Regulation effect of osteoblasts towards osteocytes by silk fibroin encapsulation

doi:10.1007/s11706-022-0617-5

Front. Mater. Sci.

2022, Vol. 16

Issue (4) : 220617 https://doi.org/10.1007/s11706-022-0617-5

RESEARCH ARTICLE

Regulation effect of osteoblasts towards osteocytes by silk fibroin encapsulation

Dandan LUO^1,^2,³, Rui ZHANG^1,², Shibo WANG^1,², M. Zubair IQBAL^1,², Ruibo ZHAO^1,²(

), Xiangdong KONG^1,²

¹. Institute of Smart Biomedical Materials, School of Materials Science and Engineering, Zhejiang Sci-Tech University, Hangzhou 310018, China
². Zhejiang-Mauritius Joint Research Center for Biomaterials and Tissue Engineering, Zhejiang Sci-Tech University, Hangzhou 310018, China
³. School of Textile Science and Engineering, Zhejiang Sci-Tech University, Hangzhou 310018, China

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Abstract

Herein, the rational design micromilieus involved silk fibroin (SF)-based materials have been used to encapsulate the osteoblasts, forming an extracellular coated shell on the cells, which exhibited the high potential to shift the regulation of osteoblasts to osteocytes by encapsulation cues. SF coating treated cells showed a change in cell morphology from osteoblasts-like to osteocytes-like shape compared with untreated ones. Moreover, the expression of alkaline phosphatase (ALP), collagen I (Col I) and osteocalcin (OCN) further indicated a potential approach for inducing osteoblasts regulation, which typically accelerates calcium deposition and cell calcification, presenting a key role for the SF encapsulation in controlling osteoblasts behavior. This discovery showed that SF-based cell encapsulation could be used for osteoblasts behavior regulation, which offers a great potential to modulate mammalian cells’ phenotype involving alternating surrounding cues.

Keywords cell encapsulation silk fibroin osteoblasts modulation cell differentiation cell calcification

Corresponding Author(s): Ruibo ZHAO

About author: Tongcan Cui and Yizhe Hou contributed equally to this work.

Issue Date: 19 October 2022

Cite this article:

Dandan LUO,Rui ZHANG,Shibo WANG, et al. Regulation effect of osteoblasts towards osteocytes by silk fibroin encapsulation[J]. Front. Mater. Sci., 2022, 16(4): 220617.

URL:

https://academic.hep.com.cn/foms/EN/10.1007/s11706-022-0617-5
https://academic.hep.com.cn/foms/EN/Y2022/V16/I4/220617

Tab.1 qPCR primers

Fig.1 Schematic showing electrostatic layer-by-layer encapsulation of PEI (+) and SF (?) on the negatively charged MC3T3-E1 cell surface. The cells treated after 1, 2, and 3 cyclic layer-by-layer encapsulation process (Cycles 1, 2, and 3) are denoted as coated MC3T3-E1 (Cycle 1), coated MC3T3-E1 (Cycle 2), and coated MC3T3-E1 (Cycle 3), respectively.

Fig.2 Preparation of coating treated MC3T3-E1 cells. Cell viability of MC3T3-E1 after co-culture with various concentrations of (a) SF and (b) PEI for 24 h. (c) Cell viability of MC3T3-E1 after co-culture with 50 μg·mL^?1 of PEI for various times. (d) Zeta potentials of MC3T3-E1, MC3T3-E1@PEI (after PEI treatment) and MC3T3-E1@PEI@SF (after PEI and SF treatment). (e) Brightfield microscope observation and (f) live/dead staining images of MC3T3-E1 and coating treated MC3T3-E1 cells. Green: live cell; red: dead cell.

Fig.3 (a) SEM observation and (b) CLSM observation of MC3T3-E1 and coating treated MC3T3-E1 cells. Green: SF-FITC, SF layer (labeled by FITC); red: cell membrane.

Fig.4 Cell proliferation of MC3T3-E1 and coating treated MC3T3-E1 cells. (a) Cell viability of coating treated MC3T3-E1 cells after culture for 1, 3, 7, and 14 d. (b) Brightfield microscope observation and (c) live/dead staining images of MC3T3-E1 and coating treated MC3T3-E1 cells after culture for 14 d. Green: live cell; red: dead cell. The results are the mean ± SD of triplicate experiments, and statistical significance is analyzed as compared with the untreated MC3T3-E1 group: **, p < 0.01; ***, p < 0.001.

Fig.5 Cell morphology of MC3T3-E1 and coating treated MC3T3-E1 cells after culture for 7 d. (a) Rhodamine-conjugated phalloidin (red) and DAPI (blue) staining images. Quantitative results of cell: (b) area, (c) major axis length, (d) minor axis length, and (e) aspect ratio. The statistical significance is analyzed as compared with the MC3T3-E1 group: *, p < 0.05; **, p < 0.01.

Fig.6 Osteogenic differentiation of coating treated MC3T3-E1 cells. (a) ALP activity (normalized by the total protein content) of MC3T3-E1 and coating treated MC3T3-E1 cells after culture for 3, 7, and 14 d. Relative expression levels of (b) specific genes and (c) proteins for osteogenic differentiation. The results are the mean ± SD of triplicate experiments, statistical significance is analyzed as compared with the MC3T3-E1 group: *, p < 0.05; **, p < 0.01; ***, p < 0.001.

Fig.7 ARS staining of MC3T3-E1 and coating treated MC3T3-E1 cells. (a) Brightfield microscopy observation images of MC3T3-E1 and coating treated MC3T3-E1 cells after culture for 21 d. (b) ARS staining area quantification of brightfield microscope observation images. The statistical significance is analyzed as compared with the MC3T3-E1 group: *, p < 0.05; **, p < 0.01.

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