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Biomedical subjects

A Pascual

Publications and source records attributed to A Pascual.

At least 181 records · Page 10Linked to original sources
PubMed1988

Cholera toxin affects nuclear ADP-ribosylation in GH1 cells.

Incubation of GH1 cells with cholera toxin for 24 h inhibits [32P]ADP-ribose incorporation into histones and non-histone nuclear proteins by more than 50%. The toxin produces a generalized decrease of incorporation into all protein acceptors and into the poly(ADP-ribosyl)ated components excised from chromatin after micrococcal nuclease digestion. The cellular levels of NAD were also decreased (40 to 80%) after treatment with cholera toxin. The inhibition of poly(ADP-ribosyl)ation is preceded by an increase of [32P]ADP-ribose incorporation, since incubation with the toxin for 3 h caused an increase instead of a decrease of incorporation. Incubation with dibutyryl cyclic AMP for 24 h also inhibited nuclear poly(ADP-ribosyl)ation, thus showing that the effect of cholera toxin might be mediated by cyclic AMP.

ADP Ribose Transferases↗
PubMed1988

Triiodothyronine (T3) induces neurite formation and increases synthesis of a protein related to MAP 1B in cultured cells of neuronal origin.

Neuroblastoma (N2A) cells were found to develop axon-like neurite extensions when grown in the presence of triiodothyronine (T3), while C6 cells (of glial origin) did not. Analysis of radiolabelled protein synthesis showed that, in N2A only, T3 increased the synthesis of a polypeptide corresponding in electrophoretic mobility to the microtubule-associated protein MAP 1B. Immunoblotting of total cell proteins with a monoclonal antibody confirmed that this polypeptide was immunologically related to MAP 1B. Further studies using indirect immunofluorescence with monoclonal antibodies against both tubulin and MAP 1B showed that both antigens were present in neurites. Taken together, these results suggest that T3 may control maturation of neural tissue via effects on the microtubule-associated proteins in cells of neuronal origin.

Animals↗
PubMed1988

Surface hydrophobicity and opsonic requirements of coagulase-negative staphylococci in suspension and adhering to a polymer substratum.

The opsonic requirements for phagocytosis in suspension of 38 clinical isolates of coagulase-negative staphylococci recovered from neonates with septicemia were found to be related to the degree of surface hydrophobicity of these strains. Sixteen isolates were adequately opsonized only in the presence of complement; this group was significant more hydrophobic (p less than 0.001) than the 22 strains not requiring complement for efficient uptake in suspension. Evidence showed that hydrophobic groups present on the bacterial surface may interfere with IgG opsonization. In contrast, IgG sufficed as an opsonin without complement being required for the efficient uptake of these hydrophobic strains when adhering to a polymer surface.

Bacterial Adhesion↗
PubMed1988

Mechanism of L-triiodothyronine (T3) uptake by glial C6 cells: regulation by butyrate.

The mode of entry of triiodothyronine (T3) and its regulation by butyrate was studied in cultured glial C6 cells. Uptake of [125I]T3 increases for at least 60 min in C6 cells. The amount of cell-associated radioactivity is 2- to 4-fold higher during the entire time-course in cells previously exposed to 2 mM butyrate for 48 h. Uptake was non-saturable since uptake velocity was linearly related to the extracellular hormone concentration between 0.2 and 800 nM T3 in control and butyrate-treated cells. Uptake velocity increased by more than 3-fold in the cells incubated with the fatty acid. T3 uptake was temperature dependent and the effect of butyrate was observed at the different temperatures examined. Preincubation with metabolic inhibitors did not block [125I]T3 uptake in either group, and monodansylcadaverine was also ineffective. Present results suggest that in C6 cells T3 uptake proceeds by a passive, energy-independent, non-saturable process, that is markedly affected by short-chain fatty acids. Additionally, this is the first study documenting that a natural compound directly influences the entry of thyroid hormones into cells.

Animals↗
PubMed1988

Comparison of the effects of forskolin and dibutyryl cyclic AMP in neuroblastoma cells: evidence that some of the actions of dibutyryl cyclic AMP are mediated by butyrate.

We have compared the effects of forskolin, N6,2'-O-dibutyryladenosine 3':5'-cyclic monophosphate (dibutyryl cyclic AMP, Bt2-cAMP), and butyrate on several aspects of neuroblastoma cell physiology. The morphology of Neuro 2A cells was similar after incubation with forskolin and Bt2-cAMP, which caused extensive neurite outgrowth, whereas in the presence of butyrate some rudimentary neurites were formed but they were not nearly as extensive. All compounds produced a dose-dependent inhibition of cell proliferation, but the effect of Bt2-cAMP was more marked than that caused by forskolin, thus showing that the effect of Bt2-cAMP is due partially to the butyrate released. Acetylcholinesterase activity was lower in the cells incubated with butyrate or Bt2-cAMP than in untreated cells or in forskolin-treated cells. This suggests that cyclic AMP does not play a role in the regulation of this enzyme. Bt2-cAMP produced histone acetylation, a well-known effect of butyrate in cultured cells, whereas forskolin did not affect this modification. Consequently, the levels of thyroid hormone receptor, a nuclear protein whose concentration is regulated by butyrate through changes in acetylation of chromatin proteins, were decreased in cells incubated with Bt2-cAMP or butyrate, but were unaffected by forskolin. Butyrate elevated the concentration of histone H1(0), a protein that increases in neuroblastoma cells as a result of different treatments that block cell division. The concentration of H1(0) in the cells treated with Bt2-cAMP was at a level intermediate between that found after treatment with butyrate and with forskolin. The present results clearly indicate that some of the effects of Bt2-cAMP on neuroblastoma cells can be attributed to the butyryl moiety of this compound rather than to the cyclic nucleotide itself.

Acetylation↗
PubMed1988

Evidence for the presence of nuclear 3,5,3'-triiodothyronine receptors in secondary cultures of pure rat oligodendrocytes.

It has been reported that oligodendrocytes do not contain nuclear T3 receptors, which is in apparent contradiction with the well-known effects of thyroid hormones on myelination. In this study we have reexamined the presence of receptors in this cell population, using pure rat oligodendrocyte cultures. T3 binding was also studied with the use of pure rat astrocytes as well as in mixed neuronal-glial cultures. The latter are mainly neuronal during the first days in culture and essentially glial thereafter. Binding studies carried out in intact cells demonstrated the presence of high affinity-low capacity binding sites for thyroid hormones in pure cultures of oligodendrocytes. The maximal binding capacity was 50-60 fmol/100 micrograms DNA and the dissociation constant (Kd) 0.13 nM. Pure rat astrocyte cultures also contained high affinity sites for thyroid hormones, although receptor concentrations was 2-3 times lower than in oligodendrocytes or neurons. This was confirmed in pure cultures of chick astrocytes and in neuronal-glial cultures during the astroglial period. The relative affinity of the receptor for thyroid hormone analogs was triiodothyroacetic acid = T3 greater than T4 greater than tetraiodothyroacetic acid in oligodendrocyte and astrocyte nuclei, and the sedimentation coefficient of the receptor was approximately 3.8S in both cell types. These results demonstrate that nuclear T3 receptors similar to those found in neurons and astrocytes are also present in oligodendrocytes. This suggests that the effects of thyroid hormones on myelination could result from a direct action of the hormone in the oligodendrocytes.

Animals↗
PubMed1988

Short chain fatty acids modulate nuclear receptor and extranuclear L-triiodothyronine levels in glial C6 cells by different mechanisms.

After incubation with [125]T3, the amount of radioactivity associated with the nuclear receptor and extranuclear compartment increases in glial C6 cells previously exposed to millimolar concentrations of butyrate. These actions of the fatty acid are accompanied by an inhibition of cell division, as shown by the decrease in [3H]thymidine incorporation or total DNA content per culture. Isobutyrate produced a similar effect at concentrations higher than 2 mM, whereas between 0.5-2 mM it increased the receptor content without affecting either extra-nuclear hormone or DNA. For both fatty acids there was a close parallelism between the dose-response curve for the inhibition of turnover of [3H]acetate from the histones and the increase in receptor levels. In contrast, a different dose dependence was found for the increase in extranuclear T3 levels, which suggests that the latter is caused by a mechanism other than histone acetylation. The increase in extranuclear T3 levels correlated better with the decrease in DNA and the accumulation of the chromosomal protein H10, which is thought to be related to the inhibition of cell proliferation. This suggests a relationship between the increase in extranuclear T3 and the inhibition of DNA synthesis. However, other agents, such as glucocorticoids or cAMP, which also inhibit C6 cell proliferation, were ineffective in increasing not only the receptor but also the extranuclear hormone. These findings show that the effect of short chain fatty acids on the receptor is not a consequence of the inhibition of cell replication, and that this inhibition could be related to, but is not sufficient per se for causing the increase in extranuclear T3.

Acetates↗
PubMed1987

Uptake and activity of rifapentine in human peritoneal macrophages and polymorphonuclear leukocytes.

Rifapentine uptake by human peritoneal macrophages and polymorphonuclear leukocytes was evaluated and its activity against intracellular Staphylococcus aureus and Staphylococcus epidermidis in both types of phagocytes was compared to that of rifampin, teicoplanin and clindamycin. Uptake of radiolabeled rifapentine by peritoneal macrophages (intracellular/extracellular ratio 61.4 +/- 5.8) and polymorphonuclear leukocytes (intracellular/extracellular ratio 87.6 +/- 3.9) was significantly higher than that of teicoplanin (intracellular/extracellular ratio 40.8 +/- 3.6 and 52.3 +/- 3.2 respectively) and clindamycin (intracellular/extracellular ratio 6.9 +/- 0.4 and 8.3 +/- 0.5 respectively). Rifapentine and rifampin showed the highest activity against intracellular staphylococci in both peritoneal macrophages and polymorphonuclear leukocytes. Clindamycin also showed efficient intracellular activity. In contrast, teicoplanin, which achieved high cellular levels with both types of phagocytes, failed to produce a significant reduction in viable intraphagocytic Staphylococcus epidermidis. On the basis of these findings it is suggested that rifapentine may play a future role in the treatment of certain types of staphylococcal infection.

Anti-Bacterial Agents↗
PubMed1987

alpha GPD response to the continuous infusion of thyroxine into the hypothyroid rat.

In the present work we have studied the disappearance of the mitochondrial enzyme alpha-glycerophosphate dehydrogenase (alpha GPD) after thyroidectomy, as well as the induction by the continuous infusion of physiological doses of thyroxine to hypothyroid male rats in three different tissues, the liver, kidney and heart. Rats were previously rendered hypothyroid by surgical thyroidectomy and enzyme activity was determined at different time intervals following the ablation of the gland. Levels of alpha GPD are specifically regulated in each tissue, as both its rate of disappearance after thyroidectomy and the rate of appearance during T4 treatment are different in the liver, the kidney and the heart. Such results suggest the existence of local factors which can modify the response generated by the thyroid hormone in individual tissues.

Animals↗
PubMed1987

Down-regulation of thyroid hormone nuclear receptor levels by L-triiodothyronine in cultured glial C6 cells.

L-Triiodothyronine (T3) produced a time- and dose-dependent depletion of nuclear thyroid hormone receptor levels in C6 cells, a rat glioma cell line. Receptor number diminished by 30-40% after a 48 h incubation with concentrations of T3 that saturate the nuclear receptor. The nuclear binding curve obtained in cells incubated for 48 h with T3 was shifted leftward of the curve obtained after a 3 h incubation, which indicates an apparent increase in receptor affinity after long-term incubation with T3. However, this change probably represents a further equilibration of the hormone, since the dissociation rate from the nuclei was similar in C6 cells after long- and short-term incubation with T3. The effect of T3 was further demonstrated in C6 cells incubated with short-chain fatty acids. Butyrate and isobutyrate increased receptor levels, and T3 partially decreased the response to these compounds. These findings suggest the existence of a desensitization process by which C6 glial cells would be protected against an excess of thyroid hormone.

Animals↗
PubMed1987

Effects of iopanoic acid on thyroid hormone receptor, growth hormone production, and triiodothyronine generation from thyroxine in pituitary GH1 cells.

We have studied the effect of iopanoic acid (IOP), a radiographic contrast agent which inhibits T4 to T3 conversion, on thyroid hormone nuclear receptors, GH response to T4 and T3, and T4 5'-monodeiodination in GH1 cells, a rat pituitary cell line. IOP at concentrations higher than 10 microM inhibits iodothyronine binding to the nuclear receptor without changing the dissociation constant (Kd) (0.1 nM for T3 and 1 nM for T4), and reduces the GH response to 50 nM T4, 5 nM T3, or the combined effect of T4 and glucocorticoids. These results could be explained by an inhibition of protein synthesis which was reduced by more than 50% by 50 microM IOP. By contrast, nontoxic concentrations of IOP did not change the GH response to different doses of T4 ranging from 1 nM to 50 nM. We also examined T3 generation from T4 and found that the intracellular T3 levels of cells incubated with 50 nM T4 were almost as high as those of cells incubated with 5 nM T3 which induces a full GH response. Intracellular T3 levels were markedly reduced in the cells incubated with T4 and IOP, but GH production was not reduced despite these differences in T3 levels. Additionally, more than 40% of the nuclear receptor was occupied by T3 in cells incubated with T4, whereas more than 90% was occupied by T4 in cells receiving the same amount of T4 with 5 microM IOP. Our results suggest that the effect of T4 on GH production by GH1 cells could be attributed to an important extent to the T3 generated from it, whereas when T4 monodeiodination is strongly inhibited, most of the biological activity is a result of intrinsic T4 activity.

Animals↗
PubMed1987

Glucocorticoids regulate insulin binding in a rat glial cell line.

We have examined insulin receptor regulation by glucocorticoids in the C6 rat glioma cell line. Dexamethasone decreased insulin binding to intact cell monolayers in a dose and time-dependent fashion. The effect is maximal between 48 and 72 h with 50 nM dexamethasone that decreased binding by 40-60%. The natural steroid corticosterone produced a similar effect although it was less potent, and the antiglucocorticoid 17 alpha-methyltestosterone was ineffective in lowering the receptor and partially antagonized the effect of dexamethasone. Total number of binding sites was decreased by glucocorticoids, and when analyzed with a two-site model a 3-fold increase in the dissociation constant (Kd) of the low affinity site was also observed. In the absence of protein synthesis the receptor accumulates at the cell surface, since cycloheximide produced a large increase of insulin binding. Cycloheximide totally blocked the effect of dexamethasone when both compounds were added together to the cells suggesting that protein synthesis is necessary for the effect of the glucocorticoid. By contrast, in cells pretreated with dexamethasone, cycloheximide was unable to produce an increase in cell surface receptor, showing that glucocorticoids probably deplete not only membrane receptor but also total cellular receptor.

Animals↗
PubMed1986

Modulation of thyroid hormone nuclear receptors by short-chain fatty acids in glial C6 cells. Role of histone acetylation.

We have studied the effect of butyrate and other short-chain fatty acids on thyroid hormone nuclear receptors in C6 cells, a rat glioma cell line. Exposure of C6 cells to butyrate leads to increased levels of L-triiodothyronine (T3) in the nuclear and extranuclear compartments. The rise in nuclear binding is not merely a reflection of the higher cellular hormone content, and Scatchard analysis of T3 binding to isolated nuclei reveals that butyrate increases receptor number without changing affinity. The effect on the receptor is quantitatively important: a 48-h incubation with 2 mM butyrate increases nuclear binding by 2-3-fold, and 5 mM butyrate by 3-5-fold. Other short-chain fatty acids were found to similarly influence both nuclear receptor and extranuclear T3 levels with the following potency: butyrate greater than valerate greater than propionate greater than acetate. On the contrary, ketone bodies were ineffective. Butyrate increases receptor levels by decreasing receptor degradation, since the apparent t1/2 of receptor disappearance increased by approximately 3-fold in cells incubated with 2 mM butyrate for 48 h. The regulation of receptor number might be secondary to an action of butyrate on regions of the chromatin to which the receptor associates. We then examined the effect of butyrate on histone acetylation. The fatty acid had little effect in increasing the level of multiacetylated forms of H3 and H4 histone when studied in acid-urea gels, but it markedly inhibited the turnover of [3H] acetate from the histone fraction. There was a striking similarity in the dose-response of butyrate for increasing receptor levels and inhibiting histone deacetylation. Furthermore, a very close correlation between receptor levels and [3H]acetate release was also found when different short-chain fatty acids were used. We thus conclude that the effect of butyrate on the receptor could be explained by a modification of the chromatin structure of C6 cells secondary to acetylation.

Acetates↗
PubMed1986

Modulation of adherence of coagulase-negative staphylococci to Teflon catheters in vitro.

The mechanism of adherence of Staphylococcus epidermidis to commercially available catheters was studied in vitro in a quantitative assay employing 3H-labelled bacteria. It was found that adherence to Teflon catheters was significantly related to the degree of hydrophobicity of the strains. When hydrophobic groups were removed from Staphylococcus epidermidis by pepsin treatment, adhesion was almost completely abolished. Preincubation of catheters in human serum also caused a 80-90% reduction of adherence. Preincubation of Staphylococcus epidermidis in serum similarly decreased adhesion. This effect of serum was mainly due to albumin, while IgG and fibronectin were less effective. Culture of Staphylococcus epidermidis in subinhibitory concentrations (0.5 MIC) of cephalothin, clindamycin and vancomycin resulted in a 30-80% reduction in adhesion.

Anti-Bacterial Agents↗