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Journal of Peking University(Health Sciences) ›› 2019, Vol. 51 ›› Issue (1): 28-34. doi: 10.19723/j.issn.1671-167X.2019.01.006

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Effects of electrospun collagen nanofibrous matrix on the biological behavior of human dental pulp cells

Qian-li ZHANG1,Chong-yang YUAN1,Li LIU2,Shi-peng WEN2,△(),Xiao-yan WANG1,△()   

  1. 1. Department of Cariology and Endodontology, Peking University School and Hospital of Stomatology & National Clinical Research Center for Oral Diseases & National Engineering Laboratory for Digital and Material Technology of Stomatology & Beijing Key Laboratory of Digital Stomatology, Beijing 100081, China
    2. Beijing Engineering Research Centre of Advanced Elastomers, Beijing University of Chemical Technology, Beijing 100029, China
  • Received:2018-10-10 Online:2019-02-18 Published:2019-02-26
  • Contact: Shi-peng WEN,Xiao-yan WANG E-mail:wensp@mail.buct.edu.cn;wangxiaoyan@pkuss.bjmu.edu.cn
  • Supported by:
    Supported by the National Natural Science Foundation of China(51503004)

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Abstract:

Objective: To compare cell adhesion, proliferation and odontoblastic differentiation of human dental pulp cells (hDPCs) on electrospun collagen nanofibrous matrix (Col_NFM) with that on collagen flat film (Col-FF), to investigate the biological effect of collagen nanofibrous matrix on hDPCs. Methods: The surface morphology of the two different collagen scaffold was analyzed by scanning electron microscopy (SEM), and the contact angle and the swelling ratio were also measured. Then hDPCs were implanted on the two different collagen scaffolds, the cell morphology was observed using SEM and laser scanning microscope (LSM), and cell proliferation was evaluated by the CCK-8 assay. After hDPCs cultured on the two different collagen scaffold with odontoblastic medium for 14 days, the expression of odontoblastic differentiation related genes was detected by real-time PCR, and alizarin red staining was used to test the formation of mineralized nodules. Results: From the SEM figures, the fibers’ diameter of Col_NFM was (884±159) nm, and there were abundant three dimensional connected pore structures between the fibers of Col_NFM, while the surface of Col_FF was completely flat without pore structure. The contact angle at 0 s of Col_NFM was 85.03°±4.45°, and that of Col_FF was 98.98°±5.81°. The swelling ratio of Col_NFM was approximately 3 folds compared with dry weight sample, while that of Col_FF was just 1 fold. Thus Col_NFM indicated better hydrophilicity and swelling property. SEM and LSM showed that hDPCs on Col_NFM presented an irregular and highly branched phenotype, and could penetrate into the nanofibrous scaffold. In contrast, the cells were spread only on the surface of Col_FF with a spindle-shaped morphology. CCK-8 assays showed that hDPCs on Col_NFM showed higher proliferation rate than on Col_FF. After hDPCs were cultured on the two different collagen scaffolds with odontoblastic medium for 14 days, more expressions of odontoblastic differentiation related genes, such as dentin sialophosphoprotein (DSPP) and dentin matrix proten-1 (DMP1) were determined in Col_NFM group (P<0.05), and more mineralization depositions were also observed in Col_NFM group according to the results of alizarin red staining. Conclusion: Col_NFM with nanoscale microstructure achieves better hydrophilic and swelling properties than Col_FF, and hDPCs cultured with Col_NFM present higher activity on cell adhesion, proliferation and odontoblastic differentiation.

Key words: Collagen, Electrospinning, Pulp regeneration, Nanofibers, Scaffold

CLC Number: 

  • R781.3

Table 1

The chain of reaction primers"

Gene Gene sequence (5'-3')
DSPP Forward: ATATTGAGGGCTGGAATGGGGA
Reverse: TTTGTGGCTCCAGCATTGTCA
DMP1 Forward: AGGAAGTCTCGCATCTCAGAG
Reverse: TGGAGTTGCTGTTTTCTGTAGAG
GAPDH Forward: GAAGGTGAAGGTCGGAGTC
Reverse: GAGATGGTGATGGGATTTC

Figure 1

Scanning electron microscopy view of the scaffolds A,Col_NFM group; B, Col_FF group."

Figure 2

Contact angles of the scaffolds A,Col_NFM group, 0 s; B, Col_NFM group, 8 s; C, Col_FF group, 0 s; D, Col_FF group, 8 s."

Figure 3

Swelling ratio of the scaffolds *P<0.05, compared with the Col_FF group at the same time."

Figure 4

Scanning electron microscopy view of hDPCs cultured on different surfaces A, B, Col_NFM group; C, D, Col_FF group. B and D were the red box region view of A and C at high magnification."

Figure 5

Laser scanning microscope view of hDPCs cultured on different surfaces A, B, Col_NFM group; C, Col_FF group."

Figure 6

The proliferation of hDPCs on different scaffolds *P<0.05, compared with the Col_FF group at the same time."

Figure 7

Quantitative real-time PCR analysis of DSPP and DMP1 gene expression of hDPCs for 14 days *P<0.05, compared with the control group; # P<0.05, compared with the Col_FF group. Abbreviations as in Table 1."

Figure 8

Alizarin red staining of hDPCs cultured on different scaffolds at day 14"

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