基于Notch 1/NF-κB/STAT3通路探讨塞来昔布逆转NK/T细胞淋巴瘤细胞阿霉素耐药的作用机制  

Mechanism of celecoxib reverses adriamycin resistance in NK/T cell lymphoma cells by Notch 1/NF-κB/STAT3 pathway

作　　者：潘战和[1] 王馨[1] 苏安[1] 张鹏[1] 纪浩南[1] 王小梅[1] PAN Zhanhe;WANG Xin;SU An;ZHANG Peng;JI Haonan;WANG Xiaomei(Zhongshan Hospital of Xiamen University,Xiamen 361004,Fujian,China)

机构地区：[1]厦门大学附属中山医院,福建厦门361004

出　　处：《中国临床药理学与治疗学》2020年第12期1330-1336,共7页Chinese Journal of Clinical Pharmacology and Therapeutics

基　　金：福建省自然科学基金资助项目(2015J01537)。

摘　　要：目的:观察塞来昔布(celecoxib)对淋巴瘤细胞SNK6阿霉素(adriamycin)耐药性的逆转及作用机制。方法:不同浓度的塞来昔布(10、20、40、60、80、100μmol/L)作用于SNK6和SNK6/ADR细胞,噻唑蓝比色法(MTT)检测塞来昔布对SNK6及SNK6/ADR细胞的生长抑制率,筛选塞来昔布对SNK6/ADR的低毒浓度,计算半数抑制浓度(50%inhibitory concentration,IC50)值;采用低毒浓度塞来昔布联合不同浓度的阿霉素处理SNK6/ADR后,测得阿霉素联合塞来昔布后的IC50,并计算逆转倍数;选择低毒浓度塞来昔布联合阿霉素处理SNK6/ADR细胞后,应用流式细胞术测细胞凋亡情况;药物蓄积实验检测罗丹明123蓄积情况;用逆转录-聚合酶链反应(reverse transcription-polymerase chain reaction,RT-PCR)检测Notch 1、核转录因子κB(nuclear factor kappa B,NF-κB)、信号转导子和转录激活子(signal transducer and activator of transcription 3,STAT3)mRNA表达水平;用蛋白质印迹法(Western blot)检测Notch 1、NF-κB、STAT3、P-gp蛋白表达水平。结果:不同浓度塞来昔布可明显抑制SNK6、SNK6/ADR细胞的增殖,且其抑制作用随着塞来昔布浓度的增大而增加。塞来昔布对SNK6细胞的IC50为(57.54±6.89)μg/mL,对SNK6/ADR细胞的IC50为(43.39±4.38)μg/mL,阿霉素对SNK6/ADR细胞的IC50为(7.34±0.56)μg/mL,与10μmol/L塞来昔布联合作用后,阿霉素对SNK6/ADR细胞的IC50为(3.51±0.25)μg/mL(P<0.01),逆转倍数为2.09;阿霉素与10μmol/L塞来昔布联合作用后,与阿霉素组比较,SNK6/ADR细胞凋亡率显著增加(P<0.01);细胞内罗丹明123的蓄积量显著增加(P<0.01);SNK6/ADR细胞内P-gp蛋白表达显著降低(P<0.01);SNK6/ADR细胞内Notch 1、NF-κB、STAT3 mRNA和蛋白表达显著降低(P<0.01)。结论:塞来昔布可诱导NK/T细胞淋巴瘤细胞凋亡及逆转其阿霉素耐药,其可能是通过调节Notch 1/NF-κB/STAT3信号通路来实现的。AIM:To observe the mechanism of celecoxib reversal adriamycin resistance in NK/T cell lymphoma cells.METHODS:SNK6 and SNK6/ADR cells were treated with celecoxib of different concentrations(10,20,40,60,80,100μmol/L),the growth inhibition rate of SNK6 and SNK6/ADR were measured by MTT method.The IC50 and low-toxic concentration of celecoxib on SNK6/ADR cells were calculated,then SNK6/ADR cells were treated with low-toxic concentration of celecoxib combined with adriamycin,the IC50 and reverse times were calculated.The apoptosis of SNK6/ADR cells were detected by FMC.The effects of apatinib on the accumulation of rhodamine123 in SNK6/ADR cells were investigated by flow cytometry.The mRNA expressions of Notch 1,NF-κB and STAT3 were detected by RT-PCR.The protein expressions of Notch 1,NF-κB,STAT3,P-gp were detected by Western blot.RESULTS:Different concentrations of celecoxib can significantly inhibit the proliferation of SNK6 and SNK6/ADR cells.The inhibition increased with the increase of celecoxib concentration.The IC50 of celecoxib on SNK6 and SNK6/ADR cells were(57.54±6.89)μg/mL,(43.39±4.38)μg/mL,The IC50 of adriamycin on SNK6/ADR cells was(7.34±0.56)μg/mL.After adriamycin combined with celecoxib10μmol/L treat SNK6/ADR cells,the IC50 on SNK6/ADR cells was(3.51±0.25)μg/mL(P<0.01),and the reverse times was 2.09.After adriamycin combined with celecoxib,compared with the adriamycin group,the apoptosis rate of SNK6/ADR cells was significantly increased(P<0.01).The intracellular accumulation of rhodamine 123 was significantly increased(P<0.01).The protein expressions of P-gp was significantly decreased(P<0.01).The mRNA and protein expressions of Notch1,NF-κB,STAT3 were significantly decreased(P<0.01).CONCLUSION:Celecoxib can induce apoptosis and reverse adriamycin resistance in NK/T cell lymphoma cells,the mechanism may be regulating the Notch 1/NF-κB/STAT3 signaling pathway.

关 键 词：塞来昔布 NK/T细胞淋巴瘤细胞 阿霉素耐药 Notch 1/NF-κB/STAT3通路 

分 类 号：R965.2[医药卫生—药理学]