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目的:研究10%纳米羟基磷灰石聚醚醚酮(10%HA/SPEEK/PEEK)复合材料浸提液培养MG63细胞后Wnt3a/β-catenin信号通路中特征性蛋白的变化,评价复合材料的生物相容性。方法:Wnt3a持续作用于10%HA/SPEEK/PEEK浸提液培养的MG63细胞。应用免疫荧光染色技术,定性检测Wnt信号通路中β-catenin蛋白的存在。应用Western-Blotting技术相对定量检测β-catenin蛋白的多少。应用实时定量PCR技术相对定量检测信号通路下游基因c-myc、cyclin D1的表达情况。结果:免疫荧光染色和Western-Blotting实验结果显示,Wnt3a作用于10%HA/SPEEK/PEEK浸提液中培养的MG63细胞,β-catenin蛋白绿色深染,位于细胞核中,表达量显著增加。实时定量RTPCR实验结果显示Wnt3a作用于10%HA/SPEEK/PEEK浸提液中培养的MG63细胞,信号通路的下游靶向基因cmyc、cyclin D1表达显著增加,Wnt通路开放。结论:10%HA/SPEEK/PEEK对MG63细胞有良好的成骨作用,可以指导临床实验研究。
OBJECTIVE: To study the changes of characteristic proteins in Wnt3a / β-catenin signaling pathway after MG63 cells were cultured with 10% nano-hydroxyapatite polyetheretherketone (10% HA / SPEEK / PEEK) Biocompatibility. Methods: Wnt3a cells were maintained in MG63 cells cultured in 10% HA / SPEEK / PEEK extracts. Immunofluorescence staining was used to detect the presence of β-catenin protein in Wnt signaling pathway. The relative quantitative detection of β-catenin protein using Western-Blotting technology. Real-time quantitative PCR was used to detect the expression of downstream genes c-myc and cyclin D1 in the signal pathway. Results: Immunofluorescence staining and Western-Blotting showed that Wnt3a was induced in MG63 cells cultured in 10% HA / SPEEK / PEEK extracts. The protein of β-catenin was stained dark green, and located in the nucleus. The expression of Wnt3a was significantly increased. Real-time quantitative RTPCR results showed that Wnt3a was induced on MG63 cells cultured in 10% HA / SPEEK / PEEK extracts. The expression of downstream target genes cmyc and cyclin D1 were significantly increased and Wnt pathway was opened. Conclusion: 10% HA / SPEEK / PEEK has a good osteogenic effect on MG63 cells, which can guide the clinical research.