Live Fluorescent Staining Platform for Drug-Screening and Mechanism-Analysis in Zebrafish for Bone Mineralization.
Live Fluorescent Staining Platform for Drug-Screening and Mechanism-Analysis in Zebrafish for Bone Mineralization.
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DOI：
10.3390/molecules22122068
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发表时间：
2017-11-27
期刊：
影响因子：
--
通讯作者：
Hsiao CD
中科院分区：
文献类型：
--
作者：
Chen JR;Lai YH;Tsai JJ;Hsiao CD
目前，药物筛选依赖于基于细胞的实验或动物模型来确认生物效应。哺乳动物系统被认为过于耗时、昂贵和复杂，无法进行高通量药物筛选。基于细胞的体外模型和体内哺乳动物模型之间存在差距。斑马鱼是一个理想的模型，可以将临床前毒性筛选与药物开发管道联系起来。斑马鱼具有基因组高度保守、发育迅速、后代数量多、成本低、操作简单等优点，被认为是基于疾病的药物筛选的优秀动物模型。在这项研究中，斑马鱼胚胎与小分子化合物孵育，这些化合物可能会影响微板中的骨矿化。阿仑磷酸钠和多索吗啡分别作为阳性对照和阴性对照。成骨矿化程度用荧光钙黄绿素染色图像的ImageJ软件进行测量和量化。在24个被测试的化合物中，我们鉴定了2个化合物，五烷胺和BML-267，它们具有促进胚胎矿化的作用，而6个化合物，RWJ-60475，左旋咪唑，盐酸四咪唑，氰戊菊酯，NSC-663284和BML-267酯，对骨矿化有抑制作用。此外，采用实时定量聚合酶链式反应(RT-qPCR)从分子水平评价骨代谢相关的生物学途径。我们证实阿伦磷酸钠通过抑制破骨细胞相关基因提高了骨矿化水平。综上所述，我们的研究建立了一种简单的方法来筛选潜在的骨代谢药物并进行体内骨矿化的机制分析。
Currently, drug screening relies on cell-based experiments or on animal models to confirm biological effects. The mammalian system is considered too time-consuming, expensive and complex to perform high-throughput drug screening. There is a gap between in vitro cell-based models and the in vivo mammalian models. The zebrafish is an ideal model that could link preclinical toxicity screening with the drug development pipeline. Taking advantage of a highly conservative genomic, rapid development, large number of offspring, low cost and easy manipulation, zebrafish has been considered an excellent animal model for disease-based drug screening. In this study, zebrafish embryos were incubated with small molecular compounds that potentially affected bone mineralization in microplates. Two compounds of alendronate and dorsomorphin were used as positive and negative controls, respectively. The level of osteogenic mineralization was measured and quantified by using ImageJ software with fluorescent calcein-staining images. Among twenty-four tested compounds from the kinase inhibitor library, we identified two compounds, pentamidine and BML-267, which showed increased embryonic mineralization; while six compounds, RWJ-60475, levamisole HCL, tetramisole HCL, fenvalerate, NSC-663284, and BML-267ester, were inhibitory to bone mineralization. In addition, real time quantitative PCR (RT-qPCR) was performed to evaluate the biological pathways involved in bone metabolism at the molecular level. We confirmed that alendronate enhanced the level of bone mineralization by inhibiting osteoclast-related genes. In summary, our research established a simple method to screen potential bone metabolic drugs and to perform mechanism analysis for bone mineralization in vivo.
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影响因子：
2.5
作者：
Okamoto M;Yamanaka S;Yoshimoto W;Shigematsu T
通讯作者：
Shigematsu T
影响因子：
4.3
作者：
Drake, Matthew T.
通讯作者：
Drake, Matthew T.
DOI：
10.4081/ejh.2012.24
发表时间：
2012-05-25
期刊：
European journal of histochemistry : EJH
影响因子：
--
作者：
Bradaschia-Correa V;Barrence FA;Ferreira LB;Massa LF;Arana-Chavez VE
通讯作者：
Arana-Chavez VE
影响因子：
158.5
作者：
Finkelstein, JS;Hayes, A;Neer, RM
通讯作者：
Neer, RM
影响因子：
20.3
作者：
Riggs, BL;Khosla, S;Melton, LJ
通讯作者：
Melton, LJ