AMFR gene silencing inhibits the differentiation of porcine preadipocytes

• Published in: Genetics and molecular research 2016-04, Vol.15 (2)
• Main Authors: Chen, C Z, Zhu, Y N, Chai, M L, Dai, L S, Gao, Y, Jiang, H, Zhang, L J, Ding, Y, Liu, S Y, Li, Q Y, Lu, W F, Zhang, J B
• Format: Article
• Language: English
• Subjects: Adipocytes - cytology; Adipocytes - metabolism; Animals; CCAAT-Enhancer-Binding Protein-alpha - genetics; CCAAT-Enhancer-Binding Protein-alpha - metabolism; Cell Differentiation; Cells, Cultured; Gene Silencing; Kruppel-Like Transcription Factors - genetics; Kruppel-Like Transcription Factors - metabolism; PPAR gamma - genetics; PPAR gamma - metabolism; Receptors, Autocrine Motility Factor - genetics; Receptors, Autocrine Motility Factor - metabolism; Sterol Regulatory Element Binding Protein 1 - genetics; Sterol Regulatory Element Binding Protein 1 - metabolism; Sterol Regulatory Element Binding Protein 2 - genetics; Sterol Regulatory Element Binding Protein 2 - metabolism; Swine
• ISSN: 1676-5680; 1676-5680
• DOI: 10.4238/gmr.15027354

Our study clarifies the role of the autocrine motility factor receptor (AMFR) gene in porcine preadipocyte differentiation. AMFR-siRNA was transfected into porcine preadipocytes and the preadipocytes were induced to differentiation. Subsequently, qRT-PCR was conducted to examine changes in mRNA expression of a series of genes in porcine preadipocytes, including AMFR, sterol-regulatory element-binding protein-1a (SREBP1a), SREBP2, insulin-induced gene 1 (Insig1), and Insig2. Expression changes in the mRNA of genes regulating adipocyte differentiation were also analyzed using qRT-PCR, including peroxisome proliferator-activated receptor gamma (PPARγ), CCAAT/enhancer-binding protein alpha (C/EBPα), and Kruppel-like factor 2 (KLF2). Western blot analysis was conducted to examine the changes in AMFR protein expression in porcine preadipocytes. Additionally, morphological changes in differentiated porcine preadipocytes were examined by oil red O staining, and changes in optical density (OD) values were measured using an ultraviolet spectrophotometer. At 24 h after transfection with AMFR-siRNA, AMFR mRNA expression significantly reduced (P < 0.01), and AMFR protein expression markedly decreased (P < 0.05). The mRNA expression of SREBP1a, SREBP2, Insig1, and C/EBPα was significantly reduced (P < 0.01), whereas the expression of KLF2 mRNA was significantly elevated (P < 0.01). After induction of preadipocyte differentiation, the number of lipid droplets decreased in the AMFR-silenced group, and the OD value markedly reduced (P < 0.05). In addition, the expression of C/EBPα mRNA significantly decreased (P < 0.05), whereas the expression of KLF2 mRNA considerably increased (P < 0.05). Taken together, silencing of the AMFR gene inhibits the differentiation of porcine preadipocytes.