探討發酵青花菜預防高脂高果糖誘導代謝紊亂進而減緩小鼠肥胖型肌少症的影響

Abstract

許多研究表明代謝症候群與肌少症密切相關，可能成因包括發炎、脂肪堆積和胰島素阻抗等代謝失調因子。肥胖型肌少症的定義是肌肉質量和功能下降，伴隨著脂肪增加。近年來，發酵食品越來越受歡迎，植物乳桿菌 (Lactiplantibacillus plantarum) 是發酵食品中常用的菌株，也是能夠預防肥胖的益生菌。青花菜 (Brassica oleracea var. italica) 是一種十字花科蔬菜，被證實具有抗肥胖和抗發炎等作用，這些作用可能來自其富含的活性化合物，包括硫代葡萄糖苷、異硫氰酸鹽、酚類化合物、礦物質、維生素和膳食纖維等成分。先前的研究指出，以植物乳桿菌發酵青花菜後能增加其活性化合物濃度，但在動物體是否具有生理功能的研究還不是很完整。因此，本研究的目的在探討以植物乳桿菌發酵青花菜是否能透過減緩胰島素阻抗、發炎和調節腸道微生物群，進而改善肥胖型肌少症，並探討其潛在分子機制，以開發促進國人健康的保健食品。本研究使用 45% 高脂飲食結合 20% 果糖飲用水誘導小鼠代謝紊亂，以評估發酵青花菜對肥胖型肌少症的影響。動物實驗共分為五組，正常飲食組 (Normal diet, ND)、高脂高果糖組 (High fat and high fructose diet, HFFD)、5% (w/w) 未發酵高劑量青花菜組 (Unfermented high dose broccoli, UHB)、1% (w/w) 發酵低劑量青花菜組 (Fermented low dose broccoli, FLB) 和5% (w/w) 發酵高劑量青花菜組 (Fermented high dose broccoli, FHB)，實驗為期 16 週犧牲。實驗結果顯示，未發酵和發酵青花菜皆能顯著降低體重、減緩脂肪細胞肥大、減少肝中三酸甘油酯累積及脂肪和肌肉組織中促發炎細胞因子的分泌，說明未發酵和發酵青花菜皆能改善高脂高果糖誘導的肥胖及發炎反應。其中，FHB 還能顯著降低體脂肪率和血中總膽固醇含量。在血糖和胰島素阻抗指標顯示，發酵青花菜能夠顯著降低空腹血糖、禁食與未禁食血清胰島素含量及血清瘦素含量，表示發酵青花菜能有效改善高脂高果糖誘導的高血糖及胰島素阻抗。此外，未發酵和發酵青花菜能增強肌肉強度，並降低肌肉組織中三酸甘油酯含量與油滴累積。FHB 還能增加瘦體組織質量，並透過活化 IRS1/PI3K/Akt 路徑，增加 GLUT4 的蛋白質表現量，促進肌肉葡萄糖的代謝。FHB還能活化 mTOR 下游路徑，促進蛋白質的合成，進一步透過抑制 FOXO3a 轉錄因子磷酸化，減少與肌肉萎縮相關分子蛋白的產生，恢復高脂高果糖誘導的骨骼肌質量損失。在腸道菌相分析結果顯示，FHB 的腸道菌相組成趨近於 ND 組，且 FHB 能增加 Roseburia 和 Acetivibrio ethanolgignens的豐富度，這兩株菌被研究在肌少症患者的腸道菌相中豐富度降低。還觀察到 FHB 的優勢菌種 Monoglobus pectinilyticus，能夠增加腸道微生物群的豐富度，以及增加 Anaerostipes hominis 等產丁酸菌的豐富度，並顯著增加盲腸中丁酸菌的含量。綜上所述，未發酵和發酵青花菜可以減緩小鼠高脂高果糖飲食誘導代謝失調與發炎反應之情形，而發酵青花菜能透過改善胰島素阻抗與調節腸道菌相，增加骨骼肌葡萄糖代謝和肌肉蛋白質合成與抑制肌肉蛋白質降解，恢復骨骼肌質量損傷，以減緩肥胖型肌少症的症狀，期望此研究能作用於未來開發發酵青花菜功能性食品的基礎，以促進國人的健康。

Numerous studies have indicated a strong association between metabolic syndrome and sarcopenia, with factors such as inflammation, fat accumulation, and insulin resistance. Sarcopenic obesity is characterized by a decline in muscle mass and function, accompanied by increased adipose tissue. In recent years, fermented foods have gained popularity, and Lactiplantibacillus plantarum is a commonly used strain in these foods, known for its potential to prevent obesity. Broccoli (Brassica oleracea var. italica) is a cruciferous vegetable that has been shown to have anti-obesity and anti-inflammatory effects. These effects may be attributed to its rich content of active compounds, including glucosinolates, isothiocyanates, phenolic compounds, minerals, vitamins, and dietary fiber. In a previous study, fermented broccoli by Lactiplantibacillus plantarum had shown an increase in its bioactive compounds. However, the physiological impact of fermented broccoli on animals has not been investigated. Therefore, this study aims to investigate whether fermented broccoli can mitigate the occurrence of metabolic disorders induced by a high-fat and fructose diet (HFFD), thereby improving sarcopenic obesity, and to explore its potential molecular mechanisms for developing functional foods promoting health in humans. We examined the effects of a 45% high-fat diet combined with 20% fructose drinking water (HFFD)-induced metabolic disorders in mice and evaluated the impact of fermented broccoli on sarcopenic obesity. Animals were divided into five groups: normal diet (ND), high-fat high-fructose diet (HFFD), 5% (w/w) unfermented high-dose broccoli (UHB), 1% (w/w) fermented low-dose broccoli (FLB), and 5% (w/w) fermented high-dose broccoli (FHB), for a 16-week period. The current results demonstrated that both unfermented and fermented broccoli significantly reduced body weight, alleviated adipocyte hypertrophy, decreased hepatic triglyceride accumulation, and reduced pro-inflammatory cytokine secretion in adipocyte and muscle tissues, indicating that both broccoli group could improve obesity and inflammatory responses induced by HFFD. Among these, FHB notably reduced fat mass and total cholesterol levels. Regarding blood glucose and insulin resistance indicators, fermented broccoli significantly decreased fasting blood glucose, fasting and feeding serum insulin levels, and serum leptin levels, indicating that fermented broccoli can improve hyperglycemia and insulin resistance induced by HFFD. Additionally, both broccoli group enhanced muscle strength and reduced triglyceride content and oil droplet accumulation in muscle tissue. FHB was found to have the potential to improve glucose metabolism via the IRS1/PI3K/Akt/GLUT4 pathways, and activated downstream mTOR pathways for protein synthesis, while inhibiting FOXO3a-related pathways involved in muscle protein degradation, thereby restoring skeletal muscle mass loss induced by HFFD. Regarding gut microbiota, the gut microbiota composition of FHB approached ND group. FHB increased the abundance of Roseburia and Acetivibrio ethanolgignens, which are reduced in the gut microbiota of sarcopenia patients. Additionally, FHB increased the abundance of Monoglobus pectinilyticus, a predominant strain specific to FHB , and increased the abundance of butyrate-producing bacteria such as Anaerostipes hominis, significantly increasing butyrate levels in the cecum. In summary, both unfermented and fermented broccoli can mitigate metabolic disorders and inflammatory responses induced by a high-fat, high-fructose diet in mice. Fermented broccoli improves insulin resistance and regulates gut microbiota, enhances skeletal muscle glucose metabolism and protein synthesis, and inhibits muscle protein degradation, thereby alleviating symptoms of sarcopenic obesity. This suggests fermented broccoli could serve as a basis for functional foods aimed at promoting health among the Taiwanese population.