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. 2019 Jan 18;39(1):BSR20181748.
doi: 10.1042/BSR20181748. Print 2019 Jan 31.

The influence of the stiffness of GelMA substrate on the outgrowth of PC12 cells

Yibing Wu  1   2   3 , Yang Xiang  3 , Jiehua Fang  1   2 , Xiaokeng Li  1   2 , Zunwen Lin  4 , Guangli Dai  5 , Jun Yin  1   2 , Peng Wei  6 , Deming Zhang  7   2
Affiliations

The influence of the stiffness of GelMA substrate on the outgrowth of PC12 cells

Yibing Wu et al. Biosci Rep. .

Abstract

Recent studies have shown the importance of cell-substrate interaction on neurone outgrowth, where the Young's modulus of the matrix plays a crucial role on the neurite length, migration, proliferation, and morphology of neurones. In the present study, PC12 cells were selected as the representative neurone to be cultured on hydrogel substrates with different stiffness to explore the effect of substrate stiffness on the neurone outgrowth. By adjusting the concentration of gelatin methacryloyl (GelMA), the hydrogel substrates with the variation of stiffnesses (indicated by Young's modulus) from approximately 3-180 KPa were prepared. It is found that the stiffness of GelMA substrates influences neuronal outgrowth, including cell viability, adhesion, spreading, and average neurite length. Our results show a critical range of substrate's Young's modulus that support PC12 outgrowth, and modulate the cell characteristics and morphology. The present study provides an insight into the relationship between the stiffness of GelMA hydrogel substrates and PC12 cell outgrowth, and helps the design and optimization of tissue engineering scaffolds for nerve regeneration.

Keywords: PC12 cells; cell spreading; cell viability; gelatin methacryloyl; substrate Young’s modulus.

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Conflict of interest statement

The authors declare that there are no competing interests associated with the manuscript.

Figures

Figure 1
Figure 1. The schematic of the synthesis of GelMA process
(A) Synthesis process of GelMA, and (B) photocrosslinking of GelMA hydrogel.
Figure 2
Figure 2. Experimental setup of the uniaxial tensile tests
Figure 3
Figure 3. Definitaion the outgrowth characteristics and morphology of PC12 cells
(A) Live-dead staining of live cells (green) and dead cells (red) in PC12 after 3 days of culture on 10% GelMA substrate. (B) The distance from the center of the cell to the end of the neurite (the orange line) represents length of neurite (Ln). The area of cell contour (the blue grid) represents spreading area (Ss).
Figure 4
Figure 4. The Young’s modulus of GelMA hydrogels with different concentrations
Bars represent average ± S.D. **P<0.01.
Figure 5
Figure 5. The presentative fluorography of PC12 cells adhesion on the surfaces of GelMA hydrogel substrates with different concentrations
Figure 6
Figure 6. PC12 cells adhesion and viability
(A) PC12 cells adhesion on GelMA substrates with different concentrations after 24 h of inoculation. (B) Cell viability after 24, 75, and 120 h of inoculation. Bars represent average ± S.D. (*P<0.05, **P<0.01).
Figure 7
Figure 7. PC12 cells spreading area and neurite length
Cell spreading area (A) and neurite length (B) of PC12 cells after 24, 75, and 120 h of inoculation. Points represent average ± S.D. (*P<0.05, **P<0.01).
See this image and copyright information in PMC

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