Isolation and transplantation of dopaminergic neurons generated from mouse embryonic stem cells

Yoshizaki, Takahito; Inaji, Motoki; Kouike, Hiroko; Shimazaki, Takuya; Sawamoto, Kazunobu; Ando, Kiyoshi; Date, Isao; Kobayashi, Kazuto; Suhara, Tetsuya; Uchiyama, Yasuo; Okano, Hideyuki

doi:10.1016/j.neulet.2004.03.074

Cited by 78 publications

(40 citation statements)

References 19 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…To our knowledge this study is the first to establish a miRNA profile from purified dopamine neurons. The overlay of TH staining and GFP fluorescence showed that approximately 70% of the GFPpositive cells are TH-expressing dopamine neurons, which is consistent with previous studies (Hedlund et al, 2007;Yoshizaki et al, 2004). The 9-kb fragment of the rat TH promoter, including intron sequences and a poly(A) tail, could efficiently drive GFP expression in dopamine neurons differentiated from ES cells.…”

Section: Discussionsupporting

confidence: 90%

“…The lack of complete overlap of TH and GFP expression could be attributed to the size of the promoter fragment or the different half-lives of TH and GFP (Hedlund et al, 2007). However, the gene expression analysis of different dopamine markers, in conjunction with previous studies, indicates that the vast majority of sorted cells are of the dopamine lineage (Yoshizaki et al, 2004;Hedlund et al, 2007). The relative miRNA expression profiles of GFP-positive, GFPnegative and neural progenitor cells during differentiation process identified by TaqMan Low Density Assay and then confirmed by real-time PCR in our study identified several miRNAs, including miR-133b, miR-9 and miR-7, that have been reported to play a role in the central neural system, dopamine neuron development and differentiation, and have possible links to PD (Junn et al, 2009;Kim et al, 2007;Leucht et al, 2008).…”

Section: Discussionmentioning

confidence: 93%

“…The pTH-GFP construct containing the GFP reporter expressed under the 9-kb 59-flanking region of the rat TH gene was a kind gift of Kazuto Kobayashi (Jomphe et al, 2005;Sawamoto et al, 2001;Yoshizaki et al, 2004) (Fukushima Medical University School of Medicine, Japan). The CGR8 ES cells were co-transfected with the pTH-GFP plasmid and pCDNA3.1-neo plasmid containing antibiotic resistant gene to obtain the stable transfection line.…”

Section: Th Promoter Engineered Gfp-reporter Es Cells Facs Rna Isolmentioning

confidence: 99%

See 2 more Smart Citations

miR-132 regulates the differentiation of dopamine neurons by directly targeting Nurr1 expression

Yang

Wang

et al. 2012

Journal of Cell Science

141

114

View full text Add to dashboard Cite

SummaryAlthough it is well established that embryonic stem (ES) cells have the potential to differentiate into dopamine neurons, the molecular basis of this process, particularly the role of microRNAs (miRNAs), remains largely unknown. Here we report that miR-132 plays a key role in the differentiation of dopamine neurons by directly regulating the expression of Nurr1 (also known as nuclear receptor subfamily 4 group A member 2; Nr4a2). We constructed a mouse ES cell line CGR8, which stably expresses GFP under the tyrosine hydroxylase (TH) promoter, so the TH-positive neurons could be easily sorted using fluorescence-activated cell sorting (FACS). Then, we performed a miRNA array analysis on the purified TH-positive neurons and found that 45 of 585 miRNAs had more than a fivefold change in expression level during dopamine neuron differentiation. Among the 45 miRNAs, we were particularly interested in miR-132 because this miRNA has been reported to be highly expressed in neurons and to have a potential role in neurodegenerative diseases. We found that the direct downregulation of endogenous miR-132 induced by miR-132 antisense oligonucleotide (miR-132-ASO) promoted the differentiation of TH-positive neurons, whereas ectopic expression of miR-132 in ES cells reduced the number of differentiated THpositive neurons but did not change the total number of differentiated neurons. Furthermore, we identified that miR-132-ASO could substantially reverse the miR-132-mediated suppression of TH-positive neuron differentiation. Moreover, through a bioinformatics assay we identified the Nurr1 gene as a potential molecular target of miR-132. Using a luciferase-reporter assay and western blot analysis, we demonstrated that miR-132 could directly regulate the expression of Nurr1. Collectively, our data provide the first evidence that miR-132 is an important molecule regulating ES cell differentiation into dopamine neurons by directly targeting Nurr1 gene expression.

show abstract

Section: Discussionsupporting

confidence: 90%

Section: Discussionmentioning

confidence: 93%

Section: Th Promoter Engineered Gfp-reporter Es Cells Facs Rna Isolmentioning

confidence: 99%

See 1 more Smart Citation

miR-132 regulates the differentiation of dopamine neurons by directly targeting Nurr1 expression

Yang

Wang

et al. 2012

Journal of Cell Science

141

114

View full text Add to dashboard Cite

show abstract

“…The significant amelioration of hypoactivity in the present HB1.F3 cell-transplanted rats compared with vehicle-infused lesioned rats parallels our previous study demonstrating similar attenuation of abnormal spontaneous locomotor activity in 6-OHDAlesioned animals that received vitamin D3 treatment (Wang et al, 2001). Improvement in drug-induced rotations are observed after transplantation of human or mouse embryonic stem (ES) cells Ben-Hur et al, 2004;Yoshizaki et al, 2004), genetically modified cells (Kang et al, 1993;Leff et al, 1998;Chen et al, 2003), or NSCs or neural progenitor/precursor cells (Svendsen et al, 1997;Armstrong et al, 2002;Sun et al, 2003;Rafuse et al, 2005;Richardson et al, 2005;Harrower et al, 2006). Cell transplantation of rat embryonic VM cells at 2 weeks after 6-OHDA lesion did not attenuate the forelimb akinesia using the cylinder test (Dowd and Dunnett, 2004).…”

Section: Hb1f3 Cell Grafts Attenuate Parkinsonian Motor Symptomssupporting

confidence: 89%

Transplantation of Human Neural Stem Cells Exerts Neuroprotection in a Rat Model of Parkinson's Disease

Yasuhara¹,

Matsukawa²,

Hara³

et al. 2006

J. Neurosci.

260

200

View full text Add to dashboard Cite

“…They will be especially useful for isolating telencephalic neural stem/progenitor cells generated during ES cell differentiation, because the majority of neuronal cells obtained by this method possess characteristics of midbrain dopaminergic neurons, whereas cells with telencephalic character represent only a minor population (39). It has been shown that neuronal cells derived from mouse ES cells, when engrafted in 6-hydroxydopamine-treated Parkinsonian rats, are able to reverse the behavioral deficits significantly (39,40). The range of clinical applications for a pure telencephalic neural stem/progenitor cell population is tremendously large, because these cells could be used for treatment of major brain diseases such as those coupled to stroke.…”

Section: Discussionmentioning

confidence: 99%

The Sox2 Regulatory Region 2 Functions as a Neural Stem Cell-specific Enhancer in the Telencephalon

Miyagi

Nishimoto

Saito

et al. 2006

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

Sox2 is expressed at high levels in neuroepithelial stem cells and persists in neural stem/progenitor cells throughout adulthood. We showed previously that the Sox2 regulatory region 2 (SRR2) drives strong expression in these cells. Here we generated transgenic mouse strains with the ␤-geo reporter gene under the control of the SRR2 in order to examine the spatiotemporal function of this regulatory region. We show that the SRR2 functions specifically in neural stem/progenitor cells. However, unlike Nestin 2nd intronic enhancer, the SRR2 shows strong regional specificity functioning only in restricted areas of the telencephalon but not in any other portions of the central nervous system such as the spinal cord. We also show by in vitro clonogenic assay that at least some of these SRR2-functioning cells possess the hallmark properties of neural stem cells. In adult brains, we could detect strong ␤-geo expression in the subventricular zone of the lateral ventricle and along the rostral migrating stream where actively dividing cells reside. Chromatin immunoprecipitation assays reveal interactions of POU and Sox factors with SRR2 in neural stem/progenitor cells. Our data also suggest that the specific recruitment of these proteins to the SRR2 in the telencephalon defines the spatiotemporal activity of the enhancer in the developing nervous system. Neural stem cells possess the remarkable ability to self-renew and to differentiate into neurons or glia. They can be found during development of the CNS 3 as well as in regions of the adult brain where neurogenesis persists (1-4).Sox2 encodes an SRY-related high mobility group box transcription factor and is expressed in at least three types of stem cells, i.e. neural stem cells (NSC), embryonic stem (ES) cells, and trophoblast stem cells, but not in their differentiated derivatives (5-12). Gene targeting experiments have revealed a cell-autonomous requirement for Sox2 in multipotent cells in early embryonic development (8), suggesting that Sox2 may also be involved in the maintenance of the stem cell state of NSCs. However, because Sox2 null mutant mice failed to develop beyond implantation, the role of this protein in brain could not be examined by conventional knock-out analysis. Notwithstanding, studies from several laboratories have demonstrated the importance of Sox2 for NSC identity. Graham et al. (13) demonstrated that constitutive expression of Sox2 results in maintenance of the neural stem/progenitor cell state and blocks neuronal differentiation. In addition, Bylund et al. (14) showed that the Sox2 protein plays an important role in maintaining neural progenitor identity by counteracting the function of the basic helixloop-helix-containing proneural transcription factors. Most intriguingly, recent knock-down analyses with RNA interference technology have demonstrated a pivotal role for Sox2 in the conversion of oligodendrocyte progenitor cells into NSC-like cells (15), suggesting that Sox2 function is not limited to maintenance of stem cell state but is also i...

show abstract

Isolation and transplantation of dopaminergic neurons generated from mouse embryonic stem cells

Cited by 78 publications

References 19 publications

miR-132 regulates the differentiation of dopamine neurons by directly targeting Nurr1 expression

miR-132 regulates the differentiation of dopamine neurons by directly targeting Nurr1 expression

Transplantation of Human Neural Stem Cells Exerts Neuroprotection in a Rat Model of Parkinson's Disease

The Sox2 Regulatory Region 2 Functions as a Neural Stem Cell-specific Enhancer in the Telencephalon

Contact Info

Product

Resources

About