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Abstract：
BACKGROUND: Breast cancer is the most common type of cancer diagnosed in women. Finding novel therapeutic agents with significant cytotoxic action and minimal adverse impact on normal cells becomes crucial. Today, natural anticancer agents present an unconventional method of treating cancer, either as a curative or preventative agent, with considerable concern for marine organisms.
METHODS: The anticancer effect of the alcoholic extract of different Red Sea Seagrasses on MCF-7 human breast cancer cell line has been investigated. Seagrasses were collected from Wadi El Gamal, Red Sea and extracted. Qualitative HPLC analysis was performed on the extracts for the identification of their active biomarkers. This study was aimed to explore the cytotoxic impact of Thalassia hemprichii (Ehren.) and Enhalus acoroides (L.f.) Royle on MCF-7 and their mode of action. Their anti-proliferative effects on cancer cells were performed using Neutral red assay. On the other hand, their apoptotic effect and their capacity to induce cell cycle arrest were investigated by flow cytometry assay. The effect of Seagrasses on the mitochondrial membrane potential (ΔψM) was studied by using JC-1 mitochondrial membrane potential assay kit in Seagrasses treated cancer cells to Δψ Caspases 3/7activity was examined using the colorimetric method. Gene expression analysis and quantitative real time RT-PCR for the sea grasses on MCF-7 was performed. Immune-blotting technique for Bcl-2 and p53 was investigated.
RESULTS: HPLC analysis demonstrated that the extracts contained mainly flavonoids and polyphenols such as Caffeic acid, Chlorogenic acids, catechin and kaempferol that might be responsible for these anticancer effects. Seagrasses alcoholic crude extract markedly suppressed the growth and expansion of MCF-7 cells concentration-dependently with no toxicity against normal human skin fibroblast HSF. Thalassia hemprichii and Enhalus acoroides trigger mode of cell death primarily via apoptosis as confirmed by the flow cytometry. Additionally, they have ability to induce G0/S cell cycle arrest in MCF-7. The data showed the depletion in mitochondrial membrane potential (ΔψM) in the treated cells dose-dependently Caspases 3/7activities markedly increased following 24 h treatment. Finally, Gene expression analysis showed a marked reduction in Bcl-2, Survivin and CDC2 gene expression levels and a significant increase in the expression of p53 and CC2D1A as compared to control cells.
CONCLUSIONS: In summary, the Methanolic extract of seagrass, Thalassia hemperchii and Enhalus ocoroides are able to induce concentration-dependent cytotoxic effects in human MCF-7 cells through intrinsic pathway of apoptosis in MCF-7 cells. This study reveals the beneficial importance of sea grasses as a source of anticancer agents. Further in vivo study is recommended for the active isolated biomolecules.
METHODS: The anticancer effect of the alcoholic extract of different Red Sea Seagrasses on MCF-7 human breast cancer cell line has been investigated. Seagrasses were collected from Wadi El Gamal, Red Sea and extracted. Qualitative HPLC analysis was performed on the extracts for the identification of their active biomarkers. This study was aimed to explore the cytotoxic impact of Thalassia hemprichii (Ehren.) and Enhalus acoroides (L.f.) Royle on MCF-7 and their mode of action. Their anti-proliferative effects on cancer cells were performed using Neutral red assay. On the other hand, their apoptotic effect and their capacity to induce cell cycle arrest were investigated by flow cytometry assay. The effect of Seagrasses on the mitochondrial membrane potential (ΔψM) was studied by using JC-1 mitochondrial membrane potential assay kit in Seagrasses treated cancer cells to Δψ Caspases 3/7activity was examined using the colorimetric method. Gene expression analysis and quantitative real time RT-PCR for the sea grasses on MCF-7 was performed. Immune-blotting technique for Bcl-2 and p53 was investigated.
RESULTS: HPLC analysis demonstrated that the extracts contained mainly flavonoids and polyphenols such as Caffeic acid, Chlorogenic acids, catechin and kaempferol that might be responsible for these anticancer effects. Seagrasses alcoholic crude extract markedly suppressed the growth and expansion of MCF-7 cells concentration-dependently with no toxicity against normal human skin fibroblast HSF. Thalassia hemprichii and Enhalus acoroides trigger mode of cell death primarily via apoptosis as confirmed by the flow cytometry. Additionally, they have ability to induce G0/S cell cycle arrest in MCF-7. The data showed the depletion in mitochondrial membrane potential (ΔψM) in the treated cells dose-dependently Caspases 3/7activities markedly increased following 24 h treatment. Finally, Gene expression analysis showed a marked reduction in Bcl-2, Survivin and CDC2 gene expression levels and a significant increase in the expression of p53 and CC2D1A as compared to control cells.
CONCLUSIONS: In summary, the Methanolic extract of seagrass, Thalassia hemperchii and Enhalus ocoroides are able to induce concentration-dependent cytotoxic effects in human MCF-7 cells through intrinsic pathway of apoptosis in MCF-7 cells. This study reveals the beneficial importance of sea grasses as a source of anticancer agents. Further in vivo study is recommended for the active isolated biomolecules.
摘要：
背景：乳腺癌是女性最常见的癌症类型。寻找具有显著细胞毒性作用和对正常细胞的最小不利影响的新型治疗剂变得至关重要。今天,天然抗癌剂是一种非常规的治疗癌症的方法，作为治疗剂或预防剂，对海洋生物相当关注。
方法：研究了不同红海海草的醇提物对MCF-7人乳腺癌细胞系的抗癌作用。海草是从WadiElGamal收集的，红海和提取。对提取物进行定性HPLC分析以鉴定其活性生物标志物。这项研究的目的是探讨红耳藻（Ehren。)和acoroides(L.f.)Royle关于MCF-7及其作用方式。它们对癌细胞的抗增殖作用使用中性红测定法进行。另一方面,通过流式细胞术检测其凋亡效应和诱导细胞周期停滞的能力。通过在海草处理的癌细胞中使用JC-1线粒体膜电位测定试剂盒研究了海草对线粒体膜电位（ΔΦM）的影响，并使用比色法检查了Caspases3/7活性。对MCF-7上的海草进行基因表达分析和定量实时RT-PCR。研究了Bcl-2和p53的免疫印迹技术。
结果：HPLC分析表明，提取物主要含有类黄酮和多酚，如咖啡酸，绿原酸,儿茶素和山奈酚可能是这些抗癌作用的原因。海草酒精粗提物浓度依赖性地显著抑制MCF-7细胞的生长和扩增，对正常人皮肤成纤维细胞HSF没有毒性。Thalassiahemprichii和Enhalusacoroides主要通过细胞凋亡触发细胞死亡模式，如流式细胞术所证实的。此外,它们具有在MCF-7中诱导G0/S细胞周期阻滞的能力。数据显示，在24小时处理后，经处理的细胞中线粒体膜电位（ΔΦM）的消耗呈剂量依赖性，Caspases3/7活性显着增加。最后,基因表达分析显示，与对照细胞相比，Bcl-2，Survivin和CDC2基因表达水平显着降低，p53和CC2D1A的表达显着增加。
结论：总之，海草的甲醇提取物，Thalassiahemperchi和Enhalusocoroides能够通过MCF-7细胞凋亡的内在途径在人MCF-7细胞中诱导浓度依赖性细胞毒性作用。这项研究揭示了海草作为抗癌剂来源的有益重要性。建议对活性分离的生物分子进行进一步的体内研究。
方法：研究了不同红海海草的醇提物对MCF-7人乳腺癌细胞系的抗癌作用。海草是从WadiElGamal收集的，红海和提取。对提取物进行定性HPLC分析以鉴定其活性生物标志物。这项研究的目的是探讨红耳藻（Ehren。)和acoroides(L.f.)Royle关于MCF-7及其作用方式。它们对癌细胞的抗增殖作用使用中性红测定法进行。另一方面,通过流式细胞术检测其凋亡效应和诱导细胞周期停滞的能力。通过在海草处理的癌细胞中使用JC-1线粒体膜电位测定试剂盒研究了海草对线粒体膜电位（ΔΦM）的影响，并使用比色法检查了Caspases3/7活性。对MCF-7上的海草进行基因表达分析和定量实时RT-PCR。研究了Bcl-2和p53的免疫印迹技术。
结果：HPLC分析表明，提取物主要含有类黄酮和多酚，如咖啡酸，绿原酸,儿茶素和山奈酚可能是这些抗癌作用的原因。海草酒精粗提物浓度依赖性地显著抑制MCF-7细胞的生长和扩增，对正常人皮肤成纤维细胞HSF没有毒性。Thalassiahemprichii和Enhalusacoroides主要通过细胞凋亡触发细胞死亡模式，如流式细胞术所证实的。此外,它们具有在MCF-7中诱导G0/S细胞周期阻滞的能力。数据显示，在24小时处理后，经处理的细胞中线粒体膜电位（ΔΦM）的消耗呈剂量依赖性，Caspases3/7活性显着增加。最后,基因表达分析显示，与对照细胞相比，Bcl-2，Survivin和CDC2基因表达水平显着降低，p53和CC2D1A的表达显着增加。
结论：总之，海草的甲醇提取物，Thalassiahemperchi和Enhalusocoroides能够通过MCF-7细胞凋亡的内在途径在人MCF-7细胞中诱导浓度依赖性细胞毒性作用。这项研究揭示了海草作为抗癌剂来源的有益重要性。建议对活性分离的生物分子进行进一步的体内研究。
