Cell–Cell Contact Mediates Gene Expression and Fate Choice of Human Neural Stem/Progenitor Cells
Abstract
:1. Introduction
2. Materials and Methods
2.1. Experimental Design
2.2. hiPSC-NPC Culture
2.3. Plating Density and In Vitro Differentiation
2.4. Immunocytochemistry
2.5. RNA Isolation and Sequencing
2.6. Sequencing Analysis
2.6.1. GO Enrichment Analysis
2.6.2. KEGG Enrichment Analysis
2.6.3. Assessment of Cell Lineage Gene Programs from RNA-Seq Data
2.7. RT-qPCR Validation
2.8. Statistical Analysis
3. Results
3.1. Differentiation Capacity of High- vs. Low- Density hiPSC-NPCs
3.2. Plating Density Alters Transcriptional Programs in hiPSC-NPCs
3.3. Functional Enrichment Analysis of Differentially Expressed Genes
3.4. Assessment of Culture Density on Biasing Cell Lineage Specification
4. Discussion
4.1. Growth Factors and Sprouty
4.2. Differential Wnt Signaling in High/Low Cell Density
4.3. Higher Culture Density Promotes Hippo-Related Signaling
4.4. NOTCH Signaling in Higher Density
4.5. Higher Plating Density Promotes Neuronal and Prevents Astroglial Fate
Cell Density Biases Interneuron Fate
4.6. RT-qPCR Validation Corroborates Functional Immunostaining
4.7. Application to Transplantation Studies
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Conflicts of Interest
References
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PROBE (Stock; Cat No.) | Dilution | Target |
---|---|---|
β-III-Tubulin (BioLegend; MMS-435P) | 1:500 | Immature neurons |
GFAP (Abcam; ab33922) | 1:500 | Astrocytes |
CNPase (Abcam; ab6319) | 1:500 | Oligodendrocytes |
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McIntyre, W.B.; Karimzadeh, M.; Riazalhosseini, Y.; Khazaei, M.; Fehlings, M.G. Cell–Cell Contact Mediates Gene Expression and Fate Choice of Human Neural Stem/Progenitor Cells. Cells 2022, 11, 1741. https://0-doi-org.brum.beds.ac.uk/10.3390/cells11111741
McIntyre WB, Karimzadeh M, Riazalhosseini Y, Khazaei M, Fehlings MG. Cell–Cell Contact Mediates Gene Expression and Fate Choice of Human Neural Stem/Progenitor Cells. Cells. 2022; 11(11):1741. https://0-doi-org.brum.beds.ac.uk/10.3390/cells11111741
Chicago/Turabian StyleMcIntyre, William B., Mehran Karimzadeh, Yasser Riazalhosseini, Mohamad Khazaei, and Michael G. Fehlings. 2022. "Cell–Cell Contact Mediates Gene Expression and Fate Choice of Human Neural Stem/Progenitor Cells" Cells 11, no. 11: 1741. https://0-doi-org.brum.beds.ac.uk/10.3390/cells11111741