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標題: 管餵苦丁茶甲醇萃取物對皮下負載TRAMP-C1癌細胞之C57BL/6J雄鼠體內免疫反應及癌化指標變化之影響
Effects of Ilex kudingcha methanol extracts on immune responses and carcinogenic markers in male C57BL/6J mice subcutaneously loaded with TRAMP-C1 cancer cells
作者: 張曉青
Sheau-Ching Chang
關鍵字: 癌症免疫療法;苦丁茶甲醇萃取物;小鼠前列腺癌細胞TRAMP-C1;Cancer immunotherapy;Ilex kudingcha methanol extract;Mouse prostate cancer TRAMP-C1 cells
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苦丁茶 (Ilex kudingcha)為傳統天然保健飲品,具有抗氧化、抗發炎等功效。本研究室先前研究已發現苦丁茶甲醇萃取物在體外實驗中,具有免疫調節功效,且對於人列腺癌PC-3細胞有抗癌之潛力,然而,其在體內之免疫調節及抗前列腺癌效果仍不清楚,因此本研究以小鼠前列腺癌TRAMP-C1細胞進行動物體外及體內實驗,以評估苦丁茶甲醇萃取物之功效。在體外實驗模式,發現直接添加與癌症免疫療法模式下,適當濃度之苦丁茶甲醇萃取物對於小鼠前列腺癌TRAMP-C1細胞的生長有抑制效果。以不同濃度癌細胞注入皮下以建立皮下負載TRAMP-C1癌細胞之C57BL/6J雄鼠動物模式,結果發現,於皮下注射癌細胞,使C57BL/6J小鼠免疫反應傾向Th1免疫平衡;觀察小鼠腫瘤大小及腫瘤組織病理變化情形,建議以注射低濃度癌細胞 (5 × 104 cells/mouse) 經過30天之條件,最適合做為動物體內負載前列腺癌之實驗模式。
利用已建立的動物體內模式管餵苦丁茶甲醇萃取物4週,結果顯示管餵苦丁茶甲醇萃取物,使C57BL/6J雄鼠脾臟細胞及腹腔巨噬細胞細胞激素分泌,傾向抗發炎之免疫平衡。此外,苦丁茶甲醇萃取物可增加小鼠周邊血液淋巴世系細胞分布比率,增加其免疫能力。觀察腫瘤組織病理變化發現,給予苦丁茶甲醇萃取物能改善腫瘤惡化情形;癌症指標結果顯示,給予小鼠中劑量 (200 mg/kg b.w./day) 樣品能降低血清中TNF-α濃度,管餵中劑量、低劑量 (50 mg/kg b.w./day) 樣品可降低小鼠血清中前列腺癌專一性抗原PSA濃度。

Ilex kudingcha, serving as a traditional natural health drink, has antioxidant and anti-inflammatory effects etc. In our preliminary studies, Ilex kudingcha methanol extract (IKME) has immunomodulatory effects and anti-cancer potential for human prostate carcinoma PC-3 cells in vitro. However, immunomodulatory and anti-prostate cancer effects of IKME in vivo are still unclear. To unravel the puzzle, mouse prostate cancer TRAMP-C1 cells were used to investigate the effects of IKME in vitro and in vivo. In vitro, we found that IKME at an appropriate concentration had anti-cancer effects against the growth of TRAMP-C1 cells via either direct addition or cancer immunotherapy. To establish an animal model for evaluating IKME effects on TRAMP-C1 cells in vivo, C57BL/6J male mice were injected subcutaneously with different concentrations of TRAMP-C1 cells for different days. The results showed that C57BL/6J male mice subcutaneously loaded with TRAMP-C1 cells had Th1-inclination immune balance. Based on tumor sizes and pathological changes of tumor tissues in the experiment mice, it is recommended that C57BL/6J male mice that are subcutaneously injected with low dose cancer cells (5 × 104 cells/mouse) for 30 days is the most suitable animal model for subsequent experiments in vivo.
Using the established animal model, IKME was administered to the experiment animal by gavage for 4 weeks. It was found that IKME administration resulted in anti-inflammatory effects according to cytokine secretion profiles by splenocytes and peritoneal macrophages of C57BL/6J male mice subcutaneously loaded with TRAMP-C1 cancer cells, respectively. Besides, IKME administration increased the percentage of lymphoid lineage cells in peripheral blood, enhancing their immunity of the experiment animals. Observation of pathological changes in tumor tissues indicated that IKME might improve the status of tumor deterioration. Evaluation on carcinogenic markers showed that medium (200 mg/kg b.w./day ) doses of IKME administration reduced serum TNF-α concentration in the experiment mice. Medium and low dose (50 mg/kg b.w./day ) of IKME administration significantly lowered serum prostate cancer specific antigen (PSA) concentration.
In conclusion, IKME administration in vitro could inhibit the growth of mouse prostate cancer TRAMP-C1 cells through either direct addition or indirect immunotherapy. In vivo, IKME administration tended to have a Th2-inclination immune balance in C57BL/6J mice subcutaneously loaded TRAMP-C1 cancer cells. IKME administration reduced pro-inflammatory cytokine secretions in the body, but increased the distribution of peripheral blood lymphocytes. Furthermore, IKME administration might improve the necrotic status in tumor tissues via increasing the immunity of the experiment mice. IKME has the potential to resist prostate cancer in vitro and in vivo.
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