目的 探究新對葉百部堿（NTS）對人胰腺癌細胞MiaPaCa-2和PANC-1凋亡的影響及作用機制。方法 以0（對照組）、20、40、60、80、100 μg·mL⁻¹的NTS分別處理MiaPaCa-2、PANC-1細胞24、48、72 h后，采用MTT法檢測細胞存活率；分別設(shè)置對照組（0 μg·mL⁻¹）和NTS低、中、高質(zhì)量濃度（20、40、60 μg·mL⁻¹）組，采用平板克隆法檢測NTS對MiaPaCa-2、PANC-1細胞的增殖能力的影響；采用劃痕愈合實驗檢測細胞遷移能力；采用Hoechst 33258染色法觀察細胞的凋亡；采用流式細胞術(shù)檢測細胞的凋亡率、線粒體膜電位；采用Western blotting法檢測凋亡相關(guān)蛋白B細胞淋巴瘤-2（Bcl-2）、Bcl-2相關(guān)X蛋白（Bax）、活化半胱氨酸蛋白酶-3（cleaved-Caspase-3）、半胱天冬酶-9（Caspase-9）相關(guān)蛋白表達水平。結(jié)果 與對照組比較，不同質(zhì)量濃度NTS可顯著降低MiaPaCa-2、PANC-1細胞存活率（P＜0.01）、克隆形成率（P＜0.05、0.01）、劃痕愈合率（P＜0.05、0.01）、線體膜電位（P＜0.01）；顯著增加MiaPaCa-2和PANC-1細胞凋亡水平（P＜0.01）；在蛋白水平上，NTS顯著上調(diào)促凋亡相關(guān)因子cleaved-Caspase-3、Bax、Caspase-9蛋白的表達水平，下調(diào)抑制凋亡因子Bcl-2蛋白表達水平（P＜0.05、0.01）。結(jié)論 NTS可有效抑制人胰腺癌MiaPaCa-2、PANC-1細胞增殖和遷移能力，并促進其凋亡，其機制可能與下調(diào)抑制凋亡蛋白Bcl-2并上調(diào)促凋亡蛋白Bax、cleaved-Caspase-3、Caspase-9表達有關(guān)。

Objective To explore the effects and mechanisms of neotuberostemonine on the apoptosis of human pancreatic cancer cells MiaPaCa-2 and PANC-1. MethodsMiaPaCa-2 and PANC-1 cells were treated with NTS at concentrations of 0, 20, 40, 60, 80, and 100 μg·mL⁻¹ for 24, 48, and 72 h, respectively. Cell viability was detected by the MTT assay. The control group (0 μg·mL⁻¹), the low, medium, high-concentration group of NTS (20, 40, 60 μg·mL⁻¹), were set up. The effect of NTS on the proliferation ability of MiaPaCa-2 and PANC-1 cells was detected by the plate cloning method. The cell migration ability was detected by the scratch healing assay. Apoptosis was observed by Hoechst 33258 staining. The apoptosis rate and mitochondrial membrane potential were detected by flow cytometry. The expression levels of apoptosis-related proteins B-cell lymphoma-2 (Bcl-2), Bcl-2-associated X protein (Bax), activated Caspase-3 (cleaved-Caspase-3), and Caspase-9 were detected by Western blotting. Results Compared with the control group, different concentrations of NTS significantly reduced the proliferation activity (P < 0.01), colony formation rate (P < 0.05 or 0.01), scratch healing rate (P < 0.05 or 0.01), and mitochondrial membrane potential (P < 0.01) of MiaPaCa-2 and PANC-1 cells. In addition, NTS increased the apoptosis level of MiaPaCa-2 and PANC-1 cells (P < 0.01). At the protein level, NTS significantly upregulated the expression levels of apoptosis-related factors cleaved-Caspase-3, Bax, and Caspase-9 proteins, while the expression level of the apoptosis factor Bcl-2 protein was significantly downregulated (P < 0.05, 0.01). Conclusion NTS can effectively inhibit the proliferation and migration of human pancreatic cancer cells MiaPaCa-2 and PANC-1 and promote their apoptosis. The mechanism may be related to the downregulation of the anti-apoptotic protein Bcl-2 and the upregulation of the pro-apoptotic proteins Bax, cleaved-Caspase-3, and Caspase-9.

R285.5

廣西壯瑤藥重點實驗室（桂科基字［2014］32號）；壯瑤藥協(xié)同創(chuàng)新中心（桂教科研［2013］20號）；廣西一流學科中藥學（民族藥學）（桂教科研[2018]12號)