Single-nucleus RNA sequencing and lipidomics reveal characteristics of transcriptional and lipid composition in porcine longissimus dorsi muscle.

BACKGROUND: Global per capita meat consumption continues to rise, especially pork. Meat quality is influenced by the content of intramuscular fat (IMF) as a key factor. The longissimus dorsi muscle of Dahe pigs (DHM, IMF: 7.98% ± 1.96%) and Dahe black pigs (DHBM, IMF: 3.30% ± 0.64%) was studied to explore cellular heterogeneity and differentially expressed genes (DEGs) associated with IMF deposition using single-nucleus RNA sequencing (snRNA-seq). The lipid composition was then analyzed using non-targeted lipidomics. RESULTS: A total of seven cell subpopulations were identified, including myocytes, fibroblast/fibro/adipogenic progenitors (FAPs), satellite cells, endothelial cells, macrophages, pericytes, and adipocytes. Among them, FAPs and adipocytes were more focused because they could be associated with lipid deposition. 1623 DEGs in the FAPs subpopulation of DHBM were up-regulated compared with DHM, while 1535 were down-regulated. These DEGs enriched in the glycolysis/gluconeogenesis pathway. 109 DEGs were up-regulated and 806 were down-regulated in the adipocyte subpopulation of DHBM compared with DHM, which were mainly enriched in the PPAR signaling pathway and fatty acid (FA) biosynthesis. The expression level of PPARG, ABP4, LEP, and ACSL1 genes in DHM was higher than that in DHBM. Lipidomics reveals porcine lipid composition characteristics of muscle tissue. A total of 41 lipid classes and 2699 lipid species were identified in DHM and DHBM groups. The top ten relative peak areas of lipid classes in DHM and DHBM were triglyceride (TG), phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylserine (PS), diglyceride (DG), cardiolipin (CL), ceramides (Cer), Simple Glc series (Hex1Cer), sphingomyelin (phSM), and phosphatidylinositol (PI). The relative peak areas of 35 lipid species in DHM were lower than DHBM, and 28 lipid species that were higher. There was a significant increase in the TG fatty acyl chains C6:0, C17:0, and C11:4, and a significant decrease in C16:0, C18:1, C18:2, and C22:4 in DHBM (p < 0.05). CONCLUSIONS: C16:0 FA may downregulate the expression level of PPARG gene, which leads to the downregulation of fat metabolism-related genes such as ACSL, PLIN2, and FABP4 in DHBM compared with DHM. This may be the reason that the lipid deposition ability of Dahe pigs is stronger than that of Dahe black pigs, which need further investigation.

Panax notoginseng stems and leaves affect microbial community and function in cecum of duzang pigs.

Panax notoginseng is a Chinese medicine with a long history in which stems and leaves are the wastes of processing Panax notoginseng and have not been effectively utilized. The effects of diets containing Panax notoginseng stems and leaves on the cecal short-chain fatty acid (SCFA) concentration and microbiome of independent pigs were studied. Diets containing Panax notoginseng stems and leaves did not affect the concentration of SCFA in the cecal contents of Duzang pigs but affected the microbial composition and diversity. Firmicutes, Proteobacteria, and Bacteroidetes dominate in the cecal of Duzang pigs. Feeding Duzang pigs with a 10% Panax notoginseng stems and leaves diet increases the abundance of Lactobacillus, Christensenellaceae R-7 group, and Akkermansia in the cecal. We found 14 genera positively associated with acetate, and they were Lactobacillus, Ruminococcaceae UCG 005, Ruminiclostridium 6; Escherichia Shigella and Family XIII AD3011 group showed negative correlations. Solobacterium, Desulfovibrio, and Erysipelatoclostridium were positively associated with propionate. Campylobacter, Clostridium sensu stricto 11, and Angelakisella were positively associated with butyrate. In conclusion, Panax notoginseng stems and leaves could affect the cecal microbial community and functional composition of Duzang pigs. Panax notoginseng stems and leaves reduce the enrichment of lipopolysaccharide biosynthetic pathway of the cecal microbiome, which may have a positive effect on intestinal health. The higher abundance of GH25 family in Duzang pig's cecal microbiome of fed Panax notoginseng stems and leaves diet. This increase may be the reason for the microbial diversity decrease.

Comparison of differentially expressed genes in longissimus dorsi muscle of Diannan small ears, Wujin and landrace pigs using RNA-seq.

Introduction: Pig growth is an important economic trait that involves the co-regulation of multiple genes and related signaling pathways. High-throughput sequencing has become a powerful technology for establishing the transcriptome profiles and can be used to screen genome-wide differentially expressed genes (DEGs). In order to elucidate the molecular mechanism underlying muscle growth, this study adopted RNA sequencing (RNA-seq) to identify and compare DEGs at the genetic level in the longissimus dorsi muscle (LDM) between two indigenous Chinese pig breeds (Diannan small ears [DSE] pig and Wujin pig [WJ]) and one introduced pig breed (Landrace pig [LP]). Methods: Animals under study were from two Chinese indigenous pig breeds (DSE pig, n = 3; WJ pig, n = 3) and one introduced pig breed (LP, n = 3) were used for RNA sequencing (RNA-seq) to identify and compare the expression levels of DEGs in the LDM. Then, functional annotation, Gene Ontology (GO) enrichment analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis, and Protein-Protein Interaction (PPI) network analysis were performed on these DEGs. Then, functional annotation, Gene Ontology (GO) enrichment analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis, and Protein-Protein Interaction (PPI) network analysis were performed on these DEGs. Results: The results revealed that for the DSE, WJ, and LP libraries, more than 66, 65, and 71 million clean reads were generated by transcriptome sequencing, respectively. A total of 11,213 genes were identified in the LDM tissue of these pig breeds, of which 7,127 were co-expressed in the muscle tissue of the three samples. In total, 441 and 339 DEGs were identified between DSE vs. WJ and LP vs. DSE in the study, with 254, 193 up-regulated genes and 187, 193 down-regulated genes in DSE compared to WJ and LP. GO analysis and KEGG signaling pathway analysis showed that DEGs are significantly related to contractile fiber, sarcolemma, and dystrophin-associated glycoprotein complex, myofibril, sarcolemma, and myosin II complex, Glycolysis/Gluconeogenesis, Propanoate metabolism, and Pyruvate metabolism, etc. In combination with functional annotation of DEGs, key genes such as ENO3 and JUN were identified by PPI network analysis. Discussion: In conclusion, the present study revealed key genes including DES, FLNC, PSMD1, PSMD6, PSME4, PSMB4, RPL11, RPL13A, ROS23, RPS29, MYH1, MYL9, MYL12B, TPM1, TPM4, ENO3, PGK1, PKM2, GPI, and the unannotated new gene ENSSSCG00000020769 and related signaling pathways that influence the difference in muscle growth and could provide a theoretical basis for improving pig muscle growth traits in the future.