缺氧对鸡成肌细胞肌纤维类型转化作用的机制探究

doi:10.11843/j.issn.0366-6964.2024.06.012

畜牧兽医学报 ›› 2024, Vol. 55 ›› Issue (6): 2397-2408.doi: 10.11843/j.issn.0366-6964.2024.06.012

缺氧对鸡成肌细胞肌纤维类型转化作用的机制探究

谢兵红¹^,²(), 刘一帆²^,³, 薛夫光¹, 单艳菊², 屠云洁², 姬改革², 巨晓军², 束婧婷²^,*(), 吴红翔¹^,*()

1. 江西农业大学动物科学技术学院，动物健康安全生产南昌重点实验室，南昌 330000
2. 江苏省家禽科学研究所，江苏省家禽遗传育种重点实验室，扬州 225125
3. 和盛食品集团有限公司，泰州 225501

收稿日期:2024-01-02 出版日期:2024-06-23 发布日期:2024-06-28
通讯作者: 束婧婷,吴红翔 E-mail:binghong1@qq.com;shujingting@163.com;2014035038@qq.com
作者简介:谢兵红(1997-)，女，江西丰城人，硕士生，主要从事家禽肉品质调控机理研究，E-mail: binghong1@qq.com
基金资助:
lanmu(bianhao)

The Mechanism of Effect of Hypoxia on Myofiber Type Transformation in Chicken Myoblasts

Binghong XIE¹^,²(), Yifan LIU²^,³, Fuguang XUE¹, Yanju SHAN², Yunjie TU², Gaige JI², Xiaojun JU², Jingting SHU²^,*(), Hongxiang WU¹^,*()

1. Nanchang Key Laboratory of Animal Health and Safety Production, College of Animal Science and Technology, Jiangxi Agricultural University, Nanchang 330000, China
2. Key Laboratory for Poultry Genetics and Breeding of Jiangsu Province, Jiangsu Institute of Poultry Science, Yangzhou 225125, China
3. Hesheng Food Group Co., Ltd, Taizhou 225501, China

Received:2024-01-02 Online:2024-06-23 Published:2024-06-28
Contact: Jingting SHU, Hongxiang WU E-mail:binghong1@qq.com;shujingting@163.com;2014035038@qq.com

摘要/Abstract

摘要：

旨在利用氯化钴(CoCl₂)模拟肌细胞缺氧，并结合转录组测序分析缺氧对肌纤维类型及相关基因的影响。本试验以鸡原代成肌细胞为研究对象，使用不同浓度氯化钴(0、50、100、200和400 μmol ·L^-1)处理细胞，每组设3个重复孔。通过检测缺氧标志基因表达和观察细胞形态筛选氯化钴浓度，确定最适宜浓度用于建模。随后，将收集的细胞通过Illumina测序检测缺氧组和对照组鸡成肌细胞的转录表达谱，并分析差异基因和富集到的信号通路。结果表明，200 μmol ·L^-1氯化钴最适合构建鸡胚成肌细胞缺氧模型，通过转录组测序发现，在24和48 h时，缺氧组和对照组分别筛选出1 474个和2 028个差异表达基因，其中有959个差异基因在两个时间点共有。GO和KEGG分析发现，在缺氧环境下，鸡成肌细胞的信号受体活性调节、质膜组成成分、神经活性配体受体作用、PI3K-AKT、肌肉收缩等信号通路关键因子发生明显表达变化。测序和qRT-PCR结果一致显示，缺氧处理后慢肌纤维特异性基因如MYH7B、CSRP2显著上调，而快肌纤维特异性基因如MYH1F、MYH1D、TNNI2、SOX6显著下调，表明缺氧能够诱导鸡成肌细胞快肌纤维向慢肌纤维转换。本研究初步揭示了缺氧对鸡成肌细胞肌纤维类型转换的机制，并提供相关基因表达数据库，为未来家禽肉质和哺乳动物缺氧研究提供了重要参考。

关键词: 缺氧, 鸡成肌细胞, 肌纤维类型转换

Abstract:

The cobalt chloride (CoCl₂) was utilized in this study to simulate hypoxia in muscle cells and employed transcriptomic sequencing to analyze the effects of hypoxia on muscle fiber type and genes expression that related to muscle fiber types. Primary chicken myoblasts were treated with different concentrations of CoCl₂ (0, 50, 100, 200 and 400 μmol ·L^-1), with 3 repeat wells in each group. The CoCl₂ concentration was screened by detecting the expression of hypoxia marker genes and observing cell morphology to determine the most suitable concentration for modeling. Subsequently, the collected cell was used to detect the gene expression profile of chicken myoblast in hypoxia treatment and control treatment by Illumina technology and assess differential gene expression and enriched signaling pathways between treatments. The results indicated that 200 μmol ·L^-1 CoCl₂ was the most suitable concentration for constructing the hypoxic model in chicken embryonic myogenic cells. Transcriptomic results showed a total of 1 474 and 2 028 differentially expressed genes in the hypoxia treatment compared to the control group at 24 and 48 h, respectively. Interestingly, 959 genes showed differential expression in both time points.GO and KEGG analyses revealed significant alterations in key factors related to signaling pathways such as regulation of signaling receptor activity, integral component of plasma membrane, neuroactive ligand-receptor interaction, PI3K-AKT signaling pathway and muscle contraction in chicken muscle cells under hypoxic conditions. Consistent findings from sequencing and real-time quantitative polymerase chain reaction (qRT-PCR) demonstrated that slow muscle fiber-specific genes such as MYH7B and CSRP2 were significantly upregulated after hypoxia treatment, while fast muscle fiber-specific genes including MYH1F, MYH1D, TNNI2, and SOX6 were significantly downregulated, which indicated a transition of chicken muscle cells from fast to slow muscle fibers was induced by hypoxia. This study provides preliminary insights into the mechanisms underlying the hypoxia-induced transition of muscle fiber types in chicken myogenic cells and offers a valuable gene expression database for future research on poultry meat quality and hypoxia-related studies in mammals.

Key words: hypoxia, chicken myoblasts, muscle fiber type transformation

中图分类号:

S831

谢兵红, 刘一帆, 薛夫光, 单艳菊, 屠云洁, 姬改革, 巨晓军, 束婧婷, 吴红翔. 缺氧对鸡成肌细胞肌纤维类型转化作用的机制探究[J]. 畜牧兽医学报, 2024, 55(6): 2397-2408.

Binghong XIE, Yifan LIU, Fuguang XUE, Yanju SHAN, Yunjie TU, Gaige JI, Xiaojun JU, Jingting SHU, Hongxiang WU. The Mechanism of Effect of Hypoxia on Myofiber Type Transformation in Chicken Myoblasts[J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(6): 2397-2408.

图/表 7

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基因 Gene	引物序列(5′→3′) Primer sequence	产物长度/bp Product length	退火温度/℃ Tm
MYH1F	F：CTTGTGGATGGTTGTTCGCA； R：CCTCCTGTTCCAGCACAAAC	191	56
MYH7B	F：CAGCAGCCATTCCAGATGAC； R：TCTTGGCCAGACGTTCATCT	171	52
HIF1A	F：CGTGTAAAGGCGTGCAAAAC； R：CTGTTGCCTTGTATGGGAGC	170	60
VEGFA	F：AGTCTACGAACGCAGCTTCT； R：ACAGGGACACATTCTAGGCC	156	60
CSRP2	F：ATGGACAGGGGAGAGAGACT； R：TTTCTCCGCCGCATAAACAG	153	60
CSRP3	F：CCTCCACACCAACTAACCCT； R：TTCCCACAAATAGCACAGCG	154	60
MYBPC2	F：CCTCATCATCAACGAAGCGG； R：TGGATCCTCTTGCTGGGTTT	240	60
MYH1D	F：CACTTCTCCCTGGTGCACTA； R：AGACCCCTTCTTCTTGCCTC	207	60
MYH1E	F：AGCAAGTGGACGATCTGGAA； R：TTTGCTCTGGATCTGGCTGA	194	60
PRKAG3	F：CACAAGCGCATCCTCAAGTT； R：TGACCGGCAGCATTAACAAC	203	60
SOX6	F：GCTTTCCCTGACATGCACAA； R：AGGTACGTTTTGGTCGAGGT	169	60
TNNI2	F：GCAAAACTACCTGGCAGAGC； R：CCCTCAGGTCAAACAGCTTC	191	60
VEGFC	F：CGAACAAAACGCTGTGATGC； R：TTGTCTCTGAAAGCCCCACA	214	60
MYOG	F：GAGGGGCTTTGGAGGAGAAG； R：TTCACCTTCTTCAGCCTCCG	154	60
PGM1	F：AGTTTTGGTACTGGCTCCGA； R：CACAAAGGAGCGGTCAAACA	234	60
PPARGC1A	F：AAAGACGTCCCTGCTCTGAA； R：TGGCTTGCAGGTGATTTGTC	153	60

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缺氧对鸡成肌细胞肌纤维类型转化作用的机制探究

The Mechanism of Effect of Hypoxia on Myofiber Type Transformation in Chicken Myoblasts

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