Int J Stomatol ›› 2020, Vol. 47 ›› Issue (3): 278-285.doi: 10.7518/gjkq.2020061

• Original Articles • Previous Articles     Next Articles

Effects of inorganic bovine bone treated with low temperature argon-oxygen plasma on the adhesion, proliferation, and differentiation of MC3T3-E1 cells

Ma Kai,Li Hao,Zhao Hongmei,Wang Yongliang,Liu Jie,Bai Na()   

  1. Dept. of Prosthodontics, The Affiliated Hospital of Qingdao University, School of Stomatology of Qingdao University, Qingdao 266003, China
  • Received:2019-10-22 Revised:2020-01-06 Online:2020-05-01 Published:2020-05-08
  • Contact: Na Bai
  • Supported by:
    National Natural Science Foundation of China(81500882)


Objective To study the adhesion, proliferation, and differentiation of mouse embryonic osteoblast MC3T3-E1 on inorganic bovine bone treated with low temperature argon-oxygen plasma. Methods After the surface activation of inorganic bovine bone by low temperature argon-oxygen plasma, the surface morphology of inorganic bovine bone before and after treatment was observed by scanning electron microscopy (SEM), and the surface elemental composition was detected by X-ray photoelectron spectroscopy. The MC3T3-E1 cells were inoculated on the surface of inorganic bovine bone treated with low temperature argon-oxygen plasma, and the cell adhesion morphology was observed by SEM. The proliferation of cells on days 1, 3, and 5 was detected using the CCK-8 method, and the differentiation of cells on days 7 and 14 was detected using the alkaline phosphatase (ALP) method. The control group comprised the inorganic bovine bone without treatment. Results No significant change was observed in the surface morphology of the inorganic bovine bone of the control and the argon-oxygen groups. Relative to the composition of the material elements in the control group, the argon-oxygen group had decreased surface C element and increased O, Ca, and P elements. Under SEM, the cell adhesion of the argon-oxygen group was more complete, and the cell extended pseudopodia. The cell proliferation test results showed that the number of cell proliferation in the argon-oxygen group was significantly higher than that in the control group at days 1, 3, and 5. The cell differentiation results showed that the ALP activity in the argon-oxygen group was higher than that in control group at day 14. Conclusion The inorganic bovine bone treated with low temperature argon-oxygen plasma can promote the adhesion, proliferation, and differentiation of mouse embryonic osteoblast cells.

Key words: low temperature plasma, inorganic bovine bone, mouse embryonic osteoblast MC3T3-E1, guided bone regeneration

CLC Number: 

  • Q25


Fig 1

Schematic diagram of low temperature argon-oxygen plasma system"

Fig 2

The relationship between treatment time of low temperature ar-gon-oxygen plasma system and surface temperature of materials"

Fig 3

The excessive temperature caused the fracture of the internal connecting structure of the inorganic bone SEM × 2 000"

Fig 4

The surface morphology of the experimental group and control group SEM"

Tab 1

XPS analysis of the percentage of bone surface elements in two groups %"

组别 C1s O1s P2p Ca2p
对照 50.92 37.05 5.94 6.10
实验 43.09 43.11 6.72 7.08

Fig 5

XPS detected changes in bone surface elements in the two groups"

Fig 6

The percentage of surface elements in the experimental group after immediate and 2 days of exposure"

Fig 7

MC3T3-E1 cell morphology on the surface of the control group and the experimental group SEM"

Fig 8

Percentage of adherent surface area of osteoblasts in the experi-mental group and control group"

Fig 9

Proliferation of MC3T3-E1 cells"

Fig 10

ALP activity of MC3T3-E1 cells"

[1] Salamanca E, Pan YH, Tsai AI , et al. Enhancement of osteoblastic-like cell activity by glow discharge plasma surface modified hydroxyapatite/β-tricalcium phosphate bone substitute[J]. Materials (Basel), 2017,10(12):E1347.
[2] Tanaka M, Haniu H, Kamanaka T , et al. Physico-chemical, in vitro, and in vivo evaluation of a 3D unidirectional porous hydroxyapatite scaffold for bone regeneration[J]. Materials (Basel), 2017,10(1):E33.
doi: 10.3390/ma10010033 pmid: 28772390
[3] Amerio P, Vianale G, Reale M , et al. The effect of deproteinized bovine bone on osteoblast growth factors and proinflammatory cytokine production[J]. Clin Oral Implants Res, 2010,21(6):650-655.
doi: 10.1111/j.1600-0501.2009.01881.x pmid: 20666792
[4] Huh JB, Kim SE, Song SK , et al. The effect of im-mobilization of heparin and bone morphogenic pro-tein-2 to bovine bone substitute on osteoblast-like cell’s function[J]. J Adv Prosthodont, 2011,3(3):145-151.
doi: 10.4047/jap.2011.3.3.145 pmid: 22053246
[5] Rolvien T, Barbeck M, Wenisch S , et al. Cellular mechanisms responsible for success and failure of bone substitute materials[J]. Int J Mol Sci, 2018,19(10):E2893.
doi: 10.3390/ijms19102893 pmid: 30249051
[6] Moriguchi Y, Lee DS, Chijimatsu R , et al. Impact of non-thermal plasma surface modification on porous calcium hydroxyapatite ceramics for bone regenera-tion[J]. PLoS One, 2018,13(3):e0194303.
doi: 10.1371/journal.pone.0194303 pmid: 29538457
[7] Yang J, Pu Y, Miao DG , et al. Fabrication of durably superhydrophobic cotton fabrics by atmospheric pressure plasma treatment with a siloxane precursor[J]. Polymers (Basel), 2018,10(4):E460.
doi: 10.3390/polym10040460 pmid: 30966495
[8] Schmitt C, Lutz R, Doering H , et al. Bio-Oss® blocks combined with BMP-2 and VEGF for the regenera-tion of bony defects and vertical augmentation[J]. Clin Oral Implants Res, 2013,24(4):450-460.
doi: 10.1111/j.1600-0501.2011.02351.x pmid: 22092937
[9] 高媛媛, 周子谦, 柳慧芬 , 等. RGD多肽修饰的无机小牛骨粉对MC3T3-E1细胞黏附、增殖及分化的影响[J]. 口腔医学研究, 2015,31(4):354-358.
Gao YY, Zhou ZQ, Liu HF , et al. Adhesion, prolife-ration and differentiation of MC3T3-E1 cell on bovine bone substitute modified with RGD peptide[J]. J Oral Sci Res, 2015,31(4):354-358.
[10] Kong MG, Kroesen G, Morfill G , et al. Plasma me-dicine: an introductory review[J]. New J Phys, 2009,11(11):115012.
[11] Kim JH, Lee MA, Han GJ , et al. Plasma in dentistry: a review of basic concepts and applications in den-tistry[J]. Acta Odontol Scand, 2014,72(1):1-12.
doi: 10.3109/00016357.2013.795660 pmid: 24354926
[12] Duske K, Koban I, Kindel E , et al. Atmospheric plasma enhances wettability and cell spreading on dental implant metals[J]. J Clin Periodontol, 2012,39(4):400-407.
doi: 10.1111/j.1600-051X.2012.01853.x pmid: 22324415
[13] Seon GM, Seo HJ, Kwon SY , et al. Titanium surface modification by using microwave-induced argon plasma in various conditions to enhance osteoblast biocompatibility[J]. Biomater Res, 2015,19:13.
doi: 10.1186/s40824-015-0034-2 pmid: 26331083
[14] Choi SH, Jeong WS, Cha JY , et al. Corrigendum: time-dependent effects of ultraviolet and nonthermal atmospheric pressure plasma on the biological ac-tivity of titanium[J]. Sci Rep, 2016,6:36430.
doi: 10.1038/srep36430 pmid: 27834351
[15] Bárdos L, Baránková H . Cold atmospheric plasma: sources, processes, and applications[J]. Thin Solid Films, 2010,518(23):6705-6713.
doi: 10.1016/j.tsf.2010.07.044
[16] Lee EJ, Kwon JS, Uhm SH , et al. The effects of non-thermal atmospheric pressure plasma jet on cellular activity at SLA-treated titanium surfaces[J]. Curr Appl Phys, 2013,13:S36-S41.
[17] Hayashi R, Ueno T, Migita S , et al. Hydrocarbon deposition attenuates osteoblast activity on titanium[J]. J Dent Res, 2014,93(7):698-703.
doi: 10.1177/0022034514536578 pmid: 24868012
[18] Pham PV . Cleaning of graphene surfaces by low-pressure air plasma[J]. R Soc Open Sci, 2018,5(5):172395.
doi: 10.1098/rsos.172395 pmid: 29892425
[19] França R, Samani TD, Bayade G , et al. Nanoscale surface characterization of biphasic calcium pho-sphate, with comparisons to calcium hydroxyapatite and β-tricalcium phosphate bioceramics[J]. J Colloid Interface Sci, 2014,420:182-188.
doi: 10.1016/j.jcis.2013.12.055 pmid: 24559717
[20] 马楚凡, 李冬梅, 李贺军 , 等. 微弧氧化方法在钛表面注入钙磷离子及对成骨细胞早期附着的影响[J]. 第一军医大学学报, 2005,25(1):62-65.
Ma CF, Li DM, Li HJ , et al. Modification of titanium surface with calcium and phosphorus ions using micro-arc oxidation and its effect on osteoblast atta-chment[J]. J First Mil Med Univ, 2005,25(1):62-65.
[21] 王卫卫 . 低温等离子体对纯钛生物活化后的表面分析[D]. 青岛: 青岛大学, 2019.
Wang WW . Surface analysis of pure titanium bioac-tivation after low temperature plasma treatment[D]. Qingdao: Qingdao University, 2019.
[22] 谢艳婷, 李绍杰, 顾舒扬 , 等. 冷常压等离子体处理对纯钛表面性能和成骨细胞增殖迁移的影响研究[J]. 中国实用口腔科杂志, 2018,11(11):674-678, 683.
Xie YT, Li SJ, Gu SY , et al. Study on the effects of cold atmospheric plasma on property of pure titanium surface and proliferation and migration of osteoblasts[J]. Chin J Pract Stomatol, 2018,11(11):674-678, 683.
[23] Shen H, Hu XX, Yang F , et al. Combining oxygen plasma treatment with anchorage of cationized gelatin for enhancing cell affinity of poly(lactide-co-glycolide)[J]. Biomaterials, 2007,28(29):4219-4230.
doi: 10.1016/j.biomaterials.2007.06.004 pmid: 17618682
[24] Wang HY, Kwok DT, Wang W , et al. Osteoblast behavior on polytetrafluoroethylene modified by long pulse, high frequency oxygen plasma immersion ion implantation[J]. Biomaterials, 2010,31(3):413-419.
doi: 10.1016/j.biomaterials.2009.09.066 pmid: 19811820
[25] Mwale F, Wang HT, Nelea V , et al. The effect of glow discharge plasma surface modification of polymers on the osteogenic differentiation of committed human mesenchymal stem cells[J]. Biomaterials, 2006,27(10):2258-2264.
doi: 10.1016/j.biomaterials.2005.11.006 pmid: 16313952
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