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P Brachet

Publications and source records attributed to P Brachet.

At least 19 recordsLinked to original sources
PubMed1995

Expression of the nerve growth factor gene is controlled by the microtubule network.

Colchicine, nocodazol, and vinblastine, three microtubule-disrupting drugs, were shown to increase the levels of both nerve growth factor (NGF) mRNA and cell-secreted NGF protein in L929 cells, with levels of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) or amyloid precursor protein (APP) mRNAs remaining unaffected. Northern blot analysis demonstrated that colchicine also increased NGF mRNA levels in rat primary astrocytes and mouse skin fibroblasts. The specificity of the effects observed was assessed by the fact that the microtubule-stabilizing agent Taxotere, a semisynthetic compound structurally related to taxol, suppressed the effects of colchicine, whereas lumicolchicine, a colchicine derivative that has no action on the microtubule network, had no influence on NGF expression. Likewise, the disruption of the microfilament network by cytochalasin B did not increase NGF mRNA levels in L929 cells. Furthermore, the increase in NGF gene expression observed following microtubule disruption depended on a cascade of events involving at least one protein kinase, which is not down-regulated by phorbol ester, and on a pertussis toxin sensitive step. These results support the concept that tubulin and/or the microtubule cytoskeleton play an active role in the regulation of the NGF gene.

Amyloid beta-Protein Precursor↗
PubMed1995

Transport and steady-state accumulation of putrescine in brush-border membrane vesicles of rabbit small intestine.

Absorption of polyamines from the lumen is essential for cell proliferation in small intestine but also in other rapidly growing body tissues and tumors. Intestinal uptake of polyamines is thought to involve one or more transport systems, but the characteristics of these systems have not yet been clearly elucidated. Because high levels of putrescine have been identified in intestinal lumen, we explored kinetic, physiochemical, and structural features of uptake of this diamine across rabbit intestinal brush-border membrane vesicles (IBBMV) prepared by CaCl2 or MgCl2 precipitation procedure. Initial rates of putrescine influx were measured during 5-min incubations at 25 or 37 degrees C (optimal temperature) for concentrations of 0.45-145 microM. At both temperatures, kinetics of putrescine transport fitted a model with a single Michaelis-Menten uptake component plus a nonsaturable uptake component. At 37 degrees C, the kinetic parameters for the saturable component of putrescine uptake, Km,app and Vmax,app, were 16.8 +/- 4.7 microM and 19.9 +/- 2.8 pmol.mg protein-1.min-1, respectively. The value of the constant for the nonsaturable component of putrescine uptake (P = 0.45 +/- 0.06 x 10(-8) l.mg protein-1.s-1) suggested this component represented essentially nonspecific binding of putrescine to IBBMV. Cadaverine, spermidine, and spermine were competitive inhibitors of putrescine transport, with inhibition constants equal to 47, 117, and 219 microM, respectively. When effects of a variety of alkyldiamines and structural analogues of polyamines (1 mM) on influx of 5.6 microM putrescine were compared, cadaverine, methylglyoxal bis(guanylhydrazone) (MGBG), and cyclic derivatives of MGBG were found to exhibit the highest inhibitory potencies.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Expression of 25(OH) vitamin D3 24-hydroxylase gene in glial cells.

The expression of the 25(OH) vitamin D3 24-hydroxylase gene was studied in C6 glioma and rat primary glial cell culture. The expression of the 25(OH)D3 24-hydroxylase gene was not detected in C6 glioma or glial cells cultured in a serum-free medium. However, the 25(OH)D3 24-hydroxylase mRNA was induced in a dose-dependent manner in cells treated with 1,25(OH)2D3. These findings provide further evidence for an involvement of vitamin D3 metabolites in brain function.

25-Hydroxyvitamin D3 1-alpha-Hydroxylase↗
PubMed1992

Activation of nerve growth factor synthesis in primary glial cells by phorbol 12-myristate 13-acetate: role of protein kinase C.

Phorbol 12-myristate 13-acetate (PMA) induces a dramatic production of nerve growth factor (NGF) in primary cultures of newborn mouse astrocytes maintained in a serum-free medium. This stimulation is dose-dependent and a maximal effect on the levels of cell-secreted factor was observed at a concentration of 10 nM. At this concentration, the promoting effect of PMA appears much more important than that elicited by 10% fetal calf serum (FCS) under the same culture conditions. PMA acts primarily on the accumulation of NGF mRNA, which was detected by northern blot analysis after 6 h of treatment. This accumulation may be totally or partially prevented when PMA-treated glial cells are concomitantly exposed to the protein kinase inhibitors H-7, H-9, and to a lesser degree, HA-1004. The known specificity of these inhibitors agrees with the possibility that protein kinase C (PKC), which constitutes so far the sole known target of PMA, represents a key element involved in the stimulation of NGF gene. The role of PKC is further supported by the observation that alpha phorbol didecanoate, which has no activity on PKC, is depleted of effect on the synthesis of NGF. Likewise, 1,2-dioctanoylglycerol (1,2-DOG) has a weak, but significant promoting action on the production of NGF, unlike the 1,3-isomer which is not active on PKC. Finally, a treatment of 15 min with 100 nM PMA is sufficient to stimulate the cells, suggesting that the activation phase of PKC, rather than its down regulation, constitutes an important trigger leading to an increased expression of the NGF gene.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Pertussis toxin provides evidence for two independent signalling pathways leading to the activation of the nerve growth factor gene.

Increased expression of the nerve growth factor (NGF) gene may be obtained by treating L929 fibroblasts with serum, phorbol 12-myristate 13-acetate (PMA), or 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3). The possible involvement of GTP-binding proteins (G proteins) in these regulatory events was monitored by exposing the cells to pertussis toxin (PT), a compound known to inactivate several types of G proteins by ADP ribosylation. Measurements of the pool of NGF mRNA by Northern blot analysis, and quantification of the factor secreted by the cells with a double-site ELISA assay, indicate that pretreatment with PT decreases by about 60% the effect of serum on the levels of NGF transcript and secreted factor. This effect is accompanied by a corresponding decrease of the expression of c-fos gene, which takes place soon after the addition of serum to the cells. In contrast, PT had no effect on the basal level of NGF mRNA found in cells maintained in serum-free medium or in cells stimulated with PMA or 1,25-(OH)2D3. These results indicate that some serum factor(s) acts via plasma membrane receptors able to interact with PT-sensitive G proteins to modulate NGF gene expression. In contrast, 1,25-(OH)2D3 appears to mediate its action through a different signalling pathway, which is likely to require its cytosolic receptor, and is independent of PT-sensitive G protein and c-fos induction.(ABSTRACT TRUNCATED AT 250 WORDS)

Blotting, Northern↗
PubMed1992

Transport of putrescine in the isolated rabbit intestine.

The transepithelial fluxes of putrescine were studied in sections of the three segments of rabbit intestine mounted in an Ussing chamber. The ileum exhibited the highest mucosal-to-serosal (Jms) and serosal-to-mucosal (Jsm) unidirectional fluxes of 1 mumol/l [3H]putrescine. Putrescine net flux (Jnet = Jms - Jsm) was deduced to be positive through the duodenum (Jnet = 53.40 +/- 14.30 pmol h-1 cm-2), not significantly different from zero through the jejunum (Jnet = 8.90 +/- 19.20 pmol h-1 cm-2) and negative through the ileum (Jnet = -34.30 +/- 13.80 pmol h-1 cm-2). Increasing putrescine concentration up to 10 mmol/l led to an increase in Jms, Jsm and Jnet without affecting the transport polarity in the ileum. The tissue retention of putrescine after 100 min was higher by the serosal side than by the mucosal side of the ileum. In parallel experiments, isolated pieces of ileum accumulated putrescine to a five- to sixfold concentration gradient by a ouabain-inhibitable mechanism. In contrast with arginine and in order of decreasing potency, putrescine, cystamine (a transglutaminase inhibitor), spermidine and spermine (1 mmol/l) reduced both unidirectional fluxes of putrescine across the ileum in the Ussing chamber. The latter effectors, except spermine, and N,N-dimethylcasein (1 mg/ml) led to an important, if not complete, suppression of putrescine secretion by the ileum, while the calmodulin antagonist melittin (0.3 micrograms/ml) reversed the polarity of polyamine transport, suggesting the involvement of transglutaminase in putrescine transport. We conclude that the heterogeneous pattern of putrescine transport along the small-intestinal epithelium constitutes an important feature of the regulation of polyamine concentrations in this tissue.

Animals↗
PubMed1992

Regional differences for the D-amino acid oxidase-catalysed oxidation of D-methionine in chicken small intestine.

1. Enzymic oxidation of D-[1-14C]methionine (D-met) to 2-keto-4-methylthiobutanoate (KMB) has been determined using 100,000 g supernatants prepared from chicken tissue homogenates. 2. The small intestinal mucosa contains substantial oxidative activity towards D-met, which represents about one-half and one-tenth the hepatic and renal activity, respectively. 3. KMB is poorly decarboxylated and rather transaminated to L-met. 4. The specific activity for D-met oxidation in the duodenal mucosa is 1.5- and 4.0-fold than in the jejunal and ileal mucosa, respectively. 5. The intestinal D-met-oxidizing activity is dramatically altered by the D-amino acid oxidase specific inhibitor benzoate.

Animals↗
PubMed1992

Thyroid hormone promotes transient nerve growth factor synthesis in rat cerebellar neuroblasts.

Primary cultures of cerebellum from 5-day-old rats indicated that proliferating neuroblasts synthesize and release nerve growth factor (NGF). Since NGF promotes DNA synthesis in these cells, our findings demonstrate that the early developing cerebellum is a suitable physiological model for studying the autocrine mitogenic action of NGF. Thyroid deficiency led to a greater reduction in the NGF content of the cerebellum than of the olfactory bulbs or hippocampus. Cerebellar NGF mRNA was also very sensitive to hormone deprivation. Physiological amounts of thyroid hormone stimulated both the mitotic activity and NGF production of cultured cerebellar neuroblasts. A lack of thyroid hormone is known to markedly alter cell formation in the cerebellum where postnatal neurogenesis is highly significant, in contrast to the olfactory bulbs and hippocampus. Taken together, these results suggest that the hormonal control of cell formation in the cerebellum is, at least partly, mediated by the autocrine mitogenic action of NGF. The thyroid hormone could temporally regulate the transient NGF synthesis by cerebellar neuroblasts directly and/or through its ontogenetic action, and hence all the NGF-dependent trophic effects.

Animals↗
PubMed1992

Putrescine uptake by rabbit intestinal brush-border membrane vesicles.

The transport of putrescine in rabbit intestinal brush-border membrane vesicles is a time-dependent, Na+ and H+ gradient-independent slow process, reaching a high equilibrium level from 1 hr onwards. An approximately 12-fold concentration gradient of 5.4 microM putrescine was generated in the vesicles after 3 h of incubation at 25 degrees C, suggesting a substantial binding of the polyamine to membrane components. Putrescine uptake did not appear to occur via a polyamine-H+ exchange pathway, but the activity of transport was strongly reduced when the medium pH was lowered from 7.5 to 6.0. Although it was inhibited in the presence of SCN-, putrescine uptake was found neither to involve a preferred anion nor to be sensitive to changes in the membrane potential. Putrescine accumulation was substantially stimulated in the presence of a 10 mM concentration of the divalent cations Ca2+, Mg2+ or Ba2+, unlike the monovalent cations K+, Na+ or Li+. Despite the presence of a Ca(2+)-dependent transglutaminase activity, vesicular accumulation does not appear to result from covalent binding of putrescine to membrane peptides. Inhibition of the uptake by polyamines, unlike choline and dibasic amino acids, shows that putrescine may bind noncovalently to other specific anionic sites.

Animals↗
PubMed1991

MC903, an analogue of 1,25-dihydroxyvitamin D3, increases the synthesis of nerve growth factor.

The effect of MC903, an analogue of 1,25-dihydroxyvitamin D3, on the expression of the nerve growth factor (NGF) gene has been studied in L cells. MC903 induces an increase in both NGF mRNA and protein with a time course similar to that obtained with 1,25-dihydroxyvitamin D3. This finding points to the potential importance of 1,25-dihydroxyvitamin D3 derivatives in the treatment of NGF-sensitive disorders.

Animals↗
PubMed1991

1,25-Dihydroxyvitamin D3 is a potent inducer of nerve growth factor synthesis.

1,25-Dihydroxyvitamin D3 (1,25-(OH)2D3), a metabolically active form of vitamin D, is shown to increase in a dose-dependent manner the cellular pool of NGF mRNA in murine L-929 fibroblasts cultured in a serum-free medium. This effect can be detected as early as 3 hours after 1,25-(OH)2D3 addition and persists for at least 28 hours. It is accompanied by an enhancement of the amount of NGF protein secreted in the culture medium. Since the proto-oncogene c-fos appears involved in the regulation of the NGF gene (Mocchetti et al.: Proceedings of the National Academy of Sciences of the United States of America 86: 3871-895, 1989; Hengerer et al: Proceedings of the National Academy of Sciences of the United States of America 87:3899-3903, 1990), the effect of 1,25-(OH)2D3 on c-fos expression was analysed and compared to that elicited by other inducers of the NGF gene, serum (Wion et al: FEBS Letters 189:37-41, 1985) and phorbol 12-myristate 13-acetate (PMA) (Wion et al: FEBS Letters 262:42-44, 1990). Addition of serum or PMA to L-929 cells was rapidly followed by a transient activation of the c-fos gene. In contrast, c-fos transcripts remained undetected in the presence of 1,25-(OH)2D3. The failure to find any evidence of c-fos expression suggests that 1,25-(OH)2D3 could enhance the pool of NGF mRNA by a mechanism independent of the c-fos pathway.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Differential effect of sunflower seed oil on hepatic and renal D-amino acid oxidase in the rat.

1. The specific activity of hepatic and renal peroxisomal D-amino acid oxidase (D-AAOX) was measured in rats fed diets containing various quantities of vegetable oil. 2. Increasing the amount of dietary sunflower seed oil (SSO) from 10 to 25% (w/w) reduced the specific activity of hepatic D-AAOX by up to 30% after 10 days. 3. In both tissues, the enzyme activity was moderately decreased during the first two-day period after administration of the 25% SSO diet was begun. Unlike hepatic D-AAOX, renal D-AAOX returned to its baseline level in the kidney after the third day. 4. In contrast to SSO, hydrogenated coconut oil (HCO) did not evoke alterations of D-AAOX activity. 5. The activity levels of another peroxisomal enzyme, L-2-hydroxy acid oxidase (L-HAOX), in the liver of rats fed the high-SSO diet vs those fed the control diet were similar. 6. The subcellular distribution of D-AAOX and L-HAOX was not altered in the liver of rats fed the 25% SSO diet during the 10-day period.

Alcohol Oxidoreductases↗
PubMed1991

Nerve growth factor-induced neuronal differentiation is accompanied by differential splicing of beta-amyloid precursor mRNAs in the PC12 cell line.

The effect of the neurotrophic factor nerve growth factor (NGF) on the expression of the beta-amyloid gene has been studied in the clonal nerve cell line PC12. The neuronal differentiation of PC12 cells in the presence of NGF was accompanied by a shift in the ratio of beta-amyloid precursor protein (APP) transcripts. In particular there was reduced expression of the transcript coding for the longest precursor form (APP770) and a concomitant increase in the shortest (APP695) transcript following NGF treatment.

Amyloid beta-Protein Precursor↗
PubMed1991

Antagonistic effects of dexamethasone and 1,25-dihydroxyvitamin D3 on the synthesis of nerve growth factor.

Dexamethasone is known to decrease the pool of nerve growth factor (NGF) mRNA in various experimental systems. The negative regulatory effect of the glucocorticoid was first observed in mouse fibroblast-like L929 cells, and was subsequently reported to take place in many experimental systems, including in vivo following sciatic nerve injury. Conversely, another steroid hormone, 1,25-dihydroxy-vitamin D3 (1,25-(OH)2D3) was recently reported to promote NGF synthesis in mouse L929 cells. The present work was undertaken to investigate the effect of the concomitant addition of both steroids to L929 cells. Measurements of NGF mRNA and assays of the mature protein secreted by the cells provide evidence that the negative regulation exerted by dexamethasone may be counteracted in a dose-dependent manner by the positive action of 1,25-(OH)2D3, and vice versa. Therefore, the expression of the NGF gene can be regulated in a subtle way by the balance between the two steroids. It may be expected on the basis of these observations that in tissues that are responsive to both hormones, administration of 1,25-(OH)2D3 should be able to reverse the down-regulation of NGF synthesis elicited by glucocorticoids.

Animals↗
PubMed1990

Nerve growth factor (NGF), proNGF and NGF receptor-like immunoreactivity in BB rat retina.

Nerve growth factor (NGF) and NGF precursor (proNGF)-like material were localized immunohistochemically in diabetic and non-diabetic adult rat retina using affinity-purified immunoglobulins to mature NGF and to synthetic peptides that reproduce sequences of the proNGF. Immunoreactivity for NGF and proNGF was detected in retinal ganglion cells, Müller cells, retinal pigment epithelium and areas adjacent to the internal limiting membrane. Immunolocalization of the NGF receptor using a monoclonal antibody (MC192) revealed positive staining of retinal pigment epithelium and Müller cells but not of ganglion cells. The staining intensity and distribution of NGF, proNGF, and NGF receptor-like immunoreactivity were similar in retinas of diabetic rats. These results suggest that NGF is synthesized endogenously in the retina by both neuronal and non-neuronal cell constituents, and that diabetes has no detectable effect on NGF/NGF receptor distribution and possibly on NGF biosynthesis.

Animals↗
PubMed1990

Purification and some characteristics of chicken liver L-2-hydroxyacid oxidase A.

The isozyme A of L-2-hydroxyacid oxidase is a peroxisomal flavoenzyme that catalyzes the oxidation of short-chain aliphatic L-2-hydroxyacids in many tissues of higher organisms. A new purification procedure allowed us to obtain a 1400-fold purified enzyme from chicken liver. The N-terminal amino acid of the polypeptide chain was found to be blocked as that of spinach glycolate oxidase, contrastingly with that of rat kidney isozyme B. Its amino acid composition was comparable to that of other known L-2-hydroxyacid oxidases. Despite different substrate specificity, some immunological identity was observed between chicken liver L-2-hydroxyacid isozyme A and rat kidney isozyme B.

Alcohol Oxidoreductases↗
PubMed1990

Phorbol 12-myristate 13-acetate (PMA) increases the expression of the nerve growth factor (NGF) gene in mouse L-929 fibroblasts.

The rise of the NGF mRNA pool which takes place following exposure of L-929 fibroblasts to serum was prevented in the presence of 5 microM K-252a, a compound which inhibits several species of protein kinase activities. To characterize further this phenomenon, L-929 cells growing in a serum-free medium were exposed to cyclic nucleotide analogs, to a divalent cation ionophore or to the phorbol ester PMA. Only this latter compound induced an enhancement of the NGF mRNA pool, suggesting an involvement of protein kinase C in the upregulation of the NGF transcripts. The effects of PMA or serum also require a synthesis of protein since the level of NGF transcripts remained stable in the presence of cycloheximide.

Animals↗