■萎缩性骨不连是最困难的骨折并发症之一。导致萎缩性骨不连的细胞因子知之甚少,间充质干细胞(MSCs)被认为是骨形成的关键贡献者。本研究旨在从增殖的角度表征从萎缩性骨不连(AN-MSCs)纤维化组织中分离的MSCs,分化潜力,衰老,和旁分泌功能。
■从四个29-37岁的供体获得人萎缩性纤维化组织,用于分离AN-MSCs,和供者匹配的骨髓从髂骨获得,用于分离MSCs(IC-MSCs)作为对照。第3代的AN-MSC或IC-MSC用于以下评价。使用流式细胞术评估在两个细胞上表达的表面标志物。比较研究了两种细胞长达11天的增殖。成骨之后,软骨形成,或成脂肪诱导,使用谱系特异性染色和谱系特异性基因表达对AN-MSCs或IC-MSCs的多谱系分化进行了比较评估.应用酶联免疫吸附测定(ELISA)评估AN-MSCs或IC-MSCs的旁分泌功能。使用衰老相关β-半乳糖苷酶(SA-β-gal)染色评估AN-MSC或IC-MSC的细胞衰老。
■来自四个供体的AN-MSCs或IC-MSCs表现出MSCs的形态学和免疫表型特征,MSCs标记物的表达和造血标记物的阴性表达。总的来说,AN-MSC显示与IC-MSC相似的增殖和成脂能力。相比之下,与AN-MSC相比,IC-MSC显示出明显更高的成骨和软骨形成能力。此外,IC-MSCs的培养基中含有显著较高水平的VEGF,TGF-β1,PDGF-BB,和IGF-1比AN-MSCs的培养基。最后,AN-MSC比IC-MSC更倾向于细胞衰老。
■体外,AN-MSCs在增殖和成脂能力方面与IC-MSCs相似,但在成骨和软骨形成能力方面不如IC-MSCs,旁分泌功能,和抗衰老。
UNASSIGNED: Atrophic nonunion is one of the most difficult complications of fracture. The cellular factors that contribute to atrophic nonunion are poorly understood, and mesenchymal stem cells (MSCs) are recognized as the key contributor to bone formation. This study aimed to characterize the MSCs isolated from the fibrotic tissue of atrophic nonunion (AN-MSCs) from the perspective of proliferation, differentiation potential, senescence, and paracrine function.
UNASSIGNED: Human atrophic fibrotic tissue was obtained from four donors aged 29-37 for isolating AN-MSCs, and donor-matched bone marrow acquired from the iliac crest for isolating MSCs (IC-MSCs) as control. The AN-MSCs or IC-MSCs in passage 3 were applied for the following evaluations. The surface markers expressed on the two cells were evaluated using flow cytometry. The proliferation of the two cells for up to 11 days was comparatively investigated. After osteogenic, chondrogenic, or adipogenic induction, multi-lineage differentiation of AN-MSCs or IC-MSCs was comparatively evaluated using lineage-specific stains and lineage-specific gene expression. Enzyme-linked immunosorbent assay (ELISA) assessment was applied to evaluate the paracrine function of AN-MSCs or IC-MSCs. Cellular senescence of AN-MSCs or IC-MSCs was evaluated using senescence-associated β-galactosidase (SA-β-gal) staining.
UNASSIGNED: AN-MSCs or IC-MSCs from the four donors showed morphologic and immunophenotypic characteristics of MSCs, with the expression of MSCs markers and negative expression of hematopoietic markers. In general, AN-MSCs showed similar proliferation and adipogenic capacity with IC-MSCs. In contrast, IC-MSCs showed significantly higher osteogenic and chondrogenic capacity compared to AN-MSCs. Moreover, the culture medium of IC-MSCs contains significantly higher levels of VEGF, TGF-β1, PDGF-BB, and IGF-1 than the culture medium of AN-MSCs. Lastly, the AN-MSCs are more prone to cellular senescence than the IC-MSCs.
UNASSIGNED: In-vitro, AN-MSCs were similar to IC-MSCs in proliferation and adipogenic capacity, but inferior to IC-MSCs in osteogenic and chondrogenic capacity, paracrine function, and anti-senescence.