Genetic and Epigenetic Evaluation of Human Spermatogonial Stem Cells Isolated by MACS in Different Two and Three-Dimensional Culture Systems
Objective: Epigenetic and genetic changes have important roles in stem cell achievements. Accordingly, the aim of thisstudy is the evaluation of the epigenetic and genetic alterations of different culture systems, considering their efficacy inpropagating human spermatogonial stem cells isolated by m...
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| Format: | Article |
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Royan Institute (ACECR), Tehran
2022-08-01
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| Series: | Cell Journal |
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| Online Access: | https://www.celljournal.org/article_254338_55181b6a9c70771e7b59ccbddb700665.pdf |
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| author | Maria Zahiri Mansoureh Movahedin Seyed Javad Mowla Mehrdad Noruzinia Morteza Koruji Mohammad Reza Nowroozi Fatemeh Asgari |
| author_facet | Maria Zahiri Mansoureh Movahedin Seyed Javad Mowla Mehrdad Noruzinia Morteza Koruji Mohammad Reza Nowroozi Fatemeh Asgari |
| author_sort | Maria Zahiri |
| collection | DOAJ |
| description | Objective: Epigenetic and genetic changes have important roles in stem cell achievements. Accordingly, the aim of thisstudy is the evaluation of the epigenetic and genetic alterations of different culture systems, considering their efficacy inpropagating human spermatogonial stem cells isolated by magnetic-activated cell sorting (MACS).Materials and Methods: In this experimental study, obstructive azoospermia (OA) patient-derived spermatogonial cells were divided into two groups. The MACS enriched and non-enriched spermatogonial stem cells (SSCs) were cultured in the control and treated groups; co-culture of SSCs with Sertoli cells of men with OA, co-culture of SSCs with healthy Sertoli cells of fertile men, the culture of SSCs on PLA nanofiber and culture of testicular cell suspension. Gene-specific methylation by MSP, expression of pluripotency (NANOG, C-MYC and OCT-4), and germ cells specific genes (Integrin α6, Integrin β1, PLZF) evaluated. Cultured SSCs from the optimized group were transplanted into the recipient azoospermic mouse.Results: The use of MACS for the purification of human stem cells was effective at about 69% with the culture of the testicular suspension, being the best culture system. Upon purification, the germ-specific gene expression was significantly higher in testicular cell suspension and treated groups (P≤0.05). During the culture time, gene-specific methylation patterns of the examined genes did not show any changes. Our data from transplantation indicated the homing of the donor-derived cells and the presence of human functional sperm.Conclusion: Our in vivo and in vitro results confirmed that culture of testicular cell suspension and selection ofspermatogonial cells could be effective ways for purification and enrichment of the functional human spermatogonial cells. The epigenetic patterns showed that the specific methylation of the evaluated genes at this stage remained constant with no alteration throughout the entire culture systems over time. |
| first_indexed | 2024-04-12T04:38:28Z |
| format | Article |
| id | doaj.art-f60507745a194e2c81919e26e816cbbe |
| institution | Directory Open Access Journal |
| issn | 2228-5806 2228-5814 |
| language | English |
| last_indexed | 2024-04-12T04:38:28Z |
| publishDate | 2022-08-01 |
| publisher | Royan Institute (ACECR), Tehran |
| record_format | Article |
| series | Cell Journal |
| spelling | doaj.art-f60507745a194e2c81919e26e816cbbe2022-12-22T03:47:44ZengRoyan Institute (ACECR), TehranCell Journal2228-58062228-58142022-08-0124848149010.22074/cellj.2022.7888254338Genetic and Epigenetic Evaluation of Human Spermatogonial Stem Cells Isolated by MACS in Different Two and Three-Dimensional Culture SystemsMaria Zahiri0Mansoureh Movahedin1Seyed Javad Mowla2Mehrdad Noruzinia3Morteza Koruji4Mohammad Reza Nowroozi5Fatemeh Asgari6Anatomical Science Department, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, IranAnatomical Science Department, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, IranDepartment of Molecular Genetics, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, IranDepartment of Medical Genetics, School of Medicine, Tarbiat Modares University, Tehran, IranDepartment of Anatomical Sciences, School of Medicine, Iran University of Medical Sciences, Tehran, IranDepartment of Urology, Uro-Oncology Research Center, Tehran University of Medical Sciences, Tehran, IranClinical Research Development Unit of Nekouei-Hedayati-Forghani Hospital, Qom University of Medical Sciences, Qom, IranObjective: Epigenetic and genetic changes have important roles in stem cell achievements. Accordingly, the aim of thisstudy is the evaluation of the epigenetic and genetic alterations of different culture systems, considering their efficacy inpropagating human spermatogonial stem cells isolated by magnetic-activated cell sorting (MACS).Materials and Methods: In this experimental study, obstructive azoospermia (OA) patient-derived spermatogonial cells were divided into two groups. The MACS enriched and non-enriched spermatogonial stem cells (SSCs) were cultured in the control and treated groups; co-culture of SSCs with Sertoli cells of men with OA, co-culture of SSCs with healthy Sertoli cells of fertile men, the culture of SSCs on PLA nanofiber and culture of testicular cell suspension. Gene-specific methylation by MSP, expression of pluripotency (NANOG, C-MYC and OCT-4), and germ cells specific genes (Integrin α6, Integrin β1, PLZF) evaluated. Cultured SSCs from the optimized group were transplanted into the recipient azoospermic mouse.Results: The use of MACS for the purification of human stem cells was effective at about 69% with the culture of the testicular suspension, being the best culture system. Upon purification, the germ-specific gene expression was significantly higher in testicular cell suspension and treated groups (P≤0.05). During the culture time, gene-specific methylation patterns of the examined genes did not show any changes. Our data from transplantation indicated the homing of the donor-derived cells and the presence of human functional sperm.Conclusion: Our in vivo and in vitro results confirmed that culture of testicular cell suspension and selection ofspermatogonial cells could be effective ways for purification and enrichment of the functional human spermatogonial cells. The epigenetic patterns showed that the specific methylation of the evaluated genes at this stage remained constant with no alteration throughout the entire culture systems over time.https://www.celljournal.org/article_254338_55181b6a9c70771e7b59ccbddb700665.pdfazoospermiagenetic and epigeneticspermatogonial stem cells |
| spellingShingle | Maria Zahiri Mansoureh Movahedin Seyed Javad Mowla Mehrdad Noruzinia Morteza Koruji Mohammad Reza Nowroozi Fatemeh Asgari Genetic and Epigenetic Evaluation of Human Spermatogonial Stem Cells Isolated by MACS in Different Two and Three-Dimensional Culture Systems Cell Journal azoospermia genetic and epigenetic spermatogonial stem cells |
| title | Genetic and Epigenetic Evaluation of Human Spermatogonial Stem Cells Isolated by MACS in Different Two and Three-Dimensional Culture Systems |
| title_full | Genetic and Epigenetic Evaluation of Human Spermatogonial Stem Cells Isolated by MACS in Different Two and Three-Dimensional Culture Systems |
| title_fullStr | Genetic and Epigenetic Evaluation of Human Spermatogonial Stem Cells Isolated by MACS in Different Two and Three-Dimensional Culture Systems |
| title_full_unstemmed | Genetic and Epigenetic Evaluation of Human Spermatogonial Stem Cells Isolated by MACS in Different Two and Three-Dimensional Culture Systems |
| title_short | Genetic and Epigenetic Evaluation of Human Spermatogonial Stem Cells Isolated by MACS in Different Two and Three-Dimensional Culture Systems |
| title_sort | genetic and epigenetic evaluation of human spermatogonial stem cells isolated by macs in different two and three dimensional culture systems |
| topic | azoospermia genetic and epigenetic spermatogonial stem cells |
| url | https://www.celljournal.org/article_254338_55181b6a9c70771e7b59ccbddb700665.pdf |
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