Tissue-engineered bone constructed by icariin-induced periosteal cells repairs bone defects

doi:10.3969/j.issn.2095-4344.0704

Abstract

Abstract:

BACKGROUND: Although icariin has a clear role in inducing differentiation of bone marrow stromal cells, no studies have reported on its use in proliferation of periosteal cells in animal experiments. Three-dimensional polycaprolactone scaffold is a new scaffold, and it has the characteristics of biodegradability, proper pore size, and good histocompatibility.

OBJECTIVE: To investigate the effect of icariin on the proliferation of periosteal cells, to construct periosteal cells and three-dimensional scaffold composite tissue-engineered bone and study its therapeutic effect on radical defects in rabbits, so as to provide a new way for the treatment of bone defects .

METHODS: The mandibular periosteum was obtained from the rabbit and the primary cells were obtained by tissue explant method. The passage 3 cells were planted on 24-well plates. In experimental group, icariin at 10^-4, 10^-3and 10^-2 mol/L were added into the corresponding cell culture wells; equal volume of PBS was added to control group; positive control group was treated with 10 μg/L basic fibroblast growth factor. The cell proliferation in each group was measured by MTT method on the 2^nd, 4^th, 6^th and 8^th days, respectively. The three-dimensional polycaprolactone scaffolds, 10^-2mol/L icariin and 10⁷/L periosteum cells were placed in cell culture plates and cultured in vitro to construct the cell-scaffold complex. After the rabbit radical defect model was established, the model rabbits were randomized into three groups (n=15 per group), followed by implanted with scaffold complex (experimental group), polycaprolactone scaffold (polycaprolactone group) or nothing (control group). Pathological sections were taken at the 2^nd, 4^th and 8^th weeks respectively for hematoxylin-eosin staining to observe the bone and count the number of osteoblasts. The X-ray films of the radius were obtained at the 4^th week.

RESULTS AND CONCLUSION: (1) The proliferation of periosteal cells induced by different concentrations of icariin, especially 10^-2 mol/L icariin, was better than that in the control group (P < 0.05), Therefore, the optimal concentration of icariin was 10^-2mol/L. (2) At 4 weeks after transplantation, in the experimental group, there were more osterocytes and less microvessels; in the polycaprolactone group, there were more new vessels, less woven newly born bone and osterocytes; in the control group, the defect region was filled with more granulation tissue and less osterocytes. At the 8^th week, complete healing in the defect region was observed in the cell-scaffold complex group, while partial healing in the polycaprolactone scaffold group. In the control group, however, fibroblasts and scar tissues were visible in the defect region, with presence of closed medullary cavity and nonunion. The number of osteocytes in the experimental group was significantly higher than that in the other two groups (P < 0.05). X-ray examination at the 4^th week showed that the defect region in the experimental group achieved healing; in the polycaprolactone group, the callus formed and defect region connected partly; and there was a clear gap in the defect region of the control group. Our findings indicate that icariin at different concentrations can promote the proliferation of periosteal cells and the concentration of 10^-2mol/L has the greatest role. Icariin can induce proliferation of periosteal cells in the three-dimensional polycaprolactone scaffold, which contributes to bone repair.

中国组织工程研究杂志出版内容重点：生物材料；骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性；组织工程

Key words: Phytoestrogens, Cell Proliferation, Tissue Scaffolds, Tissue Engineering

CLC Number:

R318

Zhong Xiu-xia, Luo Mei-lan. Tissue-engineered bone constructed by icariin-induced periosteal cells repairs bone defects[J]. Chinese Journal of Tissue Engineering Research, 2018, 22(10): 1477-1482.

Figures/Tables 6

References

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[21]张钰,陈鹏,张登军,等.淫羊藿苷对大鼠坐骨神经离断后修复与再生的影响[J].中华手外科杂志,2016, 32(1):59-61.

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[23]贺宪,孔畅,曾巧,等.淫羊藿苷对骨性关节炎家兔BMSCs增殖及软骨定向分化影响的实验研究[J].湖南中医杂志, 2017,33(3): 140-143.

[24]李利生,史源泉,龚其海.淫羊藿苷抗尿酸钠诱导的大鼠急性痛风性关节炎作用[J].中国实验方剂学杂志,2017,23:15-18.

[25]Marta C, Rodolfo M, Laura T, et al.Dental pulp stem cells for bone tissue engineering: a review of the current literature and a look to the future.Regen Med. 2018. doi: 10.2217/rme-2017-0112.

[26]Tellado SF, Chiera S, Bonani W, et al.Heparin functionalization increases retention of TGF-β2 and GDF5 on biphasic silk fibroin scaffolds for tendon/ligament-to-bone tissue engineering.Acta Biomater. 2018. pii: S1742-7061(18)30137-5.

[27]Fiume E, Barberi J, Verné E, et al. Bioactive Glasses: From Parent 45S5 Composition to Scaffold-Assisted Tissue-Healing Therapies.J Funct Biomater. 2018;9(1). pii: E24.

[28]Dalgic AD, Alshemary AZ, Tezcaner A, et al.Silicate-doped nano-hydroxyapatite/graphene oxide composite reinforced fibrous scaffolds for bone tissue engineering.J Biomater Appl. 2018:885328218763665.

[29]Hoffmann A, Leonards H, Tobies N, et al.New stereolithographic resin providing functional surfaces for biocompatible three-dimensional printing.J Tissue Eng. 2017; 8:2041731417744485.

[30]Shpichka A, Koroleva A, Kuznetsova D, et al.Fabrication and Handling of 3D Scaffolds Based on Polymers and Decellularized Tissues.Adv Exp Med Biol. 2017;1035:71-81.

[31]张峻玮,陆海涛,杨宇明,等.低氧培养对骨膜细胞成软骨分化的影响[J],中华实验外科杂志.2016,33(5):1281-1283.

[32]姜文涛,梅伟,王庆德,等.淫羊藿苷对成骨细胞增殖、分化及矿化的影响[J].中华实验外科杂志,2017, 34(03):446-448.

Lee CJ, Moon SJ, Jeong JH, et al.Kaempferol targeting on the fibroblast growth factor receptor 3-ribosomal S6 kinase 2 signaling axis prevents the development of rheumatoid arthritis. Cell Death Dis. 2018;9(3):401.[1]Barg A, Bettin CC, Burstein AH, et al. Early Clinical and Radiographic Outcomes of Trabecular Metal Total Ankle Replacement Using a Transfibular Approach.J Bone Joint Surg Am. 2018;100(6):505-515.

[2]Jiang H, Cheng P, Li D, et al.Novel standardized massive bone defect model in rats employing an internal eight-hole stainless steel plate for bone tissue engineering.J Tissue Eng Regen Med. 2018. doi: 10.1002/term.

[3]Kessler S, He K, Ukia MW. Bone morphogeneric protein2 accelerates osteointegration and remodelling of sovent-dehydrated bone substitutes. Arch Orthop Trauma Surg.2004;2124(2006):2410-2414.

[4]uttappan S, Anitha A, Minsha MG, et al. BMP2 expressing genetically engineered mesenchymal stem cells on composite fibrous scaffolds for enhanced bone regeneration in segmental defects.Mater Sci Eng C Mater Biol Appl. 2018; 85:239-248.

[5]吴涛,徐俊昌,南开辉,等.淫羊藿苷促进羊骨髓间充质干细胞的增殖和成骨分化[J].中国组织工程研究与临床康复, 2009,13(19): 3725-3729.

[6]Zhao J, Ohba S, Shinkai M, et al. Icariin induces osteogenic differentiation in vitro in a BMP- and Runx2-dependent manner. Biochem Biophys Res Commun. 2008;369: 444-448

[7]Saghebasl S, Davaran S, Rahbarghazi R, et al.Synthesis and in vitro evaluation of thermosensitive hydrogel scaffolds based on (PNIPAAm-PCL-PEG-PCL-PNIPAAm)/Gelatin and (PCL-PEG-PCL)/Gelatin for use in cartilage tissue engineering. J Biomater Sci Polym Ed. 2018:1-22..

[8]Ozkan O, Sasmazel HT. Antibacterial Performance of PCL-Chitosan Core-Shell Scaffolds. J Nanosci Nanotechnol. 2018;18(4):2415-2421.

[9]Nuntanaranont T, Promboot T, Sutapreyasri S.Effect of expanded bone marrow-derived osteoprogenitor cells seeded into polycaprolactone/tricalcium phosphate scaffolds in new bone regeneration of rabbit mandibular defects.J Mater Sci Mater Med. 2018;29(3):24.

[10]Saveleva MS, Ivanov AN, Kurtukova MO, et al.Hybrid PCL/CaCO<sub>3</sub> scaffolds with capabilities of carrying biologically active molecules: Synthesis, loading and in vivo applications.Mater Sci Eng C Mater Biol Appl. 2018; 85:57-67.

[11]李婧.骨膜来源细胞在骨组织工程应用中的研究进展[J].中国美容整形外科杂志,2015,8:496-499.

[12]邢飞,彭静,陈龙.等.组织工程骨膜包被双相陶瓷磷酸钙修复兔尺骨缺损[J].中国组织工程研究, 2016, 20(47):7006-7013.

[13]Park HC, Son YB, Lee SL, et al. Effects of Osteogenic-Conditioned Medium from Human Periosteum-Derived Cells on Osteoclast Differentiation.Int J Med Sci. 2017;14(13):1389-1401.

[14]Yoon DK, Park JS, Rho GJ, et al. The involvement of histone methylation in osteoblastic differentiation of human periosteum-derived cells cultured in vitro under hypoxic conditions. Cell Biochem Funct. 2017;35(7):441-452.

[15]陈苏平,李平. ESW对兔成骨细胞增殖影响的实验研究[J].当代医学,2012,18(4):150-152.

[16]张黎声,韩小晶,罗志荣,等.淫羊藿苷对大鼠骨髓间充质干细胞迁移作用的影响[J].中国中医药信息杂志,2017, 24(2):44-48.

[17]李军,宋光明,孙明林,等.淫羊藿苷对高重力下成骨细胞MC3T3-E1增殖与凋亡的影响[J].中国医药导报,2017, 14(6): 19-23.

[18]张温花,张文超,于远东.淫羊藿苷对前列腺癌细胞株活力、迁移与侵袭的作用[J].中国病理生理杂志,2017, 33(6):1017-1020.

[19]马婷,王丽娜,李子坚,等.淫羊藿苷抗肿瘤作用的研究进展[J].现代肿瘤医学,2017,25(9):1505-1508.

[20]戴娜桑.中药提取物淫羊藿苷作为骨质疏松症抑制剂的试验研究[J].玉林师范学院学报,2016,37(1):120-123.

[21]张钰,陈鹏,张登军,等.淫羊藿苷对大鼠坐骨神经离断后修复与再生的影响[J].中华手外科杂志,2016, 32(1):59-61.

[22]陈澜.淫羊藿苷对大鼠创伤性脑损伤后白细胞介素1β和肿瘤坏死因子α的作用[J].中国新药与临床杂志,2017, 36:65-68.

[23]贺宪,孔畅,曾巧,等.淫羊藿苷对骨性关节炎家兔BMSCs增殖及软骨定向分化影响的实验研究[J].湖南中医杂志, 2017,33(3): 140-143.

[24]李利生,史源泉,龚其海.淫羊藿苷抗尿酸钠诱导的大鼠急性痛风性关节炎作用[J].中国实验方剂学杂志,2017,23:15-18.

[25]Marta C, Rodolfo M, Laura T, et al.Dental pulp stem cells for bone tissue engineering: a review of the current literature and a look to the future.Regen Med. 2018. doi: 10.2217/rme-2017-0112.

[26]Tellado SF, Chiera S, Bonani W, et al.Heparin functionalization increases retention of TGF-β2 and GDF5 on biphasic silk fibroin scaffolds for tendon/ligament-to-bone tissue engineering.Acta Biomater. 2018. pii: S1742-7061(18)30137-5.

[27]Fiume E, Barberi J, Verné E, et al. Bioactive Glasses: From Parent 45S5 Composition to Scaffold-Assisted Tissue-Healing Therapies.J Funct Biomater. 2018;9(1). pii: E24.

[28]Dalgic AD, Alshemary AZ, Tezcaner A, et al.Silicate-doped nano-hydroxyapatite/graphene oxide composite reinforced fibrous scaffolds for bone tissue engineering.J Biomater Appl. 2018:885328218763665.

[29]Hoffmann A, Leonards H, Tobies N, et al.New stereolithographic resin providing functional surfaces for biocompatible three-dimensional printing.J Tissue Eng. 2017; 8:2041731417744485.

[30]Shpichka A, Koroleva A, Kuznetsova D, et al.Fabrication and Handling of 3D Scaffolds Based on Polymers and Decellularized Tissues.Adv Exp Med Biol. 2017;1035:71-81.

[31]张峻玮,陆海涛,杨宇明,等.低氧培养对骨膜细胞成软骨分化的影响[J],中华实验外科杂志.2016,33(5):1281-1283.

[32]姜文涛,梅伟,王庆德,等.淫羊藿苷对成骨细胞增殖、分化及矿化的影响[J].中华实验外科杂志,2017, 34(03):446-448.

Lee CJ, Moon SJ, Jeong JH, et al.Kaempferol targeting on the fibroblast growth factor receptor 3-ribosomal S6 kinase 2 signaling axis prevents the development of rheumatoid arthritis. Cell Death Dis. 2018;9(3):401.