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E Z Amri

Publications and source records attributed to E Z Amri.

At least 19 recordsLinked to original sources
PubMed2002

PPARgamma-dependent and PPARgamma-independent effects on the development of adipose cells from embryonic stem cells.

Peroxisome proliferator-activated receptor (PPAR) gamma was shown to be required for adipocyte formation both in vivo and in vitro. However, the role of PPARgamma in the initial steps of adipose cell development was not distinguished from its role in the terminal steps. We now show that PPARgamma is expressed early in embryoid bodies (EBs) derived from embryonic stem cells and in E.8.5 mouse embryos. Addition of a specific ligand for PPARgamma in developing EBs over-expressing PPARgamma did not commit stem cells towards the adipose lineage. In differentiated PPARgamma(-/-) EBs, only markers characteristic of preadipocytes were found to be expressed. PPARdelta is present in EBs but did not compensate for the lack of PPARgamma in terminal differentiation. Taken together, these results favor a critical PPARgamma-independent phase culminating in preadipocyte formation that precedes a PPARgamma-dependent phase in the development of adipose cells from pluripotent stem cells.

Adipocytes↗
PubMed2001

Structural and functional characterization of the mouse fatty acid translocase promoter: activation during adipose differentiation.

Fatty acid translocase (FAT/CD36) is a cell-surface glycoprotein that functions as a receptor/transporter for long-chain fatty acids (LCFAs), and interacts with other protein and lipid ligands. FAT/CD36 is expressed by various cell types, including platelets, monocytes/macrophages and endothelial cells, and tissues with an active LCFA metabolism, such as adipose, small intestine and heart. FAT/CD36 expression is induced during adipose cell differentiation and is transcriptionally up-regulated by LCFAs and thiazolidinediones in pre-adipocytes via a peroxisome-proliferator-activated receptor (PPAR)-mediated process. We isolated and analysed the murine FAT/CD36 promoter employing C(2)C(12)N cells directed to differentiate to either adipose or muscle. Transient transfection studies revealed that the 309 bp upstream from the start of exon 1 confer adipose specific activity. Sequence analysis of this DNA fragment revealed the presence of two imperfect direct repeat-1 elements. Electrophoretic mobility-shift assay demonstrated that these elements were peroxisome-proliferator-responsive elements (PPREs). Mutagenesis and transfection experiments indicated that both PPREs co-operate to drive strong promoter activity in adipose cells. We conclude that murine FAT/CD36 expression in adipose tissue is dependent upon transcriptional activation via PPARs through binding to two PPREs located at -245 to -233 bp and -120 to -108 bp from the transcription start site.

Adipocytes↗
PubMed2001

Inhibition of myogenesis enables adipogenic trans-differentiation in the C2C12 myogenic cell line.

C2C12 cells are a well-established model system for studying myogenesis. We examined whether inhibiting the process of myogenesis via expression of dominant negative (DN) mitogen-activated protein kinase kinase-3 (MKK3) facilitated the trans-differentiation of these cells into adipocytes. Cells expressing DN MKK3 respond to rosiglitazone, resulting in adipocyte formation. The effects of rosiglitazone appear to be potentiated through peroxisome proliferator activating receptor-gamma. This trans-differentiation is inhibited by the use of the phosphoinositide-3 (PI3) kinase inhibitor, LY294002. These results indicate that preventing myogenesis through expression of DN MKK3 facilitates adipocytic trans-differentiation, and involves PI3 kinase signalling.

Adipocytes↗
PubMed1999

Long chain fatty acids as modulators of gene transcription in preadipose cells.

During the last years, it has been clearly established that long-chain fatty acids act as modulators of gene expression in various tissues, such as adipose tissue, intestine and liver. This transcriptional action of fatty acids explains in part adaptation mechanisms of tissues to nutritional changes and especially to high-fat diets by increasing expression of proteins involved in lipid catabolism in liver and fatty acid uptake and utilization in other tissues. It is now clearly demonstrated that some of these transcriptional effects of fatty acids are mediated by activation of specific nuclear hormone receptors, called peroxisome proliferator-activated receptors (PPARs). These findings will be discussed with a special reference to control of gene expression in preadipocytes and adipose tissue development.

3T3 Cells↗
PubMed1997

Trans-differentiation of myoblasts to adipoblasts: triggering effects of fatty acids and thiazolidinediones.

Long-chain fatty acids (LCFA) and thiazolidinediones are potent activators of differentiation of preadipose cells. These adipogenic effects are, at least in part, mediated by nuclear receptors of the peroxisome proliferator-activated receptor (PPAR) subfamily. This report describes the effects of these agents on the differentiation pathway of myoblasts. Exposure of C2C12 myoblasts to LCFA or thiazolidinediones prevents the formation of multinucleated myotubes and the expression of specific muscle markers, leading in parallel to the expression of a typical adipose differentiation program. Similar transdifferentiation also occurs in mouse muscle satellite cells maintained in primary cell culture. These observations indicate that PPAR activators, such as LCFA or thiazolidinediones, convert the differentiation pathway of myoblasts into that of adipoblasts. This phenomenon could explain the appearance of adipocytes into muscle which occurs in some pathological states characterized by an increase of fatty acid disposal, such as obesity or mitochondrial myopathy.

Adipocytes↗
PubMed1996

Expression of the CD36 homolog (FAT) in fibroblast cells: effects on fatty acid transport.

An adipocyte membrane glycoprotein, (FAT), homologous to human CD36, has been previously implicated in the binding/transport of long-chain fatty acids. It bound reactive derivatives of long-chain fatty acids and binding was specific and associated with significant inhibition of fatty acid uptake. Tissue distribution of the protein and regulation of its expression were also consistent with its postulated role. In this report, we have examined the effects of FAT expression on rates and properties of fatty acid uptake by Ob17PY fibroblasts lacking the protein. Three clones (P21, P22, and P25) were selected based on FAT mRNA and protein levels. Cell surface labeling could be demonstrated with the anti-CD36 antibody FITC-OKM5. In line with this, the major fraction of immunoreactive FAT was associated with the plasma membrane fraction. Assays of oleate and/or palmitate uptake demonstrated higher rates in the three FAT-expressing clones, compared to cells transfected with the empty vector. Clone P21, which had the highest protein levels on Western blots, exhibited the largest increase in transport rates. Fatty acid uptake in FAT-expressing P21 cells reflected two components, a phloretin-sensitive high-affinity saturable component with a Km of 0.004 microM and a basal phloretin-insensitive component that was a linear function of unbound fatty acid. P21 cells incorporated more exogenous fatty acid into phospholipids, indicating that binding of fatty acids was followed by their transfer into the cell and that both processes were increased by FAT expression. The data support the interpretation that FAT/CD36 functions as a high-affinity membrane receptor/transporter for long-chain fatty acids.

Adipocytes↗
PubMed1996

Fatty acids regulate the expression of lipoprotein lipase gene and activity in preadipose and adipose cells.

During fasting, a reduction in lipoprotein lipase (LPL) activity has been observed in rat fat pad with no change in enzyme mass, whereas LPL mRNA and synthesis are increased, suggesting that insulin and/or fatty acids (FA) regulate LPL activity post-translationaly [Doolittle, Ben-Zeev, Elovson, Martin and Kirchgessner (1990) J. Biol. Chem. 265, 4570-4577]. To examine the role of FA, either preadipose Ob1771 cells or Ob1771 and 3T3-F442A adipose cells were exposed to long-chain FA and to 2-bromopalmitate, a non-metabolized FA. A rapid (2-8 h) and dose-dependent increase (up to 6-fold) in LPL mRNA occurred, primarily due to increased transcription, which is accompanied by a decrease (down to 4-fold) in LPL cellular activity. Under these conditions, secretion of active LPL was nearly abolished. Removal of FA led to full recovery of LPL activity. LPL gene expression in 3T3-C2 fibroblasts was not affected by FA treatment. However fatty acid-activated receptor transfected-3T3-C2 cells, which show FA responsiveness, had increased LPL gene expression upon FA addition. LPL synthesis and cellular content appeared unaffected by FA treatment, whereas secretion of LPL was inhibited. These results indicate that FA regulate the post-translational processing of LPL. It is proposed that the regulation of LPL activity by FA is important with regard to the fine-tuning of FA entry into adipocytes during fasting/feeding periods.

3T3 Cells↗
PubMed1995

Thiazolidinediones and fatty acids convert myogenic cells into adipose-like cells.

Fatty acids and thiazolidinediones act as potent activators of the adipose differentiation program in established preadipose cell lines. In this report, the effects of these agents on the differentiation pathway of myoblasts have been investigated. Exposure of C2C12N myoblasts (a subclone of the C2C12 cell line) to thiazolidinediones or fatty acids prevents the expression of myogenin, alpha-actin, and creatine kinase, thus abolishing the formation of multinucleated myotubes. These treatments lead in parallel to the expression of a typical adipose differentiation program including acquisition of adipocyte morphology and activation of adipose-related genes. A similar transition toward the adipose differentiation pathway also occurs in mouse muscle satellite cells maintained in primary culture. Thiazolidinediones exert their adipogenic effects only in non-terminally differentiated myoblasts; myotubes are insensitive to the compounds. Continuous exposure to inducers after growth arrest is not required to maintain the adipose phenotype, but proliferation of adipose-like C2C12N cells leads to a complete reversion toward undifferentiated cells able to undergo either myogenic or adipogenic differentiation depending on the composition of culture medium. These results indicate that adipogenic inducers, such as thiazolidinediones or fatty acids, specifically convert the differentiation pathway of myoblasts into that of adipoblasts.

5,8,11,14-Eicosatetraynoic Acid↗
PubMed1995

Cloning of a protein that mediates transcriptional effects of fatty acids in preadipocytes. Homology to peroxisome proliferator-activated receptors.

Exposure of preadipocytes to long chain fatty acids induces expression of several gene markers of adipocyte differentiation. This report describes the cloning, from a preadipocyte library, of a cDNA encoding a fatty acid-activated receptor, FAAR. The cDNA had the characteristics and ligand-binding domains of nuclear hormone receptors and encoded a 440 amino acid protein related to peroxisome proliferator-activated receptors, PPAR. The deduced protein sequence was 88% homologous to that of hNUC I, isolated from human osteosarcoma cells. FAAR mRNA was abundant in adipose tissue, intestine, brain, heart, and skeletal muscles and less abundant in kidney, liver, testis, and spleen. The mRNA was undetectable in growing Ob1771 and 3T3-F442A preadipocytes, was strongly induced early during differentiation, and was increased by fatty acid. Transcription assays using hybrid receptor showed strong stimulation by fatty acid and weaker induction by fibrates. Transfection of 3T3-C2 fibroblasts, with a FAAR expression vector, conferred fatty acid inducibility of the adipocyte lipid-binding protein and the fatty acid transporter. Transcriptional induction of these genes exhibited inducer specificity identical to that described in preadipocytes. In summary, the data indicate that FAAR is likely a mediator of fatty acid transcriptional effects in preadipocytes.

Adipocytes↗
PubMed1994

Expression cloning in K+ transport defective yeast and distribution of HBP1, a new putative HMG transcriptional regulator.

The rat HBP1 cDNA was cloned by its capacity to suppress the potassium transport-defective phenotype of mutant Saccharomyces cerevisiae cells. HBP1 cDNA encodes a 513 amino acids protein which, unexpectedly, does not share any homology with K+ transporters or K+ channels. However, a search in protein databases reveals that HBP1 contains a putative DNA-binding domain called HMG-box. Northern blot analysis shows that HBP1 is expressed in a variety of tissues and that in adipocyte and myogenic cell lines, its expression is directly related to differentiation. Taken together, the results suggest that the rat HBP1 is a new member of the HMG class of transcriptional regulators involved in cell differentiation pathways.

Adipocytes↗
PubMed1994

Retinoids are positive effectors of adipose cell differentiation.

Retinoids, especially all-trans retinoic acid (t-RA), have been reported in the last decade to inhibit the differentiation of preadipose cells. In those studies, however, the concentrations of t-RA were supraphysiological (0.1-10 microM range). In contrast we show that, when present at concentrations below or close to the Kd values of retinoic acid receptors, retinoids behave as potent adipogenic hormones (1 pM to 10 nM range). As shown by the use of specific ligands for each RAR subtype, these positive effects on adipose differentiation involve in particular the RAR alpha subtype, and have been observed in Ob17 cells exposed to serum-supplemented or serum-free medium, and in rat preadipocytes exposed to serum-free medium. Among the two classes of retinoid acid receptors (RARs) and retinoid X receptors (RXRs), RAR alpha, RAR gamma, RXR alpha and RXR beta mRNAs could be detected in growing adipoblasts and were found to be increased in committed preadipocytes and differentiated cells upon retinoid treatment. Like other adipogenic hormones, retinoids were only effective in the terminal differentiation process leading from preadipocytes to adipocytes.

Adipocytes↗
PubMed1994

Evidence for a common mechanism of action for fatty acids and thiazolidinedione antidiabetic agents on gene expression in preadipose cells.

In diabetic rodents, thiazolidinediones are able to improve insulin sensitivity of target tissues and to reverse, at least partially, the diabetic state. The effects of these drugs on phenotypic expression in various tissues, including adipose tissue, have been reported. We report here that a new thiazolidinedione compound, BRL 49653, exerts, in preadipose cells, potent effects on the expression of genes encoding proteins involved in fatty acid metabolism. These effects of BRL 49653 in Ob 1771 preadipose cells are similar, in terms of kinetics, reversibility, specificity of genes affected, and requirement for protein synthesis, to those already described for natural or nonmetabolizable fatty acids. Moreover, when used at submaximally effective concentrations, BRL49653 and 2-bromopalmitate act in an additive manner to induce gene expression in preadipose cells, but this additivity of effects is lost when one of the compounds is used at a maximally effective concentration. These observations, suggesting similar mechanisms of action for thiazolidinediones and fatty acids, are strongly supported by the demonstration that (i) both molecules activate, in a heterogolous trans-activation assay, the same nuclear receptor of the steroid/thyroid hormone nuclear receptor superfamily and (ii) transfection of 3T3-C2 fibroblasts with an expression vector for this nuclear receptor confers thiazolidinedione inducibility of adipocyte lipid-binding protein gene expression.

3T3 Cells↗
PubMed1994

Fatty acids as signal transducing molecules: involvement in the differentiation of preadipose to adipose cells.

Fatty acids are important metabolic substrates for adipose tissue and act, in preadipose cells, as potent inducers of various proteins directly involved in their metabolism. We have investigated the long-term effects of fatty acids on the conversion process of preadipose Ob1771 cells to adipose cells. Chronic exposure of cells to palmitate led, in a dose-dependent manner, to a strong stimulation of cell differentiation; this effect was confined to terminal events whereas fatty acids did not affect expression of early genes related to commitment of adipoblasts to preadipose cells. Adipogenic action of fatty acids did not require their metabolism as 2-bromopalmitate, which is not metabolized by preadipose cells, was more effective than palmitate in inducing differentiation. The critical role of fatty acids occurred during the first 3 days of the differentiation process and led subsequently to an increase in the number of differentiated cells by means of enhancement of post-confluent mitoses and over-expression of terminal differentiation-related genes. These results thus provide, at the molecular level, a potential link observed in vivo between an increase in fatty acid supply induced by high-fat or high-carbohydrate diets and the hyperplastic development of adipose tissue.

Adipocytes↗
PubMed1993

Cloning of a rat adipocyte membrane protein implicated in binding or transport of long-chain fatty acids that is induced during preadipocyte differentiation. Homology with human CD36.

A cDNA for an adipocyte membrane protein, implicated in the transport of long-chain fatty acids, was isolated by screening with a synthetic oligonucleotide derived from the amino terminal sequence of the protein. The 88-kDa adipocyte membrane protein was previously identified by covalent labeling with N-sulfosuccinimidyl esters of long-chain fatty acids which irreversibly inhibited fatty acid transport by 75% (Harmon, C. M., and Abumrad, N.A. (1993) J. Membr. Biol. 124, 261-268). The cDNA (FAT, 2432 base pairs (bp)) contained 70 bp of 5'-untranslated sequence, an open reading frame encoding a 472-amino acid protein with a predicted molecular mass of 52466, and 940 bp of 3'-untranslated sequence with two polyadenylation signal sequences but with no polyadenylation tail. The deduced protein sequence predicted two transmembrane segments and 10 potential N-linked glycosylation sites. Extensive glycosylation most likely explains why the molecular mass of the isolated protein (88 kDa) is different from that deduced from the cDNA sequence (53 kDa). The sequence of FAT is 85% homologous with that of glycoprotein IV (CD36) identified in human platelets and in lactating mammary epithelium. Consistent with this, a polyclonal antibody against CD36 reacted with adipocyte plasma membranes and detected a single band at 88 kDa. Northern blot analysis of RNA obtained from rat adipose tissue and probed with the cDNA identified two major transcripts of 4.8 and 2.9 kilobases which were abundant in heart, intestine, fat, muscle, and testis. The mRNAs were not detectable in cultured adipose cell lines (Ob1771, 3T3F442A) at the fibroblastic stage but were strongly induced during the differentiation process and by treatment of preadipocytes with dexamethasone, conditions that were also associated with an increase in oleate transport. In contrast, the fibroblastic cell lines 3T3-C2 and L929, which do not differentiate, did not express the mRNAs at all stages of culture. The data suggest that FAT and CD36 belong to a family of proteins that bind/transport long-chain fatty acids or function as regulators of these processes.

Adipose Tissue↗
PubMed1993

Cloning of alpha 2 chain of type VI collagen and expression during mouse development.

We have previously described the molecular cloning of a cDNA probe which detects a 6 kb mRNA termed pOb24. pOb24 mRNA appeared to be a marker of the preadipose state both in vitro and in vivo. A pOb24 genomic fragment was isolated and used to screen cDNA libraries in order to isolate the full-length pOb24 cDNA and to identify the corresponding protein. The screening yielded a new cDNA clone which detected a 3.7 kb mRNA species in addition to the 6 kb mRNA species. Sequences at the 3' end of the 6 kb and 3.7 kb mRNAs indicate that both mRNAs are generated from the same gene through the use of two different polyadenylation sites. The protein encoded by the 3.7 kb mRNA appeared to be homologous to the human alpha 2 chain of type VI collagen (A2COL6). The expression of the A2COL6 gene was not confined to adipose tissue; mRNA species can be detected in ovaries, adrenal glands and lungs but not in liver and skeletal muscle. The expression appeared specific for initial phase(s) of cell differentiation since it is parallel to that of the MyoD1 gene during muscle embryogenesis in vivo. In the myogenic C2C12 cell line, the A2COL6 gene exhibited the same regulation as MyoD1 and myogenin genes. These results indicate that A2COL6 gene expression is a marker of the preadipose state, but may also be a marker of other differentiation programmes such as that of muscle.

Amino Acid Sequence↗
PubMed1991

Regulation of adipose cell differentiation. I. Fatty acids are inducers of the aP2 gene expression.

The regulation of the expression of adipose-related genes, i.e., aP2, adipsin, and glycerophosphate dehydrogenase (GPDH) by growth hormone (GH) and polyamines, as well as the role of fatty acids, have been investigated in polyamine-dependent Ob1754 cells and Ob1771 preadipose cells. Growth hormone acts as an obligatory hormone for adipsin and GPDH gene expression but its presence is not required for the expression of the aP2 gene. In fully differentiated Ob1771 cells, impairment of fatty acid synthesis by glucose deprivation leads to an inhibition of the aP2 gene expression, whereas the expression of adipsin and GPDH genes remains unaffected. Supplementation of the culture medium with fatty acids prevents the decrease of aP2 gene expression, and this effect appears primarily due to an increase in the transcriptional level of aP2 gene. The induction of aP2 gene has been examined in early committed, lipid-free Ob1771 cells in which fatty acid synthesis is very low despite glucose supplementation. Long-chain fatty acids (greater than or equal to C12) are able to activate the aP2 gene. It is concluded that fatty acids or fatty acid metabolites activate the aP2 gene and subsequently modulate its expression.

Adipose Tissue↗