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为研究FoxO1与骨骼肌纤维类型之间的关系,本试验以大白猪为实验材料,利用RT-PCR和Western印迹技术,检测了FoxO1与肌纤维类型标志基因MyHCⅠ、MyHCⅡa、MyHCⅡb和MyHCⅡx在特定骨骼肌中的表达规律,以及调控肌纤维类型关键基因Mef2c和NFAT的表达,并用Wortmannin处理原代培养的猪骨骼肌成肌细胞,检测了FoxO1与肌纤维类型相关基因的表达.结果显示,FoxO1在不同骨骼肌类型中mRNA表达差异不显著(P0.05),其蛋白表达与MyHC各亚型显著相关.Wortmannin处理结果显示,在处理的第3和5d,FoxO1蛋白与MyHCⅡb,MyHCⅡx和NFAT表达显著正相关,而与MyHCⅠ,MyHCⅡa和Mef2c表达显著负相关.结果表明,FoxO1通过抑制MyHCⅠ的表达调控肌纤维类型. 相似文献
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胰岛β细胞发生去分化现象是导致其功能减退的机制之一。已有研究证明,FoxO1与β细胞去分化密切相关。然而,高糖是否可通过FoxO1诱导β细胞发生去分化目前尚未见报告。本研究通过不同浓度高糖干预MIN6细胞,采用葡萄糖刺激胰岛素分泌试验（GSIS）检测β细胞功能|实时荧光定量PCR及蛋白免疫印迹、免疫荧光方法检测高糖干预后β细胞内祖细胞标志基因、β细胞标志基因及FoxO1的表达变化。结果显示,不同浓度高糖干预β细胞后,当浓度达到35 mmol/L时,β细胞祖细胞标志基因表达明显增加。且在该浓度时,检测到β细胞标志基因表达明显降低,MIN6细胞葡萄糖刺激胰岛素分泌功能减退,磷酸化FoxO1表达减少。上述结果提示,高糖可诱导胰岛β细胞去分化的发生,其机制可能是通过FoxO1介导。 相似文献
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分别取6月龄八眉猪、长白猪和长×八杂交猪背最长肌,提取总RNA,设计并合成猪FoxO1和MyoD 引物,以β肌动蛋白作为内参,优化反应条件和体系,利用RT-PCR方法检测八眉、长白和长×八杂交猪肌肉组织中FoxO1和MyoD基因mRNA 的表达.结果表明,在不同经济类型猪品种肌肉组织中,FoxO1和MyoD基因mRNA的表达丰度均存在显著差异,FoxO1在八眉猪肌肉组织中的表达普遍高于长白猪（P<0.01）,在杂交组合长×八肌肉组织中的表达也高于长白猪（P<0.01）;而MyoD恰好相反,即在长白猪肌肉组织中的表达普遍高于八眉猪（P<0.01）,在杂交组合长×八肌肉组织中的表达也高于八眉猪（P<0.01）．结果提示,FoxO1和MyoD基因mRNA的表达在肌肉发育中存在负相关（r = 0.728 , P < 0.05）,FoxO1在肌肉组织中的上调作用,可能是造成的MyoD基因mRNA表达降低的原因之一,进而负调控肌肉的发育和骨骼肌的量． 相似文献
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八眉猪、长白猪及长×八杂交猪肌肉组织中FoxO1基因的表达 总被引：1，自引：0，他引：1
分别取6月龄八眉、长白和 (长×八)杂交猪后腿部比目鱼肌、腓肠肌和趾长伸肌, 提取总RNA, 根据人、黑猩猩及大鼠等物种FoxO1基因同源序列设计并合成引物, 以猪b-actin 基因作为内参, 优化反应条件和体系, RT-PCR 单管扩增猪FoxO1基因, 检测八眉、长白和长×八杂交猪不同类型骨骼肌中FoxO1基因mRNA的表达差异。结果表明: 在不同经济类型猪群和不同类型骨骼肌中FoxO1基因mRNA的表达丰度不同, 即在八眉 猪骨骼肌中的表达普遍高于长白猪 (P<0.01), 在杂交组合 (长×八)骨骼肌中的表达也高于长白猪 (P<0.01); 同时在以Ⅰ型纤维为主的比目鱼肌中表达丰度最低(P<0.01), 在以Ⅱb型纤维为主的趾长伸肌中表达丰度最高(P<0.01)。结果提示, FoxO1基因的表达与Ⅰ型肌纤维的含量成反比; 不同经济类型猪品种骨骼肌的发育与FoxO1基因的调控有关。 相似文献
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外源性重组人睫状神经营养因子抑制成人成肌细胞的体外分化 总被引：4，自引：0，他引：4
为探讨外源性重组人睫状神经营养因子(rhCNTF)在成肌细胞分化中的作用,实验观察了0-10 ng/mlrhCNTF对成人成肌细胞体外分化的影响。结果表明,与对照组相比,2.5-10 ng/ml rhCNTF能显著抑制成肌细胞的体外分化(P<0.01),并呈量-效依赖关系,且这种抑制作用是可逆的。Western Blot分析提示,这种抑制作用伴有成肌细胞分化期特异标志myogenin和p21表达量的显著降低(P<0.01),以及成肌细胞增殖期特异标志myf5和desmin表达量的显著增加(P<0.01)。因此可以认为,外源性rhCNTF能可逆地抑制成人成肌细胞的体外分化并保持增殖。 相似文献
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本研究旨在探究甲基转移酶METTL21C在家禽骨骼肌发育分化过程中的作用。采用实时荧光定量PCR (quantitative Real-time PCR, qPCR)检测METTL21C基因在鸡不同组织中的表达情况,绘制其组织表达谱;选取3个时间点,检测其在骨骼肌组织中的表达情况。通过酶消化法从鸡骨骼肌组织中分离得到原代细胞;将METTL21C超表达载体转染至原代鸡骨骼肌细胞,通过qPCR和Western blotting检测Pax7、MyoD、Myf5、MyoG等基因的表达水平。结果显示,METTL21C在心肌和骨骼肌组织中的表达量显著高于其他组织,在胚胎期和幼龄期骨骼肌中的表达呈上升趋势;超表达METTL21C后,成肌相关基因Pax7、MyoD、Myf5、MyoG的表达量显著升高。本研究初步发现甲基转移酶METTL21C具有促进家禽骨骼肌发育分化的作用,为骨骼肌发育的分子机理的研究及相关医学研究提供数据支持。 相似文献
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研究瘦素(leptin)对猪骨骼肌成肌细胞分化的影响, 并探讨其可能的作用机制。原代培养猪骨骼肌成肌细胞, 通过倒置显微镜定性观察细胞分化的形态学变化; 肌酸激酶(CK)活性测定法定量分析成肌细胞分化程度; 细胞免疫化学方法分析myogenin的表达情况; 免疫印迹技术(Western Blot)检测丝裂素活化蛋白激酶(mitogen activated protein kinase, MAPK)的表达变化。结果显示, 在猪骨骼肌成肌细胞分化过程中, 外源性leptin减少细胞核融合和肌管形成, 呈浓度和时间依赖性地显著降低CK活性( P < 0.05), 并显著抑制myogenin和MAPK的蛋白表达( P < 0.05)。以上结果说明, leptin抑制猪骨骼肌成肌细胞分化, 并且这种作用可能是通过激活MAPK 信号转导通路实现的。研究结果显示, leptin在骨骼肌成肌细胞分化过程中可能具有重要作用。 相似文献
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Jonathan C. Fox Judith L. Swain 《In vitro cellular & developmental biology. Animal》1993,29(3):228-230
Summary Fibroblast growth factors (FGFs) are potent inhibitors of myogenic differentiation. The recent observation that the endogenous expression of acidic and basic FGF by myogenic cells decreases coordinately with differentiation suggests a regulatory role for these growth factors in myogenesis. Inasmuch as other proteins known to influence myogenesis (e.g., MyoD1 and myogenin) activate their own expression as well as the expression of other members of their family, we hypothesized that the FGFs might be capable of similar autoregulation. We examined the effect of exogenously supplied FGF on the abundance of the mRNAs encoding acidic and basic FGF in Sol 8 myoblasts, and demonstrate that either acidic or basic FGF stimulate, through paracrine mechanisms, the accumulation of the mRNAs encoding both of these FGFs. Thus FGFs can auto- and transregulate their own expression in a manner analogous to that observed for the myogenic determination proteins. In addition, similar to that previously observed for MyoD1, both acidic and basic FGF suppress myogenin expression in myoblasts. These results suggest two mechanisms whereby endogenously produced FGFs participate in the maintenance of the undifferentiated state of myogenic cells. These data provide support for paracrine, and suggest potential autocrine, roles for FGFs in the regulation of myogenic differentiation. 相似文献
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In this experiment, we evaluated the effects of strong static magnetic fields (SMF) on the orientation of myotubes formed from a mouse-derived myoblast cell line, C2C12. Myogenic differentiation of C2C12 cells was conducted under exposure to SMF at a magnetic flux density of 0-10 T and a magnetic gradient of 0-41.7 T/m. Exposure to SMF at 10 T led to significant formation of oriented myotubes. Under the high magnetic field gradient and a high value of the product of the magnetic flux density and magnetic field gradient, myotube orientation increased as the myogenic differentiation period increased. At the 3 T exposure position, where there was a moderate magnetic flux density and moderate magnetic field gradient, myotube orientation was not observed. We demonstrated that SMF induced the formation of oriented myotubes depending on the magnetic flux density, and that a high magnetic field gradient and a high value of the product of the magnetic flux density and magnetic field gradient induced the formation of oriented myotubes 6 days after myogenic differentiation. We did not detect any effect of the static magnetic fields on myogenic differentiation or cell number. To the best of our knowledge, this is the first report to demonstrate that myotubes orient to each other under a SMF without affecting the cell number and myogenic differentiation. 相似文献
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为研究脑信号蛋白家族（Semaphorins）成员Sema7A对成肌细胞增殖和分化的影响,本文设计并合成了Sema7A基因的小干扰RNA（small interfering RNA,siRNA）,用此siRNA转染C2C12成肌细胞．通过Hoechst核染和流式细胞术检测细胞增殖情况,免疫荧光检测肌管的形成情况,real-time qPCR和Western印迹技术检测成肌标记基因的变化.结果显示,干扰Sema7A后,C2C12成肌细胞增殖减慢,处在G2和S期的细胞所占的比例明显下降,而G1期细胞的比例升高．免疫荧光检测结果显示,干扰Sema7A后,肌管的直径及MyHC+细胞所占比例均显著降低．Real-time qPCR和Western印迹结果也显示,肌肉分化标志基因MyoD、MyoG、MyHC的mRNA及蛋白质表达均下降．进一步检测Sema7A受体下游信号通路发现,干扰Sema7A后,其下游信号分子PI3K和AKT的磷酸化水平被下调．以上结果表明,Sema7A可以调节C2C12成肌细胞的增殖和分化,可能是通过其受体作用于PI3K/AKT信号通路实现的,这为进一步研究Sema7A在骨骼肌发育中的作用提供实验基础. 相似文献
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S. Watabe 《Journal of fish biology》1999,55(SA):1-18
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Vahideh Tarhriz Shirin Eyvazi Maryam Musavi Mozhgan Abasi Kazem Sharifi Hossein Ghanbarian Mohammad Saeid Hejazi 《Journal of cellular biochemistry》2019,120(11):18854-18861
Cdk9 is a serine-threonine protein kinase that has been recognized as a regulator of cardiac differentiation. Recently, we have reported that transient induction of Cdk9 using noncoding RNA targeting Cdk9 sequences results in efficient cardiac differentiation. Concerning Cdk9 regulatory roles, here, we proposed whether constant overexpression of Cdk9 might influence the differentiation of myoblast C2C12 cells into myotubes. We overexpressed Cdk9 in mouse myoblast C2C12 cells to investigate its regulatory roles on myogenic differentiation. Upon Cdk9 overexpression, the expression level of myogenic regulatory factors was determined. Moreover, the expression profile of three important myomiRs consist of miR 1, 133 and 206 was examined during the differentiation process. Although Cdk9 expression is necessary for inducing differentiation in the early stage of myogenesis, continuous Cdk9 expression inhibits differentiation by modulating myomiRs and myogenic gene expression. Our results indicate that the transient induction of Cdk9 in the early stage of differentiation is critical for myogenesis. 相似文献
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Thomas A. Linkhart Christopher H. Clegg Stephen D. Hauschka 《Journal of cellular biochemistry》1980,14(4):483-498
Regulation of the transition of mouse myoblasts from proliferation to terminal differentiation was studied with clonal density cultures of a permanent clonal myoblast cell line. In medium lacking mitogenic activity, mouse myoblasts withdraw from the cell cycle, elaborate muscle-specific gene products, and fuse to form multinucleated myotubes. Addition of a purified mitogen, fibroblast growth factor, to mitogen-depleted medium stimulates continued proliferation and prevents terminal differentiation. When mitogens are removed for increasing durations and then refed, mouse myoblasts irreversibly commit to terminal differentiation: after 2–4 h in the absence of mitogens, myoblasts withdraw from the cell cycle, elaborate muscle-specific gene products, and fuse in the presence of mitogens that have been fed back. Population kinetics of commitment determined with 3H-thymidine labeling and autoradiography suggest the following cell-cycle model for mouse myoblast commitment: (1) if mitogens are present in the extracellular environment of myoblasts in G1 of the cell cycle, the cells enter S and continue through another cell cycle; (2) if mitogens have been absent for 2 or more hours, cells in G1 do not enter S; the cells commit to differentiate, permanently withdraw from the cell cycle (will not enter S if mitogens are refed), and they subsequently elaborate acetylcholine receptors and fuse (even if mitogens are refed); (3) cells in other phases of the cell cycle continue to transit the cell cycle in the absence of mitogens until reaching the next G1. The commitment kinetics and experiments with mitotically synchronized cells suggest that the commitment “decision” is made during G1. Present results do not, however, exclude commitment of some cells in other phases of the cell cycle. 相似文献
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R. G. Mac Bride 《In vitro cellular & developmental biology. Plant》1989,25(7):617-620
Summary Addition of low concentrations of ascorbic acid (5 μg/ml to 10 μg/ml) to myogenic chick embryo cultures resulted in an early
fusion. At 30 h cultures receiving small amounts of ascorbic acid presented fusion rates 3 times that of the control. However,
control rates of fusion were not different from those of experimentals at 50 h. No such effect was seen with ascorbic acid
added at 24 h of culture, or with ascorbic acid addition to a calcium-deprived system. These findings demonstrate that the
calcium binding properties of ascorbic acid can induce precocious myogenic fusion. 相似文献