Ehlicke Franziska, Köster Natascha, Salzig Denise, Czermak Peter
Institute of Bioprocess Engineering and Pharmaceutical Technology, University of Applied Sciences Mittelhessen, Wiesenstr 14, 35390 Giessen, Germany.
Department Tissue Engineering and Regenerative Medicine, University Hospital Wuerzburg, Roentgenring 11, 97070 Wuerzburg, Germany.
Open Biomed Eng J. 2017 Jul 31;11:72-84. doi: 10.2174/1874120701711010072. eCollection 2017.
The most common cause of lower back pain is the pathological degeneration of the nucleus pulposus (NP). Promising NP regeneration strategies involving human mesenchymal stem cells (hMSCs) would require specific markers to confirm successful differentiation into the NP lineage and to distinguish the articular cartilage (AC).
We sought specific NP mRNA markers that are upregulated in native NP cells but not in dedifferentiated NP cells, undifferentiated hMSCs or chondrocytes. We also considered the suitability of non-invasive Raman spectroscopy to distinguish among these classes of cells.
We used quantitative real-time PCR and Raman spectroscopy to analyse undifferentiated hMSCs in monolayers and embedded in hydrogels, and compared the results with dedifferentiated and redifferentiated human NP and AC cells.
The redifferentiation of NP cells induced the expression of annexin A3 (), collagen type II () and proteoglycan mRNAs, whereas the redifferentiation of AC cells only induced proteoglycan expression. Redifferentiated NP cells expressed higher levels of , , paired box 1 () and mRNA than redifferentiated AC cells. Redifferentiated NP cells and undifferentiated hMSC-TERT cells expressed similar amount of mRNA, indicating that only , and are promising markers for redifferentiated NP cells. Raman spectra clearly differed among the three cell types and highlighted their differentiation status.
We recommend , and as markers to determine the success of hMSC-based differentiation to regenerate NP cells. Raman spectroscopy can be used to determine cell type and differentiation status especially in the context of clinical trials.
下背部疼痛最常见的原因是髓核（NP）的病理性退变。涉及人间充质干细胞（hMSCs）的有前景的NP再生策略需要特定标志物来确认成功分化为NP谱系并区分关节软骨（AC）。
我们寻找在天然NP细胞中上调但在去分化NP细胞、未分化hMSCs或软骨细胞中未上调的特定NP mRNA标志物。我们还考虑了非侵入性拉曼光谱区分这些细胞类型的适用性。
我们使用定量实时PCR和拉曼光谱分析单层和嵌入水凝胶中的未分化hMSCs，并将结果与去分化和再分化的人NP及AC细胞进行比较。
NP细胞的再分化诱导了膜联蛋白A3（）、II型胶原蛋白（）和蛋白聚糖mRNA的表达，而AC细胞的再分化仅诱导了蛋白聚糖表达。再分化的NP细胞比再分化的AC细胞表达更高水平的、、配对盒1（）和mRNA。再分化的NP细胞和未分化的hMSC-TERT细胞表达相似量的mRNA，表明只有、和是再分化NP细胞有前景的标志物。三种细胞类型的拉曼光谱明显不同，并突出了它们的分化状态。
我们推荐、和作为标志物来确定基于hMSC的分化以再生NP细胞是否成功。拉曼光谱可用于确定细胞类型和分化状态，特别是在临床试验背景下。
