Effects of tricalcium phosphate materials with different microstructures on the expression of non-canonical Wnt signaling pathway-related markers

doi:10.3969/j.issn.2095-4344.1960

Abstract

Abstract:

BACKGROUND: The mechanism of osteoinduction is still unclear. The scaffold material microstructure plays a key role in the process of osteoinduction by regulating the relevant bone differentiation signaling pathway.

OBJECTIVE: To investigate the effects of tricalcium phosphate with different microstructures on the expression levels of related markers in non-canonical Wnt signaling pathway in MG63 cells.

METHODS: Tricalcium phosphate materials with different microstructures (TCP-compact, TCP-middle, TCP-big) were synthesized by controlling H₂O₂ foaming and sintering temperature. The microstructure of the material was confirmed by mercury intrusion porosimetry and scanning electron microscopy. At the same time, three types of tricalcium phosphate materials with different microstructures were co-cultured with MG63 cells to detect the expression of non-canonical Wnt signaling pathway-related markers (Wnt5a, FZD1, ROR2, DKK1).

RESULTS AND CONCLUSION: The expression levels of Wnt5a, FZD1, ROR2 and DKK1 were significantly increased in MG63 cells co-cultured with microporous materials (P < 0.05). The expressions of Wnt5a, ROR2 and DKK1 were significantly different between materials with different pore sizes (P < 0.05). There was no significant difference in the expression of Wnt5a, FZD1, ROR2 and DKK1 between three material groups (P > 0.05). The material microstructure affected the expression level of non-canonical Wnt signaling pathway. The difference of the expression level of related markers was regulated by material microstructure. The larger the pore size of the material, the greater the Wnt non-canonical signaling pathway was promoted.

Key words: bone defect, osteoinduction, bone differentiation, tricalcium phosphate, microstructure, porosity, material pore size, non-canonical Wnt signaling pathway, Wnt5a, ROR2

CLC Number:

R459.9','1');return false;" target="_blank">
R459.9

R782.13

Q819

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Chen Kairui, Bao Chongyun, Liu Xian, Xiao Yu, Tan Yanlin, Liu Xiupiao, Xiong Bokai, Zhang Qiang. Effects of tricalcium phosphate materials with different microstructures on the expression of non-canonical Wnt signaling pathway-related markers[J]. Chinese Journal of Tissue Engineering Research, 2019, 23(34): 5430-5435.

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URL: https://www.cjter.com/EN/10.3969/j.issn.2095-4344.1960

https://www.cjter.com/EN/Y2019/V23/I34/5430

Figures/Tables 3

2.1 材料表征结果 X射线衍射实验显示所制备的3组材料具有相同的化学成分，同时与标准卡片对比，明确其化学成分均为纯β-磷酸三钙。扫描电镜下分别以2 000， 5 000，10 000倍观察表面形态及粗糙度，如图1示：TCP-compact材料表面较光滑，可见孔隙的孔径较小，数目也少，相反磷酸钙晶体颗粒较大；TCP-middle和TCP-big表面粗糙，可见明显的沟壑形态，具有明显的微结构；TCP-big由于表面孔隙较大在扫描电镜下形成的图像不如前2种清晰，但是其孔径及表面粗糙度差异依然可以清晰显示。利用压汞实验对3组磷酸三钙材料的微孔孔径、孔隙率进行分析测定，进一步明确材料的微结构，见表2。"

"

2.2 Wnt非经典信号通路相关基因的表达见图2。 2.2.1 材料微结构调控Wnt5a表达共培养第3天TCP-middle和TCP-big材料中Wnt5a表达水平显著上升，且孔径较大的材料促进作用更强；共培养第5天TCP-big对Wnt5a的影响仍是促进作用并且处于较高水平，但是TCP-middle组的表达量与对照组比较无显著差异(P > 0.05)；第3，5天TCP-compact组的表达呈现抑制状态；共培养第7天，各材料组与对照组无显著差异(P > 0.05)。 2.2.2 材料微结构调控ROR2表达 TCP-compact组与对照组对比显示ROR2表达水平明显的抑制，这种抑制状态随时间逐渐减弱。具有微结构的材料组(TCP-middle、TCP-big)则显示初期的轻度增加并且与材料微结构特征有关(P < 0.05)；第5天的表达水平明显增加，TCP-big在促进共培养细胞的ROR2表达时具有更强的效应(P < 0.05)。在共培养第7天，样品中的ROR2表达水平无显著差异。 2.2.3 材料微结构调控FZD1表达共培养第3天磷酸三钙材料组结果为低水平表达，材料组间没有差异。共培养第5天则表现为显著提升其表达水平，并且孔径较大的TCP-big对FZD1的表达有更积极意义。共培养第7天，材料组对FZD1的作用基本消失。 2.2.4 材料微结构调控DKK1表达具有微结构的材料对DKK1表达有明显的上调及促进作用，而致密材料则表现为一定的抑制DKK1表达。共培养第5天，TCP-middle和TCP-big均能促进其表达并且TCP-big表现更明显。整个过程中致密型磷酸三钙材料表面材料所表达的DKK1都处于抑制状态。在末期，3组间的表达量无明显差异并且仅有对照组的一半，差异有显著性意义(P < 0.05)。 "

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Effects of tricalcium phosphate materials with different microstructures on the expression of non-canonical Wnt signaling pathway-related markers

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[1]	Hua Haotian, Zhao Wenyu, Zhang Lei, Bai Wenbo, Wang Xinwei. Meta-analysis of clinical efficacy and safety of antibiotic artificial bone in the treatment of chronic osteomyelitis [J]. Chinese Journal of Tissue Engineering Research, 2021, 25(6): 970-976.
[2]	Zhang Bin, Sun Lihua, Zhang Junhua, Liu Yusan, Cui Caiyun. A modified flap immediate implant is beneficial to soft tissue reconstruction in maxillary aesthetic area [J]. Chinese Journal of Tissue Engineering Research, 2021, 25(5): 707-712.
[3]	Li Chenjie, Lü Linwei, Song Yang, Liu Jingna, Zhang Chunqiu. Measurement and statistical analysis of trabecular morphological parameters of titanium alloy peri-prosthesis under preload [J]. Chinese Journal of Tissue Engineering Research, 2021, 25(4): 516-520.
[4]	Li Xiaozhuang, Duan Hao, Wang Weizhou, Tang Zhihong, Wang Yanghao, He Fei. Application of bone tissue engineering materials in the treatment of bone defect diseases in vivo [J]. Chinese Journal of Tissue Engineering Research, 2021, 25(4): 626-631.
[5]	He Jie, Chang Qi. Biological reconstruction of large bone defects after resection of malignant tumor of extremities [J]. Chinese Journal of Tissue Engineering Research, 2021, 25(3): 420-425.
[6]	Xing Hao, Zhang Yonghong, Wang Dong. Advantages and disadvantages of repairing large-segment bone defect [J]. Chinese Journal of Tissue Engineering Research, 2021, 25(3): 426-430.
[7]	Chen Siqi, Xian Debin, Xu Rongsheng, Qin Zhongjie, Zhang Lei, Xia Delin. Effects of bone marrow mesenchymal stem cells and human umbilical vein endothelial cells combined with hydroxyapatite-tricalcium phosphate scaffolds on early angiogenesis in skull defect repair in rats [J]. Chinese Journal of Tissue Engineering Research, 2021, 25(22): 3458-3465.
[8]	Li Xinping, Cui Qiuju, Zeng Shuguang, Ran Gaoying, Zhang Zhaoqiang, Liu Xianwen, Fang Wei, Xu Shuaimei. Effect of modification of β-tricalcium phosphate/chitosan hydrogel on growth and mineralization of dental pulp stem cells [J]. Chinese Journal of Tissue Engineering Research, 2021, 25(22): 3493-3499.
[9]	Zhou Anqi, Tang Yufei, Wu Bingfeng, Xiang Lin. Designing of periosteum tissue engineering: combination of generality and individuality [J]. Chinese Journal of Tissue Engineering Research, 2021, 25(22): 3551-3557.
[10]	Zhang Zhenhua, Liu Zichen, Yu Baoqing. Status and problems of polycaprolactone and its composite materials in bone tissue engineering [J]. Chinese Journal of Tissue Engineering Research, 2021, 25(22): 3571-3577.
[11]	He Fan, Xiong Xiuli, Shan Xianfeng, Zhang Shutong, Hu Jian, Wang Xuejin. Guided bone regeneration in a small animal model of critical size craniofacial bone defects [J]. Chinese Journal of Tissue Engineering Research, 2021, 25(20): 3226-3231.
[12]	Xu Canhong, Meng Lin, Dong Panfeng, Zhan Huasong, Song Shilei. Masquelet technique in treatment of infectious nonunion: a meta-analysis [J]. Chinese Journal of Tissue Engineering Research, 2021, 25(15): 2445-2452.
[13]	Luo Yicai, Li Hao. Effect of enhanced aryl hydrocarbon receptor expression on inflammatory response and healing of alveolar bone defects in diabetic rats [J]. Chinese Journal of Tissue Engineering Research, 2021, 25(14): 2166-2171.
[14]	Chen Jufang, Tian Yulou, Hao Xin. Role and potential of adipose-derived stem cells in cranio-maxillofacial bone regeneration [J]. Chinese Journal of Tissue Engineering Research, 2021, 25(13): 2087-2096.
[15]	Meng Maohua, Li Ying, Chen Xin, Cheng Lu, Dong Qiang. Effects and mechanisms of enamel matrix derivatives on osteogenic differentiation of bone marrow mesenchymal stem cells [J]. Chinese Journal of Tissue Engineering Research, 2021, 25(13): 2108-2113.