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.

Objectives: 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 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 ( 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.