| 研究課題/領域番号 |
17K17094
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| 研究機関 | 長崎大学 |
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研究代表者 |
姜 晴 長崎大学, 医歯薬学総合研究科(歯学系), 助教 (00790007)
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| 研究期間 (年度) |
2017-04-01 – 2019-03-31
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| キーワード | 骨代謝学 |
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| 研究実績の概要 |
The involvement of 14-3-3s in bone development is poorly understood, and only a few reports have been published. To investigate the functions of Ywhag and Ywhae in bone development in detail, I established two Ywhag-/- lines and two Ywhae-/- lines by injecting each gRNA. And we used these loss of function mouse models to clarify the physiological functions of Ywhag and Ywhae in bone development.
Wild-type and Ywhag-/-, Ywhae-/- samples were collected at E15.5 and E18.5. We stained whole skeletons with alcian blue and alizarin red to identify cartilage and calcified tissues, respectively. Ywhag-/- showed similar level compared with wild-type, while size of Ywhae-/- skeletons was smaller than wild-type. Due to embryonic lethality of Ywhae-/- mice, wild-type and Ywhag-/-, Ywhae+/- samples were obtained from the mice at 4 weeks and 8 weeks of age. Body weight and bone volume of Ywhag-/- mice and Ywhae+/- mice were lower than that of wild-type mice, especially at 4-6 weeks of age.
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| 現在までの達成度 (区分) |
現在までの達成度 (区分)
2: おおむね順調に進展している
理由
Because of embryonic lethality of Ywhae-/- mice, we can not analyze Ywhae-/- mice at adult stage, so we compared wild-type and Ywhae+/- mice by microCT at 8 weeks of age. We will aslo identify the reason of dwarfism in Ywhae+/- mice.
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| 今後の研究の推進方策 |
The general functions of 14-3-3s were reported, but the functions on bone development are still unknown. This is the first study of 14-3-3s in bone development using KO mouse models. This study will greatly contribute to the elucidation of the molecular mechanisms for bone development.
We will compare Runx2 343-bp enhancer activities in primary osteoblasts from wild-type, Ywhag-/-, Ywhae-/- primary osteoblasts by reporter assay to identify the target molecules of the 14-3-3s in the activation of the Runx2 enhancer by introducing the transcription factors and cofactors, which had activated the enhancer.
As Runx2 is an important transcription factor for bone formation and homeostasis and is potentially useful for osteoporosis therapy, elucidation of the molecular mechanisms, by which 14-3-3s enhance the osteoblast-specific enhancer, will hopefully provide novel insights into the therapy of osteoporosis.
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| 次年度使用額が生じた理由 |
理由 We already have data of these KO mice condition basically, we need to research details in bone development, to examine the functions of Ywhag and Ywhae in osteoblast, osteocyte and osteoclast.
使用計画We will use knockout mice as model to identify the function of Ywhag and Ywhae, we need ELISA kit to examine TRAP5b and Osteocalcin level in serum of these KO mice, we will isolate RNA and examine mRNA level of osteoblast and osteoclast marker genes.
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