Differentiation Of Embryonal Carcinoma Cells Research Articles

Methyltransferase-inhibition interferes with neuronal differentiation of P19 embryonal carcinoma cells

We have analyzed the importance of substrate methylation by S-adenosylmethionine-dependent methyltransferases for neuronal differentiation of P19 embryonal carcinoma cells. We show that treatment of cells with methyltransferase inhibitor adenosine dialdehyde (AdOx) interferes with neuronal differentiation. Retinoic acid (RA) and AdOx co-treated cells had a decreased number of neurites and a flattened morphology compared with cells differentiated by RA. Also, the amount of neuronal class III tubulin (Tuj1) decreased from 76% to 9.6% with AdOx-treatment. Gene expression levels of wnt-1, brn-2, neuroD, and mash-1 were also down-regulated by AdOx-treatment. But AdOx-treatment did not up-regulate BMP-4 and GFAP genes. Treatment of RA decreased E-cadherin expression during neuronal differentiation. However, in AdOx/RA co-treated cells, E-cadherin expression was restored to the control level. Also, mRNA expression of N-cadherin decreased with AdOx-treatment. Taken together, these data show that methylation reactions might influence the cell-fate decision and neuronal differentiation of P19 cells.

Pharmacological properties of purinergic receptors and their effects on proliferation and induction of neuronal differentiation of P19 embryonal carcinoma cells

We have used P19 embryonal carcinoma cells as in vitro model for early neurogenesis to study ionotropic P2X and metabotropic P2Y receptor-induced Ca2+ transients and their participation in induction of proliferation and differentiation. In embryonic P19 cells, P2Y1, P2Y2 and P2X4 receptors or P2X-heteromultimers with similar P2X4 pharmacology were responsible for ATP and ATP analogue-induced Ca2+ transients. In neuronal-differentiated cells, P2Y2, P2Y6, P2X2 and possibly P2X2/P2X6 heteromeric receptors were the major mediators of the elevations in intracellular free calcium concentration [Ca2+]i. We have collected evidence for the involvement of metabotropic purinergic receptors in proliferation induction of undifferentiated and neural progenitor cells by using a BrdU-incorporation assay. ATP-, UTP-, ADP-, 2-MeS-ATP- and ADP-βS-induced proliferation in P19 cells was mediated by P2Y1 and P2Y2 receptors as judged from pharmacological profiles of receptor responses. ATP-provoked acceleration of neuronal differentiation, determined by analysis of nestin and neuron-specific enolase gene and protein expression, also resulted from P2Y1 and P2Y2 receptor activation. Proliferation- and differentiation-induction involved the activation of inositol-trisphosphate sensitive intracellular Ca2+ stores.

SOX17 directly activatesZfp202transcription during in vitro endoderm differentiation

SOX17 is a SRY-related high-mobility group (HMG) box transcription factor that is necessary for endoderm formation in multiple species. Despite its essential function during endoderm formation and differentiation, few direct targets of SOX17 are known. To identify targets of SOX17, we isolated SOX17 binding sites with a chromatin immunoprecipitation (ChIP)-cloning screen. SOX17-ChIP identified zinc finger protein 202 (Zfp202) as a direct target of SOX17 during endoderm differentiation of F9 embryonal carcinoma cells. A sequence in the first intron of Zfp202 activated transcription in differentiated F9 cells, and overexpression of Sox17 increased the transcriptional activity of this sequence. SOX17 binds to a site within this sequence in electrophoretic mobility shift assays, and mutation of this site decreases the transcriptional activation. Zfp202 is induced concomitantly with Sox17 during endoderm differentiation of F9 cells. We also show that ZFP202 represses Hnf4a, which has been reported for the human ortholog ZNF202. Identifying targets of SOX17 will help to elucidate the molecular basis of endoderm differentiation and may provide a better understanding of the role of endoderm in patterning the other germ layers.

The Role of Hypoxia in the Differentiation of P19 Embryonal Carcinoma Cells into Dopaminergic Neurons

Nervous system development at early stage is in hypoxic environment. Very little is known about the role of hypoxia in neuronal development. P19 embryonal carcinoma (EC) cells are a widely used model for studying early neuronal development. In this study we investigated the roles of hypoxia in differentiation of dopaminergic neurons derived from P19 EC cells. Results demonstrate that hypoxia increases the percentage of differentiated neurons, especially neurons of dopaminergic phenotype. To investigate the potential mechanism involved in hypoxia promoted differentiation of dopaminergic neurons, we measured the expression of hypoxia-inducible factor 1alpha (HIF-1alpha), based on its characteristic response to hypoxia. The result shows that HIF-1alpha mRNA level in P19 EC cells increases after hypoxia treatment. It is known that HIF-1alpha regulates the expression of tyrosine hydroxylase (TH) gene through binding to its promoter. Therefore, we propose that the underlying mechanism for hypoxia promoted differentiation of dopaminergic neurons was mediated by HIF-1alpha up-regulation under hypoxia.

Rex1/Zfp42 is dispensable for pluripotency in mouse ES cells

BackgroundRex1/Zfp42 has been extensively used as a marker for the undifferentiated state of pluripotent stem cells. However, its function in pluripotent stem cells including embryonic stem (ES) cells remained unclear although its involvement in visceral endoderm differentiation in F9 embryonal carcinoma (EC) cells was reported.ResultsWe showed the function of Rex1 in mouse ES cells as well as in embryos using the conventional gene targeting strategy. Our results clearly indicated that Rex1 function is dispensable for both the maintenance of pluripotency in ES cells and the development of embryos. However, Rex1-/- ES cells showed the defect to induce a subset of the marker genes of visceral endoderm, when differentiated as embryoid body, as found in EC cells.ConclusionRex1 should be regarded just as a marker of pluripotency without functional significance like the activity of alkaline phosphatase.

Immortalization of Neural Precursors When Telomerase Is Overexpressed in Embryonal Carcinomas and Stem Cells

The DNA repair enzyme telomerase maintains chromosome stability by ensuring that telomeres regenerate each time the cell divides, protecting chromosome ends. During onset of neuroectodermal differentiation in P19 embryonal carcinoma (EC) cells three independent techniques (Southern blotting, Q-FISH, and Q-PCR) revealed a catastrophic reduction in telomere length in nestin-expressing neuronal precursors even though telomerase activity remained high. Overexpressing telomerase protein (mTERT) prevented telomere collapse and the neuroepithelial precursors produced continued to divide, but deaggregated and died. Addition of FGF-2 prevented deaggregation, protected the precursors from the apoptotic event that normally accompanies onset of terminal neuronal differentiation, allowed them to evade senescence, and enabled completion of morphological differentiation. Similarly, primary embryonic stem (ES) cells overexpressing mTERT also initiated neuroectodermal differentiation efficiently, acquiring markers of neuronal precursors and mature neurons. ES precursors are normally cultured with FGF-2, and overexpression of mTERT alone was sufficient to allow them to evade senescence. However, when FGF-2 was removed in order for differentiation to be completed most neural precursors underwent apoptosis indicating that in ES cells mTERT is not sufficient allow terminal differentiation of ES neural precursors in vitro. The results demonstrate that telomerase can potentiate the transition between pluripotent stem cell and committed neuron in both EC and ES cells.

Genome-wide demethylation during neural differentiation of P19 embryonal carcinoma cells

Epigenetic regulation including DNA methylation plays an important role in several differentiation processes. We profiled global DNA methylation in the neural differentiation of P19 embryonic carcinoma cells using a microarray-based method called MIAMI. We found a genome-wide demethylation of genes. This suggests demethylation rather than methylation is important in neural differentiation.

Role of acetylcholine receptors in proliferation and differentiation of P19 embryonal carcinoma cells

Coordinated proliferation and differentiation of progenitor cells is the base for production of appropriate numbers of neurons and glia during neuronal development in order to establish normal brain functions. We have used murine embryonal carcinoma P19 cells as an in vitro model for early differentiation to study participation of nicotinic (nAChR) and muscarinic acetylcholine (mAChR) receptors in the proliferation of neural progenitor cells and their differentiation to neurons. We have previously shown that functional nicotinic acetylcholine receptors (nAChRs) already expressed in embryonic cells mediate elevations in cytosolic free calcium concentration ([Ca 2+] i) via calcium influx through nAChR channels whereas intracellular stores contribute to nAChR- and mAChR-mediated calcium fluxes in differentiated cells [Resende et al., Cell Calcium 43 (2008) 107–121]. In the present study, we have demonstrated that nicotine provoked inhibition of proliferation in embryonic cells as determined by BrdU labeling. However, in neural progenitor cells nicotine stimulated proliferation which was reversed in the presence of inhibitors of calcium mobilization from intracellular stores, indicating that liberation of intracellular calcium contributed to this proliferation induction. Muscarine induced proliferation stimulation in progenitor cells by activation of Gα q/11-coupled M 1, M 3 and M 5 receptors and intracellular calcium stores, whereas Gα i/o-protein coupled M 2 receptor activity mediated neuronal differentiation.

A Highly Efficient Method to Differentiate Smooth Muscle Cells From Human Embryonic Stem Cells

To the Editor: The molecular mechanisms and the control of smooth muscle cell (SMC) differentiation have been extensively investigated because of its therapeutic potential.1 To date, different cell types have been used to study SMC differentiation, including a variety of mouse embryonic stem cells,2 adult stem cells,3,4 and others.5 Because several fundamental differences exist between mouse and human embryonic development,6 lack of a good model system to study human SMC differentiation has hampered the progress of translating SMC knowledge to novel clinical therapies. Human embryonic stem (hES) cells provide a valuable source of cells for studying human cell differentiation and developing therapeutic potentials in regenerative medicine. Since the initial report describing the derivation of hES cells,7 a variety of studies have established in vitro differentiation strategies to several lineages. Recently, it has been demonstrated that vascular progenitors derived from hES cells could be differentiated into endothelial cells and SMCs by endothelial …

A Quantitative Proteomic Analysis of Mitochondrial Participation in P19 Cell Neuronal Differentiation

A quantitative proteomic analysis of changes in protein expression accompanying the differentiation of P19 mouse embryonal carcinoma cells into neuron-like cells using isobaric tag technology coupled with LC-MS/MS revealed protein changes reflecting withdrawal from the cell cycle accompanied by a dynamic reorganization of the cytoskeleton and an up-regulation of mitochondrial biogenesis. Further study of quantitative changes in abundance of individual proteins in a purified mitochondrial fraction showed that most mitochondrial proteins increased significantly in abundance. A set of chaperone proteins did not participate in this increase, suggesting that neuron-like cells are relatively deficient in mitochondrial chaperones. We developed a procedure to account for differences in recovery of mitochondrial proteins during purification of organelles from distinct cell or tissue sources. Proteomic data supported by RT-PCR analysis suggests that enhanced mitochondrial biogenesis during neuronal differentiation may reflect a large increase in expression of PGC-1alpha combined with down-regulation of its negative regulator, p160 Mybbp1a.

H3K27 Demethylases, at Long Last

Methylation of lysine 27 on histone H3 (H3K27me) by the Polycomb complex (PRC2) proteins is associated with gene silencing in many developmental processes. A cluster of recent papers (Agger et al., 2007; De Santa et al., 2007; Lan et al., 2007; Lee et al., 2007) identify the JmjC-domain proteins UTX and JMJD3 as H3K27-specific demethylases that remove this methyl mark, enabling the activation of genes involved in animal body patterning and the inflammatory response.

Selective repression of retinoic acid target genes by RIP140 during induced tumor cell differentiation of pluripotent human embryonal carcinoma cells

BackgroundThe use of retinoids as anti-cancer agents has been limited due to resistance and low efficacy. The dynamics of nuclear receptor coregulation are incompletely understood. Cell-and context-specific activities of nuclear receptors may be in part due to distinct coregulator complexes recruited to distinct subsets of target genes. RIP140 (also called NRIP1) is a ligand-dependent corepressor that is inducible with retinoic acid (RA). We had previously shown that RIP140 limits RA induced tumor cell differentiation of embryonal carcinoma; the pluriopotent stem cells of testicular germ cell tumors. This implies that RIP140 represses key genes required for RA-mediated tumor cell differentiation. Identification of these genes would be of considerable interest.ResultsTo begin to address this issue, microarray technology was employed to elucidate in a de novo fashion the global role of RIP140 in RA target gene regulation of embryonal carcinoma. Subclasses of genes were affected by RIP140 in distinct manners.Interestingly, approximately half of the RA-dependent genes were unaffected by RIP140. Hence, RIP140 appears to discriminate between different classes of RA target genes. In general, RIP140-dependent gene expression was consistent with RIP140 functioning to limit RA signaling and tumor cell differentiation. Few if any genes were regulated in a manner to support a role for RIP140 in "active repression". We also demonstrated that RIP140 silencing sensitizes embryonal carcinoma cells to low doses of RA.ConclusionTogether the data demonstrates that RIP140 has profound effects on RA-mediated gene expression in this cancer stem cell model. The RIP140-dependent RA target genes identified here may be particularly important in mediating RA-induced tumor cell differentiation and the findings suggest that RIP140 may be an attractive target to sensitize tumor cells to retinoid-based differentiation therapy. We discuss these data in the context of proposed models of RIP140-mediated repression.

Mechanism of acetylcholine-induced calcium signaling during neuronal differentiation of P19 embryonal carcinoma cells in vitro

Muscarinic (mAChRs) and nicotinic acetylcholine receptors (nAChRs) are involved in various physiological processes, including neuronal development. We provide evidence for expression of functional nicotinic and muscarinic receptors during differentiation of P19 carcinoma embryonic cells, as an in vitro model of early neurogenesis. We have detected expression and activity of α2–α7, β2, β4 nAChR and M1–M5 mAChR subtypes during neuronal differentiation. Nicotinic α3 and β2 mRNA transcription was induced by addition of retinoic acid to P19 cells. Gene expression of α2, α4–α7, β4 nAChR subunits decreased during initial differentiation and increased again when P19 cells underwent final maturation. Receptor response in terms of nicotinic agonist-evoked Ca2+ flux was observed in embryonic and neuronal-differentiated cells. Muscarinic receptor response, merely present in undifferentiated P19 cells, increased during neuronal differentiation. The nAChR-induced elevation of intracellular calcium ([Ca2+]i) response in undifferentiated cells was due to Ca2+ influx. In differentiated P19 neurons the nAChR-induced [Ca2+]i response was reduced following pretreatment with ryanodine, while the mAChR-induced response was unaffected indicating the contribution of Ca2+ release from ryanodine-sensitive stores to nAChR- but not mAChR-mediated Ca2+ responses. The presence of functional nAChRs in embryonic cells suggests that these receptors are involved in triggering Ca2+ waves during initial neuronal differentiation.

A new role for angiogenin in neurite growth and pathfinding: implications for amyotrophic lateral sclerosis

Mutations in human angiogenin (hANG), an angiogenic member of the RNase A superfamily, have been recently reported in patients with amyotrophic lateral sclerosis (ALS), a progressive late-onset neurodegenerative disorder. However, very little is known about the expression and subcellular distribution of ANG in the nervous system or its role in differentiation. Here we report that mouse angiogenin-1 (mAng-1) is strongly expressed in the developing nervous system during mouse embryogenesis and neuroectodermal differentiation of pluripotent P19 embryonal carcinoma cells. mAng1 is strongly expressed in motor neurons (MNs) in the spinal cord and dorsal root ganglia as well as in post-mitotic MNs derived from P19 cells. We also show for the first time that ANG expression is in the growth cones and neurites. NCI 65828, an inhibitor of the ribonucleolytic activity of hANG, affected pathfinding by P19-derived neurons but not neuronal differentiation. Our findings clearly show that ANG plays an important role in neurite pathfinding and this has implications for ALS.

P19 embryonal carcinoma cells as in vitro model for studying purinergic receptor expression and modulation of N-methyl- d-aspartate–glutamate and acetylcholine receptors during neuronal differentiation

The in vitro differentiation of P19 murine embryonal carcinoma cells to neurons resembles developmental stages which are encountered during neuronal development. Three days following induction to neuronal differentiation by retinoic acid, most cells of the P19 population lost expression of the stage specific embryonic antigen (SSEA-1) and expressed the neural progenitor cell specific antigen nestin. Beginning from day 4 of differentiation nestin expression was down-regulated, and expression of neuron-specific enolase as marker of differentiated neurons increased. The molecular mechanisms underlying neuronal differentiation are poorly understood. We have characterized the participation of purinergic ionotropic (P2X) and metabotropic (P2Y) receptors at mRNA transcription and protein levels as well as ATP-induced Ca 2+ transients during neuronal differentiation of P19 cells. Gene and protein expression of P2X 2, P2X 6, P2Y 2, and P2Y 6 receptors increased during the course of differentiation, whereas P2X 3, P2X 4, P2Y 1 and P2Y 4 receptor expression was high in embryonic P19 cells and then decreased following induction of P19 cells to differentiation. P2X 1 receptor protein expression was only detected on days 2 and 4 of differentiation. Although P2X 5 and P2X 7 mRNA transcription was present, no protein expression for this receptor subunit could be detected throughout the differentiation process. In undifferentiated cells, mainly ionotropic P2X receptors contributed to the ATP-induced Ca 2+-response. In neuronal-differentiated P19 cells, the ATP-induced Ca 2+-response was increased and the metabotropic component predominated. Purinergic receptor function is implicated to participate in neuronal maturation, as cholinergic and glutamate– N-methyl- d-aspartate (NMDA) induced calcium responses were affected when cells were differentiated in the presence of purinergic receptor antagonists pyridoxalphosphate-6-azophenyl-2′,4′-disulphonic acid (PPADS), suramin or reactive blue-2. Our data suggest that inhibition of P2Y 1 and possibly P2X 2 receptors led to a loss of NMDA receptor activity whereas blockade of possibly P2X 2 and P2Y 2 purinergic receptors during neuronal differentiation of P19 mouse led to inhibition of cholinergic receptor responses.

Connexin43 repression following epithelium-to-mesenchyme transition in embryonal carcinoma cells requires Snail1 transcription factor

Embryonic stem (ES) cells and embryonal carcinoma (EC) cells express high amounts of functional connexin43 (Cx43). During mesoderm formation and subsequent cardiac differentiation, Cx43 is initially down-regulated but is up-regulated again as the emerging cardiomyocytes mature. In this study, we investigated the regulation of Cx43 expression during early phases of differentiation in F9 and P19 EC cells. We found a striking inverse correlation between the expression of Cx43 and that of the transcriptional repressor Snail1. No clear relationship was found with Smad-interacting-protein1 (SIP1), another transcription factor inducing epithelium-to-mesenchyme transition (EMT). Promoter-reporter assays indicated Cx43 repression at the promoter level by ectopically expressed Snail1. To establish whether the Cx43 down-regulation depends on endogenous Snail1, MES-1 cells, differentiated derivatives of P19 EC, were stably transfected by an siRNA construct silencing Snail1 expression. This resulted in a mesenchyme-to-epithelium transition, which was accompanied by increased levels of Cx43 mRNA and protein and enhanced metabolic and electrical coupling. We conclude that Snail1-mediated EMT results in a Cx43 repression.

ADAM23 Plays Multiple Roles in Neuronal Differentiation of P19 Embryonal Carcinoma cells

ADAM23, belonging to ADAM (A Disintegrin And Metalloprotease) protein family, is mainly expressed in brain. P19 cells could differentiate into neuroectodermal cell lineage after cell aggregates have been induced by retinoic acid (RA). In this report, we show that the post-transcriptional and post-translational processes of ADAM23 are regulated during the differentiation of P19 cells. In P19-derived neurons, ADAM23 is polarized distributed in the proximal part. To explore the possible roles of ADAM23 during P19 cell neuronal differentiation, ADAM23-RNAi P19 cell lines were established. These transfected cells could differentiate into neurofilament-expression neurons in the absence of RA, whereas wild-type P19 cell can not. These results suggest ADAM23 may play roles in both early and later stage of neuronal differentiation.

Q2ChIP, a Quick and Quantitative Chromatin Immunoprecipitation Assay, Unravels Epigenetic Dynamics of Developmentally Regulated Genes in Human Carcinoma Cells

Chromatin immunoprecipitation (ChIP) is a key technique for studying protein-DNA interactions and mapping epigenetic histone modifications on DNA. Current ChIP protocols require extensive sample handling and large cell numbers. We developed a quick and quantitative (Q(2))ChIP assay suitable for histone and transcription factor immunoprecipitation from chromatin amounts equivalent to as few as 100 cells. DNA-protein cross-linking in suspension in presence of butyrate, elimination of background chromatin through a tube shift after washes, and a combination of cross-link reversal, protein digestion, increased antibody-bead to chromatin ratio, and DNA elution into a single step considerably improve ChIP efficiency and shorten the procedure. We used Q(2)ChIP to monitor changes in histone H3 modifications on the 5' regulatory regions of the developmentally regulated genes OCT4, NANOG, LMNA, and PAX6 in the context of retinoic-acid-mediated human embryonal carcinoma cell differentiation. Quantitative polymerase chain reaction analysis of precipitated DNA unravels biphasic heterochromatin assembly on OCT4 and NANOG, involving H3 lysine (K)9 and K27 methylation followed by H3K9 deacetylation and additional H3K27 trimethylation. Di- and trimethylation of H3K4 remain relatively unaltered. In contrast, PAX6 displays histone modifications characteristic of repressed genes with potential for activation in undifferentiated cells. PAX6 undergoes H3K9 acetylation and enhanced H3K4 trimethylation upon transcriptional activation. Q(2)ChIP of the transcription factor Oct4 demonstrates its dissociation from the NANOG promoter upon differentiation. This study is, to our knowledge, the first to reveal histone modification changes on human OCT4 and NANOG regulatory sequences. The results demonstrate ordered chromatin rearrangement on developmentally regulated promoters upon differentiation.

Neuronal differentiation of P19 embryonal carcinoma cells depends on autocrine stimulation of B2-kinin receptors

International Journal of Developmental NeuroscienceVolume 24, Issue 8 p. 560-561 Abstract [P151]: Neuronal differentiation of P19 embryonal carcinoma cells depends on autocrine stimulation of B2-kinin receptors A.H. Martins, A.H. Martins Universidade de Sao Paulo, Brazil University of Leipzig, GermanySearch for more papers by this authorR.R. Resende, R.R. Resende Universidade de Sao Paulo, Brazil University of Leipzig, GermanySearch for more papers by this authorP. Majumder, P. Majumder Universidade de Sao Paulo, Brazil University of Leipzig, GermanySearch for more papers by this authorM. Faria, M. Faria Universidade de Sao Paulo, Brazil University of Leipzig, GermanySearch for more papers by this authorD.E. Casarini, D.E. Casarini Universidade Federal de Sao Paulo, Brazil University of Leipzig, GermanySearch for more papers by this authorA. Tarnok, A. Tarnok Universidade de Sao Paulo, Brazil University of Leipzig, GermanySearch for more papers by this authorW. Colli, W. Colli Universidade de Sao Paulo, Brazil University of Leipzig, GermanySearch for more papers by this authorJ.B. Pesquero, J.B. Pesquero Universidade de Sao Paulo, Brazil University of Leipzig, GermanySearch for more papers by this authorH. Ulrich, Corresponding Author H. Ulrich n/[email protected] Universidade de Sao Paulo, Brazil University of Leipzig, GermanySearch for more papers by this author A.H. Martins, A.H. Martins Universidade de Sao Paulo, Brazil University of Leipzig, GermanySearch for more papers by this authorR.R. Resende, R.R. Resende Universidade de Sao Paulo, Brazil University of Leipzig, GermanySearch for more papers by this authorP. Majumder, P. Majumder Universidade de Sao Paulo, Brazil University of Leipzig, GermanySearch for more papers by this authorM. Faria, M. Faria Universidade de Sao Paulo, Brazil University of Leipzig, GermanySearch for more papers by this authorD.E. Casarini, D.E. Casarini Universidade Federal de Sao Paulo, Brazil University of Leipzig, GermanySearch for more papers by this authorA. Tarnok, A. Tarnok Universidade de Sao Paulo, Brazil University of Leipzig, GermanySearch for more papers by this authorW. Colli, W. Colli Universidade de Sao Paulo, Brazil University of Leipzig, GermanySearch for more papers by this authorJ.B. Pesquero, J.B. Pesquero Universidade de Sao Paulo, Brazil University of Leipzig, GermanySearch for more papers by this authorH. Ulrich, Corresponding Author H. Ulrich n/[email protected] Universidade de Sao Paulo, Brazil University of Leipzig, GermanySearch for more papers by this author First published: 16 November 2006 https://doi.org/10.1016/j.ijdevneu.2006.09.212Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat No abstract is available for this article. Volume24, Issue8ABSTRACTS TO THE 16TH BIENNIAL MEETING OF THE INTERNATIONAL SOCIETY FOR DEVELOPMENTAL NEUROSCIENCE, 24‐28 AUGUST 2006, BANFF, CANADADecember 2006Pages 560-561 RelatedInformation

Neurotransmitter receptor expression and activity during neuronal differentiation of embryonal carcinoma and stem cells: from basic research towards clinical applications

Embryonal carcinoma and embryonic stem cells have served as models to understand basic aspects of neuronal differentiation and are promising candidates for regenerative medicine. Besides being well characterized regarding the capability of embryonal carcinoma and embryonic stem cells to be precursors of different tissues, the molecular mechanisms controlling neuronal differentiation are hardly understood. Neuropeptide and neurotransmitter receptors are expressed at early stages of differentiation prior to synaptogenesis, triggering transient changes in calcium concentration and inducing neurone-specific gene expression. In vitro neuronal differentiation of embryonal carcinoma and embryonic stem cells closely resembles early neuronal development in vivo. Murine P19 EC cells are a well-characterized model for in vitro differentiation, which upon treatment with retinoic acid differentiate into neurones. Expression and activity of various receptor proteins is regulated during their differentiation. Stimulation of kinin-B2, endothelin-B, muscarinic acetylcholine, and N-methyl-D-aspartate receptors results in transient increases of intracellular free calcium concentration [Ca(2+)](i) in P19 cells undergoing neuronal differentiation, whereas embryonal cells do not respond or show a smaller change in [Ca(2+)](i) than differentiating cells. Receptor inhibition, as studied with the example of the kinin-B2 receptor, aborts neuronal maturation of P19 cells, demonstrating the crucial importance of B2 receptors during the differentiation process. Future success in obtaining desired neuronal phenotypes from pluripotent cells in vitro may offer new therapeutic perspectives for curing genetic and acquired dysfunctions of the developing and adult nervous system.