Functional analysis of lncRNAs in lipid metabolism of fat and lean line broiler embryonic livers

Abstract

As the primary site of lipogenesis in birds, the liver orchestrates avian lipid metabolism and is pivotal for fat accumulation in chickens. Lipid metabolism during the broiler embryo stage may significantly affect post-hatch growth performance, yet research on this subject remains limited. While long non-coding RNAs (lncRNAs) have been found to regulate liver lipid metabolism in post-hatch chickens, their functions during the embryonic stage remains unclear. This study revealed that, compared to lean line broiler embryos, fat line broiler embryos showed upregulated gene expression related to de novo fatty acid synthesis, glycerol-3-phosphate synthesis, triglyceride synthesis, and the degradation of both fatty acids and cholesterol. Through transcriptome analysis and functional validation, lncRNA1926 and lncRNA3223 were identified as key regulators of lipid metabolism in broiler embryo livers. Knocking down either of lncRNA1926 or lncRNA3223 significantly reduced lipid droplet accumulation, triglyceride levels, and total cholesterol levels in primary hepatocytes of broiler embryos. Our findings demonstrate distinct lipid metabolic gene expression profiles between fat and lean line broiler embryo livers, and highlight lncRNA1926 and lncRNA3223 are key regulators of lipid metabolism during the embryonic stage. This study enhances the scientific understanding of lipid metabolism regulation in chicken livers and provides a theoretical foundation for genetically improving abdominal fat traits in broilers.

Keywords: Broiler; Embryonic stage; Lipid metabolism; Liver; LncRNAs.

Fig. 1

TG and TCHO content in embryonic liver of fat and lean line broilers. (A) TG content (B) TCHO content. Note: Different capital letters indicate significant differences between embryo stages in the lean line. Different lowercase letters indicate significant differences between embryo stages in the fat line. *P < 0.05, ***P < 0.001; n = 3.

Fig. 2

Expression of genes related to the synthesis, transport, and degradation of TG in embryonic liver of fat and lean line broilers. Note: *P < 0.05, ***P < 0.001; n = 3.

Fig. 3

RT-qPCR validation of lncRNAs. (A, C, E, G, I) RT-qPCR Validation Results (B, D, F, H, J) Sequencing Results. Note: *P < 0.05, **P < 0.01; n = 3.

Fig. 4

Principal component and clustering analysis of 24 samples. (A) PCA analysis of all samples (B) Cluster analysis of all samples.

Fig. 5

Expression of lncRNA1926 and lncRNA3223 in two types of hepatocytes (A) Expression of lncRNA1926 in two types of hepatocytes (B) Expression of lncRNA3223 in two types of hepatocytes. Note: *P < 0.05, **P < 0.01; n = 3.

Fig. 6

Verification of interference effects of lncRNA1926 and lncRNA3223 Smart Silence (A) Verification of interference effect of lncRNA1926 interference fragments (B) Verification of interference effect of lncRNA3223 interference fragments. Note: * P < 0.05, **P < 0.01; n = 3.

Fig. 7

Effects of lncRNA1926 knockdown on lipid droplet deposition, TG and TCHO content in primary hepatocytes of fat line broiler embryos (A) Primary hepatocytes of fat line broiler embryos were stained and extracted colorimetrically; Ruler: 50 μm (B) Effect of knockdown lncRNA1926 on TG content in primary hepatocytes of fat line broiler embryos (C) Effect of knockdown lncRNA1926 on TCHO content in primary hepatocytes of fat line broiler embryos. Note: *P < 0.05, **P < 0.01; n = 3.

Fig. 8

Effect of lncRNA1926 knockdown on the expression of lipid metabolism-related genes in primary hepatocytes of fat line broiler embryos. Note: *P < 0.05, **P < 0.01; n = 3.

Fig. 9

Effects of lncRNA3223 knockdown on lipid droplet deposition, TG and TCHO content in primary hepatocytes of fat line broiler embryo (A) Primary hepatocytes of fat line broiler embryos were stained and extracted colorimetrically; Ruler: 50 μm (B) Effect of knockdown lncRNA3223 on triglyceride content in primary hepatocytes of fat line broiler embryos (C) Effect of knockdown lncRNA3223 on total cholesterol content in primary hepatocytes of fat line broiler embryos. Note: *P < 0.05, **P < 0.01; n = 3.

Fig. 10

Effect of lncRNA3223 knockdown on the expression of lipid metabolism-related genes in primary hepatocytes of fat line broiler embryos. Note: *P < 0.05, **P < 0.01; n = 3.