Abstract:

The aim of this study is to explore the effects of melatonin （MLT） combined with cisplatin （DDP） on the proliferation， apoptosis and invasion of cervical cancer HeLa cells. The cervical cancer HeLa cells were cultured in vitro. The effects of MLT， DDP and their combination on the proliferation of HeLa cells were detected by methyl thiazolyl tetrazolium （MTT） method. HeLa cells were divided into control group， melatonin group， cisplatin group and combination group. The melatonin group was treated with 2.5 mmol·L^-1 melatonin， cisplatin group was treated with 5 μg·mL^-1 cisplatin， and combination group was treated with 2.5 mmol·L^-1 melatonin and 5 μg·mL^-1 cisplatin. The proliferation， apoptosis and invasion of HeLa cells were detected by clone formation assay， flow cytometry， scratch assay and Transwell chamber assay， respectively. The expressions of Ki-67， PCNA， Bcl-2， Bax， N-cadherin， E-cadherin and Vimentin were detected by Western blot. MTT results showed that both MLT and DDP could significantly inhibit the proliferation of HeLa cells （P<0.05）， and the inhibition rate was significantly increased with the increase of their concentration （P<0.05）. Compared with MLT and DDP alone， their combination could significantly increased inhibition rate of HeLa cells proliferation. Compared with control group， clone formation rate， number of invaded cells per unit area， scratch healing rate， and related expression levels of Ki67， PCNA， Bcl-2 and E-cadherin proteins were decreased （P<0.05）， while apoptosis rate and relative expression levels of Bax， N-cadherin and Vimentin proteins were increased in melatonin group and cisplatin group （P<0.05）. Compared with melatonin group and cisplatin group， clone formation rate， number of invaded cells per unit area， scratch healing rate， and relative expression levels of Ki67， PCNA， Bcl-2 and E-cadherin proteins were decreased （P<0.05）， while apoptosis rate and relative expression levels of Bax， N-cadherin and Vimentin proteins were increased in combination group （P<0.05）. Results showed that， both MLT and DDP could inhibit the proliferation and invasion of HeLa cells， and promoted their apoptosis， the effects of their combination were more significant.

Key words: cervical cancer, melatonin, cisplatin, HeLa cell, cell proliferation, apoptosis

摘要：

探究了褪黑素（melatonin， MLT）联合顺铂（cisplatin， DDP）对宫颈癌HeLa细胞增殖、凋亡及侵袭的影响。体外培养宫颈癌HeLa细胞，利用四甲基偶氮唑蓝(methyl thiazolyl tetrazolium， MTT)法测定MLT、DDP及其联合使用对宫颈癌HeLa细胞增殖的影响；将HeLa细胞分为对照组、褪黑素组、顺铂组和联合组；其中褪黑素组采用2.5 mmol·L^-1的褪黑素处理细胞；顺铂组采用5 μg·mL^-1的顺铂处理细胞；联合组采用2.5 mmol·L^-1的褪黑素+5 μg·mL^-1的顺铂联合处理。采用克隆形成实验、流式细胞仪、划痕实验和Transwell小室实验分别检测各组HeLa细胞的增殖、凋亡、侵袭情况，蛋白质印迹法检测Ki-67、PCNA、Bcl-2、Bax、N钙粘蛋白（N-cadherin）、E钙粘蛋白（E-cadherin）和波形蛋白（Vimentin）蛋白表达情况。MTT结果显示MLT、DDP可以显著抑制宫颈癌HeLa细胞的增殖（P<0.05)，且随着使用浓度的升高抑制率显著升高；相比单独使用MLT、DDP，联合使用MLT和DDP可以显著升高对HeLa细胞增殖的抑制率（P<0.05)。相比对照组，褪黑素组和顺铂组细胞克隆形成率、单位面积侵袭细胞数目、划痕愈合率以及Ki67、PCNA、Bcl-2、E-cadherin蛋白相对表达水平均降低（P<0.05），细胞凋亡率和蛋白Bax、N-cadherin、Vimentin相对表达水平升高（P<0.05）。相比褪黑素组和顺铂组，联合组细胞克隆形成率、单位面积侵袭细胞数目、划痕愈合率、Ki67、PCNA、Bcl-2、E-cadherin蛋白相对表达水平均降低（P<0.05），细胞凋亡率、Bax、N-cadherin、Vimentin蛋白相对表达水平升高（P<0.05）。结果表明，MLT和DDP均可抑制宫颈癌HeLa细胞增殖和侵袭并促进其凋亡，且联合使用效果更为显著。

关键词: 宫颈癌, 褪黑素, 顺铂, HeLa细胞, 细胞增殖, 细胞凋亡