雌激素受体在机械刺激促进骨质疏松性骨折愈合过程中的作用-卵巢切除术诱导的骨质疏松性大鼠闭合骨折模型

Mechanical sensitivity of bone is the key factor of the enhancement of fracture healing through mechanical stimulation. We found that osteoporotic bone is highly responsive to mechanical signals as good as or even better than normal bone, which is different from the general belief that osteoporotic bone has reduced sensitivity to mechanical signals. My previous research work successfully demonstrated that LMHFV could accelerate osteoporotic fracture healing by enhancing callus formation, angiogenesis and mesenchymal stem cell recruitment. It was reported that ERs had multiple functions including osteogenesis and mechanical signal transduction, and this function was independent of estrogen. We have discovered that ER expression of osteoporotic fracture is delayed compared to normal fractures during the healing process, it is suggested that the ER content at the fracture site may be the key to the previously observed difference in mechanical sensitivity. The effect of mechanical stimulation on ER expression and their functions in osteoporotic fracture are unclear. By using the ovariectomy (OVX) -induced osteoporotic rat close-femoral-fracture model and in vitro research, we will study the expression of ER and other fracture healing related genes during the healing process. We also aim to investigate whether ERs are responsible for the acceleration and enhancement observed in OVX-induced osteoporotic fracture treated with LMHFV. All of that will help us to understand the mechanism of the impairment of the healing process and explore a new intervention on osteoporotic fracture healing.

骨组织机械刺激敏感性是物理治疗促进骨折愈合的关键。我们发现骨质疏松骨的机械刺激敏感性高于正常骨，与目前骨质疏松骨对机械刺激信号低敏感性的观点正好相反。我们已证实，低幅高频震动（LMHFV）可促进骨质疏松性骨折骨折端骨痂形成、血管再生及间充质干细胞（MSC）的补充，从而促进骨折愈合。雌激素受体（ER）与骨折端骨痂内成骨及机械信号传导有关，且功能不受雌激素影响。我们发现，骨折愈合过程中骨质疏松骨中ER的表达迟于正常骨，表明ER是骨折端机械信号敏感性差异的关键所在，但机械刺激对ER表达的影响及二者在骨质疏松性骨折中的相互作用尚未明确。我们拟通过卵巢切除术（OVX）诱导的骨质疏松性大鼠闭合骨折模型及体外实验，研究骨折愈合过程中ER及部分骨折愈合相关基因的表达，探索ER在LMHFV促进骨质疏松性骨折愈合过程中的功能,完善对骨质疏松性骨折愈合生理过程研究的同时，也有助于探索骨质疏松性骨折新治疗手段。

影响骨折愈合的因素包括机械力学和生物学因素，雌激素水平的降低。卵巢切除术（OVX）诱导的大鼠骨质疏松性骨折模型中，高频低幅震动（LMHFV）可有效促进骨折愈合。我们假设LMHFV对骨质疏松性骨折愈合的促进作用与ER密切相关。本研究的目的在于探索ER在LMHFV促进骨质疏松性骨折愈合过程中的具体作用。.第一部分，用正常和OVX诱导的骨质疏松性大鼠建立骨折模型，以此研究正常和骨质疏松性大鼠体内ER表达的差异。骨折术后定量检测ER基因水平的表达，并同骨骼生长相关基因的表达进行相关性研究。ER蛋白水平的表达则通过免疫组化进行量化检测。第二部分，研究ER在LMHFV促进骨质疏松性骨折愈合过程中的信号ER功能经ER调控因子ICI182,780抑制后，并与无ER功能抑制组进行对比。第三部分，在雌激素缺乏和雌激素水平维持的条件下，研究LMHFV对原始成骨性细模型中ER-α表达的影响。成骨性细胞有/无LMHFV治疗，以及雌激素和/或ICI 182,780处理后，对基因表达，细胞增殖以及成骨潜能进行综合评估。.第一部分，骨质疏松性骨折动物中，ER在骨折早期表达偏低骨折术后2周时ER表达量较低但8周时表达增加;正常动物骨折术后2周ER表达较高但8周时表达偏低。免疫组化结果与此相似，提示骨折愈合过程中骨折部位有ER出现，ER的出现与骨折愈合密切相关。骨质疏松性骨折组ER在基因及蛋白水平的表达少于且迟于正常骨折组。第二部分，经LMHFV治疗之后，骨质疏松性骨折组的ER表达水平可恢复到正常水平，同时X线形态测量和微型电脑分析也表明其骨痂形成同正常骨之间无差异。但ICI 182,780可抑制LMHFV促进骨折愈合的功能，从而进一步影响ER基因水平和蛋白水平的表达以及骨痂形成能力。第三部分，雌激素缺乏情况下，LMHFV可明显提高成骨性细胞增殖及其成骨功能。当成骨性细胞经ICI 182,780处理后，LMHFV对其所有促进作用均消失。.与正常骨骨折相比，骨质疏松性骨折愈合过程中雌激素受体延迟表达，但经LMHFV治疗后可恢复。骨痂形成功能提高过程中需要ER参与。骨质疏松性骨折在LMHFV治疗后于骨痂部位检测到ER表达恢复。