Characterization of mouse embryonic stem cell differentiation into the pancreatic lineage in vitro by transcriptional profiling, quantitative RT-PCR and immunocytochemistry

Rolletschek, Alexandra; Schroeder, Insa S.; Schulz, Herbert; Hummel, Oliver; Huebner, Norbert; Wobus, Anna M.

doi:10.1387/ijdb.082694ar

Cited by 13 publications

(10 citation statements)

References 74 publications

(115 reference statements)

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“…In an earlier study, we had differentiated murine ES cells according to a mesodermal-lineage promoting protocol and performed comparative expression analyses of undifferentiated and differentiated ES cell specimens at stage 5＋9d (6). Affymetrix gene chip-based expression profiling has provided first evidence that ES cells can be differentiated into NC/CNC-like cells as an upregulation of NC and CNC markers in differentiated specimens was observed (Rolletschek, et al, unpublished) (6). For verification and further molecular characterization, we differentiated murine ES cells of line R1 according to the mesodermal-lineage promoting protocol as described earlier (6).…”

Section: Discussionmentioning

confidence: 99%

“…To investigate the occurrence of ES cell-derived CNClike cells, expression analyses using selected CNC-associated markers (6,30) were performed ( Supplementary Table S4). Selected genes have known roles in the specification (Pax3; Fig.…”

Section: Multilineage Es Cell Progeny Express Selected Cnc-associatedmentioning

confidence: 99%

“…In one of our previous studies, murine ES cells have been differentiated until the intermediate stage 5＋9d according to a mesodermal-lineage promoting protocol (6). Affymetrix gene chip analysis, comparing undifferentiated vs differentiated ES cells at the multilineage progeny stage 5＋9d, revealed the up-regulation of transcripts known to be expressed in neural crest (NC) and cardiac neural crest (CNC) cells (Rolletschek et al, unpublished data; (6)) ( Supplementary Table S1).…”

Section: Introductionmentioning

confidence: 99%

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Molecular Characterization of Embryonic Stem Cell-Derived Cardiac Neural Crest-Like Cells Revealed a Spatiotemporal Expression of an Mlc-3 Isoform

Schmitteckert

Ziegler

Rappold

et al. 2020

IJSC

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Background and Objectives: Pluripotent embryonic stem (ES) cells represent a perfect model system for the investigation of early developmental processes. Besides their differentiation into derivatives of the three primary germ layers, they can also be differentiated into derivatives of the 'fourth' germ layer, the neural crest (NC). Due to its multipotency, extensive migration and outstanding capacity to generate a remarkable number of different cell types, the NC plays a key role in early developmental processes. Cardiac neural crest (CNC) cells are a subpopulation of the NC, which are of crucial importance for precise cardiovascular and pharyngeal glands' development. CNC-associated malformations are rare, but always severe and life-threatening. Appropriate cell models could help to unravel underlying pathomechanisms and to develop new therapeutic options for relevant heart malformations. Methods: Murine ES cells were differentiated according to a mesodermal-lineage promoting protocol. Expression profiles of ES cell-derived progeny at various differentiation stages were investigated on transcript and protein level. Results: Comparative expression profiling of murine ES cell multilineage progeny versus undifferentiated ES cells confirmed differentiation into known cell derivatives of the three primary germ layers and provided evidence that ES cells have the capacity to differentiate into NC/CNC-like cells. Applying the NC/CNC cell-specific marker, 4E9R, an unambiguous identification of ES cell-derived NC/CNC-like cells was achieved. Conclusions: Our findings will facilitate the establishment of an ES cell-derived CNC cell model for the investigation of molecular pathways during cardiac development in health and disease.

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Section: Discussionmentioning

confidence: 99%

Section: Multilineage Es Cell Progeny Express Selected Cnc-associatedmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Molecular Characterization of Embryonic Stem Cell-Derived Cardiac Neural Crest-Like Cells Revealed a Spatiotemporal Expression of an Mlc-3 Isoform

Schmitteckert

Ziegler

Rappold

et al. 2020

IJSC

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“…Western blot analysis of embryonic hearts tissue samples and several ES cell differentiation stages confirmed the presence of Lbx1 protein in our model system. Because there are progenitors from several lineages including the mesodermal as well as ectodermal lineage present at this time of ES cell differentiation [18], double immunocytochemistry using cell type-specific antibodies was performed to assign Lbx1 signal to specific phenotypes. As expected, Lbx1 immunoreactivity could be detected in skeletal muscle cell precursors and GABAergic neurons, reflecting the tissue expression pattern in mice as described previously [4, 19].…”

Section: Discussionmentioning

confidence: 99%

Transcription Factor Lbx1 Expression in Mouse Embryonic Stem Cell-Derived Phenotypes

Schmitteckert

Ziegler

Kartes

et al. 2011

Stem Cells International

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Transcription factor Lbx1 is known to play a role in the migration of muscle progenitor cells in limb buds and also in neuronal determination processes. In addition, involvement of Lbx1 in cardiac neural crest-related cardiogenesis was postulated. Here, we used mouse embryonic stem (ES) cells which have the capacity to develop into cells of all three primary germ layers. During in vitro differentiation, ES cells recapitulate cellular developmental processes and gene expression patterns of early embryogenesis. Transcript analysis revealed a significant upregulation of Lbx1 at the progenitor cell stage. Immunofluorescence staining confirmed the expression of Lbx1 in skeletal muscle cell progenitors and GABAergic neurons. To verify the presence of Lbx1 in cardiac cells, triple immunocytochemistry of ES cell-derived cardiomyocytes and a quantification assay were performed at different developmental stages. Colabeling of Lbx1 and cardiac specific markers troponin T, α-actinin, GATA4, and Nkx2.5 suggested a potential role in early myocardial development.

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“…Exhaustive microarray studies have compared gene expression in ESC-IPC derived from wild type and Pax4 + ESC at different stages of in vitro differentiation in order to attempt understand the mechanisms and factors required to produce fully functional ESC-IPC. The authors conclude that murine ESC-derived pancreatic cells are unable to complete pancreatic differentiation in vitro, as they appear to be blocked at the embryonic/fetal stage of development, as deduced by their gene expression profiles (Rolletschek, Schroeder et al 2010). To date, murine ESC-IPCs are only able to acquire full functional cell characteristics in vivo, but not in vitro, so the challenge to identify the missing factors to induce complete maturation of ESC-IPC in vitro remains.…”

Section: Differentiation Of Embryonic Stem Cells Into Insulin Producimentioning

confidence: 99%

Stem Cell Therapies for Type I Diabetes

Ciriza¹,

Manilay²

2012

Autoimmune Diseases - Contributing Factors, Specific Cases of Autoimmune Diseases, and Stem Cell and Other Therapies

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Characterization of mouse embryonic stem cell differentiation into the pancreatic lineage in vitro by transcriptional profiling, quantitative RT-PCR and immunocytochemistry

Cited by 13 publications

References 74 publications

Molecular Characterization of Embryonic Stem Cell-Derived Cardiac Neural Crest-Like Cells Revealed a Spatiotemporal Expression of an Mlc-3 Isoform

Molecular Characterization of Embryonic Stem Cell-Derived Cardiac Neural Crest-Like Cells Revealed a Spatiotemporal Expression of an Mlc-3 Isoform

Transcription Factor Lbx1 Expression in Mouse Embryonic Stem Cell-Derived Phenotypes

Stem Cell Therapies for Type I Diabetes

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