本文選題：股骨頭骨壞死 + 動脈灌注�。� 參考：《中國人民解放軍醫(yī)學院》2017年博士論文

【摘要】：一、目的:擬在研究人股骨頭骨壞死標本骨結構的基礎上,探索壞死股骨頭內(nèi)miRNA-214表達與局部成、破骨細胞活性之間的關系及潛在機制,制備腺相關病毒搭載miRNA-214抑制劑,觀察藥物調(diào)節(jié)成骨、破骨活性防治大鼠股骨頭骨壞死塌陷的效果。二、方法:①2015年3月至2016年5月期間,收集我院16例股骨頭骨壞死標本。標本進行Micro-CT掃描后,根據(jù)骨質密度的不同,將每個標本分為不同區(qū)域,分別采用病理學檢測、實時熒光定量PCR、WesternBlot等方法對骨微觀結構及成骨、破骨細胞活性進行分析。PCR檢測股骨頭骨壞死標本骨組織,觀察不同區(qū)域成骨、破骨相關基因表達情況及miRNA-214表達情況。②制備以腺相關病毒為載體的miRNA-214抑制劑藥物,評價其體外調(diào)節(jié)成骨、破骨的能力。③建立大鼠股骨頭骨壞死模型,觀察AAV-anti-miRNA-214藥物的治療效果。三、結果:①標本壞死區(qū)域miRNA-214表達顯著增高,破骨活性顯著增高,成骨活性顯著降低。而硬化區(qū)成骨活性顯著增高,破骨活性顯著降低。同時,miRNA-214在股骨頭骨壞死標本壞死區(qū)域顯著增高;②成功制備AAV-anti-miRNA-214,發(fā)現(xiàn)其能夠明顯抑制破骨細胞的功能,促進成骨細胞的功能,同時可以顯著下調(diào)miRNA-214的表達,TRAP染色結果顯示AAV-anti-miRNA-214可以顯著抑制骨髓間充質干細胞向破骨細胞分化的能力。MC3T3-E1 細胞加入AAV病毒包裹的anti-miRNA-214 , 發(fā)現(xiàn)AAV-anti-miRNA-214能夠明顯促進成骨細胞的功能,ALP染色的結果顯示成骨細胞的功能增強;③AAV-anti-miRNA-214可以促進成骨活性,抑制破骨活性,防治大鼠股骨頭骨壞死塌陷。四、結論:AAV- anti-miR-214可以促進成骨活性,抑制破骨活性,其雙向調(diào)節(jié)作用可有效防治大鼠早期股骨頭骨壞死塌陷。
[Abstract]:Objective: to explore the relationship between the expression of miRNA-214 and local osteoclast and osteoclast activity in the necrotic femoral head on the basis of studying the bone structure of the osteonecrosis specimen of human femoral head, and to prepare the adeno-associated virus carrying miRNA-214 inhibitor. To observe the effect of drug regulating osteogenesis and osteoclast activity preventing and treating osteonecrosis of femoral head in rats. Methods: from March 2015 to May 2016, 16 cases of osteonecrosis of femoral head were collected. After Micro-CT scanning, each specimen was divided into different regions according to the different bone density. The microstructure and osteogenesis of bone were detected by pathological examination and real-time fluorescence quantitative PCR Western blot. Osteoclast activity was analyzed. PCR was used to detect bone tissue of osteonecrosis specimens of femoral head. Osteogenesis, osteoclast-associated gene expression and miRNA-214 expression were observed in different regions. 2 miRNA-214 inhibitor drugs were prepared using adeno-associated virus as vector. To evaluate the ability of osteoregulation and osteoclast in vitro to establish osteonecrosis model of femoral head in rats and to observe the therapeutic effect of AAV-anti-miRNA-214 drugs. Results: the expression of miRNA-214, osteoclast activity and osteogenesis activity in the necrotic region were significantly increased and osteogenic activity was significantly decreased. However, osteogenic activity in sclerotic area was significantly increased and osteoclast activity was significantly decreased. At the same time, AAV-anti-miRNA-214 was successfully prepared by miRNA-214 in the necrotic area of osteonecrosis of femoral head. It was found that AAV-anti-miRNA-214 could significantly inhibit the function of osteoclasts and promote the function of osteoblasts. At the same time, the expression of miRNA-214 was significantly down-regulated by trap staining. The results showed that AAV-anti-miRNA-214 could significantly inhibit the ability of bone marrow mesenchymal stem cells to differentiate into osteoclasts. MC3T3-E1 cells were added to anti-miRNA-214 wrapped with AAV virus. It was found that AAV-anti-miRNA-214 could significantly promote osteoblasts. The results of ALP staining showed that the function of osteoblasts was enhanced. 3AAV-anti-miRNA-214 can promote osteogenic activity, inhibit osteoclast activity and prevent osteonecrosis of femoral head in rats. Conclusion: AAV- anti-miR-214 can promote osteogenic activity and inhibit osteoclast activity, and its bidirectional regulation can effectively prevent and cure early osteonecrosis of femoral head in rats.
【學位授予單位】：中國人民解放軍醫(yī)學院
【學位級別】：博士
【學位授予年份】：2017
【分類號】：R681.8

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