The First Affiliated Hospital, Fujian Medical University, Fuzhou 350005, China; Department of Chemistry, Beijing Key Laboratory of Microanalytical Methods and Instrumentation, The Key Laboratory of Bioorganic Phosphorus Chemistry & Chemical Biology, Tsinghua University, Beijing 100084, China.
Department of Chemistry, Beijing Key Laboratory of Microanalytical Methods and Instrumentation, The Key Laboratory of Bioorganic Phosphorus Chemistry & Chemical Biology, Tsinghua University, Beijing 100084, China.
Talanta. 2018 Feb 1;178:362-368. doi: 10.1016/j.talanta.2017.09.053. Epub 2017 Sep 21.
Osteosarcoma is the most common malignant tumour found in bones, and it has a poor prognosis. For improved therapy, it is significant to have a deep understanding of the proteomics changes in the cancer stem cells (CSCs) of osteosarcoma. Therefore, a comparative proteomics approach based on ultra-high-performance liquid chromatography coupled to an Orbitrap Fusion mass spectrometer (UHPLC-Orbitrap Fusion MS) was established to investigate the key molecular changes between CSCs and non-CSCs in human osteosarcoma HOS cells. A proteomic analysis was performed on these samples and a total of more than 6600 proteins were identified in each run. Moreover, most of the correlation coefficients between three biological repeats were more than 0.9 in each group. That finding demonstrated not only that the reproducibility of the test is good but also that the stability of this MS is excellent. A label-free quantitative method was applied to analyse differentially expressed proteins. Using the criteria of greater than 1.5-fold changes and a p value < 0.05, 124 proteins were identified as being significantly different between HOS-CSCs and non-CSCs. A pathway analysis of differentially expressed proteins by Ingenuity Pathway Analysis (IPA) revealed the potential molecular regulatory networks that may regulate CSCs. Selected differential α-actinin 4 (ACTN4) proteins were validated by Western blot assay. These findings enhance our understanding of the molecular changes in CSCs and may provide additional improvements in therapy for treating osteosarcoma. Moreover, the UHPLC-Orbitrap Fusion MS-based proteomics method is helpful in cancer research.
骨肉瘤是最常见的骨恶性肿瘤，预后较差。为了改善治疗效果，深入了解骨肉瘤肿瘤干细胞（CSC）中的蛋白质组学变化具有重要意义。因此，建立了一种基于超高效液相色谱与 Orbitrap Fusion 质谱联用（UHPLC-Orbitrap Fusion MS）的比较蛋白质组学方法，以研究人骨肉瘤 HOS 细胞中 CSC 与非 CSC 之间的关键分子变化。对这些样本进行蛋白质组学分析，每个运行中鉴定出的蛋白质超过 6600 种。此外，每组中三个生物学重复之间的大多数相关系数都大于 0.9。这一发现不仅表明该测试的重现性良好，而且还表明该 MS 的稳定性非常出色。应用无标记定量方法分析差异表达蛋白。使用大于 1.5 倍变化和 p 值 < 0.05 的标准，鉴定出 124 种蛋白在 HOS-CSC 和非 CSC 之间存在显著差异。通过 IPA 对差异表达蛋白进行通路分析，揭示了可能调节 CSC 的潜在分子调控网络。通过 Western blot 检测验证了选定的差异表达的α-辅肌动蛋白 4（ACTN4）蛋白。这些发现加深了我们对 CSC 中分子变化的理解，并可能为骨肉瘤的治疗提供额外的改善。此外，基于 UHPLC-Orbitrap Fusion MS 的蛋白质组学方法有助于癌症研究。
