NR1H3基因调控猪前体脂肪细胞分化的研究

doi:10.11843/j.issn.0366-6964.2022.07.007

摘要/Abstract

摘要： 旨在探讨NR1H3基因在猪脂肪组织中的发育性表达规律及对猪前体脂肪细胞成脂分化的影响，以确定其在脂肪沉积过程中的主要功能。本研究采用qRT-PCR方法检测30、90、240日龄马身猪皮下脂肪组织中NR1H3的发育性表达规律；采集5日龄杜长大仔猪背部脂肪组织，分离猪前体脂肪细胞；通过细胞免疫荧光技术检测细胞中Adiponectin含量以鉴定细胞的纯度；构建猪NR1H3基因的过表达载体，设计NR1H3 siRNA序列，分别转染分离得到的猪前体脂肪细胞，采用qRT-PCR、Western blot和油红O染色等方法检测过表达和干扰效率及它们对成脂分化关键基因表达的影响。结果表明，马身猪皮下脂肪组织中NR1H3的表达量随日龄增加呈上升趋势，30日龄时表达量最低，240日龄时表达量最高，差异极显著(P<0.01)。与对照组相比，猪前体脂肪细胞中过表达NR1H3，脂肪细胞的脂滴数明显增多，下游靶标SREBP-1c和ChREBP的表达量极显著提高(P<0.01)，且成脂关键基因FAS、C/EBPβ、PPARγ和FABP4的mRNA表达量极显著提高(P<0.01)，促进成脂过程；相反，干扰NR1H3基因，脂滴数明显减少，下游靶标及成脂关键基因的mRNA表达量极显著下调(P<0.01)，抑制成脂过程。本研究表明，NR1H3基因是猪前体脂肪细胞成脂分化的正调节剂，通过影响其下游靶标SREBP-1c和ChREBP的表达而影响成脂分化，研究结果对阐明猪脂肪沉积的分子机理、改善肉质品质有重要意义。

关键词: 猪, NR1H3, 猪前体脂肪细胞, 成脂分化

Abstract: The purpose of this study was to investigate the developmental expression pattern of NR1H3 gene in subcutaneous adipose tissues of pigs at different ages and the effects of NR1H3 gene on adipogenic differentiation of porcine preadipocytes, and determine its primary function in fat deposition. qRT-PCR was used to detect the expression of NR1H3 in subcutaneous adipose tissues of Mashen pigs aged 30, 90 and 240 days. Porcine preadipocytes were isolated from dorsal subcutaneous adipose tissues of 5-day-old DLY (Duroc×(Landrace×Yorksire)) piglets. Cellular immunofluorescence technique was used to detect the Adiponectin protein content in the cells to identify the purity of preadipocytes. The over-expression vector of porcine NR1H3 gene was constructed and the NR1H3 siRNA sequences were designed, and then both of them were transfected into porcine preadipocytes. The overexpression and interference efficiency and their effects on expression of key genes related to adipogenic differentiation were investigated by qRT-PCR, Western blot and Oil red O staining methods. The results showed that the expression of NR1H3 in subcutaneous adipose tissues increased as the age increased, the expression level was the highest at 240 days of age and the lowest at 30 days of age, with extremely significant difference between them (P<0.01). Compared with negative control (NC), over-expression (OE) of NR1H3 significantly increased the number of lipid droplets in adipocytes, the expression levels of downstream targets SREBP-1c and ChREBP extremely significantly increased (P<0.01), and the mRNA expression levels of key adipogenic genes FAS, C/EBPβ, PPARγ and FABP4 extremely significantly increased (P<0.01). On the contrary, interfering with NR1H3 significantly reduced the number of lipid droplets. The mRNA expression of downstream target genes and key adipogenic genes extremely significantly reduced (P<0.01), which inhibited the adipogenesis process. The results showed that NR1H3 gene was a positive regulator of the adipogenic differen-tiation of porcine preadipocytes, and affected the differentiation of porcine preadipocytes by regulating the expression of its downstream targets SREBP-1c and ChREBP. These results are of great significance to elucidate the molecular mechanism of fat deposition and improve meat quality.

Key words: pig, NR1H3, porcine preadipocyte, adipogenic differentiation

中图分类号:

S828.2

史明月, 张雪莲, 杨晓奋, 牛瑾, 邢建东, 路畅, 高鹏飞, 郭晓红, 李步高, 曹果清. NR1H3基因调控猪前体脂肪细胞分化的研究[J]. 畜牧兽医学报, 2022, 53(7): 2094-2103.

SHI Mingyue, ZHANG Xuelian, YANG Xiaofen, NIU Jin, XING Jiandong, LU Chang, GAO Pengfei, GUO Xiaohong, LI Bugao, CAO Guoqing. Study on NR1H3 Gene Regulating Differentiation of Porcine Preadipocyte[J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(7): 2094-2103.

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