title: In vitro differentiation process of human Wharton's jelly mesenchymal stem cells to male germ cells in the presence of gonadal and non-gonadal conditioned media with retinoic acid.
abstract: Human umbilical Wharton's jelly-derived mesenchymal stem cells (HWJMSCs) are the best candidate to get plentiful stem cells and differentiate them to germ cells under appropriate conditions to treat infertility. We sought to determine under which conditions HWJMSCs could form male germ cells in vitro. So, HWJMSCs were differentiated to male germ cells under a mixture of bone morphogenetic protein-4 (BMP-4) and testicular and placental culture condition (TCC and PCC) medium followed by retinoic acid for 21 d. In the present study, the HWJMSCs were obtained from Wharton's jelly of umbilical cords of male neonates delivered by cesarean section. At the third passage, mesenchymal stem cell markers and differentiation to osteocytes and adipocytes were investigated. Then, HWJMSCs were induced to differentiate into male germ cells in the presence of BMP-4, all-trans retinoic acid, PCC, and TCC for 21 d. The profile of c-Kit, DDX4, Piwil2, and Dazl gene expression was evaluated by qPCR and ICC. Data was analyzed by ANOVA test. After 3 wk of treatment with different reagents, the morphology of these spindle-like cells changed to shiny clusters and germ cell-specific markers in mRNA were upregulated in both TCC + retinoic acid (RA) and BMP-4 + RA. Induction of HWJMSCs with TCC in the presence of RA resulted in significant upregulation (P ≤ 0.05) of all germ cell-specific genes (c-Kit 2.6795 ± 0.75, DDX4 4.3188 ± 1.18, Piwil2 4.9962 ± 1.55, Dazl 6.1199 ± 0.78) compared to control and PCC + RA. Our results indicated that TCC and RA are involved in human germ cell development. Moreover, BMP signaling also induced differentiation. Our findings provide a novel effective approach for generation of germ cells in vitro and studying the interaction of germ cells with their niche. Our work represents an essential step toward gaining knowledge of the molecular properties of HWJMSCs in the field of cell therapy. We demonstrated that under a suitable situation, HWJMSCs have the ability to differentiate into germ cells and this provides an excellent pattern to study infertility cause and cure.
authors.affiliation: Department of Anatomy, Faculty of Medicine, Tehran University of Medical Sciences, P.O. Box 13145-784, Tehran, Iran. Famidi@sina.tums.ac.ir.
authors.forename: Fardin
authors.initials: F
authors.lastname: Amidi
authors.affiliation: Department of Anatomy, Faculty of Medicine, Tehran University of Medical Sciences, P.O. Box 13145-784, Tehran, Iran.
authors.forename: Nahid
authors.initials: N
authors.lastname: Ataie Nejad
authors.affiliation: Department of Obstetrics and Gynecology, Shariati Hospital, Tehran University of Medical Sciences, Tehran, Iran.
authors.forename: Marziyeh
authors.initials: M
authors.lastname: Agha Hoseini
authors.affiliation: Institute for Molecular Medicine and Cell Therapy, Düsseldorf, Germany and GENEOCELL, Advanced Molecular & Cellular Technologies, Tehran, Iran.
authors.forename: Karim
authors.initials: K
authors.lastname: Nayernia
authors.affiliation: Department of Anatomical Sciences, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran.
authors.forename: Zohreh
authors.initials: Z
authors.lastname: Mazaheri
authors.affiliation: Department of Anatomy, Faculty of Medicine, Tehran University of Medical Sciences, P.O. Box 13145-784, Tehran, Iran.
authors.forename: Nazila
authors.initials: N
authors.lastname: Yamini
authors.affiliation: Department of Anatomy, Faculty of Medicine, Semnan University of Medical Sciences, Semnan, Iran.
authors.forename: Sara
authors.initials: S
authors.lastname: Saeednia
mesh_terms: D060565:Argonaute Proteins / Q000096:biosynthesis; D055415:Bone Morphogenetic Protein 4 / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects*; D017077:Culture Media, Conditioned; D053487:DEAD-box RNA Helicases / Q000096:biosynthesis; D005260:Female; D006801:Humans; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D010920:Placenta / Q000166:cytology; D016133:Polymerase Chain Reaction; D011247:Pregnancy; D019009:Proto-Oncogene Proteins c-kit / Q000096:biosynthesis; D012333:RNA, Messenger / Q000096:biosynthesis; D016601:RNA-Binding Proteins / Q000096:biosynthesis; D015398:Signal Transduction / Q000187:drug effects; D013094:Spermatozoa / Q000166:cytology*; D013737:Testis / Q000166:cytology; D014020:Tissue Extracts / Q000494:pharmacology*; D014212:Tretinoin / Q000494:pharmacology*; D014470:Umbilical Cord / Q000166:cytology; D059631:Wharton Jelly / Q000166:cytology*
chemical_list: D060565:Argonaute Proteins; C525809:BMP4 protein, human; D055415:Bone Morphogenetic Protein 4; D017077:Culture Media, Conditioned; C101358:DAZL protein, human; C486868:PIWIL2 protein, human; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D014020:Tissue Extracts; D014212:Tretinoin; D019009:Proto-Oncogene Proteins c-kit; C498288:DDX4 protein, human; D053487:DEAD-box RNA Helicases
keywords: BMP-4; Differentiation; Fetal testis; Germ cells; Mesenchymal stem cells; Placenta; Retinoic acid; Wharton’s jelly