Abstract
Cell-cell interaction as one of the niche signals plays an important role in the balance of stem cell quiescence and proliferation or differentiation. In order to address the effect and the possible mechanisms of cell-cell connection on neural stem/progenitor cells (NSCs/NPCs) proliferation and differentiation, upon passaging, NSCs/NPCs were either dissociated into single cell as usual (named Group I) or mechanically triturated into a mixture of single cell and small cell clusters containing direct cell-cell connections (named Group II). Then the biological behaviors including proliferation and differentiation of NSCs/NPCs were observed. Moreover, the expression of gap junction channel, neurotrophic factors and the phosphorylation status of MAPK signals were compared to investigate the possible mechanisms. Our results showed that, in comparison to the counterparts in Group I, NSCs/NPCs in Group II survived well with preferable neuronal differentiation. In coincidence with this, the expression of connexin 45 (Cx45), as well as brain derived neurotrophic factor (BDNF) and neurotrophin 3 (NT-3) in Group II were significantly higher than those in Group I. Phosphorylation of ERK1/2 and JNK2 were significantly upregulated in Group II too, while no change was found about p38. Furthermore, the differences of NSCs/NPCs biological behaviors between Group I and II completely disappeared when ERK and JNK phosphorylation were inhibited. These results indicated that cell-cell connection in Group II enhanced NSCs/NPCs survival, proliferation and neuronal differentiation through upregulating the expression of gap junction and neurotrophic factors. MAPK signals- ERK and JNK might contribute to the enhancement. Efforts for maintaining the direct cell-cell connection are worth making to provide more favorable niches for NSCs/NPCs survival, proliferation and neuronal differentiation.
Highlights
The neurobiological behaviors of neural stem/progenitor cells (NSCs/NPCs), such as dormancy, proliferation or differentiation are affected by the environment where they are located (Bjornsson et al, 2015; Reinhard et al, 2016)
NSCs/NPCs were isolated from cerebral cortex of rat embryos on embryonic day 14 (E14) to 15 (E15) and cultured in serum-free growth medium following the protocol of Gage et al (1995) and optimized in our lab (Lu et al, 2011, 2013)
Neurospheres gradually formed in both groups (Figure 2C), neurospheres in Group II were significantly larger, indicating the quicker growth of NSCs/NPCs (Figure 2D)
Summary
The neurobiological behaviors of neural stem/progenitor cells (NSCs/NPCs), such as dormancy, proliferation or differentiation are affected by the environment where they are located (Bjornsson et al, 2015; Reinhard et al, 2016). Loss of the 3-D specific niche signals, growing on flat and hard glass or plastic substrates leads to the dramatic change of NSCs/NPCs behaviors (Pampaloni et al, 2007; Saha et al, 2008; Justice et al, 2009). Taking all these into account, it indicated the spatial relationship between NSCs/NPCs and their neighbor cells are critical for cell in vitro growth. In the previous in vitro study approaches, the interactions of cells with one another and with the resulting extracellular microenvironments changes had not been properly addressed (Solozobova et al, 2012)
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