藏猪脂肪细胞产热分子特征研究

doi:10.16431/j.cnki.1671-7236.2025.10.025

摘要/Abstract

摘要： 【目的】探究藏猪白色脂肪细胞(white adipocytes)和米色脂肪细胞(beige adipocytes)在产热激活前后的基因表达差异及其相关信号通路，为脂肪产热机制及仔猪抗寒研究提供理论依据。【方法】从藏猪脂肪组织中分离基质血管组分(SVF)细胞，分别诱导分化为白色脂肪细胞和米色脂肪细胞。用5 μmol/L毛喉素(FSK)处理24 h激活产热，试验共设置5组：未经处理的SVF细胞组(C)、白色脂肪细胞组(W)、FSK处理的白色脂肪细胞组(WF)、米色脂肪细胞组(B)、FSK处理的米色脂肪细胞组(BF)。采用Illumina HiSeq平台进行转录组测序，通过DESeq2筛选差异表达基因。结合GO功能和KEGG通路富集分析差异表达基因的功能及代谢通路，通过实时荧光定量PCR验证关键基因表达。整合藏猪体内冷刺激数据与米色脂肪细胞体外产热模型(BF vs B)，筛选共表达的产热相关基因和转录因子。【结果】转录组测序结果显示，FSK处理显著改变了白色脂肪细胞和米色脂肪细胞的基因表达谱。主成分分析(PCA)和热图显示，FSK处理前后细胞基因表达模式明显分离，其中米色脂肪细胞对FSK的响应更强烈。米色脂肪细胞经FSK处理后共有1 514个基因上调表达、1 544个基因下调表达；白色脂肪细胞经FSK处理后共有815个基因上调表达、1 096个基因下调表达。GO和KEGG富集结果显示，米色脂肪细胞的上调表达基因显著富集于脂代谢(如脂肪酸氧化)、cAMP信号通路和免疫相关通路，且富集分值高于白色脂肪细胞。与处理前相比，FSK处理后两种脂肪细胞共有485个上调和525个下调共同差异表达基因，上调基因富集于cAMP、AMPK和PPAR等产热相关通路，下调基因富集于脂肪合成通路(如不饱和脂肪酸生物合成)。实时荧光定量PCR结果显示，FSK处理后白色脂肪细胞中KLF11和PDK4基因表达量均极显著高于处理前(P＜0.01)。整合藏猪体内冷刺激脂肪组织与米色脂肪细胞转录组测序数据，筛选出81个体内外上调的共同表达差异基因，包括PPARGC1A、NCOA3等7个转录因子，这些转录因子与线粒体生物合成、脂质代谢重编程等过程相关，提示了其在产热调控中的潜在作用。【结论】本试验通过分析FSK处理藏猪脂肪细胞的转录组数据，揭示了脂肪细胞产热过程中的关键信号通路和基因。在体外转录组数据中筛选出了KLF11、PDK4等潜在调控产热的基因，结合体内转录组数据共筛选出7个潜在调控脂肪组织产热的转录因子。研究结果为改良猪的抗寒和脂肪沉积性状提供了潜在靶点。

关键词: 藏猪; 米色脂肪细胞; 白色脂肪细胞; 产热; 转录组测序

Abstract: 【Objective】 This study aimed to investigate the gene expression differences and associated signaling pathways in white adipocytes and beige adipocytes of Tibetan pigs before and after thermogenic activation,providing a theoretical basis for understanding adipocyte thermogenesis mechanisms and improving cold resistance in piglets.【Method】 Stromal vascular fraction (SVF) cells were isolated from the adipose tissue of Tibetan pigs and induced to differentiate into white adipocytes and beige adipocytes.Thermogenesis was activated by treating with 5 μmol/L forskolin (FSK) for 24 hours.The experiment comprised five groups:Untreated SVF cells group (C), white adipocytes group (W),FSK-treated white adipocytes group (WF), beige adipocytes group (B),and FSK-treated beige adipocytes group (BF).RNA-Seq was performed using the Illumina HiSeq platform.Differentially expressed genes were identified using DESeq2.Functional and metabolic pathway analyses of differentially expressed genes were conducted using GO and KEGG,and the expression of key genes was validated by Real-time quantitative PCR.Additionally,data from cold-stimulated Tibetan pigs were integrated with the in vitro beige adipocyte thermogenesis model (BF vs B) to identify co-expressed thermogenesis-related genes and transcription factors.【Result】 The transcriptome sequencing results demonstrated that FSK treatment significantly altered the gene expression profiles of both white adipocytes and beige adipocytes.Principal component analysis (PCA) and heatmaps revealed a marked separation in gene expression patterns before and after FSK treatment.Among them,beige adipocytes exhibited a stronger response to FSK treatment compared to white adipocytes.After treatment with FSK on beige adipocytes,a total of 1 514 genes were up-regulated and 1 544 genes were down-regulated.After treatment with FSK,a total of 815 genes were up-regulated and 1 096 genes were down-regulated in white adipocytes.GO and KEGG analyses indicated that the upregulated genes in beige adipocytes were significantly enriched in lipid metabolism pathways (fatty acid oxidation),cAMP signaling pathways,and immune-related pathways,and the enrichment score was higher than that of white adipocytes.Compared with the results before treatment after FSK treatment,there were a total of 485 up-regulated and 525 down-regulated differentially expressed genes in the two types of adipocytes.The up-regulated genes were enriched in thermogenic pathways such as cAMP,AMPK and PPAR,while the down-regulated genes were enriched in adiposynthetic pathways (such as unsaturated fatty acid biosynthesis).The results of Real-time quantitative PCR showed that the expression levels of KLF11 and PDK4 genes in white adipocytes after FSK treatment were extremely significantly upregulated (P＜0.01).By integrating the transcriptome sequencing data of cold-stimulated adipose tissue in Tibetan pigs and beige adipocytes,81 up-regulated co-expressed differentially expressed gene both in vivo and in vitro . There were 7 transcription factors such as PPARGC1A and NCOA3，and these transcription factors were related to processes such as mitochondrial biosynthesis and lipid metabolism reprogramming,suggesting their potential role in thermogenesis regulation.【Conclusion】 In this experiment,the transcriptome data of Tibetan pigs adipocytes were processed by FSK,revealing the key signaling pathways and genes in the thermogenesis process of adipocytes.Genes such as KLF11 and PDK4 that potentially regulated thermogenesis were screened out from the in vitro transcriptome data.Combined with the in vivo transcriptome data,7 transcription factors that potentially regulated thermogenesis in adipose tissue were screened out.The results provided potential targets for improving the cold resistance and fat deposition characteristics of pigs.

Key words: Tibetan pigs; beige adipocytes; white adipocytes; thermogenesis; RNA-Seq

中图分类号:

S813.3

韩煦, 彭渝, 张立兰, 刘田侠, 李红强, 陶聪. 藏猪脂肪细胞产热分子特征研究[J]. 中国畜牧兽医, 2025, 52(10): 4796-4808.

HAN Xu, PENG Yu, ZHANG Lilan, LIU Tianxia, LI Hongqiang, TAO Cong. Study on the Molecular Characteristics of Thermogenesis in Tibetan Pig Adipocytes[J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(10): 4796-4808.

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