探討黑豆與薏仁共同發酵搭配藻類對 C57BL/6J 小鼠延緩老化之功效

Abstract

根據過去研究，老化是組織與器官隨著時間而喪失功能的過程，其主要歸因於細胞的 DNA 因自由基破壞、發炎壓力、蛋白質異常摺疊、端粒限制等而受損造成細胞週期停滯，然而這些停止複製的細胞並不會自行凋亡或是被免疫細胞有效率的清除，造成其堆積在組織器官中，持續表現不正常的分泌表徵，進而影響到其他正常細胞，最後造成器官的功能喪失而導致老化相關疾病，如心血管疾病、第二型糖尿病、肌少症及失智症等。根據內政部統計處，我國 65 歲以上老年人口已達到15.28％，正式邁入世界衛生組織定義之「高齡社會」，顯示衰老相關議題之重要性，延緩老化之功能性食品的開發也漸漸被重視。黑豆為大豆的一種，除了含有豐富的優質蛋白質外，更已經被許多研究發現其具有抗發炎、抗氧化、抑制癌症等功效；薏仁屬於全榖雜糧類，自古就在傳統中草藥中被廣泛使用，目前研究指出其具有清除自由基、調節脂質代謝及調節腸道菌相等效果；藻類具有豐富膳食纖維與蛋白質含量，同時亦能提供大量維生素與礦物質，近來也發現其具有抗氧化與預防心血管疾病等功能。本實驗使用高營養價值的黑豆與薏仁進行發酵以增加其適口性、生物利用率與營養豐富度，並加入藻類粉末增添其色彩與風味的同時更提高營養密度；目的為探討黑豆與薏仁共同發酵並搭配藻類 (BAA) 能否改善小鼠老化情形。動物實驗以自然老化 72 週齡與 28 週齡之 C57BL/6 雄性與雌性小鼠，將 BAA 冷凍乾燥後分別以 2% 與 6% 的比例混入飼料中餵食共 14 週。結果顯示，BAA 能顯著降低小鼠之老化指標 GLB1、p16INK4A (p < 0.05)，並且也能降低肝臟中發炎物質 MCP-1、IL-6 與發炎相關路徑蛋白 p-PI3K/PI3K、p-Akt/Akt、p-NF-κB/NF-κB 之比值 (p < 0.05)，肝臟中氧化壓力指標 MDA 與 8-oxodG 之含量也有顯著改善 (p < 0.05)；此外，BAA 之介入在雄性小鼠中也觀察到性腺脂肪減少 (p < 0.05)、肌肉含量增加 (p < 0.01) 與 T-CHO/HDL 比率降低 (p < 0.001) 等明顯改善情形，而雌性小鼠中則發現肝臟重量減輕 (p < 0.05) 與肌肉含量增加 (p < 0.05) 的顯著變化。而在腸道菌相分析中則發現 Peptococcaceae 科與Akkermansiaceae 科 Akkermansia 屬等有益菌種之比例上升， Candidatus Saccharimonas 屬、Ruminococcaceae 科、Tannerellaceae 科中 Parabacteroides 屬與 Enterobacteriaceae 科等菌種之比例降低，使高齡小鼠腸道菌相組成更接近於年輕小鼠之狀態，而維持腸道健康與抑制腸道發炎情形。綜上所述，黑豆與薏仁共同發酵並搭配藻類 (BAA) 能透過降低發炎反應和氧化壓力進而達到延緩老化之功效，同時也能調節腸道菌相至較年輕之組成，並使健康相關數值明顯改善，而有改善老化相關指標之作用。

According to research, aging is the progressive loss of tissue and organ function over time. The main causes are reactive oxygen species, inflammatory stress, abnormal protein folding, and telomere shortening, which may lead to cell cycle arrest and cell senescence. The accumulation of senescent cells leads to abnormal secretion phenotype, exacerbating age associated diseases like cardiovascular disease, diabetes, sarcopenia, dementia, etc. According to ministry of the interior, the population above 65 years old in Taiwan has raised to 15.28% last year, which is definded as “aged society” by WHO (World Health Organization). As the elderly population increases, researchers have paid great attention to food that are effective to postpone aging or improve health span recently. Black bean is a kind of soy bean, and was reported to exert cancer prevention, anti-inflammatory and anti-oxidative effects. Adlay, a Chinese traditional medicine, has been studied having functions to regulate lipid metabolism, adjust gut microbiota, and decrease free radicals. Algae is rich in dietary fiber, high value protein, minerals and vitamins, and also has anti-oxidation effects and can prevent cardiovascular diseases. The main goal of this study is to develop a functional food that can postpone and attenuate age associated health issues to improve healthiness in the elder during aging periods. Therefore, our sample (BAA) is composed of black bean and adlay, processed by fermentation to increase its digestibility. In addition to the above, adding algae powder to enhance its taste and smell along with intensive nutrient. We supplemented BAA into normal diet at the percentage of 2 % and 6 % respectively, while 72 and 28 weeks old C57BL/6J mice were fed for fourteen weeks. Results show that BAA can significantly decrease the senescent markers GLB1 and p16INK4A, and also attenuate pro-inflammatory cytokines, MCP-1 and IL6, as well as PI3K/Akt pathway protein p-PI3K/PI3K, p-Akt/Akt, and p-NF-κB/NF-κB ratio. Furthermore, the oxidative stress markers, MDA and 8-oxodG significantly decrease as well. Additionally, we revealed improvement in muscle mass, perigonadal fat, and T-CHO/HDL ratio in male mice, as well as liver weight and muscle mass in female mice during the experiment. In addition to the beneficial effects above, the gut microbiota composition changed as well. Including rising abundances in Peptococcaceae, Akkermansiaceae families, and Akkermansia genus; and decreasing abundances in Ruminococcaceae, Tannerellaceae, Enterobacteriaceae families, and Candidatus Saccharimonas, Parabacteroides genus. After the intervention of BAA, the gut microbiota mentioned above altered, which was much closer to the composition of young mice rather than elder mice. In conclusion, BAA can delay aging via anti-inflammatory and anti-oxidative effects, and also improve aging related markers and gut microbiota composition as well.