【摘要】： 目的(1)培養(yǎng)轉染所需兔骨髓間充質干細胞(bone marrow mesenchymal stem cells, BMSCs)。(2)通過采用陽性脂質體pcDNA3.1(-)介導的轉染到BMSCs方法,獲得穩(wěn)定轉染的BMSCs細胞。(3)使用免疫組化、WesternBlot法檢測pcDNA3.1-hBMP2在兔BMSCs中蛋白的表達情況。 方法(1)貼壁法培養(yǎng)擴增所需BMSCs并從形態(tài)學、流式細胞儀測量表面標記物。(2)將已成功構建的pcDNA3.1-hBMP2重組質粒進行體外靶細胞轉染并使用G418篩選穩(wěn)定表達細胞。(3)通過免疫組化、WesternBlot法檢測其在BMSCs中的蛋白表達。 結果(1)經培養(yǎng),擴增兔骨髓間充質干細胞,P2、P3可作為hBMP2轉染的靶細胞。(2)用陽離子脂質體轉染pcDNA3.1-hBMP2的MSCs經G418篩選后,經免疫組化、WesternBlot法檢測,轉染pcDNA3.1-hBMP2后的兔骨髓間充質干細胞內有大量hBMP2 mRNA的轉錄和相關蛋白的表達,而未轉染的骨髓間充質干細胞未見陽性表達。 結論(1)兔骨髓間充質干細胞可作為hBMP2轉染的靶細胞。(2)本實驗成功構建hBMP2真核表達質粒并在兔骨髓間充質干細胞中得到相關表達。(3)本研究為進一步研究BMP2基因轉染MSCs的體外增殖并穩(wěn)定表達和在動物試驗中來加速牽張成骨新骨形成的實驗奠定了基礎。
[Abstract]:Objective (1) to culture rabbit bone marrow mesenchymal stem cells (bone marrow mesenchymal stem cells, BMSCs). (2) for transfection to BMSCs by using positive liposome pcDNA3.1 (-)-mediated transfection. Stable transfected BMSCs cells were obtained. (3) the expression of pcDNA3.1-hBMP2 in rabbit BMSCs was detected by immunohistochemistry and Western blot. Methods (1) BMSCs was cultured and amplified by adherent method. Flow cytometry was used to measure the surface markers. (2) the successfully constructed pcDNA3.1-hBMP2 recombinant plasmid was transfected into target cells in vitro and the stable expression cells were screened by G418. (3) the protein expression in BMSCs was detected by immunohistochemistry and Western Blot. Results (1) cultured rabbit bone marrow mesenchymal stem cells (BMSCs) could be used as target cells for hBMP2 transfection. (2) MSCs transfected with pcDNA3.1-hBMP2 by cationic liposome was screened by G418 and detected by immunohistochemistry with Western blot. A large number of hBMP2 mRNA transcripts and related proteins were expressed in rabbit bone marrow mesenchymal stem cells after pcDNA3.1-hBMP2 transfection, but no positive expression was found in untransfected bone marrow mesenchymal stem cells. Conclusion (1) Rabbit bone marrow mesenchymal stem cells can be used as target cells for hBMP2 transfection. (2) the eukaryotic expression plasmid of hBMP2 was successfully constructed and expressed in rabbit bone marrow mesenchymal stem cells. (3) in order to further study the transfection of BMP2 gene in rabbit bone marrow mesenchymal stem cells. The proliferation and stable expression of MSCs in vitro and the experiment of accelerating the formation of new bone in distraction osteogenesis were established in animal experiments.
【學位授予單位】：廣西醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2010
【分類號】：R329

【引證文獻】

相關碩士學位論文 前1條

1 高曉燕;金葡液在兔下頜骨牽張成骨中作用的研究[D];遼寧醫(yī)學院;2011年

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本文編號：2146702