BMP9在骨形成中的分子机制及其信号途径的研究
发布时间：2018-12-18 18:29
【摘要】： BMPs属于TGF-β超家族的一员,在胚胎发育及骨骼形成中扮演着重要的角色。虽然如今只有几种BMP被证实有促进骨形成的作用并应用于临床(主要为BMP2及BMP7),但并未有报道确其认为具有最强成骨作用的BMP。本课题从两个方面着手,一方面是从体内实验探索BMP9的特殊的信号转导通路:另一方面从体外实验去证明BMP9是最强的成骨诱导因子。 第一部分是通过基因芯片、Real-time PCR、Western-Blotting等方法探索BMP9是否遵循传统经典BMP信号传导通路介导骨形成的理论机制。比较BMP2、9作用下小鼠多潜能干细胞C3H10中BMP信号传导通路核心基因Smads的表达差异;同时采用RNA干扰的方法沉默Smad4表达以减弱经典BMP信号传导系统的作用,比较BMP2、9作用下诱导C3H10体外成骨的差异。结果是与BMP2作用下C3H10中BMP信号传导通路核心基因Smads表达皆升高相比,BMP9刺激下上述基因无明显表达上调;Smad4基因表达沉默后,BMP9诱导干细胞体外成骨的作用无明显抑制。BMP9并不完全遵循传统经典BMP信号传导通路而发挥诱导成骨的生物学作用。 第二部分是比较腺病毒介导的人骨形态发生蛋白一9(Adv-hBMP9)和Adv-hBMP2纳米羟基磷灰石/聚酰胺复合物在损伤部位的骨缺损重建修复效果。新西兰白兔36只,随机分成三组,制成双侧桡骨中段13mm骨缺损模型。A组:植入Adv-hBMP9+纳米羟基磷灰石/聚酰胺骨;B组:植入Adv-hBMP2+纳米羟基磷灰石/聚酰胺骨;C组:注入Adv-GFP+羟基磷灰石/聚酰胺骨。在2,4,8,12,16周各时相点分别进行大体观察、x线照片、组织学切片。结果是BMP9组骨缺损完全修复,BMP2组骨缺损部分修复,而GFP组骨缺损修复明显欠佳。重组腺病毒介导的BMP9对桡骨骨缺损后的成骨修复作用强于BMP2。我们的研究为BMP9的成骨中的作用机制及信号途径打下一定基础,为临床上利用BMP9基因治疗骨骼系统相关疾病提供更强的骨诱导因子。
[Abstract]:BMPs is a member of TGF- 尾 superfamily and plays an important role in embryonic development and bone formation. Although only a few types of BMP have been proven to promote bone formation and have been used in clinical applications (mainly BMP2 and BMP7), there are no reports of BMP. that it believes has the strongest osteogenic effect. In this study, two aspects are involved, one is to explore the special signal transduction pathway of BMP9 in vivo, and the other is to prove that BMP9 is the strongest osteogenic factor in vitro. The first part is to explore whether BMP9 follows the traditional classical BMP signaling pathway to mediate bone formation through gene chip, Real-time PCR,Western-Blotting and other methods. To compare the expression of Smads, the core gene of BMP signal transduction pathway, in C3H10 of mouse multipotent stem cells induced by BMP2,9. At the same time, RNA interference was used to silence the expression of Smad4 in order to attenuate the role of classical BMP signal transduction system, and to compare the difference of C3H10 osteogenesis induced by BMP2,9 in vitro. The results showed that the expression of core gene Smads of BMP signal transduction pathway in C3H10 was higher than that in C3H10 induced by BMP2, but the expression of these genes was not up-regulated under BMP9 stimulation. After the expression of Smad4 gene was silenced, BMP9 did not inhibit the osteogenesis of stem cells in vitro. BMP9 did not follow the classical BMP signal transduction pathway and played a biological role in inducing osteogenesis. The second part is to compare the effect of adenovirus-mediated human bone morphogenetic protein-9 (Adv-hBMP9) and Adv-hBMP2 nano-hydroxyapatite / polyamide composite in the reconstruction of bone defect. Thirty-six New Zealand white rabbits were randomly divided into three groups to make bilateral radial 13mm bone defect model: group A: implantation of Adv-hBMP9 nano-hydroxyapatite / polyamide bone, group B: implantation of Adv-hBMP2 nano-hydroxyapatite / polyamide bone; Group C: Adv-GFP hydroxyapatite / polyamide bone was injected. Gross observation, X-ray photograph and histological section were carried out at each time point of 2 ~ 4 ~ 8 ~ 12 ~ 12 ~ (th) weeks. The results showed that the bone defects were repaired completely in BMP9 group, partially in BMP2 group, but not in GFP group. The effect of recombinant adenovirus-mediated BMP9 on bone repair after radial bone defect is stronger than that of BMP2.. Our study lays a foundation for the mechanism and signal pathway of BMP9 osteogenesis and provides a stronger bone inducer for the clinical treatment of skeletal system related diseases with BMP9 gene.
【学位授予单位】：重庆医科大学
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R346
[Abstract]:BMPs is a member of TGF- 尾 superfamily and plays an important role in embryonic development and bone formation. Although only a few types of BMP have been proven to promote bone formation and have been used in clinical applications (mainly BMP2 and BMP7), there are no reports of BMP. that it believes has the strongest osteogenic effect. In this study, two aspects are involved, one is to explore the special signal transduction pathway of BMP9 in vivo, and the other is to prove that BMP9 is the strongest osteogenic factor in vitro. The first part is to explore whether BMP9 follows the traditional classical BMP signaling pathway to mediate bone formation through gene chip, Real-time PCR,Western-Blotting and other methods. To compare the expression of Smads, the core gene of BMP signal transduction pathway, in C3H10 of mouse multipotent stem cells induced by BMP2,9. At the same time, RNA interference was used to silence the expression of Smad4 in order to attenuate the role of classical BMP signal transduction system, and to compare the difference of C3H10 osteogenesis induced by BMP2,9 in vitro. The results showed that the expression of core gene Smads of BMP signal transduction pathway in C3H10 was higher than that in C3H10 induced by BMP2, but the expression of these genes was not up-regulated under BMP9 stimulation. After the expression of Smad4 gene was silenced, BMP9 did not inhibit the osteogenesis of stem cells in vitro. BMP9 did not follow the classical BMP signal transduction pathway and played a biological role in inducing osteogenesis. The second part is to compare the effect of adenovirus-mediated human bone morphogenetic protein-9 (Adv-hBMP9) and Adv-hBMP2 nano-hydroxyapatite / polyamide composite in the reconstruction of bone defect. Thirty-six New Zealand white rabbits were randomly divided into three groups to make bilateral radial 13mm bone defect model: group A: implantation of Adv-hBMP9 nano-hydroxyapatite / polyamide bone, group B: implantation of Adv-hBMP2 nano-hydroxyapatite / polyamide bone; Group C: Adv-GFP hydroxyapatite / polyamide bone was injected. Gross observation, X-ray photograph and histological section were carried out at each time point of 2 ~ 4 ~ 8 ~ 12 ~ 12 ~ (th) weeks. The results showed that the bone defects were repaired completely in BMP9 group, partially in BMP2 group, but not in GFP group. The effect of recombinant adenovirus-mediated BMP9 on bone repair after radial bone defect is stronger than that of BMP2.. Our study lays a foundation for the mechanism and signal pathway of BMP9 osteogenesis and provides a stronger bone inducer for the clinical treatment of skeletal system related diseases with BMP9 gene.
【学位授予单位】：重庆医科大学
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R346
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