Isolation and Identification of Bovine Preadipocytes and Screening of MicroRNAs Associated with Adipogenesis

Animals

Zhihui Zhao¹  Ming Gao²  Xiuqi Zhang²  Jiaqi Mi²  Runjun Yang²  Xiang Yu²  Lixin Xia²  Xibi Fang²  Steffen Maak³  Elke Albrecht³ 

[1] College of Agriculture, Guangdong Ocean University, Zhanjiang 524088, Guangdong Province, China;College of Animal Sciences, Jilin University, Changchun 130062, Jilin Province, China;Institute of Muscle Biology and Growth, Leibniz Institute for Farm Animal Biology (FBN), 18196 Dummerstorf, Germany;

关键词: bovine preadipocyte;    differentiation;    microRNA;    lipid metabolism;   

DOI  :  10.3390/ani10050818

来源: DOAJ

【 摘 要 】

The elucidation of the mechanisms of preadipocyte differentiation and fat accumulation in adipocytes is a major work in beef cattle breeding. As important post-transcriptional regulators, microRNAs (miRNAs) take part in cell proliferation, differentiation, apoptosis, and fat metabolism through binding seed sites of targeting mRNAs. The aim of this study was to isolate and identify bovine preadipocytes and screen miRNAs associated with adipogenesis. Bovine preadipocytes were isolated from subcutaneous fatty tissue and induced to differentiate into adipocytes. Verification of preadipocytes and adipocytes was performed by qRT-PCR (real-time quantitative reverse transcription PCR), Oil Red O staining, and immunofluorescence staining. Total RNA was extracted for small RNA sequencing. The sequencing data showed that 131 miRNAs were highly expressed in adipocytes, and 119 miRNAs were highly expressed in preadipocytes. Stem–loop qPCR (stem–loop quantitative real-time PCR) results showed that the expression patterns of 11 miRNAs were consistent with the sequencing results (miR-149-5p, miR-24-3p, miR-199a-5p, miR-33a, etc.). According to KEGG pathway and Gene Ontology (GO) analyses, multiple predicted target genes were associated with lipid metabolism. In summary, this study provides a protocol of isolating bovine preadipocytes and screening various differently expressed miRNAs during preadipocyte differentiation.

【 授权许可】

Unknown