Spontaneous spheroids from alveolar bone-derived mesenchymal stromal cells maintain pluripotency of stem cells by regulating hypoxia-inducible factors
Biol. Res
;
56: 17-17, 2023. ilus, tab, graf
Artículo
en Inglés
| LILACS
| ID: biblio-1439484
ABSTRACT
BACKGROUND: Spontaneous spheroid culture is a novel three-dimensional (3D) culture strategy for the rapid and efficient selection of progenitor cells. The objectives of this study are to investigate the pluripotency and differentiation capability of spontaneous spheroids from alveolar bone-derived mesenchymal stromal cells (AB-MSCs); compare the advantages of spontaneous spheroids to those of mechanical spheroids; and explore the mechanisms of stemness enhancement during spheroid formation from two-dimensional (2D) cultured cells. METHODS: AB-MSCs were isolated from the alveolar bones of C57BL/6 J mice. Spontaneous spheroids formed in low-adherence specific culture plates. The stemness, proliferation, and multi-differentiation capacities of spheroids and monolayer cultures were investigated by reverse transcription quantitative polymerase chain reaction (RT-qPCR), immunofluorescence, alkaline phosphatase (ALP) activity, and oil-red O staining. The pluripotency difference between the spontaneous and mechanical spheroids was analyzed using RT-qPCR. Hypoxia-inducible factor (HIFs) inhibition experiments were performed to explore the mechanisms of stemness maintenance in AB-MSC spheroids. RESULTS: AB-MSCs successfully formed spontaneous spheroids after 24 h. AB-MSC spheroids were positive for MSC markers and pluripotency markers (Oct4, KLF4, Sox2, and cMyc). Spheroids showed higher Ki67 expression and lower Caspase3 expression at 24 h. Under the corresponding conditions, the spheroids were successfully differentiated into osteogenic and adipogenic lineages. AB-MSC spheroids can induce neural-like cells after neurogenic differentiation. Higher expression of osteogenic markers, adipogenic markers, and neurogenic markers (NF-M, NeuN, and GFAP) was found in spheroids than in the monolayer. Spontaneous spheroids exhibited higher stemness than mechanical spheroids did. HIF-1α and HIF-2α were remarkably upregulated in spheroids. After HIF-1/2α-specific inhibition, spheroid formation was significantly reduced. Moreover, the expression of the pluripotency genes was suppressed. CONCLUSIONS: Spontaneous spheroids from AB-MSCs enhance stemness and pluripotency. HIF-1/2α plays an important role in the stemness regulation of spheroids. AB-MSC spheroids exhibit excellent multi-differentiation capability, which may be a potent therapy for craniomaxillofacial tissue regeneration.
Texto completo:
Disponible
Índice:
LILACS (Américas)
Asunto principal:
Esferoides Celulares
/
Células Madre Mesenquimatosas
Límite:
Animales
Idioma:
Inglés
Revista:
Biol. Res
Asunto de la revista:
Biologia
Año:
2023
Tipo del documento:
Artículo
País de afiliación:
China
Institución/País de afiliación:
Fudan University/CN
/
Shanghai Jing'an District Dental Clinic/CN
/
Shanghai University of Medicine and Health Sciences Affiliated Zhoupu Hospita/CN
/
Tongji University School of Medicine/CN
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