李彗禎謝逸憲LEE, HUEI-JANEHSIEH, YI-HSIEN李怡靜LI, YI-CHINGYI-CHINGLI醫學研究所碩士班2026-10-292030-08-2120252025-06-24https://handle.ncl.edu.tw/11296/g8e2cwhttps://scholars.csmu.edu.tw/handle/123456789/36751骨骼重塑 (bone remodeling)由蝕骨細胞 (osteoclast, OC)進行再吸收作用 (resorption)及成骨細胞 (osteoblast, OB)形成新骨骼 (formation)反覆循環。OB與OC活性失衡則容易引起骨質疏鬆 (osteoporosis, OP)等疾病。高脂飲食 (high fat diet, HFD)及高血壓 (hypertension)與OP風險增加有關，機轉含括增加促發炎細胞激素 (proinflammatory cytokine)。益生菌 (probiotics)被認為有助於維持腸道菌相 (gut microbiota)，並具抗發炎、抗氧化等功能。本實驗目的為測定益生菌對餵食高脂飲食的高血壓大鼠減緩OP之作用。實驗模式為以試管實驗測試11種益生菌菌體及菌培養液的抗氧化能力，另外以五週齡的自發性高血壓大鼠 (spontaneous hypertention rat, SHR) (N = 6)餵食12週的HFD及益生菌，並以棕櫚酸 (palmitic acid, PA)處理類成骨細胞MG-63模擬高脂狀態，加入經Caco-2腸細胞反應之益生菌代謝物，測定對細胞的作用。結果顯示，益生菌的菌體及菌液都有抗氧化能力。SHR大鼠在實驗結束後，餵食HFD的SHR組並未顯著增加體重，但與SHR組相比，B1、L6、L7組的體重顯著下降 (p < 0.05)。SHR組的收縮壓及舒張壓皆顯著高於未誘導組，但僅有B1組有顯著降低舒張壓 (p < 0.05)。SHR組的血清總膽固醇顯著高於正常組，11種益生菌組都顯著降低總膽固醇。股骨的電腦斷層結果顯示，SHR組降低BMD (bone mineral density)、BV/TV (bone volume/total volume)、Tb.N (trabecular number)、Tb.Th (trabecular thickness)，且增加Tb.Sp (trabecular separation)。以BMD做為評估指標，與SHR組相比，B1、B2、B3、L1、L3、L4、L6、L7、LC組顯著改善骨質疏鬆。MG-63以PA及經腸細胞反應之益生菌代謝物 (probiotic metabolites)處理後收集並進行mRNA檢測，結果顯示，PA處理類成骨細胞顯著減少骨礦化基因type I collagen (COLI)表現。在蛋白檢測中則發現PA處理引起alkaline phosphatase (ALP)蛋白減少，L1-L6組顯著增加ALP。綜整研究成果，雖然MG-63細胞實驗結果無法完全對應動物實驗，但仍可由動物實驗結果得知益生菌可以改善高脂高血壓引起的骨質疏鬆，有鑑於高脂高血壓可能因增加氧化壓力而引起OP，因此，未來可進一步測定益生菌抗氧化特性與OP的關聯，並選擇更適當的細胞模型進行機轉探討。Bone remodeling is a dynamic and continuous process involving the resorption of bone by osteoclasts (OCs) and the formation of new bone by osteoblasts (OBs). Disruption of the balance between OB and OC activity can lead to pathological conditions such as osteoporosis (OP). Emerging evidence suggests that a high-fat diet (HFD) and hypertension are associated with an increased risk of OP, potentially mediated by enhanced proinflammatory cytokine production and oxidative stress. Probiotics have been proposed as a potential intervention due to their ability to modulate the gut microbiota and their documented anti-inflammatory and antioxidant properties. The objective of this study was to investigate the potential protective effects of probiotics against OP in hypertensive rats fed an HFD. This study utilized both in vitro and in vivo experimental approaches. In vitro, the antioxidant capacity of 11 probiotic strains and their culture supernatants was evaluated using free radical scavenging assays. In vivo, five-week-old spontaneously hypertensive rats (SHR) (N = 6) were administered an HFD and selected probiotics for a duration of 12 weeks. To simulate a high-fat condition at the cellular level, MG-63 pre-osteoblasts were treated with palmitic acid (PA). Subsequently, the effects of probiotic metabolites, processed through Caco-2 intestinal epithelial cells were assessed. Both probiotics and their culture supernatants are antioxidant. At the conclusion of the animal experiment, HFD-fed SHR group did not result in a significant increase in body weight. However, when compared to the SHR group, the body weights of rats in the B1, L6, and L7groups were significantly reduced (p < 0.05). Both systolic and diastolic blood pressure levels were significantly elevated in the SHR group relative to the non-induced control group, yet only B1 exhibited a significant antihypertensive effect (p < 0.05). In terms of serum total cholesterol, the SHR group showed a marked increase compared to the normal control, while all 11 probiotic strains significantly reduced cholesterol levels. Assessment of bone mineral density (BMD) via micro-computed tomography (micro-CT) of the femur revealed that probiotics B1, B2, B3, L1, L3, L4, L6, L7, and LC significantly attenuated HFD-induced bone loss. At the molecular level, qRT-PCR analysis showed that PA treatment significantly reduced the expression of the bone matrix gene type I collagen (COLI) in MG-63 cells. Western blot analysis further revealed that PA reduced alkaline phosphatase (ALP) protein levels. However, treatment with L1-L6 significantly restored ALP levels. In summary, although the in vitro findings using MG-63 cells did not entirely align with the in vivo results, the animal experiments demonstrated that probiotics could ameliorate HFD- and hypertension-induced osteoporosis. Given the role of oxidative stress in OP pathogenesis under these conditions, future research should further elucidate the relationship between the antioxidant capacities of probiotics and bone health, and explore the underlying mechanisms using more physiologically relevant cellular models.益生菌骨質疏鬆高血壓高脂飲食骨礦化成骨細胞probioticsosteoporosishypertensionhigh fat dietbone mineralizationosteoblastthesis