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Organ culture of seminiferous tubules using a modified soft agar culture system

BACKGROUND: In-vitro spermatogenesis in mammalian species is considered an important topic in reproductive biology. New strategies for achieving a complete version of spermatogenesis ex vivo have been conducted using an organ culture method or culture of testicular cells in a three-dimensional soft...

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Autores principales: Gholami, Keykavos, Pourmand, Gholamreza, Koruji, Morteza, Ashouri, Sepideh, Abbasi, Mehdi
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6158910/
https://www.ncbi.nlm.nih.gov/pubmed/30257723
http://dx.doi.org/10.1186/s13287-018-0997-8
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author Gholami, Keykavos
Pourmand, Gholamreza
Koruji, Morteza
Ashouri, Sepideh
Abbasi, Mehdi
author_facet Gholami, Keykavos
Pourmand, Gholamreza
Koruji, Morteza
Ashouri, Sepideh
Abbasi, Mehdi
author_sort Gholami, Keykavos
collection PubMed
description BACKGROUND: In-vitro spermatogenesis in mammalian species is considered an important topic in reproductive biology. New strategies for achieving a complete version of spermatogenesis ex vivo have been conducted using an organ culture method or culture of testicular cells in a three-dimensional soft agar culture system (SACS). The aim of this study was to develop a new method that supports spermatogenesis to the meiotic phase and morphologically mature spermatozoa through the culture of testicular cells and seminiferous tubules (STs) in a modified SACS, respectively. METHODS: First, enzymatically dissociated testicular cells and mechanically dissociated STs of neonatal mice were separately embedded in agarose and then placed on the flat surface of agarose gel half-soaked in the medium to continue culture with a gas-liquid interphase method. RESULTS: Following 40 days of culture, the meiotic (Scp3) and post-meiotic (Acr) gene expression in aggregates and STs was confirmed by real-time polymerase chain reaction. These results were complemented by immunohistochemistry. The presence of morphologically mature spermatozoa in the frozen sections of STs was demonstrated with hematoxylin and eosin staining. We observed Plzf- or Integrin α6-positive spermatogonia in both cultures after 40 days, indicating the potency of the culture system for both self-renewal and differentiation. CONCLUSIONS: This technique can be used as a valuable approach for performing research on spermatogenesis and translating it into the human clinical setting.
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spelling pubmed-61589102018-10-01 Organ culture of seminiferous tubules using a modified soft agar culture system Gholami, Keykavos Pourmand, Gholamreza Koruji, Morteza Ashouri, Sepideh Abbasi, Mehdi Stem Cell Res Ther Research BACKGROUND: In-vitro spermatogenesis in mammalian species is considered an important topic in reproductive biology. New strategies for achieving a complete version of spermatogenesis ex vivo have been conducted using an organ culture method or culture of testicular cells in a three-dimensional soft agar culture system (SACS). The aim of this study was to develop a new method that supports spermatogenesis to the meiotic phase and morphologically mature spermatozoa through the culture of testicular cells and seminiferous tubules (STs) in a modified SACS, respectively. METHODS: First, enzymatically dissociated testicular cells and mechanically dissociated STs of neonatal mice were separately embedded in agarose and then placed on the flat surface of agarose gel half-soaked in the medium to continue culture with a gas-liquid interphase method. RESULTS: Following 40 days of culture, the meiotic (Scp3) and post-meiotic (Acr) gene expression in aggregates and STs was confirmed by real-time polymerase chain reaction. These results were complemented by immunohistochemistry. The presence of morphologically mature spermatozoa in the frozen sections of STs was demonstrated with hematoxylin and eosin staining. We observed Plzf- or Integrin α6-positive spermatogonia in both cultures after 40 days, indicating the potency of the culture system for both self-renewal and differentiation. CONCLUSIONS: This technique can be used as a valuable approach for performing research on spermatogenesis and translating it into the human clinical setting. BioMed Central 2018-09-26 /pmc/articles/PMC6158910/ /pubmed/30257723 http://dx.doi.org/10.1186/s13287-018-0997-8 Text en © The Author(s). 2018 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
spellingShingle Research
Gholami, Keykavos
Pourmand, Gholamreza
Koruji, Morteza
Ashouri, Sepideh
Abbasi, Mehdi
Organ culture of seminiferous tubules using a modified soft agar culture system
title Organ culture of seminiferous tubules using a modified soft agar culture system
title_full Organ culture of seminiferous tubules using a modified soft agar culture system
title_fullStr Organ culture of seminiferous tubules using a modified soft agar culture system
title_full_unstemmed Organ culture of seminiferous tubules using a modified soft agar culture system
title_short Organ culture of seminiferous tubules using a modified soft agar culture system
title_sort organ culture of seminiferous tubules using a modified soft agar culture system
topic Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6158910/
https://www.ncbi.nlm.nih.gov/pubmed/30257723
http://dx.doi.org/10.1186/s13287-018-0997-8
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