PDF(Pubmed)
Abstract：
The lack of appropriate mesenchymal stem cells (MSCs) selection methods has given the challenges for standardized harvesting, processing, and phenotyping procedures of MSCs. Genetic engineering coupled with high-throughput proteomic studies of MSC surface markers arises as a promising strategy to identify stem cell-specific markers. However, the technical limitations are the key factors making it less suitable to provide an appropriate starting material for the screening platform. A more accurate, easily accessible approach is required to solve the issues.
This study established a high-throughput screening strategy with forward versus side scatter gating to identify the adipogenesis-associated markers of bone marrow-derived MSCs (BMSCs) and tonsil-derived MSCs (TMSCs). We classified the MSC-derived adipogenic differentiated cells into two clusters: lipid-rich cells as side scatter (SSC)-high population and lipid-poor cells as SSC-low population. By screening the expression of 242 cell surface proteins, we identified the surface markers which exclusively found in lipid-rich subpopulation as the specific markers for BMSCs and TMSCs.
High-throughput screening of the expression of 242 cell surface proteins indicated that CD49f and CD146 were specific for BMSCs and TMSCs. Subsequent immunostaining confirmed the consistent specific expression of CD49f and CD146 and in BMSCs and TMSCs. Enrichment of MSCs by CD49f and CD146 surface markers demonstrated that the simultaneous expression of CD49f and CD146 is required for adipogenesis and osteogenesis of mesenchymal stem cells. Furthermore, the fate decision of MSCs from different sources is regulated by distinct responses of cells to differentiation stimulations despite sharing a common CD49f+CD146+ immunophenotype.
We established an accurate, robust, transgene-free method for screening adipogenesis associated cell surface proteins. This provided a valuable tool to investigate MSC-specific markers. Additionally, we showed a possible crosstalk between CD49f and CD146 modulates the adipogenesis of MSCs.
This study established a high-throughput screening strategy with forward versus side scatter gating to identify the adipogenesis-associated markers of bone marrow-derived MSCs (BMSCs) and tonsil-derived MSCs (TMSCs). We classified the MSC-derived adipogenic differentiated cells into two clusters: lipid-rich cells as side scatter (SSC)-high population and lipid-poor cells as SSC-low population. By screening the expression of 242 cell surface proteins, we identified the surface markers which exclusively found in lipid-rich subpopulation as the specific markers for BMSCs and TMSCs.
High-throughput screening of the expression of 242 cell surface proteins indicated that CD49f and CD146 were specific for BMSCs and TMSCs. Subsequent immunostaining confirmed the consistent specific expression of CD49f and CD146 and in BMSCs and TMSCs. Enrichment of MSCs by CD49f and CD146 surface markers demonstrated that the simultaneous expression of CD49f and CD146 is required for adipogenesis and osteogenesis of mesenchymal stem cells. Furthermore, the fate decision of MSCs from different sources is regulated by distinct responses of cells to differentiation stimulations despite sharing a common CD49f+CD146+ immunophenotype.
We established an accurate, robust, transgene-free method for screening adipogenesis associated cell surface proteins. This provided a valuable tool to investigate MSC-specific markers. Additionally, we showed a possible crosstalk between CD49f and CD146 modulates the adipogenesis of MSCs.
摘要：
背景：缺乏适当的间充质干细胞（MSC）选择方法给标准化收获带来了挑战，processing,和MSCs的表型程序。与MSC表面标记的高通量蛋白质组学研究相结合的基因工程作为鉴定干细胞特异性标记的有希望的策略出现。然而,技术限制是使其不太适合为筛选平台提供合适的起始材料的关键因素。更准确的，需要容易获得的方法来解决问题。
方法：本研究建立了一种高通量筛选策略，正向与侧向散射门控，以鉴定骨髓来源的MSCs（BMSCs）和扁桃体来源的MSCs（TMSCs）的脂肪形成相关标志物。我们将MSC衍生的脂肪分化细胞分为两个簇：富含脂质的细胞为侧向散射（SSC）高群体，而贫脂细胞为SSC低群体。通过筛选表达242种细胞表面蛋白,我们确定了仅在富含脂质的亚群中发现的表面标志物作为BMSCs和TMSCs的特异性标志物。
结果：高通量筛选242种细胞表面蛋白的表达表明，CD49f和CD146对BMSCs和TMSCs具有特异性。随后的免疫染色证实了CD49f和CD146在BMSCs和TMSCs中的一致特异性表达。通过CD49f和CD146表面标记富集MSCs表明，CD49f和CD146的同时表达是间充质干细胞的脂肪形成和成骨所必需的。此外，尽管具有共同的CD49f+CD146+免疫表型,但来自不同来源的MSCs的命运决定受细胞对分化刺激的不同反应调节.
结论：我们建立了准确的，健壮,筛选脂肪生成相关细胞表面蛋白的无转基因方法。这为研究MSC特异性标志物提供了有价值的工具。此外,我们显示CD49f和CD146之间可能的串扰调节MSCs的脂肪形成。
方法：本研究建立了一种高通量筛选策略，正向与侧向散射门控，以鉴定骨髓来源的MSCs（BMSCs）和扁桃体来源的MSCs（TMSCs）的脂肪形成相关标志物。我们将MSC衍生的脂肪分化细胞分为两个簇：富含脂质的细胞为侧向散射（SSC）高群体，而贫脂细胞为SSC低群体。通过筛选表达242种细胞表面蛋白,我们确定了仅在富含脂质的亚群中发现的表面标志物作为BMSCs和TMSCs的特异性标志物。
结果：高通量筛选242种细胞表面蛋白的表达表明，CD49f和CD146对BMSCs和TMSCs具有特异性。随后的免疫染色证实了CD49f和CD146在BMSCs和TMSCs中的一致特异性表达。通过CD49f和CD146表面标记富集MSCs表明，CD49f和CD146的同时表达是间充质干细胞的脂肪形成和成骨所必需的。此外，尽管具有共同的CD49f+CD146+免疫表型,但来自不同来源的MSCs的命运决定受细胞对分化刺激的不同反应调节.
结论：我们建立了准确的，健壮,筛选脂肪生成相关细胞表面蛋白的无转基因方法。这为研究MSC特异性标志物提供了有价值的工具。此外,我们显示CD49f和CD146之间可能的串扰调节MSCs的脂肪形成。
