Li Li, Xu Chi, Liu Pingtao, Huang Jia
Department I of Orthopedics, Jingmen No. 2 People's Hospital, Jingmen, Hubei 448000, P.R. China.
School of Health Sciences, Wuhan University, Wuhan, Hubei 430000, P.R. China.
Oncol Lett. 2017 Jul;14(1):271-275. doi: 10.3892/ol.2017.6135. Epub 2017 May 8.
The aim of this study was to evaluate the status and explore the impact of DNA methylation of gene with osteosarcoma in children. A total of 50 patients with primary osteosarcoma in children were enrolled. The expression of WNT6 protein was determined by immunohistochemical staining. The DNA methylation level of gene was evaluated by methylation-specific PCR (MSP). Human MG63 osteosarcoma cells and human normal Hfob1.19 osteoblasts were selected and cultured. Western blot analysis was utilized to measure the expression of WNT6 in the two cell lines. MSP was used to assess the status of DNA methylation of gene. The correlation between DNA methylation of gene and prognosis of osteosarcoma in children was evaluated by statistical analysis of the clinical and prognostic data. The results of the immunohistochemical assay showed that 84% (42/50) of primary osteosarcoma was WNT6 positive. MSP indicated that DNA methylation of gene was found in 9 cases (18%) of primary osteosarcoma. Western blot analysis showed that WNT6 protein expression in MG63 cells was significantly higher than that in normal human Hfob1.19 osteoblasts, whereas, the level of DNA methylation of gene in MG63 cells was significantly lower than in Hfob1.19 cells. Analysis of survival, prognosis and their correlation found that the DNA methylation level of gene was negatively correlated with the prognosis of children with osteosarcoma. In conclusion, there was a high level of gene expression in primary osteosarcoma, which was mainly due to low DNA methylation level of gene. The DNA methylation of gene was negatively correlated with the prognosis of patients with osteosarcoma in children.
本研究旨在评估儿童骨肉瘤相关基因的DNA甲基化状态，并探讨其影响。共纳入50例儿童原发性骨肉瘤患者。采用免疫组织化学染色法检测WNT6蛋白的表达。通过甲基化特异性PCR（MSP）评估相关基因的DNA甲基化水平。选取人MG63骨肉瘤细胞和人正常Hfob1.19成骨细胞进行培养。利用蛋白质免疫印迹分析检测两种细胞系中WNT6的表达。采用MSP评估相关基因的DNA甲基化状态。通过对临床和预后数据进行统计分析，评估相关基因的DNA甲基化与儿童骨肉瘤预后的相关性。免疫组织化学检测结果显示，84%（42/50）的原发性骨肉瘤WNT6呈阳性。MSP检测表明，9例（18%）原发性骨肉瘤存在相关基因的DNA甲基化。蛋白质免疫印迹分析显示，MG63细胞中WNT6蛋白表达明显高于正常人Hfob1.19成骨细胞，而MG63细胞中相关基因的DNA甲基化水平明显低于Hfob1.19细胞。生存及预后分析及其相关性研究发现，相关基因的DNA甲基化水平与儿童骨肉瘤患者的预后呈负相关。综上所述，原发性骨肉瘤中相关基因表达水平较高，主要是由于相关基因的DNA甲基化水平较低。相关基因的DNA甲基化与儿童骨肉瘤患者的预后呈负相关。
