- 學位論文
以db/db糖尿病小鼠模式探討調節血糖與穩定血糖之研究：以Akkermansia muciniphila菌與高壓加工果汁為例
Effects of Akkermansia muciniphila and high pressure processed fruit juices on blood glucose regulation and stabling in db/db diabetic mice
摘要
糖尿病之定義為一種慢性的代謝異常疾病，主要原因是由於體內胰島素缺乏或功能不全，而對醣類的利用及調節能力下降。台灣地區糖尿病型態以第二型糖尿病 (type 2 diabetic mellitus, T2DM) 為主，約占罹患率的95%左右，若長期血糖控制不佳，容易引起相關併發症，所以，如何穩定血糖成為近年重視與努力的健康議題之一。因此，本研究利用機能性素材探討其在調節血糖之作用，期望將其開發成具穩定血糖之保健食品。內容主要分成兩部分：(一) 利用db/db糖尿病小鼠模式探討Akkermansia muciniphila (A. muciniphila) 調節血糖及腸道菌相之功能 (二) 以 db/db 糖尿病小鼠模式探討高壓加工果汁在穩定血糖之研究 (一) 本研究以自行分離之A. muciniphila活菌與死菌作為研究介入之細菌，探討其是否具有調節血糖之作用，並探討其對腸道菌相變化之影響。使用5週齡C57BL/KSJ正常小鼠與db/db糖尿病小鼠，將動物隨機分組 (n=7/組)，分別有：(1) 控制組 (wild-type, WT)、(2) db/db糖尿病小鼠 (diabetes mellitus, DM)、(3) db/db糖尿病小鼠給予A. muciniphila活菌 (Akkermansia muciniphila-live, DM+AKK-L)、(4) db/db糖尿病小鼠給予A. muciniphila死菌 (Akkermansia muciniphila-dead, DM+AKK-D)，以及 (5) db/db糖尿病小鼠給予AKK標準菌株ATCC BAA-835 (Akkermansia muciniphila-positive, DM+AKK-P)，每隻小鼠每天給予1x108 CFU/mouse/day菌數，實驗為期八週。由實驗結果顯示，給予A. muciniphila各組別之最終體重皆顯著增加，並且顯著增加棕色脂肪組織含量。給予A. muciniphila各組之空腹血糖，皆顯著低於DM組。給予DM+AKK-L組可顯著降低db/db糖尿病小鼠之血清中總膽固醇與低密度脂蛋白膽固醇。在短鏈脂肪酸部分，給予DM+AKK-L組可顯著增加糞便乙酸之含量；給予DM+AKK-D組，可顯著增加糞便丁酸之含量。綜合以上結果得知，給予A. muciniphila菌具有調節血糖之潛力，其中AKK-L之作用最佳。 (二) 本研究以db/db糖尿病小鼠模式探討紅甘蔗、白甘蔗和鳳梨高壓加工 (600 MPa/ 5 min) 果汁對於穩定血糖之作用。將雄性 db/db 糖尿病小鼠 (n=6/組)與C57BL/KSJ正常小鼠 (n=8/組) 分為8組，分別給予單純葡萄糖水不同種類之未高壓與高壓加工之果汁，包括：正常小鼠 (wild type, WT)、db/db糖尿病小鼠 (Diabetes Mellitus, DM) 以及分別給予未高壓與高壓加工之紅甘蔗汁 (red sugarcane juice, RSJ)、白甘蔗汁 (white sugarcane juice, WSJ)、鳳梨汁 (pineapple juice, PJ) 之組別，待糖尿病小鼠空腹血糖達 300 mg/dL後，進行一次性口服葡萄糖耐受試驗 (Oral glucose tolerance test, OGTT)。由實驗結果得知，相較於給予葡萄糖水之DM組別，給予高壓加工之白甘蔗汁、紅甘蔗汁與鳳梨汁之組別，可顯著降低OGTT曲線下面積，以及抑制果汁中α-葡萄糖苷酶和蔗糖轉化酶之活性。綜合上述結果可知，經高壓加工之果汁可抑制果汁酵素，延遲葡萄糖分解，從而延緩小腸對葡萄糖之吸收，具有穩定血糖之作用。
並列摘要
Diabetes is a group of metabolic diseases characterized by hyperglycemia resulting from defects in insulin secretion, insulin action, or both. In Taiwan, Type 2 diabetes (T2DM) is the most common form of DM, which accounts for 90% to 95% of all diabetic patients. In addition, people with T2DM are often accompanied by complications, such as cardiovascular diseases, diabetic neuropathy, nephropathy, and retinopathy. T2DM has become an observably global public health problem. Therefore, this study uses functional ingredient supplements to explore their role in regulating blood sugar. The study designs are as follow: (part I) Anti- hyperglycemia effects of Akkermansia muciniphila in db/db diabetic mice; (part II) Effects of high pressure processed fruit juice on blood glucose stabling in db/db diabetic mice. In part I, we investigated the hypoglycemia potential of Akkermansia muciniphila live and dead strains isolated from healthy in db/db diabetic mice. To explore their role in regulating blood sugar and impact on the gut microbiota. Using 5-week-old male db/db diabetic mice and C57BL/KSJ wild type mice, the animals were randomly divided into groups (n=7/group), including: (1) wild type (WT), (2) db/db diabetic mice (DM), (3) Akkermansia muciniphila-live (DM+AKK-L), (4) Akkermansia muciniphila-dead (DM+AKK-D), and (5) Akkermansia muciniphila-positive ATCC BAA-835 (DM+AKK-P) groups, A. muciniphila (1x108 CFU/mice/ day) supplementation by daily gavage for 8 weeks. The results showed that AKK-L, AKK-D and AKK-P strains significantly increased the final body weight and the weight of brown adipose tissue in db/db diabetic mice. The fasting blood glucose of each AKK strains group was significantly decreased than the DM group. Treatment with A. muciniphila was significantly reduces the serum total cholesterol concentrations and LDL-C in db/db diabetic mice. The effects of The result of fecal short-chain fatty acids showed that AKK-L significantly increased acetic acid in feces; AKK-D significantly increased butyric acid in feces of db/db diabetic mice. These results indicated that A. muciniphila strains has the potential to regulate the blood sugar of db/db diabetic mice. Moreover, AKK-L was exhibited optimal therapeutic efficacy against hyperglycemia. In part II, we used the db/db diabetic mice to explore the effect of unprocessed and high hydrostatic pressure (HHP) processed juice (600 MPa/ 5 min) of red sugarcane, white sugarcane, and pineapple on stabilizing blood sugar. Using male db/db diabetic mice (n=6/group) and C57BL/KSJ wild type mice (n=8/group) into 8 groups, then give them glcouse and different types of unprocessed and high hydrostatic pressure processed juice, including: wild type (WT), db/db diabetic mice (DM), and the unprocessed and high hydrostatic pressure processed red sugarcane juice (RSJ), white sugarcane juice (WSJ), and pineapple juice (PJ). After the fasting blood glucose of db/db mice reached to 300 mg/dL in db/db diabetic mice, will start to conduct oral glucose tolerance test (OGTT). The results showed that compared with the group DM group, the group of high hydrostatic pressure processed white sugarcane juice, red sugarcane juice, and pineapple juice were significantly decreased the OGTT total areas under the curve compared with the group DM group, and could inhibit the activities of α-glucosidase and sucrose invertase in juices. In conclusion, high hydrostatic pressure process inhibited the activities of sucrose invertase and α-glucosidase then maintaining sugar molecules as polysaccharides, which delayed the absorption of glucose by the small intestine, has the effect of stabilizing blood sugar.