Evaluation of differentiation of the umbilical cord blood-derived mesenchymal stem cells into male germ-line cells in two-dimensional and three-dimensional scaffolds containing naringin

Articles in Press

Document Type : Original Article

Authors

1 Department of Anatomical Sciences, Tarbiat Modarres University, Tehran, Iran

2 Institute for Regenerative Medicine (IREM), University of Zurich, Zurich, Switzerland

3 Student Research Committee, School of Medicine, Shahroud University of Medical Sciences, Shahroud, Iran

4 Clinical Research Development Unit, Imam Hossein Hospital, Shahroud University of Medical Sciences, Shahroud, Iran

5 Center for Advanced Materials and Structures, School of Science and Technology, the University of Georgia, Tbilisi 0171, Georgia

6 Student Research Committee, School of Dentistry, Golestan University of Medical Sciences, Gorgan, Iran

7 Department of Tissue Engineering and Biomaterials, School of Advanced Medical Sciences and Technologies, Hamadan University of Medical Sciences, Hamadan, Iran

8 Regenerative Medicine Research Center, Shahroud University of Medical Sciences, Shahroud, Iran

9 Department of Tissue Engineering, School of Medicine, Shahroud University of Medical Sciences, Shahroud, Iran

10.22038/ijbms.2026.88928.19199

Abstract

Objective(s): This study aimed to investigate the differentiation potential of human umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs) into male germ-like cells using a three-dimensional poly (L-lactic acid)/gelatin nanofiber (PLLA/GNF) scaffold incorporated with naringin (Nar).
Materials and Methods: The composite scaffold was fabricated by freeze-drying a PLLA solution containing 1% Nar and electrospun GNFs. Scaffold characterization included morphological analysis, porosity assessment, drug release profiling, and biodegradation testing. UCB-MSCs were isolated and characterized for surface markers (CD44⁺/CD90⁺/CD34⁻/CD45⁻) via flow cytometry. Cells were seeded onto scaffolds and differentiated using BMP4 (25 ng/ml) alone or in combination with retinoic acid (10⁻⁶ M). Evaluations included the MTT assay for viability, immunofluorescence for germ cell markers (PLZF, DAZL, OCT4), and in vivo biocompatibility assessed by subcutaneous implantation in rat models.
Results: The scaffold demonstrated optimal physical properties, with a porosity of 81.13 ± 2.22% and an average pore size of 108.31 ± 9.29 μm. Nar release profile showed sustained delivery with cumulative release of 38.5 ± 5.72% over 14 days. Biodegradation rate reached 51.07 ± 5.64% after 14 days. MTT assay revealed significantly enhanced cell proliferation in Nar-containing scaffolds at both 24 and 72 hr (P< 0.05). Immunofluorescence analysis demonstrated markedly higher expression of germ cell markers in 3D cultures compared to 2D controls. In vivo evaluation confirmed excellent biocompatibility with minimal inflammatory response.
Conclusion: The 3D PLLA/GNF/Nar scaffold creates a favorable microenvironment that effectively promotes UCB-MSC differentiation into male germ-like cells, representing a promising platform for reproductive tissue engineering applications.

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Main Subjects

References

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Available Online from 06 January 2026

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