Saeideh Aran ¹ Mohammad Ghasem Golmohammadi ² Mohsen Sagha ^2* Kamran Ghaedi ^3*

• ¹ Department of Plant and Animal Biology, Faculty of Biotechnology, University of Isfahan, Isfahan, Iran
• ² Research Laboratory for Embryology and Stem Cells, Department of Anatomical Sciences, School of Medicine, Ardabil University of Medical Sciences, Ardabil, Iran., ardabil, Iran
• ³ Department of Cell and Molecular Biology and Microbiology, Faculty of Biological Science and Technology, University of Isfahan, Isfahan, Iran, Isfahan, Iran

Neurosphere culture is widely used to expand neural stem and progenitor cells (NSPCs) of the nervous system. Understanding the identity of NSPCs, such as the principals involved in spatiotemporal patterning, will improve our chances of using NSPCs for neurodevelopmental and brain repair studies with the ability to direct NSPCs toward distinct fates. Some reports indicate that aging can affect the nature of NSPCs over time. Therefore, in this study, we aimed to investigate how the initial neural patterning of developing NSPCs changes over time.In this research, it was presented the evidence of changing neural patterning potential in the nervous system over time. Thus, the embryonic and adult-derived NSPCs for cardinal characteristics were analyzed, and then the expression of candidate genes related to neural patterning using real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) was evaluated at various stages of embryonic (E14 and E18), neonatal, and adult brains. Finally, it was assessed the effect of cell attachment and passage on the initial neural patterning of NSPCs.The analysis of gene expression revealed that although temporal patterning is maintained in vitro, but it shows a decrease over time. Embryonic NSPCs exhibited the highest potential for retaining regional identity compared to neonatal and adult NSPCs. Additionally, it was found that culture conditions, such as cell passaging and attachment status, could affect the initial neural patterning potential, resulting in a decrease over time. Our study demonstrates that patterning potential decreases over time and aging imposes restrictions on preliminary neural patterning. These results emphasize the significance of patterning in the nervous system and the close relationship between it and fate determination, raising questions about the application of aged NSPCs in treatment of neurodegenerative diseases.