台灣土肉桂葉精油與其活性成分枷羅木醇及肉桂醛對potassium oxonate和尿酸誘發高血尿酸小鼠之肝、腎保護作用探討

The effect of the leaf essential oil of Cinnamomum osmophloeum Kanehira and its active ingredients linalool and cinnamaldehyde on liver and renal protection in hyperuricemia mice induced by potassium oxonate and uric acid

麥庭瑜(Ting-Yu Mai)

指導教授 ： 劉承慈

中山醫學大學/健康管理學院/營養學系碩士班/碩士(2017年)

摘要

目前常見的高血尿酸動物模式之一是以含有potassium oxonate和尿酸之飲食所誘發，已知此模式會影響實驗動物腎臟轉運蛋白之表現量，但尚不清楚此模式是否影響肝、腎功能及誘發其發炎性反應，因此，本研究目的為觀察此模式下是否對於肝、腎功能指標及其發炎性反應之影響，並進一步探討此模式灌食枷羅木醇型土肉桂葉精油（TC）及其活性成分枷羅木醇（linalool, Lin）和肉桂醛（cinnamaldehyde, Cin）之作用，並與目前臨床藥物allopurinol和colchicine相比較。實驗以雄性C57BL/6小鼠隨機分為十組，其中九組餵食高尿酸飲食，其餘一組則餵食一般飲食共56天，在開始誘發高血尿酸後第42天起，每隔一日分別灌食：載劑玉米油（2 ml/kgBW）、土肉桂葉精油（13 mg/kg BW）、枷羅木醇合併肉桂醛（5.2 mg/kg和0.9 mg/kg BW）、低劑量枷羅木醇（2.6 mg/kg BW）、高劑量枷羅木醇（5.2 mg/kg BW）、低劑量肉桂醛（0.45 mg/kg BW）、高劑量肉桂醛（0.9 mg/kg BW）、allopurinol（5 mg/kg BW）、colchicine（1.5 mg/kg BW），正常飲食之控制組則灌食載劑玉米油（2 ml/kgBW），共灌食八次，於最後一次灌食結束後，禁食12小時犧牲收集血液、腎臟與肝臟備用於分析。結果顯示，在長期給予含2 %（w/w）potassium oxonate和3 %（w/w）尿酸之飲食的模式下，與正常飲食組小鼠相較，雖未顯著影響小鼠肝功能指數ALT、AST，但造成肝臟中nitrate/nitrite含量增加並促進xanthine oxidase活性上升（P<0.05），且也提升小鼠血液BUN值及腎臟中nitrate/nitrite含量（P<0.05）。然而，未發現肝、腎中發炎激素IL-1β和TNF-α的含量或腎中TLR2/TLR4、NLRP3發炎相關訊號路徑分子蛋白質表現量受到此模式的影響。另一方面，此模式可抑制腎臟尿酸轉運蛋白OAT1 mRNA表現量（P<0.05），與灌食載劑組相較，TC可降低肝臟xanthine oxidase活性且TC及其活性成分Lin與Cin亦可降低肝指數AST，而allopurinol也有降低AST的效果（P<0.05），但colchicine則無。另外也發現Cin可降低高尿酸飲食下之BUN值（P<0.05），且colchicine也有降低BUN的效果（P<0.05），但allopurinol則無。TC及Lin和Cin不但降低肝、腎nitrate/nitrite含量，且也降低肝、腎中發炎激素IL-1β及/或TNF-α之含量，其改善這些數值的幅度與allopurinol及colchicine的效果相當，且與降低腎中TLR2、MYD88、MD2、NLRP3、ASC和caspase-1 p20蛋白質表現量及caspase-1活性有關。在腎臟尿酸轉運蛋白表現量方面，Lin和Cin顯著降低URAT1和增加OAT1、OAT3轉運蛋白mRNA表現量，且顯著降低URAT1和增加OAT3蛋白質表現量，顯示此兩種活性成分有助於尿酸排泄，但TC未影響這些尿酸轉運蛋白之表現。總言之，利用此高尿酸飲食模式，本研究顯示Lin與Cin提供抗發炎及促進血尿酸排除之作用，適用於預防高血尿酸相關肝、腎病症，而TC亦可用於防治高血尿酸相關之肝、腎發炎性損傷。

關鍵字

高尿酸血症 ； potassium oxonate ； 尿酸 ； 土肉桂葉精油 ； 枷羅木醇 ； 肉桂醛 ； 尿酸轉運蛋白 ； TLR2 ； TLR4 ； NLRP3 ； 促發炎激素

並列摘要

One of the most common hyperuricemic animal models is induced by a diet containing potassium oxonate and uric acid. It is known that this model will affect the expression of urate transporter in kidney of the experimental animals, but it is not clear whether this model affects the function and inflammatory response of liver or kidney. The aim of the present study is to investigate the effects of this model and the intervention with Cinnamomum osmophloeum Kanehira (TC) and its active components, linalool (Lin) and cinnamaldehyde (Cin) on the function and inflammatory response of liver and kidney. Positive control was intervened with either allopurinol or colchicines. Male C57BL/6 mice were randomly assigned to ten groups, of which nine groups were fed the high uric acid (HU) diet containing 2% (w/w) potassium oxonate and 3% (w/w) uric acid, and the rest group was fed a general diet for 56 days. On the day when treated with HU diet for 42 days, mice were intervened with corn oil (2 ml/kgBW), TC (13 mg/kgBW), Lin (5.2 mg/kg) plus Cin 0.9 mg/kg BW), Lin (2.6 or 5.2 mg/kgBW), Cin (0.45 or 0.9 mg/kgBW), allopurinol (5 mg/kgBW) or colchicine (1.5 mg/kgBW) by gavage every other day until day 56. Subsequently, all mice were fasting overnight and then sacrificed by CO2. The result showed that the persistent treatment with the HU diet, compared with the normal diet, significantly increased the content of nitrate/nitrite and the activity of xanthine oxidase in the liver (P<0.05), although not significantly affecting the activity of ALT, AST in peripheral blood (P>0.05). Moreover, HU diet increased BUN value and nitrate/nitrite content in the kidney (P<0.05). However, the expression of IL-1β and TNF-α in the liver and kidney, or the expression of TLR2/TLR4 and NLRP3-related signaling pathways in the kidney were not affected by HU diet. On the other hand, HU diet inhibited the expression of uric acid transporter OAT1 mRNA (P<0.05). Compared with the corn oil group, TC reduced the activity of liver xanthine oxidase. TC and its active components Lin and Cin also reduced peripheral AST activity. Allopurinol reduced peripheral AST activity as well (P<0.05), but colchicine did not. It was found that Cin reversed HU diet induced elevation of BUN (P<0.05), colchicine but not allopurinol showed to be effective on reducing HU diet-induced BUN levels as well (P<0.05). Among mice received HU diet, compared with mice intervened with corn oil, TC, Lin and Cin not only reduced the content of nitrate/nitrite, but also the content of IL-1β and/or TNF-α in liver and kidney and to an extent similar to that by allopurinol and colchicines. This can be attributable to decreased expression of TLR2, MYD88, MD2, NLRP3, ASC and caspase-1 p20 protein and caspase-1 activity in kidney. In terms of uric acid transporter expression in the kidney, Lin and Cin significantly reduced the expression of URAT1 mRNA and protein, while significantly increased the expression of OAT1 and OAT3 mRNA and OAT3 protein, indicating the capability of both active ingredients of TC to promote uric acid excretion. Nevertheless, TC was not found to affect the expression of either mRNA or protein of these uric acid transporters. In conclusion, this study shows that in mice received HU diet, TC and its active components Lin and Cin possess protective effect on liver and kidney through anti-inflammatory effect. In addition, Lin and Cin are also protective through the uricosuric effect.

並列關鍵字

hyperuricemia ； potassium oxonate ； uric acid ； Cinnamomum osmophloeum Kanehira ； linalool ； cinnamaldehyde ； uric acid transporter ； TLR2 ； TLR4 ； NLRP3 ； proinflammatory cytokine

參考文獻

Chuang, S. Y., Lee, S. C., Hsieh, Y. T., & Pan, W. H. (2011). Trends in hyperuricemia and gout prevalence: Nutrition and Health Survey in Taiwan from 1993-1996 to 2005-2008. Asia Pacific journal of clinical nutrition, 20(2), 301-308.

王詩芸. (2015). 台灣土肉桂葉精油與活性成分枷羅木醇及肉桂醛對於 LPS 誘發系統性發炎反應小鼠之影響. 中山醫學大學營養學研究所學位論文, 1-147.

何坤益. (2006). 台灣產土肉桂, 山肉桂, 及胡氏肉桂 (樟科) 之遺傳變異與分類地位. Bio Formosa, 41(2), 93-102.

翁碩駿, 徐國雄, 唐德成, & 吳明儒. (2012). 以腎臟科醫師的觀點看高尿酸血症. 腎臟與透析, 24(3), 149-154.

劉婉君, 洪啓智, 張哲銘, 黃尚志, & 陳鴻鈞. (2010). 尿酸與腎臟病. 內科學誌, 21(3), 197-203.