紫花前胡素经Nox1/Akt信号通路对口腔鳞癌细胞增殖及侵袭的抑制作用

Inhibition Effect of Decursin on Proliferation and Invasion of Oral Squamous Cell Carcinoma Cells via Nox1/Akt Signaling Pathway

目的探讨紫花前胡素通过Nox1/Akt信号通路抑制口腔鳞癌细胞增殖及侵袭的作用机制。方法SCC-25组(A组)及紫花前胡素低、中、高剂量组(B1组、B2组、B3组)均取10 mL SCC-25细胞(密度为5×106/mL),置有10%胎牛血清的DMEM培养基中,分别加入紫花前胡素0,20.0,40.0,80.0μg/mL,置CO2培养箱(37℃、5%CO2、2%O2、93%N2)培养72 h。采用四氮唑盐(MTT)比色法、结晶紫染色、MilliCell室、流式细胞仪、逆转录-定量聚合酶链反应(RT-qPCR)法、酶联免疫吸附(ELISA)法测定细胞增殖及凋亡水平、单克隆形成数、穿膜数、Nox1 mRNA及P-Akt mRNA表达水平,以及超氧化物歧化酶(SOD)、丙二醛(MDA)、谷胱甘肽过氧化物酶(GSH-Px)水平。结果B1组、B2组、B3组吸光度(OD)值、存活率水平、克隆形成数、穿膜数、Nox1 mRNA水平、P-Akt mRNA水平、MDA水平明显低于A组(P<0.05),且与剂量呈正相关;B1组、B2组、B3组细胞凋亡率水平、SOD及GSH-Px水平明显高于A组(P<0.05),且与剂量呈正相关。结论紫花前胡素能抑制口腔鳞癌细胞增殖、侵袭,并促进其凋亡,其机制与紫花前胡素抑制Nox1活性降低Akt的磷酸化水平,进而降低其氧化应激水平有关。

Objective To investigate the mechanism of decursin in inhibiting the proliferation and invasion of oral squamous cell carcinoma cells via Nox1/Akt signaling pathway.Methods In the SCC-25 cell group(group A)and low,medium and high dosage decursin groups(group B1,group B2 and group B3),10 mL SCC-25 cell(the density was 5×106/mL)was put into DMEM medium with 10%fetal bovine serum,then 0,20.0,40.0,80.0μg/mL decursin was added into DMEM medium respectively,and the DMEM medium was cultured in CO2 incubator(37℃,5%CO2,2%O2,93%N2)for 72 h.The cell proliferation and apoptosis,the number of monoclonal formation,the number of membrane penetration,the expression levels of Nox1 mRNA and P-Akt mRNA,the levels of superoxide dismutase(SOD),malondialdehyde(MDA)and glutathione peroxidase(GSH-Px)were detected by methyl thiazolyl tetrazolium(MTT)assay,crystal violet staining,MilliCell chamber,flow cytometry,reverse transcription-quantitative polymerase chain reaction(RT-qPCR)method and enzyme-linked immunosorbent assay(ELISA).Results The levels of optical density(OD),survival rate,the number of clone formation and number of membrane penetration,the expression levels of Nox1 mRNA and P-Akt mRNA and the level of MDA in group B1,group B2 and group B3 were lower than those in group A(P<0.05),and those positively correlated with the dosage.The apoptosis rate,levels of SOD and GSH-Px in group B1,group B2 and group B3 were higher than those in group A(P<0.05),and those positively correlated with the dosage.Conclusion Decursin can inhibit the proliferation and invasion,and promote the apoptosis of oral squamous cell carcinoma cells.Its mechanism is related to the inhibition of Nox1 activity by decursin,which reduces the phosphorylation level of Akt and then reduces the oxidative stress level.