scholarly journals Promotion of In Vitro Hair Cell-like Cell Differentiation from Human Embryonic Stem Cells through the Regulation of Notch Signaling

Metabolites ◽  
2021 ◽  
Vol 11 (12) ◽  
pp. 873
Author(s):  
Fengjiao Chen ◽  
Ying Yang ◽  
Jianling Chen ◽  
Zihua Tang ◽  
Qian Peng ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Differentiation ◽  
Hair Cell ◽  
Hair Cells ◽  
Embryonic Stem ◽  
Temporal Expression ◽  
Supporting Cells ◽  
Temporal Expression Pattern ◽  
Notch Ligands

The Notch signaling pathway plays an important role in otic neurogenesis by regulating the differentiation of inner ear hair cells and supporting cells. Notch-regulated differentiation is required for the regeneration of hair cells in the inner ear. The temporal expression pattern of Notch ligands and receptors during in vitro hair cell-like cell differentiation from human embryonic stem cells (hESCs) was detected by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Subsequently, pAJ-U6-shRNA-CMV-Puro/GFP recombinant lentiviral vectors encoding short hairpin RNAs were used to silence JAG-1, JAG-2, and DLL-1, according to the temporal expression pattern of Notch ligands. Then, the effect of each ligand on the in vitro differentiation of hair cells was examined by RT-PCR, immunofluorescence, and scanning electron microscopy (SEM). The results showed that the individual deletion of JAG-2 or DLL-1 had no significant effect on the differentiation of hair cell-like cells. However, the simultaneous inhibition of both DLL-1 and JAG-2 increased the number of hair cell-like cells and decreased the number of supporting cells. JAG-2 and DLL-1 may have a synergistic role in in vitro hair cell differentiation.

Promotion of in Vitro Hair Cell-like Cell Differentiation from Human Embryonic Stem Cells through the Regulation of Notch Signaling

2021 ◽  
Author(s):  
Fengjiao Chen ◽  
Ying Yang ◽  
Jianling Chen ◽  
Zihua Tang ◽  
Qian Peng ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Hair Cell ◽  
Notch Signaling ◽  
Hair Cells ◽  
Embryonic Stem ◽  
In Vitro Differentiation ◽  
Supporting Cells ◽  
Notch Ligands

Abstract Background Notch signaling mediates the committed induced differentiation of ear sensory cells and promotes the formation of a precise arrangement of mosaics between hair cells and supporting cells. Embryonic stem cells (ESCs) are pluripotent stem cells which have the potential to differentiate into cell lines through three germ layers. Therefore, it is necessary to study the effects of regulating Notch receptors and ligand expression on the in vitro differentiation equilibrium of hair cells and supporting cells from ESCs.Methods and Results The temporal ex-pression pattern of Notch ligands and receptors during in vitro hair cell-like cell differentia-tion from human embryonic stem cells (hESCs) was detected by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Subsequently, pAJ-U6-shRNA-CMV-Puro/GFP recombinant lentiviral vectors, encoding short hairpin RNAs, were used to silence JAG-1, JAG-2, and DLL-1, according to the temporal expression pattern of Notch ligands. Then the effect of each ligand on the in vitro differentiation of hair cells was examined by RT-PCR, immunofluorescence, and scanning electron microscopy (SEM).Conclusions Results showed that JAG-1 played an important role in regulating hESC differentiation to otic progenitors. The individual deletion of JAG-2 or DLL-1 had no significant effect on the differentiation of hair cell-like cells. Although the simultaneous inhibition of both DLL-1 and JAG-2 could increase the number of hair cell-like cells, it decreased the number of supporting cells.

scholarly journals A Critical E-box in Barhl1 3′ Enhancer Is Essential for Auditory Hair Cell Differentiation

Cells ◽  
2019 ◽  
Vol 8 (5) ◽  
pp. 458 ◽  
Author(s):  
Kun Hou ◽  
Hui Jiang ◽  
Md. Rezaul Karim ◽  
Chao Zhong ◽  
Zhouwen Xu ◽  
...  
Keyword(s):  
Cell Differentiation ◽  
Hair Cell ◽  
Hair Cells ◽  
Embryonic Stem ◽  
Cell Development ◽  
Homologous Gene ◽  
Auditory Hair Cells ◽  
E Box ◽  
Hair Cell Differentiation

Barhl1, a mouse homologous gene of Drosophila BarH class homeobox genes, is highly expressed within the inner ear and crucial for the long-term maintenance of auditory hair cells that mediate hearing and balance, yet little is known about the molecular events underlying Barhl1 regulation and function in hair cells. In this study, through data mining and in vitro report assay, we firstly identified Barhl1 as a direct target gene of Atoh1 and one E-box (E3) in Barhl1 3’ enhancer is crucial for Atoh1-mediated Barhl1 activation. Then we generated a mouse embryonic stem cell (mESC) line carrying disruptions on this E3 site E-box (CAGCTG) using CRISPR/Cas9 technology and this E3 mutated mESC line is further subjected to an efficient stepwise hair cell differentiation strategy in vitro. Disruptions on this E3 site caused dramatic loss of Barhl1 expression and significantly reduced the number of induced hair cell-like cells, while no affections on the differentiation toward early primitive ectoderm-like cells and otic progenitors. Finally, through RNA-seq profiling and gene ontology (GO) enrichment analysis, we found that this E3 box was indispensable for Barhl1 expression to maintain hair cell development and normal functions. We also compared the transcriptional profiles of induced cells from CDS mutated and E3 mutated mESCs, respectively, and got very consistent results except the Barhl1 transcript itself. These observations indicated that Atoh1-mediated Barhl1 expression could have important roles during auditory hair cell development. In brief, our findings delineate the detail molecular mechanism of Barhl1 expression regulation in auditory hair cell differentiation.

scholarly journals In Vitro Germ Cell Differentiation from Cynomolgus Monkey Embryonic Stem Cells

PLoS ONE ◽  
2009 ◽  
Vol 4 (4) ◽  
pp. e5338 ◽  
Author(s):  
Kaori Yamauchi ◽  
Kouichi Hasegawa ◽  
Shinichiro Chuma ◽  
Norio Nakatsuji ◽  
Hirofumi Suemori
Keyword(s):  
Stem Cells ◽  
Cell Differentiation ◽  
Embryonic Stem Cells ◽  
Germ Cell ◽  
Cynomolgus Monkey ◽  
Embryonic Stem ◽  
Germ Cell Differentiation

Role of Mael in early oogenesis and during germ-cell differentiation from embryonic stem cells in mice in vitro

Zygote ◽  
2013 ◽  
Vol 22 (4) ◽  
pp. 513-520 ◽  
Author(s):  
I. Bahena ◽  
E. Xu ◽  
M. Betancourt ◽  
E. Casas ◽  
Y. Ducolomb ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Differentiation ◽  
Embryonic Stem Cells ◽  
Germ Cell ◽  
Embryonic Stem ◽  
Oocyte Growth ◽  
Germ Cell Differentiation ◽  
Rnai Technology

SummaryIn a previous study, we have identified a set of conserved spermatogenic genes whose expression is restricted to testis and ovary and that are developmentally regulated. One of these genes, the transcription factor Mael, has been reported to play an essential role in mouse spermatogenesis. Nevertheless, the role of Mael in mouse oogenesis has not been defined. In order to analyse the role of Mael in mouse oogenesis, the expression of this gene was blocked during early oogenesis in mouse in vitro using RNAi technology. In addition, the role of Mael during differentiation of embryonic stem cells (ESC) into germ cells in vitro was analysed. Results show that downregulation of Mael by a specific short interfering RNA disrupted fetal oocyte growth and differentiation in fetal ovary explants in culture and the expression of several germ-cell markers in ESC during their differentiation. These results suggest that there is an important role for Mael in early oogenesis and during germ-cell differentiation from embryonic stem cells in mouse in vitro.

scholarly journals Toward a better definition of hematopoietic progenitors suitable for B cell differentiation

PLoS ONE ◽  
2020 ◽  
Vol 15 (12) ◽  
pp. e0243769
Author(s):  
Florian Dubois ◽  
Anne Gaignerie ◽  
Léa Flippe ◽  
Jean-Marie Heslan ◽  
Laurent Tesson ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Differentiation ◽  
B Cells ◽  
B Cell ◽  
Embryonic Stem ◽  
B Cell Differentiation ◽  
Hematopoietic Progenitors ◽  
Cell Based Therapy ◽  
Definition Of

The success of inducing human pluripotent stem cells (hIPSC) offers new opportunities for cell-based therapy. Since B cells exert roles as effector and as regulator of immune responses in different clinical settings, we were interested in generating B cells from hIPSC. We differentiated human embryonic stem cells (hESC) and hIPSC into B cells onto OP9 and MS-5 stromal cells successively. We overcame issues in generating CD34+CD43+ hematopoietic progenitors with appropriate cytokine conditions and emphasized the difficulties to generate proper hematopoietic progenitors. We highlight CD31intCD45int phenotype as a possible marker of hematopoietic progenitors suitable for B cell differentiation. Defining precisely proper lymphoid progenitors will improve the study of their lineage commitment and the signals needed during the in vitro process.

In vitro induction and differentiation of newborn guinea pig hippocampus neural stem cells into cells resembling inner hair cells, using artificial perilymph

2011 ◽  
Vol 125 (8) ◽  
pp. 771-775 ◽  
Author(s):  
Y Wang ◽  
M-M Dong
Keyword(s):  
Stem Cells ◽  
Neural Stem Cells ◽  
Hair Cell ◽  
Hair Cells ◽  
Guinea Pigs ◽  
Bovine Serum ◽  
Fetal Bovine Serum ◽  
Specific Antibodies ◽  
Fetal Bovine

AbstractObjective:To investigate whether artificial perilymph can induce neural stem cells, derived from the hippocampus of newborn guinea pigs, to differentiate into inner ear hair cells, in vitro.Methods:Primary neural stem cells derived from the hippocampus of newborn guinea pigs were incubated in medium containing either 10 per cent fetal bovine serum or 5, 10 or 15 per cent artificial perilymph, for three weeks. Differentiated cells were identified using immunofluorescence, Western blot and scanning electron microscopy.Results:Both fetal bovine serum and artificial perilymph induced the neural stem cells to differentiate into cells with hair-cell-specific antibodies.Conclusion:Neural stem cells can survive in both fetal bovine serum and artificial perilymph, and within these media can differentiate into cells with hair-cell-specific antibodies. This provides an experimental basis for transplantation of neural stem cells into the inner ear.

scholarly journals Lin28 Inhibits the Differentiation from Mouse Embryonic Stem Cells to Glial Lineage Cells through Upregulation of Yap1

2021 ◽  
Vol 2021 ◽  
pp. 1-12
Author(s):  
Juan Luo ◽  
Hailin Zou ◽  
Liang Deng ◽  
Xiang Sun ◽  
Ping Yuan ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Differentiation ◽  
Embryonic Stem Cells ◽  
Glial Cell ◽  
Rna Binding ◽  
Embryonic Stem ◽  
Mouse Embryonic Stem Cells ◽  
Neuronal Marker ◽  
Glial Lineage

The RNA-binding protein Lin28 regulates neurogliogenesis in mammals, independently of the let-7 microRNA. However, the detailed regulatory mechanism remains obscured. Here, we established Lin28a or Lin28b overexpression mouse embryonic stem cells (ESCs) and found that these cells expressed similar levels of the core pluripotent factors, such as Oct4 and Sox2, and increased Yap1 but decreased lineage-specific markers compared to the control ESCs. Further differentiation of these ESCs to neuronal and glial lineage cells revealed that Lin28a/b overexpression did not affect the expression of neuronal marker βIII-tubulin, but dramatically inhibited the glial lineage markers, such as Gfap and Mbp. Interestingly, overexpression of Yap1 in mouse ESCs phenocopied Lin28a/b overexpression ESCs by showing defect in glial cell differentiation. Inhibition of Yap1/Tead-mediated transcription with verteporfin partially rescued the differentiation defect of Lin28a/b overexpression ESCs. Mechanistically, we demonstrated that Lin28 can directly bind to Yap1 mRNA, and the induction of Yap1 by Lin28a in mESCs is independent of Let7. Taken together, our results unravel a novel Lin28-Yap1 regulatory axis during mESC to glial lineage cell differentiation, which may shed light on glial cell generation in vitro.

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