| 摘要: | 骨質疏鬆症亦屬於一種骨骼老化的現象，主要的原因是骨骼中的鈣質常在不知不覺中流失，因此發現通常骨質已經流失相當程度。目前為止，臨床上治療骨質疏鬆症的方式包括鈣的補充、維生素D 投與促進鈣質的吸收、或利用雌性激素或bisphosphonates 類的投與而抑制蝕骨細胞的作用以降低骨質受到破壞的速度。許多的癌症會誘發大量的骨溶蝕作用而引起嚴重的骨質疏鬆。而這些也會造成病人產生高血鈣、疼痛和骨折。除此，由於產生骨溶蝕時會放出骨骼基質的生長因子，因此會反向的刺激腫瘤的生長，造成癌症惡化而縮短病人存活時間。目前研究證實癌細胞分泌细胞核因子κB受体活化因子配基 (receptor activator of nuclear factor kappa B ligand, RANKL)，促進蝕骨細胞的成熟和分化是癌症誘發骨溶蝕最主要的原因，因此若能有效抑制癌細胞分泌RANKL 或是抑制RANKL 對蝕骨細胞成熟的活性，必能減緩癌症造成的骨溶蝕作用，同時解除病人疼痛、高血鈣和易骨折的問題。本研究對於芝麻(Sesamun indicum)的活性成分芝麻酚(Sesamol) 刺激人類骨肉瘤細胞MG63分化形成成骨細胞的效果進行評估。證實Sesamol能刺激MG 63細胞骨鈣素(osteocalcin, OCN)與骨形成指標血鹼性磷酸酶(alkaline phosphatase，ALP) 基因表達，並增強早期分化指標ALP的活性。Sesamol抑制細胞中轉錄因子Runx2基因表達，使骨保護素(osteoprotegerin, OPG)基因轉錄與其蛋白質含量下調。Sesamol激活MG63成骨細胞的BMP/Smad1/5/8 pathway與p44/42 mitogen-activated protein kinase (MAPK) pathway，促進骨橋蛋白(osteopontin, OPN)與RANKL蛋白質的水平。刺激MG63成骨細胞後期分化作用的礦化作用(mineralization)進行。我們進一步評估Sesamol抑制老鼠巨噬細胞株RAW264.7被RANKL誘導，進行分化形成蝕骨細胞的效果。結果證實芝麻酚可以早期抑制蝕骨前趨細胞RAW264.7被後期的RANKL誘導發生蝕骨分化作用。明顯提前抑制細胞中抗酒石酸酸性磷酸酶 (tartrate-resistant acidic phosphatase, TRAP)、組織蛋白酶K (cathepsin K, CTSK)等基因轉錄物被RANKL誘導表達，能顯著抑制RANKL 誘導TRAP的活性。抑制RANKL 誘導磷酸化p38-MAPK 絲裂原活化蛋白激酶和細胞外信號調節激酶(ERK)，減少RANKL 誘導表現破骨細胞最重要的轉錄因子NFATc1。研究結果也顯示Sesamol對於RANKL誘導NF-κB2 (p100/p52)活化而轉位進入細胞核的作用並無明顯抑制的效果。推論芝麻酚對成骨細的影響可透過BMP/SMAD-indepedent 路徑，活化 p38 與p44/42 MAPK pathway，促進ALP、OCN與OPN作用，刺激MG63成骨細胞分化進行礦化作用。而Sesamol抑制蝕骨的效果主要藉由抑制P38與ERK1/2等MAPK蛋白激酶路徑的活化，降低表現NFATc1並抑制破骨細胞分化。因此具有應用於治療骨吸收過度的相關疾病的潛能。本研究將分子細胞生物學理論與傳統中草藥作用相互配合，從傳統中草藥開發出有效的抗骨質疏鬆症藥物並深入地探索其抗骨質疏鬆症之活性機轉。
Osteoporosis is a disease characterized by low bone mass caused by an imbalance between bone resorption and bone formation. Bone homeostasis requires balanced interplay between osteoblasts and osteoclasts. In this study, we investigated the effects of sesamol, isolated from sesame (Sesamun indicum) in bone cells. First to investigate the osteoblast differentiation of Sesamol in a human osteoblast-like cell lines, MG 63. Sesamol increased the marker of osteoblastic maturation (ALP activity). Low-dose Sesamol also increased the osteoblast differentiation marker OCN gene expression. In addition, Sesamol increased osteoblastgenesis from osteoblast-like cells via BMP/SMAD1/5/8 pathway phosophorylation, phospho-P38 and Phospho-p44/42 MAPK pathway activation. In contrast, Sesamol decreased the transcript level of transcription factor Runx2 and osteoprotegerin (OPG) in dose-dependent manner. Sesamol increased the osteopontin (OPN), RANKL protein level and enhanced mineralization. On other hand, we found that sesamol markedly inhibited the receptor activator of RANKL-induced osteoclastic differentiation from RAW264.7 macrophage cells. However, RANKL-induced p38 and ERK but not JNK phosphorylation was attenuated by sesamol. The inhibitory effect of Sesamol resulted in clearly decrease of osteoclast-specific genes like matrix metalloproteinase 9 (MMP9), tartrate-resistant acid phosphatase (TRAP), and cathepsin K. Finally, Sesamol slight decreased the transcriptional activity of NFATc1, the master regulator of osteoclast differentiation. Our data suggest that the sesamol increase osteoblastgenesis through BMP/SMAD1/5/8 and MAPK pathway from human osteosarcoma cell lines MG-63. In addition, we demonstrate that sesamol inhibit osteoclastogenesis from macrophage cells via attenuation of RANKL-induced p38 and ERK activation. The data indicate that Sesamol has potential for use in treatment of diseases involving abnormal bone lysis such as osteoporosis. |
