Department of Microbiology and Immunology, Medical University of Silesia in Katowice, Jordana 19, 41 808 Zabrze, Poland.
Int J Oncol. 2011 Apr;38(4):941-53. doi: 10.3892/ijo.2011.930. Epub 2011 Feb 1.
Prostate cancer represents an ideal disease for chemopreventive intervention. Propolis possesses immuno-modulatory, anti-tumour and chemopreventive properties. The tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) is an important endogenous anti-cancer agent that induces apoptosis selectively in tumour cells. However, some cancer cells are resistant to TRAIL-mediated apoptosis. Naturally occurring phenolic and polyphenolic compounds sensitize TRAIL-resistant cancer cells and augment the apoptotic activity of TRAIL. The ethanolic extract of Brazilian green propolis (EEP) is rich in phenolic components. Our in vitro results indicate the potential targets in the TRAIL-induced apoptotic pathway for the cancer chemopreventive activity of Brazilian propolis. We examined the cytotoxic and apoptotic effects of Brazilian EEP and its bioactive components in combination with TRAIL on LNCaP prostate cancer cells. The chemical composition of Brazilian green propolis was determined by high performance liquid chromatography-diode array detection. The cytotoxicity was measured by 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl-tetrazolium and lactate dehydrogenase assays. Apoptosis was detected using annexin V-FITC by flow cytometry and fluorescence microscopy. The mitochondrial membrane potential (∆Ψm) was evaluated using DePsipher staining by fluorescence microscopy. Flow cytometry was used to analyse death receptor (TRAIL-R1 and TRAIL-R2) expression in LNCaP cells. The inhibition of nuclear factor-κB (NF-κB) (p65) activation in cancer cells was confirmed by the ELISA-based TransAM NF-κB kit. The LNCaP cells were shown to be resistant to TRAIL-induced apoptosis. Our study demonstrates that EEP sensitizes TRAIL-resistant prostate cancer cells. The main phenolic components detected in Brazilian green propolis are artepillin C, quercetin, kaempferol and p-coumaric acid. Brazilian propolis and its bioactive components markedly augmented TRAIL-mediated apoptosis and cytotoxicity in prostate cancer cells. Brazilian EEP enhanced the expression of TRAIL-R2 and the activity of NF-κB in LNCaP cells. The co-treatment of prostate cancer cells with 100 ng/ml TRAIL and 50 µg/ml EEP increased the percentage of apoptotic cells to 65.8 ± 1.2% and caused a significant disruption of ∆Ψm in LNCaP cells. We show that Brazilian EEP helped cells overcome TRAIL resistance by engaging both intrinsic and extrinsic apoptotic pathways and regulating NF-κB activity. The data demonstrate the important role of Brazilian green propolis and its bioactive compounds in prostate cancer chemoprevention through the enhancement of TRAIL-mediated apoptosis.
前列腺癌是化学预防干预的理想疾病。蜂胶具有免疫调节、抗肿瘤和化学预防作用。肿瘤坏死因子相关凋亡诱导配体（TRAIL）是一种重要的内源性抗癌剂，可选择性诱导肿瘤细胞凋亡。然而，一些癌细胞对 TRAIL 介导的凋亡具有抗性。天然存在的酚类和多酚类化合物使 TRAIL 耐药的癌细胞敏感，并增强 TRAIL 的凋亡活性。巴西绿蜂胶的乙醇提取物富含酚类成分。我们的体外结果表明，巴西蜂胶的抗癌化学预防活性可能针对 TRAIL 诱导的凋亡途径中的潜在靶标。我们研究了巴西 EEP 及其生物活性成分与 TRAIL 联合对 LNCaP 前列腺癌细胞的细胞毒性和凋亡作用。采用高效液相色谱-二极管阵列检测法测定巴西绿蜂胶的化学成分。通过 3-[4,5-二甲基噻唑-2-基]-2,5-二苯基四唑和乳酸脱氢酶测定法测定细胞毒性。通过流式细胞术和荧光显微镜检测用 annexin V-FITC 检测凋亡。通过荧光显微镜用 DePsipher 染色评估线粒体膜电位（∆Ψm）。通过流式细胞术分析 LNCaP 细胞中的死亡受体（TRAIL-R1 和 TRAIL-R2）表达。通过基于 ELISA 的 TransAM NF-κB 试剂盒证实了对癌细胞中核因子-κB（NF-κB）（p65）激活的抑制。结果表明，LNCaP 细胞对 TRAIL 诱导的凋亡具有抗性。我们的研究表明，EEP 使 TRAIL 耐药的前列腺癌细胞敏感。在巴西绿蜂胶中检测到的主要酚类成分是 artepillin C、槲皮素、山奈酚和对香豆酸。巴西蜂胶及其生物活性成分显著增强了前列腺癌细胞中 TRAIL 介导的凋亡和细胞毒性。巴西 EEP 增强了 LNCaP 细胞中 TRAIL-R2 的表达和 NF-κB 的活性。用 100ng/ml TRAIL 和 50μg/ml EEP 共同处理前列腺癌细胞，使凋亡细胞的百分比增加到 65.8±1.2%，并导致 LNCaP 细胞中 ∆Ψm 显著破坏。我们表明，巴西 EEP 通过参与内在和外在凋亡途径以及调节 NF-κB 活性来帮助细胞克服 TRAIL 耐药性。数据表明，巴西绿蜂胶及其生物活性化合物通过增强 TRAIL 介导的凋亡在前列腺癌的化学预防中发挥重要作用。
