Simvastatin預防大鼠卵巢切除引發之骨質流失及增加骨生成蛋白之表現

Simvastatin Prevents Estrogen-Deficient Osteopenia and Increases Osteogenic Proteins in Ovariectomized Rats

陳宜慧(Yi-Hui Chen)

指導教授 ： 何美泠 ； 張瑞根

高雄醫學大學/醫學院/醫學研究所/碩士(2005年)

摘要

摘要
婦女停經後所引起之骨質疏鬆症是一種常見且重要的骨疾病。停經後，由於雌性素的缺乏導致骨替換率(Bone Turnover)增加，進而影響骨重塑之平衡，最終導致骨質的流失及增加骨折的機率。骨生成細胞分化的過程中，Type I Collagen, Bone Morphogenetic Protein 2 (BMP2)以及Osteocalcin等為重要的促進骨生成之蛋白質。過去學者的研究指出，Statins類藥物可以促使卵巢切除後之大鼠骨生成作用增加。然而，Statins類藥物影響骨生成作用的機轉是尚未清楚的。因此，本研究目的是探討大鼠在雌性素缺乏或正常的狀態下，Simvastatin在骨組織原位上對Type I collagen, BMP2, Osteocalcin等骨生成蛋白之表現的影響，以了解其作用機制。
本研究使用57隻三個月大雌性Sprague-Dawley大鼠，給予偽手術(Sham Operation)或卵巢切除術(Ovariectomy)後，分別給予10mg, 20mg以及30mg/kg/day Simvastatin持續灌食六週。實驗前以及實驗後分別以Dual Energy X-Ray Absorbsiometry(DEXA)來測定骨密度(Bone Mineral Density)的改變。Simvastatin治療六週後犧牲大鼠，取L2脊椎骨、遠端股骨和近端脛骨，製作成5μm厚度之H/E染色切片，觀察組織型態的改變，並量化分析骨小樑面積百分比。最後由免疫染色觀察Type I Collagen, BMP2, Osteocalcin等骨生成蛋白在骨組織的免疫定位(Immunolocalization)之變化。結果顯示卵巢切除組於術後六週，不論骨密度、組織型態學以及骨小樑面積百分比相較於偽手術組顯著的減少。大鼠卵巢切除後，給予 Simvastatin 20mg/kg/day或30mg/kg/day為期六週，其骨密度、組織型態學以及骨小樑面積百分比均有顯著性的增加。更進一步的在骨組織中觀察到Simvastatin治療組之Type I Collagen, BMP2以及Osteocalcin等骨生成蛋白的表現皆明顯地比偽手術或卵巢切除組表現增加。
綜合我們的研究結果顯示卵巢切除術後六週，骨密度及骨小樑面積均有意義的減少。但若手術後立即給予Simvastatin藥物治療，可以減緩雌性素缺乏引起之骨量不足(Osteopenia)。而且，其骨組織的中骨生成蛋白(Type I Collagen, BMP2, Osteocalcin)之表現皆明顯地增加。因此，我們認為Simvastatin可能藉由增加骨生成蛋白的表現而減緩因雌性素缺乏所誘發的骨質疏鬆症。然而，大鼠偽手術後給予Simvastatin藥物治療，並無顯著地影響骨密度及骨小樑的面積，但在骨組織定位上，骨生成蛋白有顯著增加。顯示對於正常骨重塑之大鼠，Simvastatin藥物在骨生成作用的影響較不明顯。

關鍵字

骨量不足 ； 雌性素缺乏 ； Simvastatin ； 骨生成蛋白

並列摘要

Abstract
Postmenopausal osteoporosis is one of the most important chronic bone diseases. After menopause, the decrease of estrogen results in an increase in bone turnover and a negative bone remodeling balance. The bone loss of postmenopausal women increases the risk of bone fracture. Previous reports indicated that statins promoted bone formation in ovariectomized rats. Type I collagen, BMP2 and osteocalcin are important osteogenic genes to induce osteogenesis. In this study, we investigated the effect of simvastatin on the expressions of type I collagen, BMP2 and osteocalcin in bone tissue of normal and estrogen deficient rats.
In this study, 57 three-month-old female Sprague-Dawley rats were used. Rats were treated with simvastatin 0, 10, 20 and 30 mg/kg/day for 6 weeks after ovariectomy (OVX) or sham operation (as the control). Bone mineral density (BMD) was measured by using a dual energy X-ray absorbsiometry (DEXA) right before operation and 6 weeks after operation. Bone sections were prepared from L2 vertebral body, proximal femur and distal tibia. Histomorphometic study and immunohistochemistry were performed to localize the osteogenic proteins on bone sections.
Our results showed that BMD and bone volume were markedly decreased in OVX group in comparison with sham group. After treatment with 20mg/kg/day or 30mg/kg/day of simvastatin for six weeks, BMD and bone volume were significantly increased in simvastatin treated OVX group in comparison with non-drug treated OVX group. Moreover, a six week treatment of neither 20mg nor 30mg of simvastatin showed significantly changed in BMD of sham operated rats. Furthermore, the expressions of osteogenic genes (type I collagen, BMP2 and osteocalcin) in bone sections were more obvious in the simvastatin treated groups than those of the sham-operation or OVX control group.
Our results indicated that OVX significantly induced osteopenia. Simvastatin was found to prevent OVX induced osteopenia. Furthermore, the result of immunolocalization in bone tissues showed that the osteogenic proteins were markedly increased by simvastatin in both sham operation and OVX groups. From these results, we suggest that simvstatin may prevent OVX induced osteoporosis through increasing the expressions of osteogenic genes. However, simvastatin showed less effect on bone formation in normal rats.

並列關鍵字

Simvastatin ； Estrogen-Deficient ； Osteopenia ； Osteogenic Proteins

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