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《天津医科大学学报》[ISSN:1006-8147/CN:12-1259/R]

卷:

31卷

期数:

2025年03期

页码:

243-250

栏目:

基础医学

出版日期:

2025-05-20

文章信息/Info

Title:

The effect and mechanism of vitamin D on the differentiation of skeletal muscle cells

文章编号:

1006-8147(2025)03-0243-08

作者:

古秋¹; 岳莹莹²; 牛文彦¹

（1.天津医科大学基础医学院免疫学系，天津300070；2.天津市第一中心医院检验科，天津 300192）

Author(s):

GU Qiu¹; YUE Yingying²; NIU Wenyan¹

（1. Department of Immunology，School of Basic Medical Sciences，Tianjin Medical University，Tianjin 300070，China；2.Department of Clinical Laboratory，Tianjin First Central Hospital，Tianjin 300192，China）

关键词:

骨骼肌; 分化; 维生素D; Wnt/ β-catenin

Keywords:

skeletal muscle; differentiation; Vitamin D; Wnt/β-catenin

分类号:

R392.1

DOI:

10.20135/j.issn.1006-8147.2025.03.0243

文献标志码:

A

摘要:

目的：探讨维生素D对骨骼肌细胞分化的影响及其分子机制。方法：用不同浓度的1，25-二羟维生素D3（VD3）孵育C2C12小鼠骨骼肌细胞，选择合适的药物浓度进行后续实验。将C2C12细胞分为对照（control）组、Wnt/β-catenin信号通路激动剂（SKL2001）组、VD3组和VD3+SKL2001组。qPCR检测维生素D受体（VDR）、肌球蛋白重链（MyHC）、肌生成素（MyoG）、肌生成决定因子（MyoD）、β-连环蛋白（β-catenin）、无翅型MMTV整合位点家族成员11（Wnt11）、轴抑制蛋白2（Axin2）和Fermintin家族同源蛋2（Fermt2）mRNA水平；细胞免疫荧光检测MyHC和MyoG。倒置显微镜观察各组细胞形态。免疫印迹检测各组VDR、MyHC、MyoG、MyoD和β-catenin蛋白水平。结果：与control组相比，VD3组VDR的mRNA水平显著升高（t=11.50，P<0.001），MyHC、MyoG、MyoD、Wnt11、Axin2和Fermt2的mRNA水平显著降低（t=9.849、8.620、17.88、4.069、6.953、4.685，均P<0.01），β-catenin的mRNA水平无明显变化（t=2.657，P>0.05）；与control组相比，VD3组MyHC和MyoG阳性的肌管数量减少，细胞融合受损。显微镜观察结果显示，与control组相比，SKL2001组肌管数量增多，肌管长度增加，VD3组细胞融合受损、肌管数量减少、肌管长度缩短，与VD3组相比，VD3+SKL2001组肌管数量增多，肌管长度增加；免疫印迹结果显示，与control组相比，SKL2001组MyHC、MyoG和MyoD蛋白水平升高（F=27.91、133.6、33.25，均 P<0.05），VD3组VDR蛋白水平上升（F=118.0，P<0.001），MyHC、MyoG和MyoD蛋白水平降低（F=27.91、133.6、33.25，均 P<0.05）；与VD3组相比，VD3+ SKL2001组MyHC、MyoG和MyoD蛋白水平升高（F=27.91、133.6、33.25，均P<0.05）；与control组相比，SKL2001组胞质中β-catenin蛋白水平降低（F=64.00，P<0.05），胞核中β-catenin蛋白水平显著升高（F=32.89，P<0.05）；VD3组胞质中β-catenin蛋白水平上调（F=64.00，P<0.001），细胞核中β-catenin蛋白水平下调（F=32.89，P<0.05）；与VD3组相比，VD3+SKL2001组胞质中β-catenin蛋白水平降低（F=64.00，P<0.05），胞核中β-catenin蛋白水平显著升高（F=32.89，P<0.05）。结论：维生素D可能通过抑制Wnt/β-catenin信号通路，抑制C2C12骨骼肌细胞分化。

Abstract:

Objective： To investigate the effect of vitamin D on the differentiation of skeletal muscle cells and its potential molecular mechanism. Methods：The C2C12 mouse skeletal muscle cells were incubated with different concentrations of 1，25-dihydroxyvitamin D3 （VD3），and appropriate drug concentrations were selected for follow-up experiments. The C2C12 cells were divided into control group，Wnt/β-catenin signaling pathway agonists （SKL2001） group，VD3 group and VD3+SKL2001 group. The mRNA levels of vitamin D receptor （VDR），myosin heavy chain （MyHC），myogenin （MyoG），myogenic determing factor （MyoD），β-catenin，wingless-type MMTV integration site family member 11 （Wnt11），Axis inhibitor 2 （Axin2） and fermitin family member 2 （Fermt2） were detected by qPCR. Cellular immunofluorescence was used to detect MyHC and MyoG. The cell morphology of all groups was observed by inverted microscope. The protein levels of VDR，MyHC，MyoG，MyoD and β-catenin were detected by Western blotting. Results：Compared with con-trol group，the mRNA level of VDR in VD3 group was significantly increased （t=11.50，P<0.001），while the mRNA levels of MyHC，MyoG，MyoD，Wnt11，Axin2 and Fermt2 were decreased （t=9.849，8.620，17.88，4.069，6.953，4.685，all P<0.01）. The mRNA level of β-catenin remained stable （t=2.657，P>0.05）. Compared with control group，the numbers of MyHC and MyoG positive myotubes were decreased，and cell fusion were impaired in VD3 group. Microscopic observation showed that compared with the control group，the numbers and length of myotubes were increased in SKL2001 group，while cell fusion was impaired，the numbers and length of myotubes were decreased in VD3 group. Compared with VD3 group，the numbers and length of myotubes were increased in VD3+SKL2001 group. Western blotting showed that the protein levels of MyHC，MyoG and MyoD in SKL2001 group were significantly increased （F=27.91， 133.6，33.25， 32.89，all P<0.05） compared with control group. The protein levels of MyHC，MyoG and MyoD in VD3 group were significantly decreased （F=27.91，133.6，33.25，all P<0.05），whereas the protein level of VDR was significantly increased （F=118.0，P<0.001）. Compared with VD3 group， the protein levels of MyHC，MyoG and MyoD in VD3+SKL2001 group were significantly increased （F=27.91，133.6，33.25， all P<0.05）. Compared with control group，β-catenin protein level in cytoplasm of SKL2001 group was de-creased （F=64.00，P<0.05）， and β-catenin protein level in nucleus was significantly increased （F=32.89，P<0.05）. The level of β-catenin protein in the cytoplasm of VD3 group was increased （F=64.00，P<0.001），and the level of β-catenin protein in the nucleus was decreased （F=32.89，P<0.05） compared with control group. Compared with VD3 group，the level of β-catenin protein in cytoplasm was decreased （F=64.00，P<0.05） in VD3+SKL2001 group，and the level of β-catenin protein in nucleus was significantly increased （F=32.89，P<0.05）. Conclusion：Vitamin D may inhibit the differentiation of C2C12 skeletal muscle cell by suppressing the Wnt/β-catenin signal pathway.

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备注/Memo

备注/Memo:

基金项目: 国家自然科学基金面上项目（82270856，81870547）
作者简介: 古秋（1999-），女，硕士在读，研究方向：免疫学；
通信作者：牛文彦，E-mail：wniu@tmu.edu.cn。

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更新日期/Last Update: 2025-06-01