-operated rats was studied. Preparations purified 6 h after 70% hepatectomy exhibited an increased Vmax of uridine uptake (3.7 vs. 1.4 pmol/mg prot/3 s) without any change in Km (6 microM). Incubation of the vesicles in the presence of monensin decreased uridine uptake although the differences between both experimental groups remained identical. It is concluded that uridine transport is induced early after partial hepatectomy by a mechanism which does not involve changes in the transmembrane Na+ gradient. This is the first evidence in favor of modulation of nucleoside transport into liver cells.

Animals↗
PubMed1993

Alanine uptake by liver of mid-lactating rats.

L-Alanine transport in liver plasma membrane vesicle preparations from fed virgin and 15-day-lactating rats was studied. Lactation was found to induce a decrease of the maximal rate (Vmax) of a high-capacity-low-affinity component of the Na(+)-dependent L-alanine uptake. However, a high-affinity-low-capacity agency was significantly induced in lactating-rat livers. L-Alanine uptake was differentially inhibited by other amino acids in those preparations from lactating rats, and showed different sensitivity to Li+ as a cosubstrate instead of Na+ and to inhibition by sulfhydryl modifying reagents (N-ethylmaleimide [NEM] and p-chloromercuribenzosulfonate [PCMBS]). All of these observations taken together suggest that system A is upregulated in lactating-rat livers, thus resulting in a different contribution of both agencies A and ASC to the total Na(+)-dependent alanine transport into liver plasma membrane vesicles. This was demonstrated using the analogue alpha-methyl-aminoisobutyric acid (MeAIB), a specific system A substrate. L-Alanine uptake rates, as calculated from plasma membrane enzyme marker recoveries, were also enhanced in the physiologic range of alanine concentrations in blood. Our results prove that the physiologic adaptation to lactation involves modulation of system A activity in the liver.

4-Chloromercuribenzenesulfonate↗
PubMed1993

Influence of the contrast sensitivity function on the reaction time.

The reaction time (RT) vs spatial frequency (SF) curve is determined, using gratings from 1 to 40 c/deg, at seven different contrast levels between 0.95 and 0.02. The form of the RT/SF function: (a) replicated the inverse of the contrast sensitivity function (CSF) at near threshold contrast levels; (b) behaved differently at higher contrasts, exhibiting two branches at contrast close to 1. The interpretation is that there are two factors determining this function: (1) the transition from the operation of fast transient channels at low SF to the operation of slow sustained channels at high SF, the transition taking place within a narrow SF band close to 6 or 8 c/deg (depending on the subject) and (2) the contrast attenuation by the optical and neural transfer function, operating throughout the SF range. At high contrasts, the effect of the first factor can be clearly observed, because the effect of the second factor does not change with spatial frequency except in a region where the RT/SF function changes rapidly. At lower contrasts, however, the second factor becomes increasingly relevant while the first becomes less and less observable.

Contrast Sensitivity↗