【作者】 刘哲；

【导师】 于太永；

【作者基本信息】 西北农林科技大学 ， 动物遗传育种与繁殖， 2025， 硕士

【摘要】 随着我国人民生活水平不断提高,对猪肉品质的要求也越来越高。国外引进品种生长速度快、饲料转化率高,但其肉品质却不能满足人们日益增长的品质要求。随着对优质肉类的需求持续上升,改善肉质已成为一个重要的研究重点。肌内脂肪(Intramuscular fat,IMF)含量是评估肉质的关键指标,在提高肉的风味、嫩度和多汁性方面起着关键作用。有研究表明,纤维脂肪生成祖细胞(Fibro-adipogenic progenitors,FAPs)是产生肌内脂肪的重要来源。然而,FAPs细胞群对肌内脂肪沉积的调控机制仍不甚明确。传统的bulk RNA-seq往往是从整个组织或细胞群体的水平进行研究,对细胞异质性问题不能很好的解释,而单核RNA测序技术可以帮助我们从单个细胞的维度来探究骨骼肌细胞群的组成差异。因此,本研究以中国地方猪种汉江黑猪(高IMF含量)与汉中白猪(低IMF含量)为研究对象,利用单核RNA测序和代谢组学等技术,解析其在肉品质、肌细胞群和FAPs组成方面的差异及其潜在机制。主要研究结果如下:(1)汉江黑猪与汉中白猪肉品质的比较分析:本研究探究了汉江黑猪与汉中白猪在肉质性状、氨基酸代谢物组成和脂肪酸代谢物组成方面的差异。结果表明:在肉质性状方面,汉江黑猪的肌内脂肪显著高于汉中白猪,其分别为4.24±1.33%、2.42±0.74%;在氨基酸代谢物组成方面,汉江黑猪中含有更多有益健康的氨基酸及其衍生物,包括甘氨酸、5-羟基赖氨酸和L-半胱氨酸磺酸等;在脂肪酸代谢物组成方面,汉江黑猪中含有较多的多不饱和脂肪酸,如C18:1(n-9)、C18:1(n-12)T、C18:1(n-9)T、C18:2(n-6)T等。总之,相比于汉中白猪,汉江黑猪的肉质更鲜嫩、风味更浓郁,且营养价值更高。(2)猪骨骼肌单细胞转录图谱的构建:本研究构建了汉江黑猪与汉中白猪背最长肌单细胞图谱,旨在进一步探究其细胞群组成差异对肉品质的影响。研究表明,汉江黑猪中FAPs和脂肪细胞比例更高,而肌纤维比例较低。功能富集分析发现,汉江黑猪中与脂肪酸合成、脂质储存相关的通路显著富集,进一步说明汉江黑猪的肌内脂肪沉积能力强于汉中白猪。(3)影响猪肌内脂肪沉积关键基因的挖掘:通过探究IMF沉积相关的细胞亚群,发现FAPs中的细胞亚群LPL~+FAPs(LPL~+,COL4A1~+),其可能是影响猪肌内脂肪沉积的关键细胞亚群;在LPL~+FAPs中,利用关键节点基因和差异基因等共定位发现,HMCN2、COL1A1和NRXN1可能是影响IMF的关键基因。(4)猪骨骼肌细胞亚群间通讯机制的探究:通过探究汉江黑猪与汉中白猪骨骼肌细胞间的互作关系,发现汉江黑猪中存在更多的通信信号途径和更强大的通信信号强度。此外,汉江黑猪的肌源性谱系细胞群比汉中白猪更具增殖能力,如PCDH、NCAM、EGF和ADGRL等与细胞粘附,增殖和分化相关通路的表达高于汉中白猪。在汉江黑猪中FAPs细胞通过NRXN1与DAG1相互作用调节脂肪细胞的功能,而这一特定的互作模式在汉中白猪中并未观察到,这可能是影响肌内脂肪沉积的关键分子机制。综上所述,本研究探究了汉江黑猪与汉中白猪在肉品质方面的差异,构建了猪背最长肌的单细胞图谱,并发现了可能影响猪肌内脂肪沉积的关键细胞亚群和关键基因。本研究为猪IMF沉积的调控机制提供了新见解,为猪肉品质的遗传改良提供了新靶标。更多还原

【Abstract】 With the continuous improvement of living standards,the demand for high-quality meat is increasing.Foreign pig breeds,which are characterized by rapid growth and high feed conversion rates,fail to meet the growing requirements for meat quality.As the demand for high-quality meat rises,improving meat quality has become a significant research focus.Intramuscular fat(IMF)content is a key indicator for assessing meat quality and plays a crucial role in enhancing flavor,tenderness,and juiciness.Research has shown that fibro-adipogenic progenitors(FAPs)are an important source of intramuscular fat deposition.However,the regulatory mechanisms of FAPs in IMF deposition remain unclear.Bulk RNA-seq,which is conducted at the level of whole tissues or cell populations,cannot effectively address cellular heterogeneity.In contrast,single-nucleus RNA sequencing could help us investigate the compositional differences of skeletal muscle cell populations at the single-cell level.Therefore,this study used Chinese local pig breeds Hanjiang Black Pig(HJB)(high IMF content)and Hanzhong White Pig(HZW)(low IMF content)as research objects,and used sn RNA-seq and metabolomics techniques,the differences in meat quality,muscle cell populations and FAPs composition and their potential mechanisms were analyzed.The main research results are as follows:(1)Comparative analysis of the quality of Hanjiang black pig and Hanzhong white pig:this study explored the differences in meat quality,amino acid metabolite composition,and fatty acid metabolite composition between HJB pigs and HZW pigs.The results showed that the intramuscular fat of HJB pigs was significantly higher than that of HZW pigs,which were 4.24±1.33%and 2.42±0.74%respectively;HJB pigs contained more health-promoting amino acids and their derivatives,including Glycylglycine,5-Hydroxylysine,and L-Cysteinesulfinic acid;HJB pigs contained more polyunsaturated fatty acids,such as C18:1(n-9),C18:1(n-12)T,C18:1(n-9)T,C18:2(n-6)T etc.These differences in composition give HJB meat a unique flavor and more comprehensive nutritional components.(2)Construction of single-cell atlas of pig skeletal muscle:this study constructed single-cell atlas of the longissimus dorsi muscle of HJB pigs and HZW pigs to further explore the impact of differences in their cell population composition on meat quality.The study showed that the proportion of FAPs and adipocytes in HJB pigs was higher,while the proportion of muscle fibers was lower.Functional enrichment analysis found that pathways related to fatty acid synthesis and lipid storage were significantly enriched in HJB pigs,further indicating that the ability of HJB pigs to deposit intramuscular fat is stronger than that of HZW pigs.(3)Identification of key genes affecting intramuscular fat deposition in pigs:by exploring the cell subpopulations related to IMF deposition,we found that there is a cell subpopulation LPL~+FAPs(LPL~+,COL4A1~+)in FAPs,which may be the key cell subpopulation affecting pig intramuscular fat deposition;in LPL~+FAPs,using co-localization of key node genes and differentially expressed genes,we found that HMCN2,COL1A1 and NRXN1 may be the key genes affecting IMF.(4)Communication between pig skeletal muscle cell subpopulations:by exploring the communication relationship between skeletal muscle cells of different pig breeds,it was found that there were more communication signal pathways and stronger communication signal intensity in HJB pigs.In addition,the myogenic lineage of the HJB group was more proliferative than that of the HZW group.For example,the expression of pathways related to cell adhesion,proliferation and differentiation,such as PCDH,NCAM,EGF and ADGRL,was higher than that of the HZW.Ligand-receptor interaction analysis further showed that FAPs in HJB pigs regulated the function of adipocytes through the interaction between NRXN1 and DAG1,while this specific interaction pattern was not observed in HZW pigs,which may be the key molecular mechanism affecting IMF deposition.In addition,molecular and cellular verification revealed that NRXN1 has the function of inhibiting the proliferation and differentiation of porcine intramuscular adipocytes.This study constructed a single-cell map of the longissimus dorsi muscle of pigs and discovered key genes that may affect pig intramuscular fat deposition.This study provides new insights into the regulatory mechanism of IMF deposition in pigs and offers new targets for genetic improvement of meat quality.更多还原