miR-144-5p在不同体型双峰驼血浆外泌体中的表达及靶基因验证

doi:10.16431/j.cnki.1671-7236.2025.03.005

Abstract

Abstract: 【Objective】 The aim of this study was to explore the potential regulatory mechanisms of miR-144-5p in lipid metabolism within plasma exosomes of Bactrian camels.【Method】 The exosomes were separated from two groups of Bactrian camels with different levels of fat and lean using ultracentrifugation in this study.The morphology of the exosomes was observed by transmission electron microscopy and identified using nanoparticle tracking analysis.Total exosomal RNA was extracted for small RNA sequencing,and the differentially expressed miRNA in plasma exosomes from two groups of Bactrian camels was selected.TargetScan and miRWalk online software were used to predict candidate target gene of differentially expressed miRNA,followed by GO function and KEGG pathway enrichment analysis of the target genes using DAVID and KEGG bioinformatics systems.The wild-type vectors containing target sites and mutant expression plasmids were constructed.The transfection of miR-144-5p minicis with 293T cells was done for dual-luciferase reporter assay to validate the relationship between miR-144-5p and its candidate gene,exploring its mechanism of action in the regulation of lipid metabolism.【Result】 The results revealed that plasma exosomes from Bactrian camels exhibited cup-shaped or round morphology with a diameter of approximately 100 nm,with a particle size distribution ranging between 30 to 150 nm.A total of 40 differentially expressed miRNAs were identified between the two groups,including significant differential expression of miR-144-5p (P＜0.01).The sequence of miR-144-5p was highly conserved among different species.PGC-1α was predicted as its target gene,mainly involved in processes like cholesterol balance and lipid phosphorylation,enriched in glycerolipid metabolism,cholesterol metabolism,and atherosclerosis signaling pathways.miR-144-5p significantly inhibited the relative luciferase activity of the wild-type PGC-1α-3'-UTR vector (P＜0.01),while it had no effect on the mutant PGC-1α-3'-UTR vector compared to the negative control group at 48 hours after transfection (P＞0.05),indicating that miR-144-5p inhibits PGC-1α gene expression by binding to the gene’s seed sequence.【Conclusion】 This study revealed the expression differences of miR-144-5p in plasma exosomes of Bactrian camels with different body types and confirmed its important role in lipid metabolism by regulating PGC-1α.

Key words: Bactrian camels; exosome; miR-144-5p; lipid metabolism; target gene; dual-luciferase reporter assay system

CLC Number:

S852.2

Siriguleng, YU Wen, JIANG Xiaowei, LI Ziyi, JIN Junjian, BAI Haoyu. The Expression of miR-144-5p in Plasma Exosomes of Different Body Types of Bactrian Camels and the Verification of Target Gene[J]. China Animal Husbandry and Veterinary Medicine, 2025, 52(3): 1022-1032.

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The Expression of miR-144-5p in Plasma Exosomes of Different Body Types of Bactrian Camels and the Verification of Target Gene

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