The influence of 3D-printed polycaprolactone scaffolds coated with platelet-rich plasma on the biological functions of dental pulp cells

doi:10.3877/cma.j.issn.1674-1366.2017.03.004

Abstract

Abstract:

Objective

To study the influence of different 3D-printed polycaprolactone scaffolds coated with platelet-rich plasma on the adhesion, proliferation and differentiation of dental pulp cells (DPCs) .

Methods

DPCs were seeded on the scaffold of three groups. Cell attachment, proliferation and ALP activity were evaluated with immunofluorescence staining, CCK-8 assay and ALP kit, respectively. The expression of osteogenic genes was determined with RT-PCR.

Results

More cell attachment was found on the freeze-dried PRP-PCL scaffold (1999.33/field) than the rest (1043.33 and 843.00/field, F= 19.36, P<0.01) . In the cell proliferation test, the amount of cells was increased in all groups but there was no statistically significant difference between groups of the same day. The number of migrated cells at the bottom of the transwell chamber in the freeze-dried PRP-PCL scaffold group (279.75/field) was significantly higher than that of the gelatinous PRP-PCL scaffold group (167.00/field) and the bare PRP-PCL scaffold group (134.75/field) (F_f-b= 7.45, F_g-b= 1.88, P<0.01) . There was no significant difference in the migrated cells between the gelatinous PRP-PCL scaffold and bare PRP-PCL scaffold. The ALP activity of the freeze-dried PRP-PCL scaffold (ALP_{7 d}= 12.57 U/gprot, ALP_{14 d}= 23.20 U/gprot, ALP_{21 d}= 58.98 U/gprot) was significantly higher than that of the gelatinous PRP-PCL scaffold (ALP_{7 d}= 6.65 U/gprot, ALP_{14 d}= 13.35 U/gprot, ALP_{21 d}= 47.83 U/gprot) and the bare PCL scaffold (ALP_{7 d}= 5.93 U/gprot、ALP_{14 d}= 15.56 U/gprot、ALP_{21 d}= 53.74 U/gprot) on 7, 14 and 21 d (F_{7 d}= 3.20, P_{7 d}<0.01; F_{14 d}= 5.34, P_{14 d}<0.01; F_{21 d}= 6.04, P_{21 d}<0.01) (F_{7 d}= 3.60, P_{7 d}= 0.04; F_{14 d}= 4.14, P_{14 d}<0.01; F_{21 d}= 2.84, P_{21 d}= 0.01) . However, no statistically significant difference was found in the ALP activity between the gelatinous PRP-PCL scaffold and the bare PCL scaffold except for that on 21 d. Except for the expression of OCN by DPSCs on the gelatinous PRP-PCL scaffold on 14 d, the expression of RUNX2 and OCN by DPSCs on freeze-dried PRP-PCL scaffold (OCN_{7 d}= 4.67, RUNX2_{7 d}= 2.32, RUNX2_{14 d}= 5.88) and gelatinous PRP-PCL scaffold (OCN_{7 d}= 2.60, RUNX2_{7 d}= 2.23, RUNX2_{14 d}= 4.67) on 7 and 14 d were significantly higher than that on the bare scaffold (OCN_{7 d}= 1.00, RUNX2_{7 d}= 1.00, RUNX2_{14 d}= 1.00) (F_{7 d}= 11.1, P_{7 d}<0.01; F_{7 d}= 3.20, P_{7 d}= 0.04; F_{14 d}= 11.80, P_{14 d}<0.01) . The expression of OCN on7 d and RUNX2 on 14 d on the freeze-dried PRP-PCL scaffold (OCN_{7 d}= 4.67) was significantly higher than that of gelatinous PRP-PCL scaffold (OCN_{7 d}= 2.60, F= 6.26, P<0.01) . However, no statistical significance was found in the expression of OCN on 14 d between the freeze-dried PRP-PCL scaffolds and the gelatinous PRP-PCL scaffolds.

Conclusion

The freeze-dried PRP-PCL scaffolds were more favorable for the adhesion, proliferation and differentiation of DPCs compared with the gelatinous PRP-PCL scaffolds.

Key words: Platelet-rich plasma, Printing, Three-Dimensional, Polycaprolactone, Tissue scaffold, Dental pulp cells, Biological functions

Junda Li, Meilin Chen, Xiaoying Wei, Yishan Hao, Jinming Wang. The influence of 3D-printed polycaprolactone scaffolds coated with platelet-rich plasma on the biological functions of dental pulp cells[J]. Chinese Journal of Stomatological Research(Electronic Edition), 2017, 11(03): 149-156.

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Figures/Tables 5

References 20

[1]	Yao Q, Wei B, Liu N，et al. Chondrogenic regeneration using bone marrow clots and a porous polycaprolactone-hydroxyapatite scaffold by three-dimensional printing[J]. Tissue Eng Part A，2014，7-8(21):1388-1397.
[2]	Betsch M, Schneppendahl J, Thuns S，et al. Bone marrow aspiration concentrate and platelet rich plasma for osteochondral repair in a porcine osteochondral defect model[J]. PLoS One，2013，8(8):e71602.
[3]	Gronthos S, Mankani M, Brahim J，et al. Postnatal human dental pulp stem cells（DPSCs）in vitro and in vivo[J]. Proc of the Natl Acad Sci U S A，2000，97(25):13625-13630.
[4]	Pati F, Song TH, Rijal G，et al. Ornamenting 3D printed scaffolds with cell-laid extracellular matrix for bone tissue regeneration[J]. Biomaterials，2015(37):230-241.
[5]	Yoon H, Kim GH, Koh YH. A micro-scale surface-structured PCL scaffold fabricated by a 3D plotter and a chemical blowing agent[J]. J Biomater Sci Polym Ed，2010，21(2):159-170.
[6]	Tarafder S, Koch A, Jun Y，et al. Micro-precise spatiotemporal delivery system embedded in 3D printing for complex tissue regeneration[J]. Biofabrication，2016，8(2):25003.
[7]	Jang CH, Cho YB, Choi CH，et al. Effect of umbilical cord serum coated 3D PCL/alginate scaffold for mastoid obliteration [J]. Int J Pediatr Otorhinolaryngol，2014，78(7):1061-1065.
[8]	Wang H, Wu G, Zhang J，et al. Osteogenic effect of controlled released rhBMP-2 in 3D printed porous hydroxyapatite scaffold [J]. Colloids Surf B Biointerfaces，2016(141):491-498.
[9]	Jeong CG, Hollister SJ. A comparison of the influence of material on in vitro cartilage tissue engineering with PCL，PGS，and POC 3D scaffold architecture seeded with chondrocytes[J]. Biomaterials，2010，31(15):4304-4312.
[10]	Jo S, Kang SM, Park SA，et al. Enhanced adhesion of preosteoblasts inside 3D PCL scaffolds by polydopamine coating and mineralization[J]. Macromol Biosci，2013，13(10):1389-1395.
[11]	Kang SW, Bae JH, Park SA，et al. Combination therapy with BMP-2 and BMSCs enhances bone healing efficacy of PCL scaffold fabricated using the 3D plotting system in a large segmental defect model[J]. Biotechnol Lett，2012，34(7):1375-1384.
[12]	Reed GL. Platelet secretory mechanisms[J]. Semin Thromb Hemost，2004，30(4):441-450.
[13]	Chang SH, Hsu YM, Wang YJ，et al. Fabrication of pre-determined shape of bone segment with collagen-hydroxyapatite scaffold and autogenous platelet-rich plasma[J]. J Mater Sci Mater Med，2009，20(1):23-31.
[14]	Lee JH, Nam J, Kim HJ，et al. Comparison of three different methods for effective introduction of platelet-rich plasma on PLGA woven mesh[J]. Biomed Mater，2015，10(2):25002.
[15]	Zhong D, Wang CG, Yin K，et al. In vivo ossification of a scaffold combining β-tricalcium phosphate and platelet-rich plasma[J]. Exp Ther Med，2014，8(5):1381-1388.
[16]	Pietramaggiori G, Kaipainen A, Czeczuga JM，et al. Freeze-dried platelet-rich plasma shows beneficial healing properties in chronic wounds[J]. Wound Repair Regen，2006，14(5):573-580.
[17]	Nakatani Y, Agata H, Sumita Y，et al. Efficacy of freeze-dried platelet-rich plasma in bone engineering[J]. Arch Oral Biol，2017(73):172-178.
[18]	Sell SA, Wolfe PS, Ericksen JJ，et al. Incorporating platelet-rich plasma into electrospun scaffolds for tissue engineering applications[J]. Tissue Eng Part A，2011，17(21-22):2723-2737.
[19]	胡军红，谢建红，陆玲，等.标本溶血对血清酶测定影响的研究[J].吉林医学，2014，35(1):115-116.
[20]	郑铁生，叶立新，薛锦，等.不同激活型缓冲体系对碱性磷酸酶活性测定的影响[J].临床检验杂志，2005，23(6):423-424.

组别	支架数	A ₄₅₀
组别	支架数	1d	3d	5d	7d
冻干组	3	0.17 ± 0.03	0.48 ± 0.06	0.61 ± 0.02	0.97 ± 0.03
凝胶组	3	0.17 ± 0.04	0.38 ± 0.11	0.59 ± 0.13	0.96 ± 0.17
单纯支架组	3	0.27 ± 0.02	0.58 ± 0.33	0.81 ± 0.05	1.28 ± 0.15
F值	?	3.14	1.73	2.67	4.22
P值	?	0.15	0.28	0.18	0.10

组别	支架数	A ₄₅₀
组别	支架数	1d	3d	5d	7d
冻干组	3	0.17 ± 0.03	0.48 ± 0.06	0.61 ± 0.02	0.97 ± 0.03
凝胶组	3	0.17 ± 0.04	0.38 ± 0.11	0.59 ± 0.13	0.96 ± 0.17
单纯支架组	3	0.27 ± 0.02	0.58 ± 0.33	0.81 ± 0.05	1.28 ± 0.15
F值	?	3.14	1.73	2.67	4.22
P值	?	0.15	0.28	0.18	0.10

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