Division of Molecular Biology, School of Life Sciences, Faculty of Medicine, Tottori University, Yonago, Tottori 683-8503, Japan.
Bone. 2012 Sep;51(3):369-75. doi: 10.1016/j.bone.2012.06.012. Epub 2012 Jun 23.
Osteocalcin is a major noncollagenous protein component of bone extracellular matrix, synthesized and secreted exclusively by osteoblastic cells during the late stage of maturation. We introduced a 10kb human osteocalcin enhancer/promoter (OC)-luciferase (Luc) construct into a hairless mouse line. Examination of tissue RNAs from these transgenic mice showed a predominant restriction of Luc mRNA expression to bone-associated tissues. Immunohistochemical staining of calvaria tissue sections revealed the localization of Luc protein to osteoblasts. Utilizing in vivo bioluminescence imaging, supplementation of 1α,25-dihydroxyvitamin D(3) increased Luc activity throughout the skeleton, consistent with in vitro transient transfection studies in osteoblast-like cells. Moreover, we observed an abrupt decrease in bioluminescence activity as the mice reached puberty, and a further decrease gradually thereafter. Using a radius skeletal repair model, we observed enhanced bioluminescence at the fracture site in both young (14-22 weeks old) and aged (50-66 weeks old) mice. However, peak bioluminescence was delayed in aged mice compared with young mice, suggesting retarded osteocalcin expression with aging. Our in vivo imaging system may contribute to the therapy and prevention of various bone metabolic disorders through its effective monitoring of the bone formation process.
骨钙素是骨细胞外基质的主要非胶原蛋白成分，由成骨细胞在成熟后期特异性合成和分泌。我们将一个 10kb 人类骨钙素增强子/启动子（OC）-荧光素酶（Luc）构建体导入无毛鼠系。对这些转基因小鼠的组织 RNA 进行检查显示，Luc mRNA 的表达主要局限于与骨相关的组织。对颅骨组织切片进行免疫组织化学染色显示 Luc 蛋白定位于成骨细胞。利用体内生物发光成像，补充 1α,25-二羟维生素 D(3)可增加整个骨骼的 Luc 活性，与成骨样细胞的体外瞬时转染研究一致。此外，我们观察到随着小鼠进入青春期，生物发光活性急剧下降，此后逐渐下降。使用桡骨骨骼修复模型，我们观察到年轻（14-22 周龄）和老年（50-66 周龄）小鼠的骨折部位的生物发光增强。然而，与年轻小鼠相比，老年小鼠的峰值生物发光延迟，表明随着年龄的增长，骨钙素表达减慢。我们的体内成像系统可能通过有效监测骨形成过程，有助于治疗和预防各种骨代谢紊乱。
