【摘要】 该文探讨人皮肤成纤维细胞(NCF)和脐带间充质干细胞(hUC-MSCs)分泌蛋白的差异,从而初步阐明脐带间充质干细胞分泌蛋白对于治疗退行性疾病的可能分子机理。基于串联质谱(LC-MS/MS)的方法,收集用无血清培养基培养48 h的NCF和hUC-MSCs上清样品,进行TMT(tandem mass tags for relative and absolute quantitation)检测,从而对蛋白质酶解之后的肽段进行同位素标记,之后对分泌蛋白的表达差异进行精确的鉴定和定量。通过生物信息学分析,建立NCF和hUC-MSCs分泌蛋白谱图,并通过GO(gene ontology)对这些分泌蛋白进行功能分析,对于在蛋白分子互作图中处于中心位置的CCL2、COL4A1、TGFβ1、SERPINE1、SEMA7A我们进行了Real-time PCR定量分析。通过TMT以及生物信息学分析,该文鉴定出1 000多种共有蛋白,其中有704种蛋白在hUC-MSCs中的表达量比NCF中高。通过Real-time PCR,我们证实了CCL2、COL4A1在hUC-MSCs中的分泌量的确高于其在NCF中的分泌量,且差异具有显著性。通过检索文献以及Uniprot的相应数据库数据,该文发现,这两种分子在炎症的消除方面起到十分重要的作用,而炎症反应在帕金森病等退行性疾病的发生与发展过程中起到重要作用。通过对NCF和hUC-MSCs培养基中分泌的蛋白进行检测分析,该研究为探讨脐带间充质干细胞分泌蛋白治疗神经退行性疾病的可能分子机制提供启示。更多还原

【Abstract】 To elucidate the probable mechanism of hUC-MSCs (umbilical cord mesenchymal stem cells) secretory proteins in the treatment of neurodegenerative diseases,this study investigated the differences in secretory proteins between NCF (human skin fibroblasts) and hUC-MSCs.Based on LC-MS/MS (tandem mass spectrometry),the supernatant samples of hUC-MSCs and NCF cultured in serum-free medium for 48 hours were collected for TMT (tandem mass tags for relative and absolute quantitation) to detect peptides after protein digestion.Isotope labeling was performed,and then the difference in expression of secretory proteins was accurately identified and quantified.The secretory proteins of NCF and hUC-MSCs were established by bioinformatics analysis,and functional analysis of these secretory proteins were performed by GO (gene ontology).Proteins at the central position were analysed by Real-time PCR,such as CCL2,COL4A1,TGFβ1,SERPINE1 and SEMA7A.More than 1 000 consensus proteins were identified and the corresponding functions were searched by Uniprot.And the paper confirmed that the secretion of CCL2 and COL4A1 in hUC-MSCs were more than the secretion in NCF.The paper find that these two molecules play an important role in the elimination of inflammation,and the inflammatory response plays an important role in the occurrence and development of degenerative diseases such as Parkinson’s disease.By analyzing the proteins secreted in NCF and hUC-MSCs culture medium,this study lays the foundation for elucidating the differences in molecular mechanisms between mesenchymal stem cells and skin fibroblasts.更多还原