title: Alteration of the miRNA expression profile in male porcine anterior pituitary cells in response to GHRH and CST and analysis of the potential roles for miRNAs in regulating GH.
abstract: OBJECTIVE
  Growth hormone releasing hormone (GHRH) is a major positive regulator of growth hormone (GH) in the anterior pituitary gland, while cortistatin's (CST) role is negative. miRNAs (microRNAs or miRs) are small RNA molecules modulating gene expression at the post-transcriptional level. However, little is known about the function of miRNAs in the regulation of GH synthesis and/or secretion. This study investigated potential functional miRNAs involved in GH secretion in the normal porcine pituitary.
  
  
  METHODS
  Primary porcine anterior pituitary cells were cultivated and then treated with 10 nmol/L GHRH and 100 nmol/L CST, respectively. The effects of GHRH and CST on GH secretion were determined using RIA. miRNA microarrays were employed to analyze miRNA expression after treatment and then differentially expressed miRNAs were screened. Bioinformatics analysis was used to analyze the potential targets in growth hormone regulation of altered miRNAs. Furthermore, functional experiments were conducted to study the function of ssc-let-7c.
  
  
  RESULTS
  GHRH significantly promoted GH secretion, while CST suppressed GH secretion. 19 and 35 differentially expressed miRNAs were identified in response to GHRH and CST treatments respectively. Verification of 5 randomly selected miRNAs by quantitative real-time PCR (qRT-PCR) showed similar changes with microarray analysis. Target analysis showed that some miRNAs may be involved in GH secretion-related pathways. Importantly, ssc-let-7c was predicted to target GH1 and GHRHR mRNA 3'untranslated regions (3'UTRs), which was supported by luciferase reporter assay. Furthermore, functional experimental results showed that ssc-let-7c was involved in GH secretion regulation, and overexpression of ssc-let-7c inhibited GH secretion in porcine anterior pituitary cells.
  
  
  CONCLUSIONS
  GHRH and CST modulated porcine pituitary cell miRNA expression. Bioinformatics analysis revealed a complicated network among differentially expressed miRNAs, GH regulation-related genes and hormones. More interestingly, ssc-let-7c inhibited both GH1 and GHRHR mRNA 3'UTR reporter vectors' luciferase activity and overexpression of ssc-let-7c led to a decrease of GH secretion.
authors.affiliation: Guangdong Provincial Key Lab of Agro-Animal Genomics and Molecular Breeding, College of Animal Science, South China Agriculture University, Wushan Road, Guangzhou 510642, China.
authors.forename: Qi-En
authors.initials: QE
authors.lastname: Qi
authors.affiliation: Guangdong Provincial Key Lab of Agro-Animal Genomics and Molecular Breeding, College of Animal Science, South China Agriculture University, Wushan Road, Guangzhou 510642, China.
authors.forename: Qian-Yun
authors.initials: QY
authors.lastname: Xi
authors.affiliation: Guangdong Provincial Key Lab of Agro-Animal Genomics and Molecular Breeding, College of Animal Science, South China Agriculture University, Wushan Road, Guangzhou 510642, China.
authors.forename: Rui-Song
authors.initials: RS
authors.lastname: Ye
authors.affiliation: Guangdong Provincial Key Lab of Agro-Animal Genomics and Molecular Breeding, College of Animal Science, South China Agriculture University, Wushan Road, Guangzhou 510642, China.
authors.forename: Ting
authors.initials: T
authors.lastname: Chen
authors.affiliation: Guangdong Provincial Key Lab of Agro-Animal Genomics and Molecular Breeding, College of Animal Science, South China Agriculture University, Wushan Road, Guangzhou 510642, China.
authors.forename: Xiao
authors.initials: X
authors.lastname: Cheng
authors.affiliation: Guangdong Provincial Key Lab of Agro-Animal Genomics and Molecular Breeding, College of Animal Science, South China Agriculture University, Wushan Road, Guangzhou 510642, China.
authors.forename: Chao-Yun
authors.initials: CY
authors.lastname: Li
authors.affiliation: Guangdong Provincial Key Lab of Agro-Animal Genomics and Molecular Breeding, College of Animal Science, South China Agriculture University, Wushan Road, Guangzhou 510642, China.
authors.forename: Xiao-Tong
authors.initials: XT
authors.lastname: Zhu
authors.affiliation: Guangdong Provincial Key Lab of Agro-Animal Genomics and Molecular Breeding, College of Animal Science, South China Agriculture University, Wushan Road, Guangzhou 510642, China.
authors.forename: Gang
authors.initials: G
authors.lastname: Shu
authors.affiliation: Guangdong Provincial Key Lab of Agro-Animal Genomics and Molecular Breeding, College of Animal Science, South China Agriculture University, Wushan Road, Guangzhou 510642, China.
authors.forename: Li-Na
authors.initials: LN
authors.lastname: Wang
authors.affiliation: Guangdong Provincial Key Lab of Agro-Animal Genomics and Molecular Breeding, College of Animal Science, South China Agriculture University, Wushan Road, Guangzhou 510642, China.
authors.forename: Qing-Yan
authors.initials: QY
authors.lastname: Jiang
authors.affiliation: Guangdong Provincial Key Lab of Agro-Animal Genomics and Molecular Breeding, College of Animal Science, South China Agriculture University, Wushan Road, Guangzhou 510642, China. Electronic address: Zhangyl@scau.edu.cn.
authors.forename: Yong-Liang
authors.initials: YL
authors.lastname: Zhang
mesh_terms: D000818:Animals; D016466:CHO Cells; D002478:Cells, Cultured; D006224:Cricetinae; D003412:Cricetulus; D005786:Gene Expression Regulation / Q000187:drug effects; D013006:Growth Hormone / Q000235:genetics* / Q000378:metabolism; D013007:Growth Hormone-Releasing Hormone / Q000494:pharmacology*; D008297:Male; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism; D046228:Microarray Analysis; D009479:Neuropeptides / Q000494:pharmacology*; D010903:Pituitary Gland, Anterior / Q000187:drug effects* / Q000378:metabolism; D013552:Swine; D059467:Transcriptome / Q000187:drug effects*
chemical_list: D035683:MicroRNAs; D009479:Neuropeptides; C099323:cortistatin; D013006:Growth Hormone; D013007:Growth Hormone-Releasing Hormone
keywords: Growth hormone; Pituitary; Porcine; miRNA