SRSF10对山羊肌内前体脂肪细胞分化的影响

doi:10.11843/j.issn.0366-6964.2022.06.011

摘要/Abstract

摘要： 旨在克隆山羊SRSF10基因序列，明确其生物学特性，并通过过表达和干扰手段阐明SRSF10对山羊肌内脂肪细胞分化的影响。本研究以简州大耳羊(Capra hircus)为试验对象，利用RT-PCR技术克隆山羊SRSF10基因序列，并进行生物信息学分析; 利用实时荧光定量PCR(real-time quantitative PCR，qPCR)技术研究其在成脂诱导分化不同阶段细胞中的表达水平; 转染pcDNA3.1-SRSF10过表达载体和SRSF10-siRNA至山羊肌内前体脂肪细胞并诱导分化，通过油红O和Bodipy染色法从形态学上明确其对脂滴积聚的影响; 通过qPCR方法检测脂肪细胞分化标志基因表达的变化。结果显示，获得山羊SRSF10基因序列为1 026 bp，其中CDS区552 bp，共编码183个氨基酸; SRSF10在诱导分化96 h的肌内脂肪细胞中表达量最高; 过表达和干扰山羊SRSF10分别促进和抑制了肌内脂肪细胞中的脂滴积聚; 过表达后基因SREBP1、PPARγ和C/EBPα的相对表达量极显著上调(P < 0.01)，干扰后分化标志基因SREBP1、PPARγ和C/EBPα相对表达水平极显著降低(P < 0.01)。结果表明，山羊SRSF10是肌内脂肪细胞分化的正调控作用因子，并且这种调控作用可能主要通过调节SREBP1、PPARγ和C/EBPα的表达来实现。

关键词: 山羊, SRSF10, 基因克隆, 表达特性, 肌内前体脂肪细胞, 细胞分化

Abstract: This experiment aimed to clone goat SRSF10 gene sequence, reveal its biological characteristics, and clarify the effect of SRSF10 on goat intramuscular adipocyte differentiation through overexpression and interference techniques. Taking Jianzhou big ear goat(Capra hircus)as the experimental object, the goat SRSF10 gene sequence was cloned by RT-PCR technique, along with bioinformatics analysis. The real-time quantitative PCR(qPCR)technique was used to investigate its expression level in adipogenic differentiation of cells at different stages. pcDNA3.1-SRSF10 overexpression vector and SRSF10-siRNA were transfected into goat intramuscular preadipocytes, followed by inducing differentiation. Oil red O and Bodipy staining were used to clarify the effect of SRSF10 on lipid droplet accumulation, and the expression changes of adipocyte differentiation marker genes were detected by qPCR technique. The results showed that the obtained goat SRSF10 gene sequence was 1 026 bp in length, the CDS region was 552 bp, encoding a total of 183 amino acids. The highest expression level of SRSF10 in goat was detected at 96 h in intramuscular adipocytes after inducing differentiation. Overexpression and interference of goat SRSF10 promoted and inhibited the accumulation of lipid droplets in intramuscular adipocytes, respectively. After overexpression of SRSF10, the relative expression of SREBP1, PPARγ and C/EBPα extremely significantly up-regulated(P < 0.01), and the relative expression of differentiation marker genes SREBP1, PPARγ and C/EBPα extremely significantly down-regu-lated after interference(P < 0.01). The results show that goat SRSF10 is a positive regulatory factor for the differentiation of intramuscular adipocytes, and this regulation may be achieved mainly through regulating the expression of SREBP1, PPARγ and C/EBPα.

Key words: goat, SRSF10, gene cloning, expression characteristics, intramuscular preadipocytes, cell differentiation

中图分类号:

S827.2

刘珂含, 王永, 李艳艳, 王重洋, 朱江江, 熊燕, 李安, 林亚秋. SRSF10对山羊肌内前体脂肪细胞分化的影响[J]. 畜牧兽医学报, 2022, 53(6): 1768-1778.

LIU Kehan, WANG Yong, LI Yanyan, WANG Chongyang, ZHU Jiangjiang, XIONG Yan, LI An, LIN Yaqiu. Effects of SRSF10 on the Differentiation of Intramuscular Preadipocytes in Goats[J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(6): 1768-1778.

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