Laboratory Study for Analysis of Intra-tumor Heterogeneity of Ovarian Cancer by Fourier Transform Infrared Spectroscopy
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摘要:
目的 研究卵巢癌异质性细胞群傅里叶变换红外光谱(fourier transform infrared spectroscopy, FTIR)的异同, 为异质性卵巢癌的临床诊断提供新的途径。 方法 对由人卵巢癌细胞系A2780所建立的高、低侵袭/迁移能力的细胞亚群(A-H和A-L)进行裸鼠体内成瘤实验和体外Transwell实验鉴定, 利用FTIR技术进行检测, 使用Omnic软件将二者的红外光谱进行对照解析。 结果 A-H与A-L相比, 细胞侵袭性更强(2.211±0.256比0.149±0.097, P=0.0003);迁移活性更高(2.412±0.298比0.337±0.079, P=0.0005);成瘤体积更大[(2.42±0.43)cm比(0.63±0.22)cm, P < 0.0001]。红外光谱图显示, A-H细胞核酸吸收峰1086和1240 cm-1强度比A-L更高(0.359±0.014比0.323±0.018, P=0.020;0.343±0.010比0.317±0.012, P=0.021);蛋白质酰胺Ⅰ带1655 cm-1的吸收峰强度更高(1.710±0.024比1.640±0.017, P=0.045);与脂类相关的2925和2853 cm-1谱带吸收峰强度更高(0.669±0.025比0.613±0.016, P=0.034;0.397±0.008比0.376±0.010, P=0.039)。A-H细胞蛋白质二级结构β-折叠和转角结构的含量比A-L细胞更高(22.50±0.54比21.30±0.50, P=0.037;50.00±1.12比47.70±0.76, P=0.029), 无规卷曲含量更低(8.50±0.82比11.50± 1.32, P=0.011)。 结论 FTIR可以快速揭示异质性卵巢癌细胞群的分子组成和结构方面的差异, 为其表征提供可靠信息, 可能为临床诊断和预后判断提供新的途径。 Abstract:Objective Our study aimed to establish a potential way of fourier transform infrared(FTIR) spectra to diagnose malignant tumors. Methods Ovarian cancer cell subclones with high(A-H) or low(A-L) malignant potential were confirmed by Matrigel invasive/migratory assay in vitro and the tumor xenograft model in vivo. FTIR spectra of cells was recorded and analyzed in the region from 600 cm-1 to 4000 cm-1. Results A-Hcells were confirmed to have higher invasive and migratory activities in vitro compared with A-L cells(2.211±0.256 vs. 0.149±0.097, P=0.0003; 2.412±0.298 vs. 0.337±0.079, P=0.0005).A-H-derived tumors were significantly larger than A-L-derived tumors(2.42 cm±0.43 cm vs. 0.63 cm±0.22 cm, P < 0.0001). Significant spectral differences between A-H and A-L cells were observed in the band intensity of 1086 cm-1 and 1240 cm-1(0.359±0.014 vs. 0.323±0.018, P=0.020; 0.343±0.010 vs. 0.317±0.012, P=0.021), as well as in that of 1655 cm-1(1.710±0.024 vs. 1.640±0.017, P=0.045), 2925 cm-1(0.669±0.025 vs.0.613±0.016, P=0.034) and 2853 cm-1(0.397±0.008 vs. 0.376±0.010, P=0.039), which indicated higher amounts and synthesis of nucleic acids, proteins and lipids in A-H cells. The amount of secondary protein structures of β-sheet and turns in A-H was higher than that in A-L(22.50±0.54 vs. 21.30±0.50, P=0.037; 50.00±1.12 vs. 47.70±0.76, P=0.029). The amount of random coil component of protein in A-H was lower than that in A-L(8.50±0.82 vs. 11.50±1.32, P=0.011). Conclusions FTIR quickly provides important and reliable information about heterogeneous ovarian cancerous cells on the difference in molecular components and structures. The findings of this study demonstrate the potential use of infrared spectroscopy in differentiating ovarian tumors at different developing stages. -
Key words:
- fourier transform infrared /
- ovarian cancer /
- tumor heterogeneity /
- clinical diagnoses
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图 2 卵巢癌细胞红外光谱的谱带指认
A.核酸;B.磷脂;C.A-H;D.A-L;E.白蛋白A-H、A-L:同图 1
图 3 蛋白质酰胺Ⅰ带的二阶导数和分峰图谱
A、C.1740~1600 cm-1区域谱带的二阶导数图谱;B、D.1740~1600 cm-1区域谱带的分峰图谱A-H、A-L:同图 1
表 1 A-H和A-L细胞裸鼠成瘤实验结果比对
细胞亚群 成瘤时间$ \left( {\overline x \pm s} \right) $ 成瘤率(%) 肿瘤大小$ \left( {\overline x \pm s} \right) $ A-H 7.7±1.9 100 2.42±0.43 A-L 23.0±2.0 20 0.63±0.22 P值 < 0.0001 0.0067 < 0.0001 A-H、A-L:同图 1 表 2 卵巢癌细胞A-H和A-L的FTIR光谱图的峰带指认
波数(cm-1) 指认 细胞成分 3300 νN-H 蛋白质 2958 νasCH3 蛋白质和脂类 2925 νasCH2 蛋白质和脂类 2875 νsCH3 蛋白质和脂类 2853 νsCH2 脂类 1741 νC=O 脂类 1655 酰胺Ⅰ 蛋白质 1547 酰胺Ⅱ 蛋白质 1460 δCH2, 等 蛋白质和脂类等 1400 δC-H, δC-O-H, 等 蛋白质和脂类等 1240 νC-N, νP-O 蛋白质、核酸和磷脂 1086 νC-O, νP-O 糖原、核酸和磷脂 A-H、A-L:同图 1;FTIR:傅里叶变换红外光谱 表 3 A-H和A-L各谱带相对强度的比较
峰强比(Ⅰ/Ⅰ) A-H A-L P值 2925/1547 0.669±0.025 0.613±0.016 0.034 2853/1547 0.397±0.008 0.376±0.010 0.039 1655/1547 1.710±0.024 1.640±0.017 0.045 1240/1547 0.343±0.010 0.317±0.012 0.021 1086/1547 0.359±0.014 0.323±0.018 0.020 A-H、A-L:同图 1 表 4 A-H和A-L在1740~1600 cm-1区域谱带各子峰的峰位和峰面积
峰位(cm-1) 峰面积(%) 指认 A-H A-L A-H A-L 1629 1628 7.00±0.37 7.30±0.33 β-折叠 1636 1636 15.50±0.62 14.00±0.24 β-折叠 22.50±0.54 21.30±0.50 β-折叠结构总含量 1643 1643 8.50±0.82 11.50±1.32 无规卷曲 1649 1649 19.00±1.10 19.30±0.79 α-螺旋 1658 1658 19.50±0.22 20.70±0.41 转角 1667 1667 16.10±0.57 15.30±0.40 转角 1676 1676 4.90±1.08 7.50±1.55 转角 1681 1682 9.60±2.44 4.20±1.22 转角 50.10±1.12 47.70±0.76 转角结构总含量 A-H、A-L:同图 1 -
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