PDF(Pubmed)
Abstract：
α‑Phellandrene (α‑PA), a natural constituent of herbs, inhibits cancer cell viability and proliferation. 5‑Fluorouracil (5‑FU) is a frequently utilized chemotherapeutic medicine for the treatment of colon cancer, which works by triggering cancer cell apoptosis. The present study examined how the combination of α‑PA and 5‑FU affects the suppression of human colon cancer cells by promoting apoptosis. The impact of this treatment on cell viability, apoptosis, and the expression levels of Bcl‑2 family members, caspase family members and mitochondria‑related molecules in HT‑29 cells was assessed by the MTT assay, immunocytochemistry, western blotting and quantitative PCR. The combination of 5‑FU and α‑PA had a synergistic inhibitory effect on cell viability, as determined by assessing the combination index value. Bax protein expression levels were higher in the 50, 100 or 250 µM α‑PA combined with 5‑FU groups compared with those in the 5‑FU alone group (P<0.05). By contrast, Bcl‑2 protein expression levels and mitochondrial membrane potential (MMP, ΔΨm) were lower in the 100 or 250 µM α‑PA combined with 5‑FU groups than those in the 5‑FU alone group (P<0.05). In addition, hexokinase‑2 (HK‑2) protein expression levels were lower in the 50, 100 or 250 µM α‑PA combined with 5‑FU groups than those in the 5‑FU alone group (P<0.05). Compared with 5‑FU alone, after HT‑29 cells were treated with 50, 100 or 250 µM α‑PA combined with 5‑FU, the mRNA expression levels of extrinsic‑induced apoptotic molecules, including caspase‑8 and Bid, were higher (P<0.05). Treatment with 50, 100 or 250 µM α‑PA combined with 5‑FU also increased the mRNA expression levels of cytochrome c, caspase‑9 and caspase‑3, regulating intrinsic apoptosis (P<0.05). These results showed that α‑PA and 5‑FU had a synergistic effect on reducing the viability of human colon cancer HT‑29 cells by inducing extrinsic and intrinsic apoptosis pathways. The mechanism by which apoptosis is induced may involve the intrinsic apoptosis pathway that activates the mitochondria‑dependent pathway, including regulating the expression levels of Bcl‑2 family members, including Bax, Bcl‑2 and Bid, regulating MMP and HK‑2 expression levels, and increasing the expression of caspase cascade molecules, including caspase‑9 and caspase‑3. In addition, it may involve the extrinsic apoptosis pathway that activates caspase‑8 and caspase‑3 leading to apoptosis.
摘要：
α-Phellandrene(α-PA),草药的天然成分，抑制癌细胞的活力和增殖。5-氟尿嘧啶（5-FU）是一种常用的化疗药物，用于治疗结肠癌，通过触发癌细胞凋亡起作用。本研究检查了α‑PA和5‑FU的组合如何通过促进细胞凋亡来影响人结肠癌细胞的抑制。这种处理对细胞活力的影响，凋亡，和Bcl-2家族成员的表达水平，通过MTT测定法评估HT-29细胞中的caspase家族成员和线粒体相关分子，免疫细胞化学,蛋白质印迹和定量PCR。5-FU和α-PA的组合对细胞活力具有协同抑制作用，通过评估组合指数值确定。50、100或250µMα‑PA联合5‑FU组的Bax蛋白表达水平高于单独5‑FU组(P<0.05)。相比之下,Bcl‑2蛋白表达水平和线粒体膜电位(MMP，在100或250µMα‑PA联合5‑FU组中，ΔWm）低于单独5‑FU组（P<0.05）。此外,50、100或250µMα‑PA联合5‑FU组的己糖激酶‑2(HK‑2)蛋白表达水平低于5‑FU单独组(P<0.05)。与仅5-FU相比，在用50、100或250µMα‑PA联合5‑FU处理HT‑29细胞后，外源性诱导的凋亡分子的mRNA表达水平，包括caspase-8和Bid，均较高(P<0.05)。用50、100或250µMα‑PA与5‑FU联合治疗也增加了细胞色素c的mRNA表达水平，caspase-9和caspase-3调节内源性细胞凋亡（P<0.05）。这些结果表明，α‑PA和5‑FU通过诱导外在和内在的凋亡途径对降低人结肠癌HT‑29细胞的活力具有协同作用。诱导凋亡的机制可能涉及激活线粒体依赖性途径的内在凋亡途径，包括调节Bcl-2家族成员的表达水平，包括Bax,Bcl‑2和投标，调节MMP和HK-2表达水平，增加胱天蛋白酶级联分子的表达，包括caspase-9和caspase-3。此外,它可能涉及激活caspase-8和caspase-3导致细胞凋亡的外源性细胞凋亡途径。
