Study on Expressions of CNR1 and FABP4 Genes in Ovine Intramuscular Preadipocytes

doi:10.13304/j.nykjdb.2021.0120

Abstract

Abstract:

To study the pattern and difference of expression of CNR1 and FABP4 during the period of proliferation and differentiation of adipocytes， the longissimus dorsi muscle tissue sourced from a 3-day-old Hu lambs was collected， and then digested by the type II collagenase to separate and observe the primary preadipocytes. The preadipocytes were cultured and the growth curve was drawn. The preadipocytes were induced by insulin， dexamethasone and 3-isobutyl-1-methylxanthine， and the morphology of mature adipocytes was also observed by oil red O staining. The expressions of CNR1 and FABP4 genes were detected using real-time PCR method at different time of the adipocytes differentiation. The results showed that the preadipocytes were successfully isolated from the longissimus dorsi muscle of lamb， and the adherent cells were shown to be spindle shape. The formation of lipid droplets were observed in the preadipocytes stained with oil red O， which showed the characteristics of adipocytes. The relative expression levels of CNR1 and FABP4 genes were lower at the early stage of differentiation， then gradually increased at the middle and late stage. There were no significant differences in relative expression level of CNR1 and FABP4 on the 2^nd， 4^th and 8^th day， but the relative expression level of FABP4 was significantly higher than that of CNR1 on the 12^th day. It showed that CNR1 and FABP4 genes showed different expression characteristics in ovine intramuscular fat cells， which provided reference data for further revealing the regulatory relationship between two genes of intramuscular fat deposition and improving ovine meat traits.

Key words: sheep, intramuscular preadipocytes, FABP4, CNR1, gene expression

摘要：

为研究绵羊脂肪细胞增殖分化过程中参与动物能量代谢调控的大麻素受体1（cannabinoid receptor 1）基因CNR1和调控脂肪沉积的脂肪酸结合蛋白4（fatty acid binding protein 4）基因FABP4的表达规律和差异，采集一只3日龄湖羊羔羊背最长肌组织，背肌组织经Ⅱ型胶原酶消化后分离观察原代前体脂肪细胞；开展前体脂肪细胞培养并绘制生长曲线；应用胰岛素、地塞米松和3-异丁基-1-甲基黄嘌呤诱导前体脂肪细胞后，采用油红O染色观察成熟脂肪细胞形态；收集不同分化时间点细胞，应用实时荧光定量PCR方法测定CNR1和FABP4基因表达量。结果表明，羔羊背最长肌组织成功分离出了前体脂肪细胞，贴壁细胞呈梭形；诱导前体脂肪细胞经油红O染色后观察到脂滴形成，具有脂肪细胞特征；CNR1和FABP4基因表达量在诱导分化早期均较低，但在诱导分化中后期表达量逐步升高；CNR1和FABP4基因表达量在诱导分化后的第2、第4和第8天均差异不显著，但FABP4基因在第12天的表达量显著高于CNR1基因。上述结果为进一步探究绵羊CNR1和FABP4基因调控关系及利用两基因改良绵羊肉品性状提供了参考数据。

关键词: 绵羊, 肌内前体脂肪细胞, FABP4, CNR1, 基因表达

CLC Number:

S826

Wei YAN, Yutao WANG, Yonghao ZHANG, Haixia LIU, Dayong HAN, Aiwen ZHU. Study on Expressions of CNR1 and FABP4 Genes in Ovine Intramuscular Preadipocytes[J]. Journal of Agricultural Science and Technology, 2022, 24(3): 95-102.

闫伟, 王玉涛, 张永浩, 刘海霞, 韩大勇, 朱爱文. 绵羊肌内前体脂肪细胞CNR1和FABP4基因表达研究[J]. 中国农业科技导报, 2022, 24(3): 95-102.

Figures/Tables 6

Table 1 Sequences of amplified primers

基因名Gene name	引物序列Primer sequence （5’-3’）	扩增片段长度 Length of amplifed fragment/bp
GAPDH	F： GCAAGTTCCACGGCACAG	249
	R： GGTTCACGCCCATCACAA
CNR1	F： GACCAAGAGCGGAGAACG	144
	R： TCAATGACGGAGAAGGGAC
FABP4	F： CATGAAAGAAGTGGGTGTGGG	150
	R： TCCTGGCCCAATTTGAAGGA

Fig. 1 Primary cultured cells of intramuscular preadipocyte in sheep （200 ×）

Fig. 2 Growth curve of intramuscular preadipocyte cells in sheep

Fig. 3 Induced differentiation of intramuscular preadipocyte in sheep （100 ×）

Fig. 4 CNR1 and FABP4 expression level of intramuscular preadipocyteNote： * and ** represent significant differences between groups at P<0.05 and P<0.01 levels， respectively.

Fig. 5 Difference of CNR1 and FABP4 ExpressionNote： * represents significant differences between groups at P<0.05 level.

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