miR-208b和miR-499在巴马香猪骨骼肌肌纤维类型转化中的作用研究

发布时间：2018-02-22 18:14

本文关键词： 巴马香猪肌纤维类型 MyHC miR-208b miR-499 AMPK PGC-1α　出处：《广西大学》2015年硕士论文　论文类型：学位论文

【摘要】：MicroRNAs是一类进化上非常保守的小的非编码RNA,在骨骼肌发育中起到了至关重要的调控作用。本实验以巴马香猪为对象,探究miR-208b和miR-499在骨骼肌肌纤维类型转化中的作用,为揭示巴马香猪优良肉质形成机制的研究奠定基础。获得的主要研究结果如下：1.miR-208b和miR-499在巴马香猪背最长肌、股二头肌、心脏中均有高丰度表达,在其他组织中的表达丰度较低。对巴马香猪不同发育时期的背最长肌进行检测,发现miR-208b和miR-499的表达量在胚胎期持续升高,出生后表达量下降。2.巴马香猪背最长肌发育过程中,MyHC Ⅰ的表达量显著性逐渐降低(P0.05),ⅡA的表达量变化不大(P0.05),ⅡX的表达量显著性逐渐升高(P0.05),ⅡB的表达量在E90开始升高。胚胎期氧化型肌纤维标志基因的mRNA所占的比例高于酵解型,表达比例在E40最高,随后下降；成年时期氧化型肌纤维标志基因的mRNA所占的比例低于酵解型。3.巴马香猪胚胎期,AMPK的表达与MyHC Ⅰ的表达呈显著正相关(r=0.9561,P0.05),PGC-1α的表达与MyHC ⅡB的表达呈显著负相关(r=-0.9410,P0.05)。4.miR-208b和miR-499的表达量均随着C2C12和小鼠肌肉卫星细胞的分化显著上升(P0.05)。5.C2C12和小鼠肌肉卫星细胞过表达miR-208b后,肌纤维形态更细长；在分化中后期显著上调MyHC Ⅰ的表达量(P0.05),显著下调MyHC ⅡB的表达量(P0.05)；分化早中期标志基因MyoG在C2C12及肌肉卫星细胞分化过程中的表达量实验组和对照组无差异(P0.05);C2C12及肌肉卫星细胞分化过程中,AMPK的表达量实验组和对照组均无显著差异(P0.05)；肌肉卫星细胞中PGC-1α的表达水平显著上调(P0.05)。6.C2C12和小鼠肌肉卫星细胞过表达miR-499后,肌纤维形态更细长；在分化中后期显著上调MyHCI的表达(P0.05),显著下调MyHC ⅡB的表达(P0.05)；MyoG基因的表达量实验组和对照组无差异(P0.05)；AMPK、PGC-1α的表达水平均显著上调(P0.05)。综上所述,miR-208b和miR-499对氧化型肌纤维的形成有促进作用,对酵解型肌纤维有抑制作用,可能通过调控AMPK、PGC-1α起作用。
[Abstract]:MicroRNAs is a very conservative small non RNA encoding a class of evolutionarily, during skeletal muscle development plays a vital role. In this experiment, Bama miniature pig as the object, to explore miR-208b and miR-499 in skeletal muscle fiber type transformation in the role of the study laid the foundation for revealing the excellent meat quality of Bama miniature pig formation mechanism. The main results are as follows: 1.miR-208b and miR-499 in Bama miniature pig longissimus muscle, unit two biceps, expressed in high abundance in the heart, expressed in other tissues. The lower of Bama miniature pigs in different developmental stages of longissimus muscle were detected and found that expression of miR-208b and miR-499 in the embryo the duration of elevated expression of.2. decreased after the birth of Bama miniature pig longissimus muscle during development, the expression of MyHC I was significantly decreased (P0.05), expression of A II (P0.05), little expression of X II Significantly increased (P0.05), the expression of B II began to increase at E90. The embryonic oxidative muscle fiber marker gene mRNA proportion is higher than that of glycolysis, expression ratio in the E40 is the highest, and then decreased; adult oxidative muscle fiber marker gene mRNA is lower than the proportion of glycolytic.3. pig embryos, there was a positive correlation between the expression of AMPK and MyHC I (r=0.9561, P0.05), negatively related to expression of PGC-1 alpha and MyHC II B (r=-0.9410, P0.05) expression of.4.miR-208b and miR-499 increased with the differentiation of C2C12 and mouse muscle satellite cells increased significantly (P0.05.5.C2C12) and mouse muscle satellite cells after overexpression of miR-208b, muscle fiber shape is more slender; in the late differentiation significantly up-regulated expression of MyHC I (P0.05), the amount of expression was significantly down regulated MyHC II B (P0.05) in the early stage; differentiation marker gene MyoG in C2C1 The amount of the experimental group and the control group had no difference in the expression of 2 and muscle satellite cells during differentiation (P0.05); C2C12 and muscle satellite cell differentiation, there were no significant differences in the expression of AMPK in the experimental group and the control group (P0.05); the expression level of PGC-1 alpha in muscle satellite cells increased significantly (P0.05) and.6.C2C12 mouse muscle satellite cells after overexpression of miR-499, muscle fiber morphology is more slender; expression in the differentiation of late significantly up-regulated the expression of MyHCI (P0.05), MyHC II B expression was significantly reduced (P0.05); the expression of MyoG gene in the experimental group and control group had no difference (P0.05); AMPK, PGC-1 protein expression significantly increase (P0.05). To sum up, miR-208b and miR-499 can promote the formation of oxidized muscle fibers, the glycolytic muscle fibers are inhibited, by regulating AMPK, PGC-1 alpha plays a role.

【学位授予单位】：广西大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S828

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