370G 對於骨骼代謝之影響

Abstract

人類的骨骼終其一生都在不斷進行汰舊換新。骨骼的代謝主要是負責分解老舊骨骼的蝕骨細胞，和負責合成新骨質的造骨細胞之間的協同來維持。當二者之間失去平衡時就會引發各種疾病，例如蝕骨細胞功能低落的佝僂病 (osteopetrosis), 以及因蝕骨細胞相較之下過於旺盛所造成的骨質疏鬆症 (osteoporosis)。實驗中所用到的化合物370G，其結構是修飾自咖啡酸苯乙酯 (Caffeic acid phenethyl ester，CAPE)。CAPE是蜂膠中的主要成分，其具有很多種生理活性，包括抗氧化、抗發炎、抑制腫瘤、抑菌、抗病毒、抗黴菌、心血管保護與神經保護等等，而對於骨頭方面可以減少因氧化壓力所造成的造骨細胞的分化抑制。本論文則探討370G對於骨骼代謝的影響。in vivo方面，本篇實驗使用3週大的公鼠，利用兩種動物模式來評估370G對其骨骼代謝的影響。其一是在其膝部埋22號針，再經此每天施以vehicle (DMSO/saline)或370G (100 nM, 500 nM , 5 uM/10ul)，連續給予一週，再過一週後犧牲之取出脛骨作切片染色分析。結果發現其海綿骨量以及TRAP-positive的組織區域皆有增加的傾向。另一個動物模式為將其坐骨神經切除後使其骨質流失，再每天以腹腔注射給予370G (1 mg/ kg)，兩週後再犧牲之取出脛骨，用DXA檢測其骨密度。結果發現給予370G後，有拮抗因坐骨神經被切除所導致的骨質流失。而in vitro方面，分別作造骨細胞和蝕骨細胞的培養，發現370G均可促進其分化及功能。而在RT-PCR的分析中，發現370G可以增加造骨細胞Osteocalcin和BDNF的mRNA的表現量。總而言之，實驗中發現370G in vivo 方面可以促進骨質的新生， in vitro方面對於造骨細胞和蝕骨細胞均有促進的效果，因此370G可以增加骨骼之代謝，並且產生骨質新生的淨效果。

The skeleton is a metabolically active organ that undergoes continuous remodeling throughout life. Bone remodeling is controlled by two equal, but opposing forces: bone formation by osteoblasts and bone destruction or resorption by osteoclasts. When they lose the equilibrium, it will lead to bone diseases such as osteopetrosis, that the rate of bone formation is higher than the rate of bone resorption; and osteoporosis, that is the imbalance of bone remodeling favoring osteoclastic bone resorption. The structure of the compound 370G uesd in this experiment is derived from the modification of caffeic acid phenethyl ester (CAPE). CAPE is the major component in propolis. CAPE has exhibited a variety of properties, including antioxidant, antiinflammatory, antitumor, antibacterial, antimicrobial, cardioprotective, and neuroprotective effect. CAPE can also reduce the oxidative stress-induced inhibition of osteoblastic differentiation. Here we investigate the effect of 370G on the metabolism of bone in vivo and in vitro. 3 week-old S.D. male rats were used. Implantation of cannula (22G) was done from the posteriolaterial side into the proximal tibial metaphysis in limbs of the rats. Vehicle (DMSO/saline) or 370G (100 nM, 500 nM, 5 uM/10ul) were locally administered into tibia through the cannula for 7 consecutive days, the rats were then sacrificed on Day 14. The tibia bone slices were used for the analysis. In Comparison with the vehicle-treated side, treatment with the 370G markedly increased bone volume and the TRAP-positive cells. In addition, sciaticotomy was performed to cause the loss of bone formation. Intraperitoneal injection of 370G (1 mg/ kg) was performed on the following day of sciaticotomy, which persists for 2 week, and the rats were then sacrificed and the tibia bone was used for the analysis of BMD by DXA. Compared with the vehicle-treated-group, 370G can reverse the bone loss from the sciaticotomy. It was found that 370G can stimulate the differentiation of both osteoclasts and osteoblasts in cell cultures. 370G causes the up regulation of the expression of osteocalcin and BDNF mRNA in primary osteoblastic culture. In conclusion, we have found that 370G can stimulate the bone formation in vivo, and increase the differentiation and/or maturation of osteoclasts and osteoblasts leading to the net effect of bone formation.