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Study on the Inhibitory Mechanism of Pomegranate Peel Polyphenols on Human Prostate Cancer PC3
Cells Based on Apoptosis and Autophagy Pathway
FENG Tian ，LIU Meng ，CHENG Lufeng ，GAO Xiaoli ，YANG Xiaojun（1. College of Food Science and
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1
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3
3
Pharmacy，Xinjiang Agricultural University，Urumqi 830052，China；2. Dept. of Pharmacy，the Affiliated
Cancer Hospital of Xinjiang Medical University，Urumqi 830000，China；3. College of Pharmacy，Xinjiang
Medical University，Urumqi 830011，China）
ABSTRACT OBJECTIVE：To study the inhibitory mechanism of pomegranate peel polyphenols（PPP）on the proliferation of
human prostate cancer PC3 cells based on autophagy and apoptosis pathway. METHODS：CCK-8 assay was used to investigate the
effects of PPP with different concentrations（25-300 μg/mL）on PC3 cell activity after culturing for 24，48，72 h，so as to screen the
drug concentration and treatment time. PC3 cells were divided into control group（complete culture medium），PPP low-，medium-
and high-concentration groups. After treated for 48 h，flow cytometry and Annexin V-FITC/PI staining were used to detect cell
cycle distribution and apoptosis of PC3 cells. Western blotting assay was used to detect the expression of apoptosis-related protein
as Bax，Bcl-2，as well as the expression of autophagy-related proteins as LC3，Beclin-1，p62，Atg12 and Atg16. RESULTS：The
culturing time was chosen as 48 h. IC50 of PPP was 110 μg/mL，and 50，100，200 μg/mL were chosen as low，medium，high
concentrations of PPP. Compared with control group，the percentage of PC3 cells at phase G0/G1 decreased significantly in PPP low-
and medium-concentration groups while increased significantly at phase S；that of PC3 cells at phase G0/G1 increased significantly
in PPP high-concentration group； while that of PC3 cells at phase G2/M decreased significantly in PPP medium- and
high-concentration groups（P＜0.05 or P＜0.01）. The apoptosis rate of PC3 cells was increased significantly in PPP groups（P＜
0.05 or P＜0.01）. Compared with control group，protein expression of anti-apoptosis protein Bcl-2 and autophagy-related promoting
protein p62 were decreased in PPP groups to different extents，while protein expression of promoting-apoptosis protein Bax as wells
as autophagy-related protein LC3-Ⅱ/LC3-Ⅰ ration and protein expression of Beclin-1，Atg5，Atg12 and Atg16 were increased to
different extents；there was statistical significance in above indexes in PPP high-concentration group and some of above indexes in
PPP low- and medium-concentration groups（P＜0.05 or P＜0.01）. CONCLUSIONS：PPP can inhibit the proliferation of human
prostate cancer PC3 cells，mechanism of which may be related to inducing autophagy and promoting apoptosis.
KEYWORDS Pomegranate peel polyphenols；Prostate cancer；PC3 cell；Apoptosis；Autophagy；Mechanism

前列腺癌是中老年男性常见的恶性肿瘤，其在我国 PPP治疗前列腺癌的药效机制奠定理论基础。
的发病率也逐渐上升，至今还是临床难以攻克的疑难之 1 材料
症 [1-2] ，因此探索更加高效便捷的治疗方法具有重要意 1.1 仪器
义。有研究表明，前列腺癌的发展与自噬有密切关 HERAcell 150i 型 CO2恒温培养箱、KS12 型生物安
[3]
系 。自噬是细胞吞噬自身细胞质蛋白或细胞器进入囊 全柜、iBright FL1000 型凝胶制备成像系统（美国 Ther-
泡，与溶酶体形成自噬溶酶体并降解其包裹的内容物的 mo Fisher Scientific 公司）；ME54E 型分析天平（上海升
[4]
过程，以此可更新细胞器并完成细胞的自身代谢 。适 亮电子科技有限公司）；4K1S 型高速冷冻离心机（美国
度自噬能让靶细胞对外界损伤作出反应，使其存活，同 Sigma公司）；SpectraMax M5型多功能酶标仪（美国Mo-
时也能够将受损的细胞器清除干净，有利于维持细胞稳 lecular Devices公司）；LSR Ⅱ型流式细胞仪（美国BD公
[5]
态 。在前列腺癌的治疗过程中，化疗药物能够激活肿 司）；Mini-Protean 1658001型电泳槽（美国Bio-Rad公司）。
[6]
瘤细胞自噬，进而诱导细胞凋亡 ；自噬也能通过对受损 1.2 药品与试剂
细胞器的降解和再利用来保护细胞器和细胞成分免受 PPP[新疆医科大学药学院自制，批号：20171224，含
损伤 ，因此自噬作用可能具有重要的临床价值。 量：82.7％（采用高效液相色谱法测定，以安石榴苷
[7]
研究表明，石榴科石榴属植物石榴（Punica grana- 计） ]由新疆医科大学药剂教研室常占瑛实验师提供。
[12]
tum L.）的果汁和果皮提取物均能抑制人前列腺癌 PC3 RPMI 1640培养基、磷酸盐缓冲液（pH 7.4）、胰蛋白
细胞系的增殖、迁移和集落形成，且果皮提取物比果汁 酶（美国Hyclone公司，批号：SH30809.01、SH30256.01B、
的效果更优 。为了探讨石榴果皮有效部位的抗肿瘤活 J170041）；胎牛血清（FBS）、青-链霉素双抗（美国 Gibco
[8]
性，本课题组开展了前期研究工作，结果发现石榴皮多 公司，批号：10099141、15140-122）；CCK-8 细胞增殖检
酚（Pomegranate peel polyphenols，PPP）对 PC3 细胞具有 测试剂盒（日本Dojindo公司，批号：KR675）；Annexin V/
一定的抑制作用，并可促进其凋亡，表现出一定的抗肿 PI 试剂盒（上海贝博生物科技公司，批号：BB-4101）；山
瘤活性 [9-11] 。基于此，本文将基于自噬与凋亡途径研究 羊抗兔Bax单克隆抗体、山羊抗兔Bcl-2单克隆抗体（武
PPP 对人前列腺癌 PC3 细胞的抑制作用机制，为阐明 汉三鹰生物技术有限公司，批号：50599-2、12789-1）；山

中国药房 2020年第31卷第16期 China Pharmacy 2020 Vol. 31 No. 16 ·1979 ·