张亚莲,舒彬,杨忠,等.低强度脉冲超声促进C2C12成肌细胞分化的作用机制研究[J].中华物理医学与康复杂志,2021,43(3):193-199
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| 低强度脉冲超声促进C2C12成肌细胞分化的作用机制研究 |
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| DOI：10.3760/cma.j.issn.0254-1424.2021.03.001 |
| 中文关键词: 低强度脉冲超声波 脂联素 脂联素受体 分化 |
| 英文关键词: Ultrasound intensity Pulsed ultrasound Adiponectin Adiponectin receptor Differentiation Myoblasts |
| 基金项目:国家自然科学基金项目（31171148） |
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| 摘要点击次数: 3195 |
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| 中文摘要: |
| 目的 观察不同功率密度低强度脉冲超声（LIPUS）对骨骼肌C2C12成肌细胞脂联素（Adiponectin）及其受体的影响，探讨LIPUS促进C2C12成肌细胞分化的潜在机制。 方法 将体外培养的C2C12成肌细胞根据有无超声干预或超声干预的功率密度随机分为对照组、U0.1组、U0.3组和U0.5组，对照组接受假LIPUS干预，U0.1组、U0.3组、U0.5组分别接受功率密度为0.1 W/cm2、0.3 W/cm2、0.5 W/cm2的LIPUS干预。干预5 d后，分别采用CCK-8法检测细胞活力，采用荧光定量逆转录-聚合酶链反应（RT-PCR）法检测脂联素、脂联素受体1（AdipoR1）和T-钙黏蛋白（T-Cadherin）mRNA水平，采用Western blot法检测脂联素、AdipoR1、T-Cadherin、腺苷酸活化蛋白激酶(AMPK)、活化型-磷酸化腺苷酸活化蛋白激酶（P-AMPK）、肌细胞生成蛋白（MYOG）和胚胎肌球蛋白重链（eMHC）的蛋白水平，并对4组细胞肌管的分化能力进行免疫荧光化学检测。 结果 LIPUS干预5 d后，U0.1、U0.3和U0.5组细胞相对活力明显高于对照组（P<0.01）。U0.3组和U0.5组脂联素、受体AdipoR1和T-Cadherin mRNA水平均显著上调，与对照组比较，差异均有统计学意义（P<0.05）。U0.3组和U0.5组脂联素、AdipoR1、T-Cadherin蛋白和AMPK磷酸化水平与对照组比较，均显著增加（P<0.05），且U0.1组和U0.5组均显著低于U0.3组，差异均有统计学意义（P<0.01）。U0.3组和U0.5组C2C12成肌细胞分化指标eMHC、MYOG蛋白水平和C2C12成肌细胞融合指数与对照组比较，均明显增加，差异均有统计学意义（P<0.05）。 结论 LIPUS可促进C2C12成肌细胞的分化，并以0.3 W/cm2、5 min/d、1 MHz，占空比为20%的LIPUS干预作用最明显，其调控机制可能与C2C12成肌细胞脂联素、受体AdipoR1和T-Cadherin的表达上调和下游AMPK磷酸化水平活化有关。 |
| 英文摘要: |
| Objective To observe the effect of low-intensity pulsed ultrasound (LIPUS) at different intensities on the expression of adiponectin and its receptors in C2C12 myoblasts, and to explore the potential mechanism by which LIPUS promotes the differentiation of C2C12 myoblasts. Methods C2C12 myoblasts cultured in vitro were randomly divided into a control group and U0.1, U0.3 and U0.5 groups. The control group received sham LIPUS exposure, while the U0.1, U0.3 and U0.5 groups were exposed to LIPUS at intensities of 0.1W/cm2, 0.3W/cm2 or 0.5W/cm2 respectively, and 1MHz for 5 min daily for 5 days. Cell viability was measured using CCK-8 assays. Fluorescence quantitative reverse transcription-polymerase chain reactions were used to detect the mRNA expression of adiponectin, adiponectin receptor 1 (adipoR1) and T-cadherin in the cells. Western blotting was employed to assess the protein expression of adiponectin, adipoR1, T-cadherin, adenosine monophosphate activated protein kinase (AMPK), activated phosphorylated adenylate-activated protein kinase (P-AMPK), embryonic myosin heavy chain (eMHC) and myogenin (MYOG). The differentiation ability of the 4 groups was measured using cell immunofluorescence chemistry. Results After the intervention the cell viability in the U0.1, U0.3 and U0.5 groups was significantly higher than in the control group. Compared with the control group, the average mRNA expression of adiponectin and the receptors of adipoR1 and T-cadherin were up-regulated significantly in the U0.3 and U0.5 groups. The average adiponectin, adipoR1 and T-cadherin protein expressions, and the AMPK phosphorylation level in the U0.3 and U0.5 groups had increased significantly compared with the control group, but all were significantly lower than in the U0.3 group. The average protein expression of eMHC and MYOG, and the C2C12 myoblast fusion indices of the U0.3 and U0.5 groups were significantly higher the control group′s averages. Conclusions LIPUS can promote the differentiation of C2C12 myoblasts. It is most effective at 0.3W/cm2, administered for 5min/d at 1MHz with a 20% duty cycle. Its regulatory mechanism may be related to up-regulation of the expression of adiponectin, the adipoR1 and T-cadherin receptors, and the activation of AMPK phosphorylation in C2C12 myoblasts. |
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