纤维细胞生长因子受体抑制剂AZD4547对SK-MES-1肺鳞癌细胞增殖和凋亡的作用

Effects of fibroblast growth factor receptor inhibitor(AZD4547)on the proliferation and apoptosis of SK-MES-1 lung cancer cells

目的探讨纤维细胞生长因子受体(FGFR)抑制剂AZD4547经磷脂酰肌醇3-激酶/蛋白激酶B(PI3K/Akt)信号通路影响肺鳞癌细胞增殖和凋亡的作用机制。方法用免疫组化检测FGFR1、PI3K、Akt和含半胱氨酸的天冬氨酸蛋白水解酶-3(Caspase-3)在肺鳞癌患者癌组织和癌旁组织中的表达。SK-MES-1细胞用5个浓度(1,10,100 nmol·L^-1以及1,10μmol·L^-1)AZD4547处理48 h,以未加AZD4547作为空白组。用CCK8检测细胞的活力,用流式细胞仪检测细胞凋亡率,用蛋白质印迹法检测凋亡相关蛋白Caspase-3、FGFR1和p-Akt的蛋白表达水平。结果不同浓度AZD4547作用48 h后,肺鳞癌细胞活力随AZD4547浓度增加而逐渐降低,呈剂量依赖方式。空白组和由低到高5个浓度AZD4547组的细胞凋亡率分别为(13.39±0.22)%,(10.08±0.17)%,(7.49±0.12)%,(4.96±0.10)%,(18.51±0.16)%和(10.25±0.20)%,其中1μmol·L^-1SK-MES-1组的早期和晚期凋亡率高于其余浓度处理的细胞。1μmol·L^-1AZD4547能明显引起Caspase-3激活。1,10μmol·L^-1 AZD4547可明显阻断FGFR1,抑制Akt的磷酸化。结论 FGFR1可能通过调控下游的Akt磷酸化从而抑制SK-MES-1细胞的增殖,并可能通过激活内源性凋亡途径激活Caspase-3引起SK-MES-1细胞凋亡。

Objective To investigate the effects and mechanism of fibroblast growth factor receptor(FGFR)inhibitor(AZD4547)on proliferation and apoptosis of lung squamous cancer cells(LSCC)via the phosphatidylinositol-3-kinase(PI3K)/proteinkinase B(Akt)signaling pathway.Methods Immunohistochemical staining was used to detect the expression of FGFR1,Akt,PI3K,and Caspase-3 in LSCC patients undergoing surgical resection and adjacent normal tissues.The SK-MES-1 cells were treated with five concentrations of AZD4547(1,10,100 nmol·L^-1,and 1,10μmol·L^-1)for 48 h,blank group without AZD4547.The CCK-8 assay was used to measure the proliferation inhibition rates.The flow cytometer was used to detect cell apoptosis rate after treatment.The Western blotting was used to measure the protein levels of Caspase-3,FGFR1,Akt and its phosphorylated form(p-Akt).Results The cell viability was reduced with the increase of AZD4547 concentration after 48 h in a dose dependent manner.The apoptosis rates in blank group and the five concentrations were(13.39±0.22)%,(10.08±0.17)%,(7.49±0.12)%,(4.96±0.10)%,(18.51±0.16)%,(10.25±0.20)%,respectively.The early and late apoptosis rates of SK-MES-1 cells treated with 1μmol·L^-1 were higher than those treated with other concentrations.The 1μmol·L^-1 AZD4547 significantly induced Caspase-3 activation.1,10μmol·L^-1 AZD4547 significantly blocked FGFR1 and inhibited Akt phosphorylation.Conclusion AZD4547 could inhibit proliferation of nasopharyngeal carcinoma cell through reducing the phosphorylation of FGFR1 and p-Akt.Additionally,AZD4547 can induce SK-MES-1 apoptosis by activating intrinsic apoptotic pathway via cleaving Caspase-3.