CAJ | 학술논문

目的观察二甲双胍对骨肉瘤细胞增殖及蛋白激酶B(Akt)信号通路中Akt、叉头框转录因子O亚族1(FoxO1)蛋白磷酸化的影响,探讨二甲双胍抑制骨肉瘤细胞增殖的作用机制.方法以骨肉瘤MG63细胞株为研究对象,采用含有10％胎牛血清的RPMI 1640培养液在体外培养至细胞融合达70％时,更换终质量浓度为50 mmol/L的不含血清的培养液继续培养,在加入实验剂量的二甲双胍后12、24、36、48、72 h分别采用细胞计数试剂盒(CCK-8)法检测细胞的增殖,收获24 h的细胞,提取总蛋白后采用Western blot检测Akt、磷酸化Akt (p-Akt)、FoxO1、磷酸化FoxO1(p-FoxOl)的表达.结果二甲双胍可显著抑制MG63细胞增殖,呈明显的时间依赖性,用药48 h后,MG63细胞内Akt、FoxO1的磷酸化水平明显下降.二甲双胍组的p-Akt和对照组的p-Akt水平分别为0.62 ±0.04和1.33±0.16(P＜0.05),二甲双胍组的p-FoxO1和对照组的p-FoxO1水平分别为0.82 ±0.02和1.71±0.05(P ＜0.05).结论二甲双胍可通过Akt/FoxOl信号通路的去磷酸化而抑制MG63细胞的增殖.
목적 관찰이갑쌍고대골육류세포증식급단백격매B(Akt)신호통로중Akt、차두광전록인자O아족1(FoxO1)단백린산화적영향,탐토이갑쌍고억제골육류세포증식적작용궤제.방법 이골육류MG63세포주위연구대상,채용함유10％태우혈청적RPMI 1640배양액재체외배양지세포융합체70％시,경환종질량농도위50 mmol/L적불함혈청적배양액계속배양,재가입실험제량적이갑쌍고후12、24、36、48、72 h분별채용세포계수시제합(CCK-8)법검측세포적증식,수획24 h적세포,제취총단백후채용Western blot검측Akt、린산화Akt (p-Akt)、FoxO1、린산화FoxO1(p-FoxOl)적표체.결과 이갑쌍고가현저억제MG63세포증식,정명현적시간의뢰성,용약48 h후,MG63세포내Akt、FoxO1적린산화수평명현하강.이갑쌍고조적p-Akt화대조조적p-Akt수평분별위0.62 ±0.04화1.33±0.16(P＜0.05),이갑쌍고조적p-FoxO1화대조조적p-FoxO1수평분별위0.82 ±0.02화1.71±0.05(P ＜0.05).결론 이갑쌍고가통과Akt/FoxOl신호통로적거린산화이억제MG63세포적증식.
Objective To investigate the effect of metformin on proliferation of osteosarcoma MG63 cells and protein kinase B (Akt) and forkhead box transcription factor O1 (FoxO1) phosphorylation in Akt signal pathway,and to explore the possible mechanism of metformin inhibiting proliferation of MG63 cells.Methods Osteosarcoma MG63 cells were cultured with RPMI 1640 supplemented with 10％ fatal bovine serun (FBS) untill the cells reached 70％ confluence,and cultured continuously with 50 mmol/L FBS-free RPMI 1640.Cell counting kit-8 (CCK-8) was used to test the proliferation of osteosarcoma cells treated with metformin at 12,24,36,48,and 72 h.After extraction of total protein,the Akt,p-Akt,FoxO1,and p-FoxO1 expression was analyzed by using Western blotting in MG63 cells at 24 h.Results The proliferation of the MG63 cells could be significantly inhibited by metformin in a time-independent manner,and protein Akt and FoxO1 phosphorylation could also be inhibited after 48 h.The p-Akt expression in metformin and control groups was 0.62 ± 0.04 and 1.33 ± 0.16 respectively (P ＜ 0.05).The p-FoxOl expression in metformin and control groups was 0.82 ±0.02 and 1.71 ±0.05 respectively (P ＜ 0.05).Conclusion Metformin can inhibit proliferation of osteosarcoma cells in vitro,which is associated with the dephosphorylation of Akt/FoxO1 signal pathway.