PRL-3基因对胶质母细胞瘤替莫唑胺敏感性的影响

Effect of PRL-3 gene on sensitivity of Temozolomide in glioblastoma

目的研究PRL-3基因在胶质母细胞瘤中的表达及其对胶质母细胞瘤替莫唑胺敏感性的影响。方法通过TCGA数据库分析PRL-3基因在胶质母细胞瘤及正常组织中表达情况,进一步选取空军军医大学第一附属医院诊治的15例胶质母细胞瘤患者术后肿瘤组织(原发性10例,复发性5例),采用免疫组织化学法检测PRL-3阳性细胞在胶质母细胞瘤中的表达情况。选取人胶质母细胞瘤LN229细胞系及替莫唑胺耐药的LN229/TMZ-R细胞系,采用qRT-PCR、Western blot检测PRL-3 mRNA及蛋白的表达情况。选取LN229、SHG44细胞系及对替莫唑胺不敏感的U87细胞系,分别设置正常组、对照组、空载质粒组、PRL-3过表达组、PRL-3下调组。比较正常组、空载质粒组、PRL-3过表达组、PRL-3下调组PRL-3 mRNA表达情况以验证PRL-3过表达及下调是否成功。然后将对照组、PRL-3过表达组、PRL-3下调组采用替莫唑胺(LN229、SHG44细胞系加入替莫唑胺100μmol/L,U87细胞系加入替莫唑胺500μmol/L)进行处理,正常组不处理,72 h后采用CCK-8法测定细胞活性。结果数据库分析显示,原发性胶质母细胞瘤中PRL-3表达高于正常组织,且随肿瘤级别增高而升高(P<0.05)。复发性胶质母细胞瘤组织中PRL-3阳性细胞免疫组织化学法评分高于原发性胶质母细胞瘤组织(P<0.05)。LN229/TMZ-R细胞系中PRL-3 mRNA表达高于LN229细胞系(P<0.05),PRL-3蛋白在LN229/TMZ-R细胞系中高表达。PRL-3过表达组PRL-3 mRNA高于正常组,PRL-3下调组PRL-3 mRNA低于正常组(P<0.05)。在LN229、SHG44、U87细胞系中,对照组细胞活性低于正常组,PRL-3过表达组细胞活性高于对照组,PRL-3下调组细胞活性低于对照组(P<0.05)。结论PRL-3基因低表达可以增强胶质母细胞瘤对替莫唑胺治疗的敏感性,有望成为协同替莫唑胺化疗的有益靶点。

Objective To investigate the expression of PRL-3 gene in glioblastoma and its effect on the sensitivity of Temozolomide in glioblastoma.Methods The expression of PRL-3 gene in glioblastoma and normal tissues was analyzed by the TCGA database,and postoperative tumor tissues of 15 patients with glioblastoma diagnosed and treated by the First Affiliated Hospital of Air Force Military Medical University(ten primary cases,five recurrent cases)were further selected,the expression of PRL-3 positive cells in primary and recurrent glioblastoma tissues was detected by immunohistochemistry.Human glioblastoma LN229 and temozolomid-resistant LN229/TMZ-R cell lines were selected to detect PRL-3 mRNA and protein expression by qRT-PCR and Western blot.LN229,SHG44,and U87 cell lines which was insensitive to Temozolomide were selected,and normal group,control group,no-load liposome group,PRL-3 overexpression group,and PRL-3 downregulation group were set,respectively.The PRL-3 mRNA expression of control group,no-load liposome group,PRL-3 overexpression group,and PRL-3 downregulation group were compared to verify the success of PRL-3 overexpression and downregulation.Then the normal group,PRL-3 overexpression group,and PRL-3 downregulation group were treated with Temozolomide(LN229 and SHG44 cell lines were treated with 100μmol/L Temozolomide,U87 cell line were treated with 500μmol/L Temozolomide),while the normal group was not treated,the cell activity was evaluated by CCK-8 method after 72 h.Results Database analysis showed that PRL-3 expression in primary glioblastoma was higher than that in normal tissues and increased with the increase of tumor grade(P<0.05).The immunohistochemical scores of PRL-3 positive cells in recurrent glioblastoma tissues was higher than that in primary glioblastoma tissues(P<0.05).The mRNA expression of PRL-3 in LN229/TMZ-R cell line was higher than that in LN229 cell line(P<0.05),and PRL-3 protein was highly expressed in LN229/TMZ-R cell line.The PRL-3 mRNA in PRL-3 overexpression group was higher than that in normal group,and the PRL-3 downregulation group was lower than that in normal group(P<0.05).In LN229,SHG44,and U87 cell lines,the cell activity of control group was lower than that of normal group,the cell activity of PRL-3 overexpression group was higher than that of control group,and the cell activity of PRL-3 downregulation group was lower than that of control group(P<0.05).Conclusion The low expression of PRL-3 gene can enhance the sensitivity of glioblastoma to Temozolomide treatment,which is expected to be a beneficial target for synergistic chemotherapy with Temozolomide.