| 摘要: | 研究顯示膳食炸油會顯著惡化小鼠結腸炎病程，但致病機轉不明，因此本研究目的在於探討飼料添加白藜蘆醇，是否有具有降低或緩和炸油惡化結腸發炎病程的功效，由抗氧化/助氧化系統、發炎反應與腸黏膜防禦系統三大方面著手，探究白藜蘆醇的效應機轉。
採用 7 週齡 C57BL/6 品系雌鼠 40 隻，隨機分成3組，包括控制組 (15% 新鮮油，C)、炸油組 (15% 炸油，O)、炸油加白藜蘆醇組 (15% 炸油 + 白藜蘆醇，R)，實驗全程供應 AIN-93G修正後的15%高油飼料，令其自由攝取。飼養 6 週後於飲用水中添加 2% (w/v) (Dextran sulfate sodium, DSS)，誘發腸道發炎，並計算疾病活動指數 (disease activity index, DAI)，連續 5 天後換成去離子水 2 天後犧牲，犧牲前禁食24小時，採集血液、肝臟與大腸，分析脂質過氧化指標TBARS (Thiobarbituric acid reactive substance)、發炎相關細胞激素、抗氧化分子、酵素活性與大腸黏膜完整性相關mRNA 基因表現。
結果顯示：(1) 生長狀況：給予2% DSS前，O組和R組的生長體重和飼料利用率顯著低於C組，給予2% DSS誘導結腸炎期間，第2~7天O組和R組的DAI指數皆顯著高於C組，顯示炸油影響生長狀況且有惡化腸炎病程發展的特性；(2) 抗氧化/助氧化系統：結腸方面，O組與R組的TAS (Total Antioxidant Status) 皆顯著高於C組；肝臟方面，脂質過氧化指標TBARS結果，O組和R組皆顯著高於C組，表示炸油餵飼顯著增加肝臟脂質過氧化；R組的維生素C含量顯著高於C組，表示炸油能耗盡肝臟中維生素E的含量，為提高維生素E的還原作用，相對的維生素C也隨之增加；O組和R組的 GST (Glutathione S-transferase)和Catalase皆顯著高於C組，O組的 SOD (Superoxide dismutase) 和Se-GPX (Se-Glutathione peroxidase) 皆顯著低於C組，R組的Se-GPX顯著低於C組；O組和R組的ALT (Alanine transaminase) 皆顯著高於C組，顯示炸油餵飼有輕微肝損傷情況。(3) 發炎反應：結腸方面，O組的血清發炎指標Hp (Haptoglobin) 皆顯著高於C組和R組，R組的IL-6有趨勢低於C組和O組，O組和R組的 MPO (Myeloperoxidase) 顯著低於C組，mRNA表現，C組的NOX1 (NADPH oxidase 1) mRNA皆顯著高於O組和R組，表示炸油有促使全身性發炎狀況。肝臟：R組的COX-2 (Cyclooxygenase-2)、IL-6 (Interleukin-6) mRNA皆顯著低於C組和O組，R組的IL-1β mRNA顯著低於C組。白藜蘆醇有減緩炸油對肝臟的促發炎細胞激素分泌及助氧化分子的基因表現。(4) 腸黏膜防禦系統：R組的MMP3 (Matrix metalloproteinases)、MMP9、MUC2 (Mucins2)、TEF3 (Trefoil factors-3)和HES1(Hairy enhance of split-1) 三組之間皆沒有顯著差異。
結論：添加白藜蘆醇無法緩和腸炎病程進展，推測（1）白藜蘆醇添加劑量太低，保護效果不顯著，（2）抗氧化失衡或加劇發炎可能不是炸油惡化腸炎病程的主因，另外白藜蘆醇在肝臟降低促發炎細胞激素的效應明顯大於結腸。
Our previous studies indicated that dietary oxidized frying oil (OFO) resulted in deterioration of dextran sulfate sodium (DSS)-induced colitis in mice. However, mechanisms for these effects remain unknown. The purpose of this study was to investigate the effect of trans-resveratrol supplementation on DSS-induced colitis in OFO-fed mice. In this study, to elucidate the possible mechanisms underlying OFO effects, we focus on three aspects including antioxidant-prooxidant system, inflammation and colonic mucosa function.
7-wk-old female C57BL/6 mice (n=40) were used in this study. They were acclimatized in our animal house were fed powder diets and water ad libitum. There were randomized into three dietary groups, including C group (15% fresh soybean oil), O group (15% OFO) and R group (15% OFO+10 mg trans-resveratrol/kg diet). After the 6-wk feeding period, mice were treated by 2% (w/v) DSS administered in drinking water for 5 days, followed by providing DSS-free water for 2 days. After the 12 h fasting period, mice were sacrificed with carbon dioxide asphyxiation. Blood, liver and colon were collected and assessed for associated parameters, such as inflammatory cytokines, antioxidant defense system, and colonic mucosal function.
Our results showed that body weight gain and feed efficiency were significantly decreased in the O and R groups than in the C group. Mice exposed to DSS to induce acute colitis, disease activity index (DAI) were significantly increased in the O and R groups than in the C group. According to DAI results, we found no significant preventive effect of resveratrol on OFO-worsen colitis. Serum total antioxidant status (TAS) levels were significantly increased in the O and R groups than in the C group. We used serum haptoglobin (Hp) to assess the state of systemic inflammation. We found that O group was significantly elevated compared with C group, but had a significantly decrease in R group compared with O group. In addition, serum alanine transaminase (ALT) activity in the O and R group had a increase compared to the C group.
Thiobarbituric acid reactive substance (TBARS) in the liver of the O group and R group were higher than those of C group. That indicated that TBARS were significantly increased in OFO-fed mice. The level of vitamin C was significantly higher in R group than that in C group. Glutathione S-transferase (GST) and catalase activities in the O group and R group were significantly increased compared to the C group, superoxide dismutase (SOD) and Se-Glutathione peroxidase (Se-GPX) activities in the O group was significantly decreased compared to the C group, the activities of Se-GPX was significantly lower in C group than that in R group. Colonic myeloperoxidase (MPO) activities were significantly decreased in the O group and R group. We conjectured that had distinctive effect of OFO on MPO expression. In the mRNA expression of liver, results showed relatve amounts of COX-2 and IL-6 mRNA in R group were decreased as compared with C group and O group, the expression of IL-1β mRNA were significantly lower in the R group than in the C group. Trans-resveratrol had a significant decrease on hepatic proinflammatory cytokines expression in DSS-induced colitis in mice. Results showed that colonic mRNA expression were no significant differences existed among 3 groups, including matrix metalloproteinases (MMP3), MMP9, mucin2 (MUC2), trefoil factors-3 (TEF3) and hairy enhance of split-1(HES1).
In conclusion, trans-resveratrol had no marked inhibitory effect on OFO-worsen DSS-induced colitis in mice. We speculated that (1) the dosage of trans-resveratrol was too low, (2) the adverse effect of OFO might be not due to either the antioxidant imbalance or aggravating inflammation. In addition, the lowing effects of trans-resveratrol on pro-inflammatory cytokines were significant in liver than in colon. |
