摘要
背景与目的:O-GlcNAc糖基转移酶(O-GlcNAc transferase,OGT)广泛参与肿瘤细胞的生物学行为,并与癌症的进展相关。分析骨肉瘤中OGT的表达水平,并观察其对骨肉瘤细胞增殖和顺铂敏感性的影响。方法:采用实时荧光定量聚合酶链反应(real-time fluorescence quantitative polymerase chain reaction,RTFQ-PCR)、蛋白质印迹法(Western blot)及免疫组织化学法分析骨肉瘤组织中OGT的表达,并观察OGT的表达与临床病理学特点及与海南医学院第一附属医院收治的患者预后的关系。通过RNA干扰技术沉默骨肉瘤细胞中OGT的表达,通过细胞计数试剂盒(cell counting kit-8,CCK-8)及5-乙炔基-2’脱氧尿嘧啶核苷(5-ethynyl-2’-deoxyuridine,EdU)分析其增殖情况。然后采用顺铂作用于骨肉瘤细胞,采用RTFQ-PCR及Western blot分析OGT mRNA及蛋白的表达情况。进一步改变骨肉瘤细胞中OGT的表达后加入顺铂,采用CCK-8及流式细胞术观察对顺铂的敏感性及骨肉瘤细胞的活力。结果:RTFQ-PCR、Western blot及免疫组织化学结果显示,骨肉瘤组织中OGT的表达水平显著提升(P<0.05),OGT的表达与患者肿瘤直径的大小及病理学分期密切相关,且高表达的患者预后差(P<0.05)。沉默骨肉瘤细胞中OGT的表达后,其增殖能力明显减弱(P<0.05),不同浓度的顺铂作用于骨肉瘤细胞后,OGT表达水平随着顺铂浓度的提升而提升。抑制骨肉瘤细胞中OGT的表达能够显著增强骨肉瘤细胞对顺铂的敏感性(P<0.05),相反过表达OGT会导致顺铂耐受。结论:OGT在骨肉瘤组织中高表达,且与患者的预后密切相关,沉默OGT的表达可以抑制骨肉瘤的生长并增强对顺铂的敏感性,OGT可能是骨肉瘤靶向治疗的潜在生物标志物。
Background and purpose:O-GlcNAc transferase(OGT)is widely involved in biological behaviors of tumor cells and associated with tumor progression.In this study,we measured expression level of OGT in osteosarcoma tissues,and evaluated its effect on proliferation of the osteosarcoma cells and their sensitivity to cisplatin.Methods:OGT expression was determined by real-time fluorescence quantitative polymerase chain reaction(RTFQ-PCR),Western blot and immunohistochemistry,and its effects on clinicopathological features and prognosis of the patients who were treated in the First Affiliated Hospital of Hainan Medical College were evaluated.Then,OGT expression in osteosarcoma cells was silenced by RNA interference to detect its effect on cell proliferation,as demonstrated by cell counting kit-8(CCK-8)assay and 5-ethynyl-2’-deoxyuridine(EdU)assay.And mRNA and protein expressions of OGT in the osteosarcoma cells were also observed after treatment with cisplatin.Furthermore,the osteosarcoma cells were treated with cisplatin after modulation of OGT expression to measure its effect on cisplatin sensitivity of the osteosarcoma cells,as depicted by CCK-8 assay and apoptosis assay using flow cytometry.Results:The results of RTFQ-PCR,Western blot and immunohistochemistry demonstrated that a significant increase in OGT expression in osteosarcoma tissues(P<0.05),which was closely related to larger size of tumor,more advanced pathological stage and poorer prognosis(P<0.05).After silencing OGT,proliferation of the osteosarcoma cells significantly decreased(P<0.05).And OGT expression increased in response to cisplatin treatment in a dose-dependent manner.Silencing OGT in osteosarcoma cells significantly enhanced their sensitivity to cisplatin(P<0.05),while overexpressing OGT led to cisplatin tolerance.Conclusion:OGT is highly expressed in osteosarcoma tissues and closely related to prognosis of the patients.Silencing OGT expression can inhibit growth of osteosarcoma and sensitize it to cisplatin.OGT may be a potential biomarker for targeted therapy of osteosarcoma.
作者
祝开忠
王挺锐
陈焕雄
陈涛
钟贞浩
ZHU Kaizhong;WANG Tingrui;CHEN Huanxiong;CHEN Tao;ZHONG Zhenhao(Department of Spinal Osteopathy Surgery,the First Affiliated Hospital of Hainan Medical College,Haikou 570102,Hainan Province,China)
出处
《中国癌症杂志》
CAS
CSCD
北大核心
2020年第9期682-688,共7页
China Oncology
基金
海南省自然科学基金(819QN365)。