MicroRNA-1285 Regulates 17β-Estradiol-Inhibited Immature Boar Sertoli Cell Proliferation via Adenosine Monophosphate-Activated Protein Kinase Activation.
This study investigated the capacity of 10 μM 17β-estradiol to inhibit immature boar Sertoli cell (SC) proliferation and the involvement of microRNA (miR)-1285 in this process. SC viability and cell cycle progression were investigated using a cell counting kit-8 and flow cytometry, respectively. Expression of AMP-activated protein kinase (AMPK), S phase kinase-associated protein 2 (Skp2), and miR-1285 was analyzed by real-time RT-PCR and Western blotting. 17β-Estradiol (10 μM) reduced SC viability and miR-1285 expression and promoted AMPK phosphorylation. A double-stranded synthetic miR-1285 mimic promoted SC viability, increased levels of ATP, and phosphorylated mammalian target of rapamycin (mTOR) and Skp2 mRNA and protein, whereas p53 and p27 expression decreased, and 17β-estradiol-mediated effects on SCs were significantly attenuated. A single-stranded synthetic miR-1285 inhibitor produced the opposite effects on these measures. Activation of AMPK inhibited SC viability, reduced levels of ATP, phosphorylated mTOR and Skp2 mRNA and protein, and increased p53 and p27 expression. An AMPK inhibitor (compound C) attenuated the effects of 17β-estradiol on SCs. This indicated that 17β-estradiol (10 μM) reduced SC proliferation by inhibiting miR-1285 and thus activating AMPK. Phosphorylated AMPK is involved in the regulation of 17β-estradiol-mediated inhibition of SC viability through increasing p53 and p27 expression and inhibiting mTOR and Skp2 expression. Our findings also implicated Skp2 as the downstream integration point of p53 and mTOR. These findings indicated that miR-1285 may represent a target for the manipulation of boar sperm production.
Chongqing Key Laboratory of Forage and Herbivore (Z.J.J., W.Y., Y.W.R., W.X.Z.), College of Animal Science and Technology, Southwest University, Chongqing 400715, China; and Genetic Engineering and Stem Cell Biology Laboratory (Z.J.J., J.D.K.), Department of Animal Biotechnology, Faculty of Biotechnology, Jeju National University, Jeju 690756, South Korea.
Zhang Jiao
ZJ
Jiao
Chongqing Key Laboratory of Forage and Herbivore (Z.J.J., W.Y., Y.W.R., W.X.Z.), College of Animal Science and Technology, Southwest University, Chongqing 400715, China; and Genetic Engineering and Stem Cell Biology Laboratory (Z.J.J., J.D.K.), Department of Animal Biotechnology, Faculty of Biotechnology, Jeju National University, Jeju 690756, South Korea.
Wang
W
Yi
Chongqing Key Laboratory of Forage and Herbivore (Z.J.J., W.Y., Y.W.R., W.X.Z.), College of Animal Science and Technology, Southwest University, Chongqing 400715, China; and Genetic Engineering and Stem Cell Biology Laboratory (Z.J.J., J.D.K.), Department of Animal Biotechnology, Faculty of Biotechnology, Jeju National University, Jeju 690756, South Korea.
Yang Wei
YW
Rong
Chongqing Key Laboratory of Forage and Herbivore (Z.J.J., W.Y., Y.W.R., W.X.Z.), College of Animal Science and Technology, Southwest University, Chongqing 400715, China; and Genetic Engineering and Stem Cell Biology Laboratory (Z.J.J., J.D.K.), Department of Animal Biotechnology, Faculty of Biotechnology, Jeju National University, Jeju 690756, South Korea.
Jeong Dong
JD
Kee
Chongqing Key Laboratory of Forage and Herbivore (Z.J.J., W.Y., Y.W.R., W.X.Z.), College of Animal Science and Technology, Southwest University, Chongqing 400715, China; and Genetic Engineering and Stem Cell Biology Laboratory (Z.J.J., J.D.K.), Department of Animal Biotechnology, Faculty of Biotechnology, Jeju National University, Jeju 690756, South Korea.
Wang Xian
WX
Zhong
D055372:AMP-Activated Protein Kinases / Q000235:genetics* / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western; D049109:Cell Proliferation / Q000187:drug effects / Q000235:genetics*; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D002478:Cells, Cultured; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000235:genetics / Q000378:metabolism; D004958:Estradiol / Q000494:pharmacology*; D005786:Gene Expression Regulation / Q000187:drug effects; D008297:Male; D035683:MicroRNAs / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D044786:S-Phase Kinase-Associated Proteins / Q000235:genetics / Q000378:metabolism; D012708:Sertoli Cells / Q000187:drug effects / Q000378:metabolism*; D013552:Swine; D058570:TOR Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism
D035683:MicroRNAs; D044786:S-Phase Kinase-Associated Proteins; D016159:Tumor Suppressor Protein p53; D050760:Cyclin-Dependent Kinase Inhibitor p27; D004958:Estradiol; D000255:Adenosine Triphosphate; D058570:TOR Serine-Threonine Kinases; D055372:AMP-Activated Protein Kinases