PTHrP不同片段在PTH和CaR双基因缺失中的代偿作用及机制研究

PTH and CaR play an important role in regulating calcium phosphorus balance and bone homeostasis. However, CaR-/-PTH-/- mice show almost normal skeleton development.Both previous studies and our preliminary experiment reveal that PTHrP may play a compensatory effect of bone formation in CaR-/-PTH-/- mice. But the mechanism of different fragments of PTHrP in the bone formation is still unclear. Therefore, it is hypothesized that PTHrP amino-terminus plays its role mediated via PTH receptor, while Nuclear localization sequence(NLS)and carboxy-terminal play its role by transfering into nuclei in enhancing osteogenesis.In order to testify the hypothesis, we will culture bone marrow mesenchymal stem cells and generate CaR-/-PTH-/-PTHrP+/- and CaR-/-PTH-/-PTHrP KI+/- mouse model using techniques of Imageology, histopathology, and cellular and molecular biology to assess whether PTHrP plays an important role, whereas PTHrP NLS and C-terminal play a key role in compensation for the deletion of PTH in CaR deficient mice, and determine whether PTHrP amino-terminus plays its role mediated via PTH receptor, whereas its NLS and carboxy-terminal plays its role by transfering into nuclei through intracrine manner in enhancing osteogenesis by stimulating the BM-MSC proliferation and differentiation into osteoblasts. Results from this study will provide the theoretical and experimental bases for clinical applications of different fragments of PTHrP in the acceleration of fracture healing and treatment of osteoporosis.

PTH和CaR在调节钙磷平衡和骨骼稳态中发挥重要作用。但是， CaR-/-PTH-/-小鼠却表现为几乎正常的骨骼发育。以往的研究和我们预实验结果均提示PTHrP可能在CaR-/-PTH-/-小鼠骨形成中发挥代偿作用。目前PTHrP不同片段在骨形成中的作用机制尚不清楚。我们提出假设：PTHrP 氨基端经PTH受体介导，而其核定位序列(NLS)和C-端则进入细胞核发挥促进骨形成作用。为验证这一假说，将通过培养骨髓间充质干细胞、建立CaR-/-PTH-/-PTHrP+/-和CaR-/-PTH-/-PTHrP KI+/-小鼠模型，从分子、细胞、组织和动物整体水平多方面验证PTHrP在CaR-/-PTH-/-小鼠的代偿作用以及NLS和C-端的关键代偿作用，明确PTHrP氨基端经PTH受体介导、而NLS和C-端经胞内分泌的作用机制。本研究将为PTHrP促进骨折愈合和防治骨质疏松的临床应用提供理论依据。

PTH 和CaR 在调节钙磷平衡和骨骼稳态中发挥重要作用。但是， CaR-/-PTH-/-小鼠却表现为几乎正常的骨骼发育。以往的研究和我们预实验结果均提示PTHrP 可能在.CaR-/-PTH-/-小鼠骨形成中发挥代偿作用。目前PTHrP 不同片段在骨形成中的作用机制尚不清楚。我们提出假设：PTHrP 氨基端经PTH 受体介导，而其核定位序列(NLS)和C-端则进入细胞核发挥促进骨形成作用。为验证这一假说，将通过培养骨髓间充质干细胞、建立CaR-/-PTH-/-PTHrP+/-和CaR-/-PTH-/-PTHrP KI+/-小鼠模型，从分子、细胞、组织和动物整体水平多方面验证PTHrP 在CaR-/-PTH-/-小鼠的代偿作用以及NLS 和C-端的关键代偿作用，明确PTHrP 氨基端经PTH 受体介导、而NLS 和C-端经胞内分泌的作用机制。结果发现：PTH具有促进骨形成调节骨转换的作用，其但依赖于钙敏感受体的存在，间歇外源性PTH注射可提升血清钙离子水平，并经CaSR介导通过调节细胞周期等机制刺激骨形成。 本研究进一步阐述了CaSR和PTH在骨形成中的相互作用及机制，从而为提高钙剂和促骨形成类药物的临床疗效提供理论和实验依据；为以CaSR基因和细胞周期信号为靶向的骨形成的药物研发提供新的视点。