Pax3/Pax7对猪肌纤维发育及类型转化的决定作用

The porcine meat quality is closely related with muscle fiber types. Whether nutrition could regulate skeletal muscular fiber types and their transformation depends on differentiations of muscluar prenatal stem cell and postnatal satellite cells. This study hypothesizes that the myogenic regulators Pax3/7 play decisive role in this biological process and aims to explore the evidences for the hypothesis.Firstly ,sows fed high or low-energy diets will be used to examine the fetal expressions of Pax3/7 and their relationships with musclular fiber types at various ages of gestation. Then, embryonic muscle stem cells and adult satellite cells will be isolated and transfected with vectors of Pax3/7 siRNA. The heterogenetic proliferation and differentiation of cells are going to be analysized by Laser Scanning Confocal Microscope and Flew Cytometry. Expressions of both Pax3/7 and several marker genes of myocytes will be detected by means of real-time PCR, western bolt and immunohistochemistry. The above evidences will indicate that Pax3/7 play a decisive role for the process that porcine embryonic musclular stem cells differentiate into distinct musclular fiber types and for the fiber type transformations resulted from differentiation and amalgamation of postnatal satellite cells. This study will be fundamental to further understanding the development and establishing nutritional manipulation of porcine meat quality.

猪肉品质与肌纤维类型密切相关。营养调控肌纤维类型与类型转化取决于胚胎期肌肉干细胞和生后卫星细胞的定向分化情况。本项目假设生肌调控因子Pax3/Pax7对这一生物学过程起着决定性调节作用,并旨在探索其正确性。项目首先检测饲喂高低能营养日粮母猪在妊娠不同时间的胎猪的骨骼肌Pax3/7表达规律，并分析其与肌纤维类型的关系；然后相继以分离培养的胚胎肌源干细胞及成体卫星细胞为材料，构建Pax3 、Pax7siRNA并转染，采用激光共聚焦、流式细胞术等方法分析细胞的增殖及分化差异，采用real-time PCR、western blot、免疫组化等技术检测Pax3/7及肌细胞标志基因的表达情况，以探明Pax3/7在猪肌肉干细胞定向分化成不同类型肌纤维及生后骨骼肌卫星细胞分化融合导致肌纤维类型转化的决定作用，为深入认识猪肉品质改善原理及建立营养调控技术奠定生理基础。

猪肉品质与肌纤维类型密切相关。营养调控肌纤维类型与类型转化取决于胚胎期肌肉干细胞和生后卫星细胞的定向分化情况。本项目假设生肌调控因子Pax3/Pax7对这一生物学过程起着决定性调节作用。项目首先检测了妊娠及生后不同时间点猪骨骼肌Pax3/7表达规律，并分析其与肌纤维类型的关系。研究发现，胚胎期，Pax7的表达与肌纤维发育和肌纤维类型间具有显著相关性；出生后，Pax3和Pax7的表达与肌纤维类型有关，且具有品种和肌肉特异性。然后，以小鼠卫星细胞、C2C12细胞及基因敲除小鼠为材料，采用激光共聚焦、流式细胞术、real-time PCR、western blot、免疫组化等技术，研究发现Pax3+卫星细胞维持更长时间自我更新和增殖能力，分化后以表达慢肌为主，而Pax7+卫星细胞更倾向分化，分化后以表达快肌为主；Pax7可激活Zac1/GPR39系统，促使CaMK-II磷酸化，抑制细胞核内β-catenin，促进快肌纤维形成。维生素D3可促进快肌生成，其机制是阻断shh信号途径，促进T3分泌，抑制Pcbp1的表达，增加Pax7表达；适当降低饲粮营养水平可提高Pax3、Myf5、MyoG和Mrf4表达量，增加育肥猪背最长肌中氧化型肌纤维、降低酵解型肌纤维表达，从而提高肌肉嫩度，改善肉质。结果证明，Pax3/7在肌纤维发育及类型转化以及营养调控中发挥决定作用。研究结果为深入认识猪肉品质改善原理及建立营养调控技术奠定了生理基础。