PDF(Pubmed)
Abstract：
Bone marrow-derived mesenchymal stromal cells (BM-MSCs) are used in regenerative medicine and related research involving immunomodulatory, anti-inflammatory, anti-fibrotic and regenerative functions. Isolation of BM-MSCs from samples obtained during total hip arthroplasty (THA) is routinely possible. Advanced age and comorbidities of the majority of patients undergoing THA limit their applicability. Our study aimed to evaluate the potential of bone marrow obtained during periacetabular osteotomy (PAO) as a novel source of BM-MSCs from young donors by analyzing cell yield and cell characteristics.
Bone samples were obtained from the anterior Os ilium or superior Os pubis during PAO and from the femoral cavity during primary THA. Isolation of bone marrow-derived mononuclear cells (BM-MNCs) was performed by density gradient centrifugation. The samples from PAO and THA patients were compared in terms of BM-MSC yield, colony formation and the proportion of BM-MSCs within the BM-MNC population using flow cytometry analysis. The cells were characterized based on the expression of BM-MSC-specific surface markers. The functionality of the cells was compared by quantifying post-thaw viability, metabolic activity, proliferation capacity, senescence-associated beta galactosidase (SA-β-gal) expression, trilineage differentiation potential and major secretome proteins.
Isolation of BM-MNCs was possible in a reliable and reproducible manner when using bone from PAO containing more than 0.24 g bone marrow. PAO patients were younger than patients of the THA group. Bone obtained during PAO contained less bone marrow and led to a lower BM-MSC number after the first cell culture passage compared to BM-MSCs obtained during THA. BM-MSCs from PAO samples are characterized by a higher proliferation capacity. This results in a higher yield in cell culture passage two, when normalized to the sample weight. BM-MSCs from PAO patients showed increased secretion of TGF-β1, TIMP2, and VEGF upon osteogenic differentiation. BM-MSCs from PAO and THA patients revealed similar results regarding the onset of SA-β-gal expression and trilineage differentiation capacity.
We suggest that bone obtained during PAO is a promising novel source for BM-MSCs from young donors. Limited absolute cell yield due to low sample weight must be considered in early cell culture passages and might be critical for the range of clinical applications possible for BM-MSCs from this source. The higher proliferation capacity and increased growth factor secretion of BM-MSCs from young donors may be beneficial for future regenerative cell therapies, in vitro models, and tissue engineering.
Bone samples were obtained from the anterior Os ilium or superior Os pubis during PAO and from the femoral cavity during primary THA. Isolation of bone marrow-derived mononuclear cells (BM-MNCs) was performed by density gradient centrifugation. The samples from PAO and THA patients were compared in terms of BM-MSC yield, colony formation and the proportion of BM-MSCs within the BM-MNC population using flow cytometry analysis. The cells were characterized based on the expression of BM-MSC-specific surface markers. The functionality of the cells was compared by quantifying post-thaw viability, metabolic activity, proliferation capacity, senescence-associated beta galactosidase (SA-β-gal) expression, trilineage differentiation potential and major secretome proteins.
Isolation of BM-MNCs was possible in a reliable and reproducible manner when using bone from PAO containing more than 0.24 g bone marrow. PAO patients were younger than patients of the THA group. Bone obtained during PAO contained less bone marrow and led to a lower BM-MSC number after the first cell culture passage compared to BM-MSCs obtained during THA. BM-MSCs from PAO samples are characterized by a higher proliferation capacity. This results in a higher yield in cell culture passage two, when normalized to the sample weight. BM-MSCs from PAO patients showed increased secretion of TGF-β1, TIMP2, and VEGF upon osteogenic differentiation. BM-MSCs from PAO and THA patients revealed similar results regarding the onset of SA-β-gal expression and trilineage differentiation capacity.
We suggest that bone obtained during PAO is a promising novel source for BM-MSCs from young donors. Limited absolute cell yield due to low sample weight must be considered in early cell culture passages and might be critical for the range of clinical applications possible for BM-MSCs from this source. The higher proliferation capacity and increased growth factor secretion of BM-MSCs from young donors may be beneficial for future regenerative cell therapies, in vitro models, and tissue engineering.
摘要：
背景：骨髓间充质基质细胞（BM-MSCs）用于再生医学和涉及免疫调节的相关研究，抗炎,抗纤维化和再生功能。从在全髋关节置换术(THA)期间获得的样品中分离BM-MSC通常是可能的。大多数接受THA的患者的高龄和合并症限制了其适用性。我们的研究旨在通过分析细胞产量和细胞特征来评估髋臼周围截骨术（PAO）中获得的骨髓作为年轻供体BM-MSCs新来源的潜力。
方法：从PAO期间的前Os或耻骨上Os和初次THA期间的股体腔获得骨样本。通过密度梯度离心进行骨髓来源的单核细胞(BM-MNC)的分离。PAO和THA患者的样本在BM-MSC产量方面进行了比较，集落形成和BM-MSC在BM-MNC群体中的比例使用流式细胞术分析。基于BM-MSC特异性表面标志物的表达来表征细胞。通过定量解冻后的活力来比较细胞的功能。代谢活动，增殖能力,衰老相关β-半乳糖苷酶（SA-β-gal）表达，三系分化潜能和主要分泌体蛋白。
结果：当使用含有超过0.24g骨髓的PAO骨时，BM-MNC的分离可能以可靠和可重复的方式进行。PAO患者比THA组患者年轻。与在THA期间获得的BM-MSC相比,在PAO期间获得的骨含有较少的骨髓并且在第一次细胞培养传代后导致较低的BM-MSC数量。来自PAO样品的BM-MSC的特征在于更高的增殖能力。这导致在细胞培养传代2中更高的产量，当归一化为样本重量时。来自PAO患者的BM-MSC在成骨分化后显示TGF-β1，TIMP2和VEGF的分泌增加。来自PAO和THA患者的BM-MSC在SA-β-gal表达和三系分化能力方面表现出相似的结果。
结论：我们建议在PAO过程中获得的骨是年轻供体BM-MSCs的有希望的新来源。在早期细胞培养传代中必须考虑由于低样品重量而导致的有限的绝对细胞产量,并且对于来自该来源的BM-MSC的可能的临床应用范围可能是关键的。来自年轻供体的BM-MSCs更高的增殖能力和增加的生长因子分泌可能有利于未来的再生细胞治疗。体外模型，和组织工程。
方法：从PAO期间的前Os或耻骨上Os和初次THA期间的股体腔获得骨样本。通过密度梯度离心进行骨髓来源的单核细胞(BM-MNC)的分离。PAO和THA患者的样本在BM-MSC产量方面进行了比较，集落形成和BM-MSC在BM-MNC群体中的比例使用流式细胞术分析。基于BM-MSC特异性表面标志物的表达来表征细胞。通过定量解冻后的活力来比较细胞的功能。代谢活动，增殖能力,衰老相关β-半乳糖苷酶（SA-β-gal）表达，三系分化潜能和主要分泌体蛋白。
结果：当使用含有超过0.24g骨髓的PAO骨时，BM-MNC的分离可能以可靠和可重复的方式进行。PAO患者比THA组患者年轻。与在THA期间获得的BM-MSC相比,在PAO期间获得的骨含有较少的骨髓并且在第一次细胞培养传代后导致较低的BM-MSC数量。来自PAO样品的BM-MSC的特征在于更高的增殖能力。这导致在细胞培养传代2中更高的产量，当归一化为样本重量时。来自PAO患者的BM-MSC在成骨分化后显示TGF-β1，TIMP2和VEGF的分泌增加。来自PAO和THA患者的BM-MSC在SA-β-gal表达和三系分化能力方面表现出相似的结果。
结论：我们建议在PAO过程中获得的骨是年轻供体BM-MSCs的有希望的新来源。在早期细胞培养传代中必须考虑由于低样品重量而导致的有限的绝对细胞产量,并且对于来自该来源的BM-MSC的可能的临床应用范围可能是关键的。来自年轻供体的BM-MSCs更高的增殖能力和增加的生长因子分泌可能有利于未来的再生细胞治疗。体外模型，和组织工程。
