Details

Autor(en) / Beteiligte

• Nikmahzar, Aghbibi
• Koruji, Morteza
• Jahanshahi, Mehrdad
• Khadivi, Farnaz
• Shabani, Maryam
• Dehghani, Sanaz
• Forouzesh, Mehdi
• Jabari, Ayob
• Feizollahi, Narjes
• Salem, Maryam
• Ghanami Gashti, Nasrin
• Abbasi, Yasaman
• Abbasi, Mehdi

Titel

Differentiation of human primary testicular cells in the presence of SCF using the organoid culture system

Ist Teil von

• Artificial organs, 2023-12, Vol.47 (12), p.1818-1830

Ort / Verlag

United States: Wiley Subscription Services, Inc

Erscheinungsjahr

2023

Quelle

Wiley Online Library

Beschreibungen/Notizen

• Purpose Development of organoids using human primary testicular cells has remained a challenge due to the complexity of the mammalian testicular cytoarchitecture and culture methods. In this study, we generated testicular organoids derived from human primary testicular cells. Then, we evaluated the effect of stem cell factor (SCF) on cell differentiation and apoptosis in the testicular organoid model. Methods The testicular cells were harvested from the three brain‐dead donors. Human spermatogonial stem cells (SSCs) were characterized using immunocytochemistry (ICC), RT‐PCR and flow cytometry. Testicular organoids were generated from primary testicular cells by hanging drop culture method and were cultured in three groups: control group, experimental group 1 (treated FSH and retinoic acid (RA)), and experimental group 2 (treated FSH, RA and SCF), for five weeks. We assessed the expression of SCP3 (Synaptonemal Complex Protein 3) as a meiotic gene, PRM2 (Protamine 2) as a post‐meiotic marker and apoptotic genes of Bax (BCL2‐Associated X Protein) and Bcl‐2 (B‐cell lymphoma 2), respectively by using RT‐qPCR. In addition, we identified the expression of PRM2 by immunohistochemistry (IHC). Results Relative expression of SCP3, PRM2 and Bcl‐2 were highest in group 2 after five weeks of culture. In contrast, BAX expression level was lower in experimental group 2 in comparison with other groups. IHC analyses indicated the highest expression of PRM2 as a postmeiotic marker in group 2 in comparison to 2D culture and control groups but not find significant differences between experimental group 1 and experimental group 2 groups. Morphological evaluations revealed that organoids are compact spherical structures and in the peripheral region composed of uncharacterized elongated fibroblast‐like cells. Conclusion Our findings revealed that the testicular organoid culture system promote the spermatogonial stem cell (SSC) differentiation, especially in presence of SCF. Developed organoids are capable of recapitulating many important properties of a stem cell niche. The testicular cells were harvested from the brain‐dead donors and human testicular organoids were generated from primary testicular cells and were cultured in a medium supplemented with stem cell factor (SCF). Our results showed that the human primary testicular could differentiated in organoid culture system, especially in the presence of SCF.