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目的:利用傅里叶变换红外光谱(Fourier transformed infrared spectrum,FT-IR)技术探讨肿瘤细胞共培养对不同分化阶段树突状细胞(dendritic cells,DCs)的脂含量和蛋白质二级结构的影响,为深入理解肿瘤的免疫逃逸机制寻找线索。方法:免疫磁珠法分离人CD14+单核细胞,以经典方法将其诱导为未成熟DCs(immature DCs,imDCs)和成熟DCs(mature DCs,mDCs),分别将imDCs和mDCs与肝癌细胞株BEL7402、红白血病细胞株K562以及人脐静脉内皮细胞(human umbilical vein endothelialcells,HUVECs)共培养48 h,正常培养的DCs作为对照。采用FT-IR技术研究不同肿瘤细胞对不同分化阶段DCs的脂含量和蛋白质二级结构的影响。结果:与正常培养的DCs相比,与肿瘤细胞BEL7402和K562共培养后imDCs和mDCs的膜磷脂成分减少(2.718±0.296,3.124±0.361vs4.855±0.324,P<0.05;2.964±0.136,3.522±0.173vs4.217±0.206,P<0.05),而总的脂含量增加(3.768±0.185,3.591±0.197vs2.487±0.212,P<0.05;4.288±0.156,4.155±0.167vs3.233±0.206,P<0.05);蛋白质α-螺旋含量减少(1.863±0.192,1.754±0.169vs2.364±0.188,P<0.05;1.124±0.133,1.016±0.107vs1.392±0.113,P<0.05),β-折叠(3.397±0.225,3.433±0.236vs2.486±0.198,P<0.05;2.646±0.209,2.591±0.216vs1.558±0.159,P<0.05)和转角含量(4.366±0.284,4.322±0.266vs3.127±0.272,P<0.05;2.675±0.221,2.627±0.235vs1.773±0.181,P<0.05)增加;并且mDCs比imDCs更容易受到肿瘤来源因素的影响。结论:与肿瘤细胞共培养能够导致mDCs和imDCs的脂含量和蛋白质二级结构发生改变,可能是肿瘤导致DCs功能损伤的结构基础。
OBJECTIVE: To investigate the effect of tumor cell co-culture on lipid content and protein secondary structure of dendritic cells (DCs) in different differentiation stages by using Fourier transform infrared spectroscopy (FT-IR) To understand the mechanism of tumor immune escape to find clues. Methods: The human CD14 + monocytes were isolated by immunomagnetic beads method and induced by immature DCs (imDCs) and mature DCs (mDCs) by classical methods. The imDCs and mDCs were respectively compared with hepatocellular carcinoma cell lines BEL7402, The erythrocytic leukemia cell line K562 and human umbilical vein endothelial cells (HUVECs) were co-cultured for 48 h. Normal cultured DCs served as controls. The effects of different tumor cells on lipid content and protein secondary structure of DCs in different stages of differentiation were studied by FT-IR. RESULTS: Compared with normal DCs, the phospholipid components of imDCs and mDCs decreased after co-cultured with tumor cells BEL7402 and K562 (2.718 ± 0.296, 3.124 ± 0.361 vs 4.855 ± 0.324, P <0.05; 2.964 ± 0.136, 3.522 ± 0.173 vs 4.217 ± 0.206, P <0.05), while the total lipids increased (3.768 ± 0.185, 3.591 ± 0.197 vs 2.487 ± 0.212, P <0.05; 4.288 ± 0.156, 4.155 ± 0.167 vs 3.233 ± 0.206, (P <0.05); protein α-helix content decreased (1.863 ± 0.192,1.754 ± 0.169vs2.364 ± 0.188, P <0.05; 1.124 ± 0.133,1.016 ± 0.107vs1.392 ± 0.113, P <0.05) (3.397 ± 0.225,3.433 ± 0.236vs2.486 ± 0.198, P <0.05; 2.646 ± 0.209,2.591 ± 0.216vs1.558 ± 0.159, P <0.05) and the content of rotation angle (4.366 ± 0.284,4.322 ± 0.266vs3.127 ± 0.272, P <0.05; 2.675 ± 0.221, 2.627 ± 0.235vs1.773 ± 0.181, P <0.05); and mDCs were more likely to be affected by tumor-derived factors than imDCs. CONCLUSION: Co-culture with tumor cells can lead to the changes of lipid content and secondary structure of proteins in mDCs and imDCs, which may be the structural basis of tumor-induced functional impairment of DCs.