目的 探究柚皮素對順鉑耐藥宮頸癌細胞（HeLa/DDP）順鉑耐藥性的影響，并探究其分子機制。方法 體外培養(yǎng)人HeLa/DDP細胞、HeLa細胞，CCK-8法檢測不同質(zhì)量濃度順鉑對HeLa/DDP、HeLa細胞增殖的影響及不同質(zhì)量濃度柚皮素對HeLa/DDP細胞增殖的影響。設置對照組、順鉑組（2 mg/L順鉑）、順鉑＋柚皮素組（2 mg/L順鉑＋60 mg/L柚皮素）和順鉑＋柚皮素＋AMPK通路抑制劑組（2 mg/L順鉑＋60 mg/L柚皮素＋50 μmol/L Compound C）；CCK-8法檢測各組HeLa/DDP細胞增殖情況；流式細胞儀檢測各組HeLa/DDP細胞凋亡情況；蛋白免疫印跡（Western blotting）法檢測各組HeLa/DDP細胞中腺苷酸活化蛋白激酶（AMPK）、p-AMPK、Bcl-2相關(guān)X蛋白（Bax）、Beclin1、微管相關(guān)蛋白1輕鏈3（LC3）Ⅰ、LC3Ⅱ蛋白表達情況。結(jié)果 順鉑對HeLa/DDP細胞、HeLa細胞的半數(shù)抑制濃度（IC₅₀）分別在4～8、1～2 mg/L，柚皮素對HeLa/DDP細胞的IC₅₀在30～60 mg/L。與對照組相比，順鉑組HeLa/DDP細胞增殖抑制率、凋亡率及細胞中p-AMPK/AMPK、Bax、Beclin1、LC3Ⅱ蛋白表達水平顯著升高（P<0.05），LC3Ⅰ蛋白表達水平顯著降低（P<0.05）；與順鉑組相比，順鉑＋柚皮素組HeLa/DDP細胞增殖抑制率、凋亡率及細胞中p-AMPK/AMPK、Bax、Beclin1、LC3Ⅱ蛋白表達水平顯著升高（P<0.05），LC3Ⅰ蛋白表達水平顯著降低（P<0.05）；與柚皮素組相比，AMPK通路抑制劑組HeLa/DDP細胞增殖抑制率、凋亡率及細胞中p-AMPK/AMPK、Bax、Beclin1、LC3Ⅱ蛋白表達水平顯著降低（P<0.05），LC3Ⅰ蛋白表達水平顯著升高（P<0.05）。結(jié)論 柚皮素可能通過激活AMPK通路介導的自噬反應，與順鉑聯(lián)用發(fā)揮對HeLa/DDP細胞增殖抑制作用及凋亡促進作用。

Objective To investigate the effect of naringenin on cisplatin resistance of cervical cancer HeLa/DDP cells and its molecular mechanism.Methods HeLa/DDP cells and HeLa cells were culturedin vitro, CCK-8 method was used to detect the effects of different concentrations of cisplatin on the proliferation of HeLa/DDP and HeLa cells, and the effects of naringenin on the proliferation of HeLa/DDP cells. HeLa/DDP cells were divided into control group, cisplatin group (2 mg/L cisplatin), cisplatin + naringenin group (2 mg/L cisplatin + 60 mg/L naringenin) and cisplatin + naringenin + AMPK pathway inhibitor group (2 mg/L cisplatin + 60 mg/L naringenin + 50 μmol/L Compound C). The proliferation of HeLa/DDP cells was detected by CCK-8 method, the apoptosis of HeLa/DDP cells was detected by flow cytometry. Western blotting (WB) method was used to detect the expression of adenosine monophosphate activated protein kinase (AMPK), p-AMPK, Bcl-2 associated X protein (Bax), Beclin1, microtubule-associated protein 1 light chain 3 (LC3) I and LC3 II protein in HeLa/DDP cells.Results The IC₅₀ value of cisplatin to HeLa/DDP cells and HeLa cells were 4—8 mg/L and 1—2 mg/L, respectively. IC₅₀ value of naringenin to HeLa/DDP cells was in the range of 30—60 mg/L. Compared with those in the control group, the proliferation inhibition rate, apoptosis rate and the protein expression levels of p-AMPK/AMPK, Bax, Beclin1 and LC3 Ⅱ in HeLa/DDP cells were significantly higher in cisplatin group(P<0.05), and the expression level of LC3 Ⅰ protein was significantly lower (P<0.05). Compared with those in cisplatin group, the proliferation inhibition rate and apoptosis rate of HeLa/DDP cells and the expression levels of p-AMPK/AMPK, Bax, Beclin1 and LC3 Ⅱ protein in cisplatin + naringenin group were significantly higher, and the expression level of LC3 Ⅰ protein was significantly lower (P<0.05). Compared with those in cisplatin + naringenin group, the proliferation inhibition rate and apoptosis rate of HeLa/DDP cells and the protein expression levels of p-AMPK/AMPK, Bax, Beclin1 and LC3 Ⅱ were significantly lower in cisplatin + naringenin + AMPK pathway inhibitor group, the expression level of LC3Ⅰ protein was significantly higher (P<0.05).Conclusion Naringenin can inhibit proliferation and promote apoptosis of HeLa/DDP cells by activating autophagy mediated by AMPK pathway.

R965

國家自然科學基金資助項目（81741147）