AbstractBackground Mesenchymal stem cells (MSCs) as a source of osteoblasts play a key role in bone repair and regeneration. Fetal mesenchymal stem cells (Fetal-MSCs) exhibit lower immunogenicity, higher potential for expansion, and higher differentiation potential into various cell lineages than adult MSCs. Herein,in vitroandin vivoevaluations were performed to study the role of human Fetal-MSCs in osteogenic differentiation and bone regeneration in comparison to that of adult bone marrow mesenchymal stem cells (BM-MSCs). Methods Fetal-MSCs were isolated from fetal tissues derived from ectopic pregnancy at eight and half and half weeks of gestation and studied for karyotypic stability, proliferation over successive passages, and expression of embryonal and mesenchymal markers using flow cytometry. The ability of BM-MSCs and Fetal-MSCs to differentiate into osteoblasts was assessed using alkaline phosphatase (ALP) and alizarin red staining(ARS). The expression of osteogenic markers such as ALP, osteopontin(OPN), RUNX2, and osteocalcin(OCN) was evaluated using quantitative reverse transcription polymerase chain reaction. mRNA sequencing was performed to identify the differences in gene expression between Fetal-MSCs and BM-MSCs. In addition, their effects were assessed in rat calvarial defect model using micro-computed tomography, immunohistochemistry, and histological examination. Results Fetal-MSCs were more efficient than BM-MSCs in promoting osteogenesis and had a greater bone regeneration capacityin vitroandin vivo. In particular, Hairy ears Y-linked (HEY)1andHEY2, which are involved in osteoblast development, showed a significantly high differential expression between Fetal-MSCs and BM-MSCs and were the key target genes in the Notch signaling pathway. Notch signaling plays a key role in the osteogenic differentiation of Fetal-MSCs. Conclusions We demonstrated for the first time that Notch signaling plays a key regulatory role in promoting the osteogenic differentiation of Fetal-MSCs. In addition, Fetal-MSCs with their strong bone formation ability may be a promising alternative to adult-derived MSCs for bone disease treatment.
