中国农业科学Issue(12)：2460-2468,9.DOI:10.3864/j.issn.0578-1752.2015.12.019
TGF-β/Smad信号通路在Follistatin调节鸭骨骼肌卫星细胞增殖过程中的作用机制
The Regulated Mechanism of Follistatin on the Proliferation Process of Duck Skeletal Muscle Satellite Cell Involved in TGF-β/Smad Signaling Pathway
摘要
Abstract
Objective]Follistatin can regulate skeletal muscle hypertrophy and fat deposition, and promote proliferation of the skeletal muscle satellite cell. The study intends to useinvitro recombination follistatin to treat the duck skeletal muscle satellite cell, which can be proved that the functional mechanism of TGF-β/smad signaling pathway is playing a role in its proliferation process.[Method] Based on differential centrifugation technology, skeletal muscle satellite cells isolated from embryonic 14-day ducks were treated by proliferation medium containing 0,1,10 and 100 ng·mL-1 follistatin, respectively, after the cells density reached 70%-80% confluence. After incubation for 36 h, the degree of skeletal muscle satellite cells proliferation were tested by CCK-8. The cells were identified by immunofluorescence staining with pax7 antibody. Real-time qPCR was performed to measure the genes of expression differntiation including proliferation marker gene PCNA, and myogenic MyoD, smad2/3, TGF-βcaused by follistatin supplement.[Result]Under the inverted microscope, ti was found that a part of duck skeletal muscle cells did not adhere to round, a of cells part of cells adhered to fusiform 12 hours of culture. Adherent cell number increased and cells slightly longer after 24 h. Cells completely adherent, and fusiform, on day 2. The cell number increased, some cells differentiated, after 3 days. The number of cells further was increased, the cells thicker, some cells started to fuse after 4 days. A small amount of cells began differentiation, and further fusion of the cell on day five. The result of immunofluorescence staining showed that pax7 in more than 95% of the skeletal muscle satellite cells was positive, and the population of cells stained with Pax7 was suggested that the group of 10 ng·mL-1 follistatin was significantly higher than the control group (P<0.01). The gene expression analysis showed that compared with the control group, the expression level of MyoD in 10 ng·mL-1 follistatin treated group was significantly decreased (P<0.05), whereas, the expression levels of PCNA, TGF-β and Smad2 in follistatin treated groups were significantly increased (P<0.05). The expression of Smad3 gene ultimately significantly increased (P<0.01). Results of detection of protein expression by Western blotting showed that compared with the control group, TGF-β protein and Smad2/3 phosphorylat ion protein levels were also significantly increased.[Conclusion]Results of study suggested that follistatin promoted the expression of PCNA gene and reduced MyoD gene expression in duck skeletal muscle satellite cell. Furthermore, follistatin raised the expression of TGF-β, Smad2/3. Generally, 10 ng·mL-1 concentration of follistatin could significantly promote proliferation of the duck skeletal muscle satellite cell, the same that TGF-β / Smad signaling pathway maybe play a important role in it.关键词
卵泡抑素(Follistatin)/骨骼肌卫星细胞/增殖/鸭Key words
Follistatin/skeletal muscle satellite cell/proliferation/duck引用本文复制引用
林凯,虞德兵,解晓东,于敏莉,李东锋,杜文兴..TGF-β/Smad信号通路在Follistatin调节鸭骨骼肌卫星细胞增殖过程中的作用机制[J].中国农业科学,2015,(12):2460-2468,9.基金项目
国家科技支撑计划（2013BAD20B05），江苏省农业科技自主创新基金（CX（11）1033） （）
