title: Expression profiling of microRNA in cryptorchid testes: miR-135a contributes to the maintenance of spermatogonial stem cells by regulating FoxO1.
abstract: OBJECTIVE
  We elucidated the mechanism of spermatogonial stem cell disturbance of cryptorchidism and investigated the expression of miRNAs and their target genes in undescended testes.
  
  
  METHODS
  Using microarray analysis we compared total miRNA expression in unilateral undescended testes with that in contralateral descended and normal testes in a rat model of cryptorchidism. The model was derived by administering flutamide to pregnant Sprague Dawley® rats. We identified mRNA targets of miRNAs by bioinformatic analysis, followed by in situ hybridization and immunohistochemistry to localize candidate miRNAs and mRNAs, respectively. We also investigated whether miRNAs could inhibit target protein expression in vitro.
  
  
  RESULTS
  Microarray analysis and subsequent quantitative reverse transcriptase-polymerase chain reaction showed that only miR-135a was expressed at a lower level in undescended testes. We identified its target as FoxO1, which is essential for stem cell maintenance. miR-135a and FoxO1 localized to spermatogonial stem cells. FoxO1 localized to the spermatogonial stem cell nucleus less frequently in undescended testes, indicating that the activity of FoxO1, which acts as a transcription factor, is altered in undescended testes. Finally, miR-135a transfection into spermatogonia in vitro resulted in down-regulation of FoxO1 expression.
  
  
  CONCLUSIONS
  In cryptorchid testes there is a decreased number of spermatogonial stem cells in which FoxO1 is activated, indicating that failure of spermatogonial stem cell maintenance results in spermatogenesis alteration. We also noted interaction between miR-135a and FoxO1, and propose that miR-135a contributes to spermatogonial stem cell maintenance through modulation of FoxO1 activity.
authors.affiliation: Department of Nephro-Urology, Graduate School of Medical Sciences, Nagoya City University, Nagoya, Japan.
authors.forename: Yoshinobu
authors.initials: Y
authors.lastname: Moritoki
authors.affiliation: Department of Nephro-Urology, Graduate School of Medical Sciences, Nagoya City University, Nagoya, Japan. Electronic address: yutaro@med.nagoya-cu.ac.jp.
authors.forename: Yutaro
authors.initials: Y
authors.lastname: Hayashi
authors.affiliation: Department of Nephro-Urology, Graduate School of Medical Sciences, Nagoya City University, Nagoya, Japan.
authors.forename: Kentaro
authors.initials: K
authors.lastname: Mizuno
authors.affiliation: Department of Nephro-Urology, Graduate School of Medical Sciences, Nagoya City University, Nagoya, Japan.
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Kamisawa
authors.affiliation: Department of Nephro-Urology, Graduate School of Medical Sciences, Nagoya City University, Nagoya, Japan.
authors.forename: Hidenori
authors.initials: H
authors.lastname: Nishio
authors.affiliation: Department of Nephro-Urology, Graduate School of Medical Sciences, Nagoya City University, Nagoya, Japan.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Kurokawa
authors.affiliation: Department of Neurobiology and Anatomy, Graduate School of Medical Sciences, Nagoya City University, Nagoya, Japan.
authors.forename: Shinya
authors.initials: S
authors.lastname: Ugawa
authors.affiliation: Department of Urology, Fukushima Medical University, Fukushima, Japan.
authors.forename: Yoshiyuki
authors.initials: Y
authors.lastname: Kojima
authors.affiliation: Department of Nephro-Urology, Graduate School of Medical Sciences, Nagoya City University, Nagoya, Japan.
authors.forename: Kenjiro
authors.initials: K
authors.lastname: Kohri
mesh_terms: D000818:Animals; D003456:Cryptorchidism / Q000235:genetics* / Q000378:metabolism*; D051858:Forkhead Transcription Factors / Q000502:physiology*; D005786:Gene Expression Regulation; D008297:Male; D035683:MicroRNAs / Q000096:biosynthesis*; D009419:Nerve Tissue Proteins / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013093:Spermatogonia / Q000166:cytology*; D013234:Stem Cells; D000071161:Forkhead Box Protein O1
chemical_list: D051858:Forkhead Transcription Factors; D035683:MicroRNAs; D009419:Nerve Tissue Proteins; C074639:Foxo1 protein, rat; D000071161:Forkhead Box Protein O1
keywords: Foxo1 protein; cryptorchidism; gene expression; rat; spermatogenesis; testis