以高脂飲食誘導肥胖大鼠模式探討Epigallocatechin gallate-quercetin-rutin複方對肥胖與腸道菌相之影響

肥胖為重要的公共衛生議題，許多慢性疾病的發生都與肥胖密切相關。高油脂飲食會造成腸道菌相的失衡，進而加劇慢性發炎與肥胖之發生。先前文獻指出，表沒食子兒茶素沒食子酸酯 (epigallocatechin gallate, EGCG)、槲皮素 (quercetin) 和芸香苷 (rutin) 對肥胖具有改善作用，加上複方型式的產品具有協同作用之特性，因此本研究以組合複方的方式開發具抗肥胖作用之 EGCG-quercetin-rutin (EQR) 複方，並以高脂飲食預先誘導Wistar大鼠為肥胖症之動物模式，評估EQR複方改善肥胖與調控腸內菌相之影響，並釐清其分子作用機轉。本實驗將60隻6週齡Wistar雄性大鼠進行預養一週後，動物隨機分成正常飲食組 (normal diet, ND) 與高脂飲食組 (high-fat diet, HFD)。其中高脂飲食組大鼠以HFD誘導四週產生肥胖症 (肥胖大鼠之體重顯著高於正常飲食大鼠10%以上，即可進行後續介入試驗)。肥胖大鼠隨機分成五組 (n=10/group)，包括：組別1：高脂飲食組 (HFD)、組別2：高脂飲食給予EQR複方低劑量組 (HFD + EQR formula-low dose, HFD + EQRF-LD)、組別3：高脂飲食給予EQR複方中劑量組 (HFD + EQR formula-medium dose, HFD + EQRF-MD)、組別4：高脂飲食給予EQR複方高劑量組 (HFD + EQR formula-high dose, HFD + EQRF-HD) 與組別5：高脂飲食給予市售健康食品正控制組 (HFD + positive control, HFD + PC) 為期四週。結果顯示，給予肥胖大鼠EQR複方後可顯著抑制高脂飲食所誘導之體重和脂肪組織重量增加，並降低肝臟總脂質含量。由肝臟基因表現結果得知，給予EQR複方可提升高脂飲食誘導肥胖大鼠之PPAR-、ACO、CPT-1和HSL基因表現，並抑制其PPAR-和FATP-1基因表現。此外，在脂肪組織基因表現上，給予EQR複方可提升高脂飲食誘導肥胖大鼠之SIRT1和CPT-1基因表現，並抑制其FAS和FATP-1基因表現。在棕色脂肪基因表現上，給予EQR複方可提升高脂飲食誘導肥胖大鼠之AMPK和PGC-1基因表現。在腸道菌相的結果部分，EQR複方可提升菌群豐富度及多樣性；在菌門層面可降低Deferribacteres之相對豐富度，並增加Fusobacteria之相對豐富度；在菌科層面可增加Fusobacteriaceae和Christensenellaceae之相對豐富度；在菌屬層面可降低Mucispirillum之相對豐富度，並增加Enterorhabdus、Christensenellaceae R-7 group、Lachnoclostridium和Parvibacter之相對豐富度；在糞便短鏈脂肪酸的部分，其可增加乙酸、丙酸、丁酸和總短鏈脂肪酸的排出。綜合以上動物實驗得知，EQR複方可改善高脂飲食所誘導大鼠肥胖症，亦可逆轉高脂飲食之菌群組成，進而達到抗肥胖之作用。

Obesity is considered an important public health issue, and contributes to several chronic diseases. Meanwhile high-fat diet (HFD) is a inducer of gut dysbiosis which can result in the promotion of chronic inflammation and obesity. Previous publications have established that epigallocatechin gallate (EGCG), quercetin, and rutin have anti-obesity action. Based on the synergistic effect of the formula, the effect of EGCG-quercetin-rutin (EQR) formula on obesity, gut microbiota, and its molecular mechanisms in HFD-induced obese Wistar rats were investigated. After one week accommodation, sixty, and six-week-old male Wistar rats were divided into normal and obese groups, and then obese rats were prefed a HFD for four weeks. After the mean body weight of the obese rats (HFD groups) were over 10% heavier than that of the non-obese rats (normal diet, ND group). The obese rats were then divided into five groups (n=10/group) according to different doses of EQR formula for four weeks. Group 1: the HFD, Group 2: HFD + EQR formula-low dose (HFD + EQRF-LD), Group 3: HFD + EQR formula-medium dose (HFD + EQRF-MD), Group 4: HFD + EQR formula-high dose (HFD + EQRF-HD), and Group 5: HFD + positive control (HFD + PC). The results showed that, the weights of body, adipose tissues, and hepatic total lipid in HFD + EQR formula groups were significantly decreased as compared with the HFD group. In the gene expressions, EQR formula groups upregulated the gene expressions of PPAR-, ACO, CPT-1, and HSL, whereas it downregulated the gene expressions of PPAR- and FATP-1 in the liver of HFD-induced obese rats. In addition, EQR formula also upregulated the gene expressions of CPT-1, SIRT1, FAS, and FATP-1 in the perirenal adipose tissue of HFD-induced obese rats. Moreover, EQR formula could upregulate the gene expression of AMPK and PGC-1 in the brown adipose tissue of HFD-induced obese rats. Furthermore, EQR formula groups increased richness and diversity of the microbiota; decreased the relative abundance of the Deferribacteres, and increased the relative abundance of the Fusobacteria at the phylum level; increased the relative abundance of the Fusobacteriaceae and Christensenellaceae at the family level; decreased the relative abundance of the Mucispirillum, and increased the relative abundance of the Enterorhabdus, Christensenellaceae R-7 group, Lachnoclostridium, and Parvibacter at the genus level. In the contents of short chain fatty acids (SCFAs), EQR formula significantly increased the concentration of fecal acetic acid, propionic acid, butyric acid, and total SCFAs in HFD-induced obese Wistar rats. These results demonstrated that EQR formula is effective in preventing HFD-induced obesity and modulating gut microbiota.