首页|酪蛋白激酶1α/色素框同源物4轴在骨肉瘤细胞增殖、转移中的调控机制

酪蛋白激酶1α/色素框同源物4轴在骨肉瘤细胞增殖、转移中的调控机制

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目的 探讨酪蛋白激酶1α(CK1α)/色素框同源物4(CBX4)轴在骨肉瘤细胞增殖、转移中的调控机制.方法 采用Lipofectamine™ 3000转染CK1α过表达载体(过表达组)及阴性对照载体(对照组)至U-2OS骨肉瘤细胞中,实时荧光定量聚合酶链反应(Real-time PCR)及蛋白质印迹法(Western blot)检测转染效果.噻唑蓝(MTT)法检测细胞活力,细胞克隆形成实验检测细胞克隆能力,Hoechst染色检测细胞凋亡,Tanswell实验检测细胞侵袭能力,Western blot检测CBX4、B淋巴细胞瘤-2基因(bcl-2)、bcl-2相关蛋白X(bax)、基质金属蛋白酶(MMP)-2、MMP-9、E-钙黏蛋白(E-cadherin)、波形蛋白(Vimentin)、Wnt3a、β-连环蛋白(β-catenin)及p-糖原合成酶激酶-3β(p-GSK-3β)蛋白表达.独立样本t检验分析组间差异性.结果 过表达组中CK1a mRNA及蛋白表达量均高于对照组(1.87±0.04 比 1.00±0.06、2.13±0.12 比 0.32±0.02,t=6.346、6.893,P<0.05).过表达组中细胞凋亡率、Wnt3a、β-catenin、bcl-2、MMP-2、MMP-9及Vimentin蛋白表达量高于对照组[(36.98±3.62)%比(4.31±0.30)%、1.02±0.06 比 0.22±0.01、0.96±0.07 比0.32±0.02、0.88±0.06 比 0.33±0.02、0.92±0.07 比 0.28±0.01、0.87±0.05 比 0.28±0.02、0.97±0.07 比 0.35±0.02,t=6.671、6.879、5.967、6.345、5.631、6.124、5.983,P<0.05].过表达组中细胞活力、细胞克隆数目、细胞侵袭数目、CBX4、p-GSK-3β、bax及E-cadherin蛋白表达量低于对照组(0.35±0.01 比 0.58±0.03、32.61±2.68 比 184.49±12.42、33.74±2.62 比 161.32±10.58、1.98±0.11 比 0.30±0.02、0.13±0.01 比 0.72±0.05、0.14±0.01 比 0.71±0.05、0.11±0.00 比0.63±0.04,t=5.897、6.894、6.692、6.672、5.872、5.652、6.126,P<0.05).结论 上调表达 CK1α可抑制U-2OS骨肉瘤细胞增殖、侵袭及转移,可能通过阻断Wnt/β-catenin信号通路实现的.
Regulatory mechanism of casein kinase 1α/chromobox homolog 4 axis on the proliferation and metastasis of osteosarcoma cells
Objective To explore regulatory mechanism of casein kinase 1α(CK1 α)/chromobox homolog 4(CBX4)axis on the proliferation and metastasis of osteosarcoma cells.Methods Transfection of CK1 α over-expression vector(over-expression group)and negative control vector(control group)into U-2OS osteosarcoma cell was used by Lipofectamine™ 3000.The transfection effect was detected by real-time fluorescence quantitative polymerase chain reaction(Real-time PCR)and Western blotting.Cell via-bility was measured by methyl thiazolyl tetrazolium(MTT)assay.Cell clone ability was detected by cell clone formation test.Cell apoptosis was detected by Hoechst staining.Cell invasion ability was detected by Transwell method.The expression of B-lymphomatoma-2(bcl-2),bcl-2 related protein X(bax),matrix metalloproteinase(MMP)-2,MMP-9,E-cadherin,Vimentin,Wnt3a,β-catenin and p-glycogen synthase kinase-3β(p-GSK-3 β)was detected by Western blotting.Inter group differences was adopted by independ-ent-samples T-test.Results The expression of CK1α mRNA and protein in over-expression group was higher than that in control group(1.87±0.04 vs.1.00±0.06,2.13±0.12 vs.0.32±0.02,t=6.346,6.893,P<0.05).Cell apoptotic rate,the expression of Wnt3a,β-catenin,bcl-2,MMP-2,MMP-9 and Vimentin protein in over-expression group was higher than that in control group[(36.98±3.62)%vs.(4.31±0.30)%,1.02±0.06 vs.0.22±0.01,0.96±0.07 vs.0.32±0.02,0.88±0.06 vs.0.33±0.02,0.92±0.07 vs.0.28±0.01,0.87±0.05 vs.0.28±0.02,0.97±0.07 vs.0.35±0.02,t=6.671,6.879,5.967,6.345,5.631,6.124,5.983,P<0.05].Cell viability,cell clone number,cell invasion number,the expression of CBX4,p-GSK-3β,bax and E-cadherin protein in over-expression group was lower than that in control group(0.35±0.01 vs.0.58±0.03,32.61±2.68 vs.184.49±12.42,33.74±2.62 vs.161.32±10.58,1.98±0.11 vs.0.30±0.02,0.13±0.01 vs.0.72±0.05,0.14±0.01 vs.0.71±0.05,0.11±0.00 vs.0.63±0.04,t=5.897,6.894,6.692,6.672,5.872,5.652,6.126,P<0.05).Conclusion Over-regulation expression of CK1α might inhibit the proliferation and metastasis of U-2OS osteosarcoma cell,which related to inhibition of Wnt/β-catenin signaling pathway.

OsteosarcomaU-2OS cellCasein kinase 1αtChromobox homolog 4Prolif-erationMetastasis

高建、张政阔、赵致忠、武壮壮

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山西医科大学第二医院骨科,太原 030001

骨肉瘤 U-2OS细胞 酪蛋白激酶1α 色素框同源物4 增殖 转移

2024

中华实验外科杂志
中华医学会

中华实验外科杂志

CSTPCD
影响因子:0.759
ISSN:1001-9030
年,卷(期):2024.41(11)