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|標題:||Studies on the quercetin metabolism and the immunomodulatory functions of alcohol-soluble and alcohol-insoluble fractions from strawberry and mulberry juices using primary immune cells
|關鍵字:||quercetin;槲皮素;strawberry juice;mulberry juice;primary immune cells;uptake and metabolism;immunomodulatory functions;草莓榨汁液;桑椹榨汁液;初代免疫細胞;攝入與代謝;免疫調節功能||出版社:||食品暨應用生物科技學系||摘要:||
The aim of this study was to evaluate the immunomodulatory functions of alcohol soluble of strawberry and mulberry juice (AS-S and AS-M), and alcohol-insoluble residue of strawberry and mulberry (AIR-S and AIR-M) using primary immune cells in vitro. AS-S and AS-M including a standard flavonoid, quercetin, were conducted to investigate their metabolism by primary immune cells. For the sakes, the experiments in this study were divided into two parts. The first part of this study was to determine the metabolism of flavonoids in AS-S and AS-M by immune cells and the immunomodulatory functions of AS-S and AS-M on the immune cells. The second part of this study was to determine the effects of AIR-S and AIR-M administration on immunomodulation and apoptosis of immune cells.
The results from the first part in this study showed that AS-S and AS-M demonstrated differential effects on primary immune cells either entering the cells or not. Fewer metabolites of quercetin quinones from AS-S and AS-M than that of standard quercetin administrated alone were detected by HPLC. The results suggest that the quercetin in AS-S and AS-M are more stable than purified quercetin. AS-S and AS-M administration significantly decreased the secretion of Th1 cytokine IFN-γ (P<0.05), however increased the secretion of Th2 cytokine IL-10 produced by lipopolysaccharide (LPS)-stimulated splenocytes. AS-S and AS-M administration did not significantly affect the inflammation status of LPS-stimulated peritoneal cells (mostly macrophages). These results suggested that the anti-inflammatory constitutes in strawberry and mulberry juice were AIR-S and AIR-M, but not AS-S and AS-M.
The second part of this study showed that AIR-S and AIR-M administration significantly increased the proliferation of splenocytes from female BALB/c mice (P<0.05). AIR-S administration significantly decreased the ratio of IL-2/IL-10 (Th1/Th2) (P<0.05) secreted by splenocytes compared to the control. AIR-S administration demonstrated Th2-prone immune responses. Though AIR-M administration significantly increased both Th1 and Th2 cytokine secretion (P<0.05), AIR-M administration significantly decreased the ratio of (IL-2+TNF-α) / (IL-4+IL-5+IL-10) (Th1/Th2) (P<0.05) secreted by splenocytes. In LPS-stimulated peritoneal cell cultures, AIR-S and AIR-M administration significantly inhibited the secretion of pro-inflammatory cytokine IL-1β and IL-6 (P<0.05). Simultaneously, AIR-S and AIR-M administration increased the secretion of anti-inflammatory cytokine IL-10 (P<0.05). Both AIR-S and AIR-M demonstrated an anti-inflammatory potential against LPS-stimulated peritoneal cells. The administration with AIR-S and AIR-M to the LPS-stimulated peritoneal cell cultures enhanced the expression of bcl-2, an anti-apoptotic protein located at mitochondria. However, AIR-S and AIR-M administration could not significantly affect the bcl-2 expression using primary splenocyte cultures.
In conclusion, the results in this study indicated that quercetin, AS-S and AS-M exerted differential immunomodulatory effects on different immune cells. Esperically, quercetin or its metabolites might modulate primary splenocytes via entering the cells. In the immunomodulatory section, AIR-S and AIR-M extracted from strawberry and mulberry juice exhibited potent anti-inflammatory and anti-apoptotic effects in vitro.
本論文之目的主要在探討經酒精分離之草莓及桑椹榨汁液中溶於酒精之萃取物 (alcohol solube of strawberry and mulberry juice, AS-S and AS-M)，及不溶於酒精之沈澱物 (alcohol-insoluble residue of strawberry and mulberry, AIR-S and AIR-M)，其對體外免疫細胞之調節作用，並以槲皮素 (quercetin) 為標準品，探討免疫細胞對AS-S及AS-M中類黃酮之代謝，因此，第一部份實驗主要在探討免疫細胞對AS-S及AS-M中類黃酮之代謝，及評估AS-S及AS-M免疫調節作用，第二部份實驗主要在評估AIR-S及AIR-M對免疫細胞之調節及其對細胞凋亡之影響。
第一部份實驗顯示，槲皮素、AS-S及AS-M在相同或不同免疫細胞中，槲皮素及類黃酮之代謝均有不同影響。實驗結果發現，AS-S及AS-M中之槲皮素均比單獨存在之槲皮素標準品穩定，不會產生許多槲皮素苯醌類代謝產物而影響細胞。在免疫調節評估部份，添加脂多醣 (lipopolysaccharide, LPS) 誘發脾臟細胞發炎下，分別添加AS-S及AS-M，均顯著降低Th1細胞激素IFN-γ之分泌量 (P<0.05)，且增加Th2細胞激素IL-10之分泌量，使免疫反應傾向Th2反應。在以LPS誘發腹腔細胞發炎下，分別添加AS-S及AS-M則沒有明顯之抗發炎趨勢，推測影響腹腔抗發炎之活性成分，可能存在於酒精沈澱物部份 (AIR-S及AIR-M)。
第二部份實驗顯示，AIR-S及AIR-M均能顯著刺激脾臟細胞增生(P<0.05)，在調節細胞激素分泌方面，AIR-S之添加顯著降低脾臟細胞細胞激素IL-2/IL-10 (Th1/Th2) 之比值 (P<0.05) ，AIR-M之添加則明顯刺激Th1及Th2細胞激素之分泌 (P<0.05) ，但 (IL-2+TNF-α) / (IL-4+IL-5+IL-10) (Th1/Th2) 之比值顯著減少 (P<0.05)；腹腔細胞培養顯示，AIR-S及AIR-M均顯著抑制LPS誘發腹腔細胞發炎所分泌之發炎型細胞激素IL-1β及IL-6 (P<0.05)，並同時增加抗發炎細胞激素IL-10之分泌量 (P<0.05)，具有良好之抗發炎能力。評估經粒線體途徑之細胞凋亡實驗顯示，AIR-S及AIR-M刺激LPS誘發發炎腹腔細胞的抗凋亡蛋白bcl-2表現，但對脾臟細胞凋亡表現則沒有明顯之作用趨勢。整體而言，AIR-S及AIR-M具有良好抑制腹腔細胞發炎之能力，具有調節脾臟細胞活化，使免疫細胞傾向Th2之免疫反應。
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