【作者】 徐丽；

【导师】 巴雪青；

【作者基本信息】 东北师范大学 ， 遗传学， 2013， 硕士

【摘要】 成肌细胞融合是一个多步骤复杂的严格调控过程，需要粘附分子和调节因子的相互作用进而对这一过程进行调节。很多研究报道分别关注了M-cadherin，MyoD和MyoG对肌细胞融合过程的调节。M-cadherin是钙粘素家族成员中专一性在骨骼肌细胞以及成肌细胞中表达的粘附分子，一般表达于肌细胞的表面，主要介导细胞与细胞之间的同型粘附。研究报道，M-cadherin与肌细胞融合过程中粘附位点的建立有关，在肌细胞之间相互粘附以及使成肌细胞之间相互融合形成肌纤维过程中具有重要的作用。MRFs家族主要包括四个成员：MyoG，MyoD，Myf5和MRF4。其中与肌细胞融合相关的调节因子被报道最多而且比较明确的是MyoG和MyoD。MyoD可以使静置的骨骼肌卫星细胞转化为成肌细胞，还能使成纤维细胞、脂肪细胞等许多种细胞转化为成肌细胞，并能促使成肌细胞融为肌管、肌纤维。其中MyoG又称肌细胞生成素，它能控制成肌细胞的起始融合，促进单核成肌细胞融合形成多核的肌纤维。虽然很多研究报道分别关注粘附分子和生肌调节因子对肌细胞融合过程的调节，但对于粘附分子和生肌调节因子是如何通过相互作用对肌融合这一过程进行调节的报道甚少，很多问题都值得思考和进一步的探究和揭示。本研究中，我们选用C2C12小鼠的骨骼肌卫星细胞，分化模型中确定6个时间点，探究了M-cadherin、MyoD和Myog在肌细胞融合过程中的基因表达调控情况。根据以上结果，我们将这几种分子的Real-time结果进行了比较，在T1时间点MyoD的表达量与MyoG和M-cadherin的表达量相比，MyoD的表达量最高，T2时间点虽然也是MyoD表达量最高，但是在T2时间点也是M-cadherin的表达量最高的时间点。这与文献报道的MyoD通过作用于M-cadherin的启动子从而调节M-cadherin的表达相一致。MyoD的相对高表达从而启动M-cadherin的表达。这预示着MyoD的表达能调节M-cadherin的表达。MyoG在T3时间点表达量最高，可能是由于在T2时间点M-cadherin的高表达使成肌细胞之间建立稳定的粘附，稳定粘附的信号转导到细胞内，促使MyoG在T3时间点高表达，使成肌细胞进一步融合，肌管不断生长。根据上述结果，将M-cadherin进行干涉掉之后确实影响MyoG的表达。从整个时间点分析加入siM-cadherin的实验组与对照组相比成肌细胞的融合受到明显的抑制，几乎不发生融合。这些实验结果初步验证了我们对肌融合过程分子调节机制的推断: MyoD活化M-cadherin的表达，从而诱导MyoG表达。希望能为肌融合作用的分子机制奠定理论基础。更多还原

【Abstract】 Cell-cell fusion is a crucial and highly regulated event, cadherins and MRFs can be adjustedfor this process. Many research reports focus on the regulation of M-cadherin, MyoD and MyoGduring myoblast fusion process. M-cadherin is a member of cadherins, which is one of fivecategories adhesion molecules, mediated adhesion between the same types of cells. M-cadherinis the only member of the cadherin superfamily whose expression is restricted in the developingskeletal muscle and correlated with differentiation of myogenic cells. In the study reported thatM-cadherin Related to the establishment of adhesion sites during myoblast fusion process,M-cadherin mutual adhesion between myoblasts as well as mutual fusion between myoblasts hasan important role in the process of forming the muscle fibers. MRFs mainly consists of fourmembers: MyoG, MyoD, Myf5, and MRF4. MyoG and MyoD are the regulation factor aboutmyoblast fusion. MyoD can make the skeletal muscle satellite cells transformed into myoblasts,and also can make Fibroblast and Fat cells transformed into myoblasts. MyoD also can makeMyoblast fuse to muscle fibers. MyoG, also known as myogenin, can control the initiation fusionof Myoblast and promote mononuclear myoblasts fusing to form multinucleated muscle fibers.MyoG played a leading role in the MRFs family.Although many researches about the regulation of M-cadherin, MyoD and MyoG during themyoblast fusion have been reported, But for adhesion molecules and myogenic regulatory factoris how the interaction of muscle fusion of the regulation of this process has rarely been reported.Many issues are worth doing further research. Therefore, in my study, choosing C2C12skeletalmuscle satellite cells of mouse in the differentiation model at6time point to researchM-cadherin, MyoD, and Myog expression of the gene transcriptional regulationin the myoblastfusion process From the above results, the Real-time results of molecular are compared. At thetime point T1, MyoD expression level is the highest, compared with the MyoG and M-cadherinexpression level. At the time point T2, MyoD expression level is still the highest, whileM-cadherin expression level reaches the highest value. This is consistent with the literature, thatMyoD can act on the promoter of M-cadherin to control M-cadherin expression. The MyoDrelatively high expression promotes M-cadherin expression. This indicates that MyoD cancontrol M-cadherin expression. The highest expression level of Myog is at the time point T3.This may be due to the high expression of M-cadherin establishing a stable adhesion betweenmyoblasts at T2. The signal of this stable adhesion is transduced into cells, which promotesmyog to achieve high expression at T3. According to the above results, when the interference isused to get rid of the M-cadherin, the MyoG expression is indeed affected. Analyzed andcompared the SIM-cadherin experimental group with the control group from the whole point-in-time, the fusion of Myoblast is inhibited; almost no integration happens. These resultsinitially validate our inference about the muscle fusion molecules adjustment mechanism: MyoDactivated the expression of M-cadherin, to induce the expression of MyoG. This article hopes tolay the theoretical foundation for the muscle fusion of the molecular mechanism.更多还原