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Enhancing Differentiation of Menstrual Blood-Derived Stem Cells Into Female Germ Cells Using a Bilayer Amniotic Membrane and Nano-Fibrous Fibroin Scaffold Publisher Pubmed



Izanlou S1 ; Afshar A2 ; Zare A3 ; Zhilisbayeva KR4 ; Bakhshalizadeh S5, 6 ; Safaei Z7 ; Sehatbakhsh S8 ; Khaledi S1 ; Asgari HR1 ; Kazemnejad S9 ; Ajami M10 ; Ajami M10 ; Dehghan Tarzjani M12 ; Najafzadeh V13 Show All Authors
Authors
  1. Izanlou S1
  2. Afshar A2
  3. Zare A3
  4. Zhilisbayeva KR4
  5. Bakhshalizadeh S5, 6
  6. Safaei Z7
  7. Sehatbakhsh S8
  8. Khaledi S1
  9. Asgari HR1
  10. Kazemnejad S9
  11. Ajami M10
  12. Ajami M10
  13. Dehghan Tarzjani M12
  14. Najafzadeh V13
  15. Kouchakian MR1
  16. Mussin NM14
  17. Kaliyev AA14
  18. Aringazina RA15
  19. Mahdipour M16, 17
  20. Shirazi R1, 18
  21. Tamadon A3, 4

Source: Tissue and Cell Published:2023


Abstract

Three-dimensional nanofiber scaffolds offer a promising method for simulating in vivo conditions within the laboratory. This study aims to investigate the influence of a bilayer amniochorionic membrane/nanofibrous fibroin scaffold on the differentiation of human menstrual blood mesenchymal stromal/stem cells (MenSCs) into female germ cells. MenSCs were isolated and assigned to four culture groups: (i) MenSCs co-cultured with granulosa cells (GCs) using the scaffold (3D-T group), (ii) MenSCs using the scaffold alone (3D-C group), (iii) MenSCs co-cultured only with GCs (2D-T group), and (iv) MenSCs without co-culture or scaffold (2D-C group). Both MenSCs and GCs were independently cultured for two weeks before co-culturing was initiated. Flow cytometry was employed to characterize MenSCs based on positive markers (CD73, CD90, and CD105) and negative markers (CD45 and CD133). Additionally, flow cytometry and immunocytochemistry were used to characterize the GCs. Differentiated MenSCs were analyzed using real-time PCR and immunostaining. The real-time PCR results demonstrated significantly higher levels of VASA expression in the 3D-T group compared to the 3D-C, 2D-T, and 2D-C groups. Similarly, the SCP3 mRNA level in the 3D-T group was notably elevated compared to the 3D-C, 2D-T, and 2D-C groups. Moreover, the expression of GDF9 was significantly higher in the 3D-T group when compared to the 3D-C, 2D-T, and 2D-C groups. Immunostaining results revealed a lack of signal for VASA, SCP3, or GDF9 markers in the 2D-T group, while some cells in the 3D-T group exhibited positive staining for all these proteins. These findings suggest that the combination of a bilayer amniochorionic membrane/nanofibrous fibroin scaffold with co-culturing GCs facilitates the differentiation of MenSCs into female germ cells. © 2023 Elsevier Ltd
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