title: Complete Human and Rat Ex Vivo Spermatogenesis from Fresh or Frozen Testicular Tissue.
abstract: Until now, complete ex vivo spermatogenesis has been reported only in the mouse. In this species, the duration of spermatogenesis is 35 days, whereas it is 54 days in the rat and 74 days in humans. We performed long-term (until 60 days) cultures of fresh or frozen rat or human seminiferous tubule segments in a bioreactor made of a hollow cylinder of chitosan hydrogel. Testicular tissues were obtained from 8- or 20-day-old male rats or from adult human subjects who had undergone hormone treatments leading to a nearly complete regression of their spermatogenesis before bilateral orchiectomy for gender reassignment. The progression of spermatogenesis was assessed by cytological analyses of the cultures; it was related to a dramatic increase in the levels of the mRNAs specifically expressed by round spermatids, Transition protein 1, Transition protein 2, and Protamine 3 in rat cultures. From 2% to 3.8% of cells were found to be haploid cells by fluorescence in situ hybridization analysis of human cultures. In this bioreactor, long-term cultures of seminiferous tubule segments from prepubertal rats or from adult men allowed completion of the spermatogenic process leading to morphologically mature spermatozoa. Further studies will need to address the way of optimizing the yield of every step of spermatogenesis by adjusting the composition of the culture medium, the geometry, and the material properties of the chitosan hydrogel bioreactors. Another essential requirement is to assess the quality of the gametes produced ex vivo by showing their ability to produce normal offspring (rat) or their biochemical normality (human).
authors: INSERM U 1208, Bron, France.
authors: Marie-Hélène
authors: MH
authors: Perrard
authors: Laboratoire Ingénierie des Matériaux Polymères, CNRS UMR 5223, Université Claude Bernard Lyon1, Université Lyon, Villeurbanne, France.
authors: Nicolas
authors: N
authors: Sereni
authors: Hospices Civils de Lyon, Université Claude Bernard Lyon 1, CRNL, CNRS UMR5292; TIGER Team, Lyon, France Service de Génétique, Laboratoire de Cytogénétique Constitutionnelle, Bron, France.
authors: Caroline
authors: C
authors: Schluth-Bolard
authors: Kallistem, Ecole Normale Supérieure de Lyon, Lyon, France.
authors: Antonine
authors: A
authors: Blondet
authors: Hospices Civils de Lyon, Université Claude Bernard Lyon 1, INSERM U 1208; Service de Médecine de la Reproduction, Hôpital Femme Mère Enfant, Bron, France.
authors: Sandrine Giscard
authors: SG
authors: D Estaing
authors: Hospices Civils de Lyon, Université Claude Bernard Lyon 1, INSERM U 1208; Service d'Hormonologie, Endocrinologie Moléculaire et Maladies Rares, Bron, France.
authors: Ingrid
authors: I
authors: Plotton
authors: Hospices Civils de Lyon. Service d'Urologie, Centre Hospitalier Lyon Sud, Chemin du Grand Revoyet, Pierre-Bénite, France.
authors: Nicolas
authors: N
authors: Morel-Journel
authors: Hospices Civils de Lyon, Université Claude Bernard Lyon 1, INSERM U 1208; Service de Médecine de la Reproduction, Hôpital Femme Mère Enfant, Bron, France.
authors: Hervé
authors: H
authors: Lejeune
authors: Laboratoire Ingénierie des Matériaux Polymères, CNRS UMR 5223, Université Claude Bernard Lyon1, Université Lyon, Villeurbanne, France.
authors: Laurent
authors: L
authors: David
authors: Kallistem, Ecole Normale Supérieure de Lyon, Lyon, France philippe.durand@ens-lyon.fr.
authors: Philippe
authors: P
authors: Durand
mesh_terms: D000328:Adult; D000818:Animals; D019149:Bioreactors; D048271:Chitosan; D015925:Cryopreservation / Q000379:methods; D003470:Culture Media; D006801:Humans; D020100:Hydrogels; D017404:In Situ Hybridization, Fluorescence; D066298:In Vitro Techniques; D008297:Male; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D012671:Seminiferous Tubules / Q000166:cytology / Q000378:metabolism; D013045:Species Specificity; D013087:Spermatids / Q000166:cytology / Q000378:metabolism; D013091:Spermatogenesis / Q000235:genetics / Q000502:physiology*; D013737:Testis / Q000166:cytology* / Q000378:metabolism
chemical_list: D003470:Culture Media; D020100:Hydrogels; D012333:RNA, Messenger; D048271:Chitosan
keywords: bioreactor; chitosan; fertility preservation; hollow cylinder; human spermatogenesis