Biomedical subjects

D Wion

Publications and source records attributed to D Wion.

54 records · Page 3Linked to original sources

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

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

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↗

PubMed1989

Levels of nerve growth factor secreted by rat primary fibroblasts and iris transplants are influenced by serum and glucocorticoids.

Previous work performed with mouse fibroblast-like L cells has shown that the level of expression of NGF gene is modulated in these transformed cells by the composition of the growth medium. Glucocorticoids were found to exert a down-regulation on NGF production, while serum stimulated the synthesis of the factor. The contrasting effects of serum and dexamethasone were further investigated in cultures of primary rat fibroblasts or in iris transplants. ELISA assays of NGF released by fibroblasts or by transplanted irides showed that both experimental systems responded to dexamethasone by a 4-5-fold decrease of the amounts of secreted factor. Half-maximal effect took place at a concentration of 3-5 X 10(-9) M, a value close to the dissociation constant of the glucocorticoid receptor in fibroblasts. The glucocorticoid did not influence the secretion of macromolecules. Assays of NGF mRNA performed at a concentration of 10(-7) M dexamethasone indicated that the steroid decreased the pool of NGF transcripts in either experimental systems. In contrast to dexamethasone, serum induced a 4-fold enhancement of the amounts of factor secreted by fibroblasts. This effect was reproduced with serum that was previously heat-treated at mild acidic pH, or with a macromolecular fraction of this heat-treated serum which contains an effector promoting NGF synthesis in L cells. The fact that promotion of NGF synthesis takes place in primary cells raises the possibility that this process may also occur in vivo, for instance following disruption of vasculature, as a part of a wound mechanism. Data collected with iris transplants provide some support to this interpretation.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1989

[Intracerebral graft of a line of chromaffin cells. Immunologic aspects and role of nerve growth factor in survival and differentiation of the graft].

The efficiency of neuronal grafts is correlated with the differentiation of the grafted cells and the connections they establish with the host tissue. The prolonged survival of a chromaffin cell line (PC12) transplanted into an immunodepressed rat brain shows that neuronal differentiation is correlated with the amount of nerve growth factor (N.G.F.) present in the grafted structure. This result is in agreement with in vitro studies. The characterization of other factors that influence the differentiation of PC12 cells in culture could lead to an increase in the efficiency of intracerebral grafts of adrenal medulla cells in Parkinson's disease.

Adrenal Gland Neoplasms↗

PubMed1988

Partial sequence of the rat heavy neurofilament polypeptide (NF-H). Identification of putative phosphorylation sites.

A 3 kb cDNA clone has previously been isolated in this laboratory corresponding to the rat heavy neurofilament polypeptide (NF-H). This clone, equivalent to approximately 70% of the total mRNA of the protein has been sequenced and shown to contain the carboxy-terminal region of the message. This contains 51 of the Lys-Ser-Pro repeat triplets which are reported to be the site of neurofilament phosphorylation. The sequence obtained was subsequently compared to those of mouse and human NF-H, showing a homology of approximately 85%. There is, however, one region which is variable between the species, this being the highly phosphorylated region of the protein containing the Lys-Ser-Pro triplet repeat.

Amino Acid Sequence↗

PubMed1988

Serum contains a macromolecular effector promoting the synthesis of nerve growth factor (NGF) in L cells.

Addition of serum to the culture medium of murine L cells increased both the cellular level of NGF mRNA and the secretion of mature factor. Stimulation of NGF production by the serum was dose-dependent and appeared mediated by some specific factor(s). After gel filtration chromatography of serum, most of the biological activity formed a major peak with an apparent MW of about 160 kDa. This promoting factor was sensitive to heat at neutral pH, but resisted after heating at pH4. An activity inducing NGF synthesis, and displaying a comparable thermal sensitivity was also detected in Cohn fraction IV of human or bovine plasma.

Animals↗

PubMed1988

Messenger RNAs of beta-amyloid precursor protein and prion protein are regulated by nerve growth factor in PC12 cells.

The effect of the neurotrophic factor NGF on the expression of two genes involved in the accumulation of amyloid deposits in neurodegenerative disorders was studied in a clonal cell line, PC12. Use of hybridization methods showed that NGF increased the cellular pool of the mRNA of the prion protein, a macromolecule known to generate fibrillary aggregates in the brain of scrapie-infected animals. Maximal levels of prion mRNA were obtained after 7 days of treatment, but a significant increase was already detectable after 48 hr of exposure to NGF. In contrast, the factor did not increase the cellular content of the transcripts coding for the precursor of the beta-amyloid peptide (APP), which participates in the formation of neuritic plaques in human brains affected by Alzheimer's disease. However, NGF caused a drop in the molecular weight of that mRNA. This change, which is likely to result from a loss of 100-200 bp, was already detected after 24 hr of treatment. These results indicate that NGF induces in target neuronal cells a quantitative and a qualitative modification of the transcription products encoding two different amyloid precursor proteins.

Amyloid↗

PubMed1987

Retinoic acid increases the expression of NGF gene in mouse L cells.

Retinoic acid (RA), the acid form of vitamin A, is shown to enhance the synthesis of nerve growth factor (NGF) in cultures of mouse L cells. Maximal stimulation was observed in cells growing in a serum-free medium supplemented with 10(-6)M RA during 48 h. The drug increased both the level of NGF mRNA and the amount of mature NGF protein secreted by the cells. RA was previously reported to increase the number of NGF receptors on some neuroblastoma cells (Haskell et al., 1987 Cell and Tiss. Res., 247, 67-73). It seems, therefore, that RA may influence nerve cell differentiation by promoting both the synthesis of the neurotrophic factor and the responsiveness of target cells.

Animals↗

PubMed1986

Isolation of a cDNA for the rat heavy neurofilament polypeptide (NF-H).

We have isolated from a rat brain lambda gt11 expression library two overlapping cDNA clones of sizes 2.5 and 3.0 kb corresponding to the heavy neurofilament polypeptide (NF-H). The 2.5 kb insert apparently represents virtually the whole of the C-terminal tail, the 3.0 kb insert also encodes the conserved epitope for the monoclonal antibody, anti-IFA. The identity of the cDNAs was established by comparison of the predicted amino acid sequence with the known partial amino acid sequence of porcine NF-H. A repeat peptide sequence that may be a multiphosphorylation site was identified in the C-terminal non-helical tail.

Amino Acid Sequence↗

PubMed1986

Molecular cloning of the avian beta-nerve growth factor gene: transcription in brain.

A chicken gene cross-hybridizing with a murine beta-nerve growth factor (beta NGF) cDNA probe was identified by Southern blot analysis and isolated from a genomic DNA library. The DNA sequence coding for the putative mature beta NGF protein was determined, providing direct evidence for the existence in birds of a neurotrophic factor sharing a high degree of sequence homology with mammalian beta NGF. In addition this gene is shown to be transcriptionally active in adult avian brain as demonstrated by Northern blot analysis.

Animals↗

PubMed1986

Dexamethasone rapidly reduces the expression of the beta-NGF gene in mouse L-929 cells.

Mouse L-929 cells were treated with dexamethasone, and the cellular levels of beta-NGF mRNA were estimated by hybridization of the RNAs with a beta-NGF cDNA probe. The results revealed that the glucocorticoid decreased specifically, in a dose-dependent manner, the pool of beta-NGF transcripts. After 4 h, L-929 cells cultured with 10(-7) M dexamethasone contained one-fifth as much beta-NGF mRNA as untreated control cells, and as little as one-tenth as much when the glucocorticoid concentration was 10(-6) M. The effect of the hormone became maximal after 8 h of treatment. Amounts of beta-NGF secreted by the cells during 24 h were measured with a two-site enzyme immunoassay. They also appeared reduced in cultures exposed to the glucocorticoid. These data indicate that dexamethasone controls negatively the expression of the beta-NGF gene in L-929 cells at some pre-translational level.

Animals↗

PubMed1985

Serum and thyroid hormones T3 and T4 regulate nerve growth factor mRNA levels in mouse L cells.

Mouse L cells synthesize and secrete a neurotrophic factor related to the beta subunit of the submaxillary gland nerve growth factor (NGF) of male mice. Use of a cDNA probe which encodes the beta-NGF mRNA demonstrated that L cells produce a transcript identical in size to that of the submaxillary gland. Moreover, target sites of restriction enzymes EcoRI, PstI and BamHI were not significantly rearranged in the beta-NGF gene locus of these cells. The abundance of the beta-NGF transcript was found to depend on culture conditions. Removal of serum depressed the cellular content of polyadenylated RNA by a factor of 1.7, and decreased specifically the pool of beta-NGF transcript by an additional factor of 4. The presence of 10(-7) M testosterone in the serum-free medium did not modify the level of beta-NGF mRNA, while addition of 10(-7) M T3 (or T4) increased this level by a factor of 1.5. These data provide the first evidence that the beta-NGF mRNA of L cells is subjected to regulation, but in a way apparently different from that described for the submaxillary gland.

Animals↗

PubMed1984

Synthesis and partial maturation of the alpha- and gamma-subunits of the mouse submaxillary gland nerve growth factor in Xenopus laevis oocytes.

Sera raised against the alpha-, beta- and gamma-subunits of the mouse 7 S NGF were used to characterize translation products coded by submaxillary gland mRNAs microinjected into Xenopus oocytes. Anti-beta NGF sera did not cross-react with any material. In contrast, the precursors of the alpha- and gamma-subunits, as well as that of renin were identified. Use of tunicamycin, and a comparison of the translation products obtained in oocytes or in the reticulocyte lysate indicated that oocytes achieved the cleavage of signal sequences, the glycosylation of the alpha- and gamma-precursors, and the subsequent secretion of the 3 proteins. In the submaxillary gland, however, the mature forms of alpha NGF, gamma NGF and renin are composed of peptides of smaller size than those produced by the oocytes. These latter appear to lack specific proteases involved in the terminal processing of the submaxillary gland proteins.

Animals↗

PubMed1983

[The neurotrophic factor nerve growth factor (NGF) synthetized by murine L-929 cells is not associated with alpha and gamma subunits].

L-929 cells secrete a neurotrophic factor which is immunologically related to the beta NGF present in the male Mouse submaxillary gland. In order to determine whether L-929 cells also synthesize the alpha and gamma subunits which are associated with the beta NGF in the gland, immunoprecipitations were performed with appropriate antisera. Our results indicate the absence of alpha and gamma subunits in the extracellular medium or in the cell extracts and suggest that the synthesis and secretion of the beta NGF by the L cells does not require these two proteins.

Animals↗

PubMedDate not supplied

Epidermal growth factor modulates the expression of vascular endothelial growth factor in the human prostate.

The growth and dissemination of tumors in the body has been associated with angiogenesis. Vascular endothelial growth factor (VEGF) is an angiogenic factor that stimulates endothelial cell growth and enhances vascular permeability. VEGF exerts its action by binding to specific cell surface receptors. Three receptors, VEGFR-1 (flt-1), VEGFR-2 (flk-1), and VEGFR-3 (flt-4) have been identified. Very little information on the coordinated expression of VEGF and its receptors in normal prostate, benign prostatic hyperplasia (BPH), and prostate carcinoma is available. Therefore, we examined the immunohistochemical localization of VEGF and its receptors in tissues derived from normal human prostate, BPH, and prostatic carcinoma. Immunostaining for VEGF was absent in the normal prostate. Epithelium lining the glands of prostate derived from patients with BPH exhibited strong immunostaining. The intensity of staining was relatively less in prostate carcinoma. It is interesting that VEGFR-1 and VEGFR-3 were strongly expressed in both stromal and epithelial tissues in normal prostate, BPH, and carcinoma. In comparison, VEGFR-2 was not localized to normal prostate and its expression in the stroma of BPH and epithelium of carcinoma was very weak. Because progression of prostate cancer is accompanied by altered expression of epidermal growth factor (EGF) and its receptor (EGFR) in malignant cells, we investigated the effect of EGF on VEGF gene expression by Northern blot analysis in 2 human prostate cancer cell lines that express EGFR. EGF greatly enhanced the expression of VEGF messenger RNA in DU145 and PC3 cell lines in a dose-dependent manner. The EGF induction of VEGF gene expression suggests a mechanism by which angiogenesis could be accelerated in BPH and prostate carcinoma.

Carcinoma↗