Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/92630
標題: 八種中草藥複合物(8KHC)抗白血病之活性分析
Analysis of the anti Leukemia Activity of the Eight Kind Herbs Compounds (8KHC)
作者: 蕭智鴻
Chih-Hong Hsiao
關鍵字: 八種中草藥複合物;白血病;細胞增殖率;8KHC;Leukemia;Cell Proliferation rates
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摘要: 
根據台灣兒童癌症基金會統計結果,白血病約佔所有兒童癌症的 31%,大多數患者生存期短,目前對白血病的治療方式是以化學藥物誘導癌細胞凋亡同時亦造成正常細胞的損害。本研究利用咸豐草(Bidens pilosa)等八種植物混合靜置 40 天後以水萃取,再經冷乾磨粉後的複合物(8KHC),為本試驗的活性分析材料,探討對小鼠與人類正常細胞的毒性與誘導人類白血病細胞分化及增殖現象等之影響。使用 8 週 ICR 公鼠以口服方式進行投藥,每天 1 次連續 28 天進行體內急性毒性試驗,計算 Lethal Dose 50% (LD 50),每天觀察兩次記錄臨床症狀、死亡情形及每週體重之變化。首先以二次蒸餾水(ddH2O)與二甲基亞碸 (DMSO) 進行劑比較試驗,於人類正常細胞SC試驗發現純水為溶解 8KHC 最適合的溶劑。選用人類白血病細胞株 K562, MOLT-4,以人類的正常細胞 SC 作為對照組,使用 MTS 試驗法測定細胞活性,以線性回歸分析影響濃度之 IC50。以細胞凋亡試驗法測定半胱天冬酶-3 (Caspase-3) 活性;以 BrdU 細胞增殖試驗法測定細胞複製所需之 BrdU量。試驗結果以 Prism V4.0 軟體進行統計分析。二種溶劑的比較發現 ddH2O 不影響人類正常細胞,而 DMSO 則會抑制人類正常細胞的活性。高劑量的 8KHC 口服急性毒性試驗結果發現,小鼠所有的臨床症狀正常,亦無任何死亡情形,平均體重亦無顯著差異。以MTS 試驗法測定細胞活性結果發現, 8KHC 可抑制白血病細胞株的活性;細胞凋亡試驗顯示 8KHC 不會直接讓白血病癌細胞走向凋亡路徑;細胞增殖試驗顯示 8KHC 部會影響正常細胞 SC 的增殖率,而可顯著抑制白血病細胞株的增殖。本研究結果顯示,8KHC 對 ICR 小鼠是安全且不具急毒性,對正常細胞的活性與增殖並無影響,且在低劑量 (1mg/ml) 即具有顯著的效果,可抑制白血病癌細胞 K562 67%, MOLT-4 40% 的活性;抑制 K562 62%, MOLT-4 61% 的細胞增殖率,與目前的化療藥物 As2O3 有相同的效果。證明 8KHC 具有顯著抑制白血病癌細胞活性的效果,由此推論,天然無毒的 8KHC 是一個可使急性白血病患者快速且有效獲得緩解的天然複合物。

The statistics of Children's Cancer Foundation of Taiwan show that 31% of childhood cancer disease is leukemia, the majority of patients has a short life expectancy, the current therapy for leukemia is adopted chemotherapy to induce cancer cells go apoptosis and cause damage to the normal cells at the same time. The purpose of this study was to investigate the anti human leukemia cell lines activity of the eight kind herbs compounds (8KHC) which were extracted from eight kind herbs by standing for 40 days, freeze drying and grinding to a fine powder. The solvent comparison test with the normal human cells with ddH2O and DMSO found that the distilled water is a suitable solvent for 8KHC application. Eight weeks ICR male mice were subjected to the oral acute toxicity test to determine the Lethal Dose 50% (LD 50) by daily feeding for 28 days. To determine the effective IC50, the human leukemia cell lines K562 and MOLT-4 were applied to 8KHC treatment with SC cell line as a control and used the MTS assay to determine cell viability. In addition, cell apoptosis was determined by Caspase-3 activity analysis; cell proliferation rate was detected by BrdU incorporation assay. All the experiments were conducted with three replications and analyzed by using Prism V4.0 software, the means with standard deviation were analyzed with 'Student's t-test'. The experiment results suggested that all clinical signs are normal, 100% survival rates, and no significant difference in body weight of ICR mice in the high dose 8KHC oral acute toxicity test. In solvent comparison, we found that ddH2O did not affect human normal cell but the DMSO suppressed human normal cell activity. The cell activity assay suggested that 8KHC can inhibit leukemia cell activity. Cell apoptosis assay shown that 8KHC did not induce the leukemia cell lines go apoptosis pathway. Analysis of cell proliferation suggested that 8KHC can inhibit leukemia cell prolifetration. In conclusions, 8KHC was safe, no acute toxicity, no negative effect on human normal cell SC, and had significant suppression in cell activity of leukemia cancer cell lines K562 67% and MOLT-4 40%; and inhibited cell proliferation rates of K562 62% and MOLT-4 61%. This study showed that 8KHC is able to effectively inhibit leukemia cells activity and cell proliferation and shown similar effect as Arsenic Trioxide (As2O3). It's suggested that the 8KHC may provide an alternative therapy for acute leukemia patients.
URI: http://hdl.handle.net/11455/92630
其他識別: U0005-0402201415252300
Rights: 同意授權瀏覽/列印電子全文服務,2017-02-05起公開。
Appears in Collections:生命科學院碩士在職專班

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