Gezer Ugur, Mert Ufuk, Ozgür Emre, Yörüker Ebru E, Holdenrieder Stefan, Dalay Nejat
Department of Basic Oncology, Oncology Institute, Istanbul University, Capa 34390, Istanbul, Turkey.
Oncol Lett. 2012 May;3(5):1095-1098. doi: 10.3892/ol.2012.600. Epub 2012 Feb 10.
Circulating DNA is present in plasma/serum, mainly complexed with histones as nucleosomes. The detection of circulating nucleosomes (cNUCs) in the peripheral blood may be a diagnostic modality for cancer-associated changes of modified histone tails in blood circulation. In the present study, we investigated the correlation between the trimethylation of H3 lysine 9 (H3K9me3) and H4 lysine 20 (H4K20me3), which are hallmarks of pericentric heterochromatin, and cNUCs in healthy subjects and patients with colorectal cancer (CRC) and multiple myeloma (MM). The plasma concentration of cNUCs was measured using the Cell-Death Detection ELISA kit. Histone methylation marks were detected using chromatin immunoprecipitation (ChIP), followed by quantitative PCR with pericentric satellite 2 as the target sequence. The results showed a high variation in the concentrations of cNUCs, with healthy subjects exhibiting the lowest levels (median 0.194), the CRC patients intermediate (median 0.25) and the MM patients the highest levels (median 0.648). However, the differences between the groups did not reach statistical significance (p>0.05). Analysis using the Pearson's correlation test revealed a significant positive correlation between the concentration of cNUCs and H3K9me3 and H4K20me3 in the whole study group (N=57, p<0.001 for both histone marks). A study of the correlation between cNUCs and histone marks in the individual study groups demonstrated the correlation between cNUCs and H3K9me3 in CRC patients to be weak (p=0.046), indicating that circulating H3K9me3 may be modified in CRC patients. The histone marks were normalized using the values of cNUCs. In agreement with the weak correlation between cNUCs and H3K9me3 in CRC patients, H3K9me3 levels (median 0.047) were lowest in this group compared with the other two groups (0.06 in healthy subjects, 0.2 in MM patients, p = not significant). For H4K20me3, the median values were 0.022 in healthy subjects, 0.052 in CRC patients and 0.056 in MM patients. In conclusion, our findings indicate a marked correlation between cNUCs and histone methyl marks.
循环DNA存在于血浆/血清中，主要与组蛋白结合形成核小体。检测外周血中的循环核小体（cNUCs）可能是一种诊断血液循环中组蛋白尾巴修饰的癌症相关变化的方法。在本研究中，我们调查了健康受试者、结直肠癌（CRC）患者和多发性骨髓瘤（MM）患者中，作为着丝粒周围异染色质标志的H3赖氨酸9（H3K9me3）和H4赖氨酸20（H4K20me3）的三甲基化与cNUCs之间的相关性。使用细胞死亡检测ELISA试剂盒测量cNUCs的血浆浓度。使用染色质免疫沉淀（ChIP）检测组蛋白甲基化标记，随后以着丝粒卫星2为靶序列进行定量PCR。结果显示，cNUCs浓度变化很大，健康受试者水平最低（中位数0.194），CRC患者居中（中位数0.25），MM患者水平最高（中位数0.648）。然而，各组之间的差异未达到统计学显著性（p>0.05）。使用Pearson相关检验分析显示，在整个研究组（N=57，两种组蛋白标记的p均<0.001）中，cNUCs浓度与H3K9me3和H4K20me3之间存在显著正相关。对各个研究组中cNUCs与组蛋白标记之间相关性的研究表明，CRC患者中cNUCs与H3K9me3之间的相关性较弱（p=0.046），表明CRC患者循环中的H3K9me3可能发生了修饰。使用cNUCs的值对组蛋白标记进行标准化。与CRC患者中cNUCs与H3K9me3之间的弱相关性一致，该组中H3Kme3水平（中位数0.047）低于其他两组（健康受试者为0.06，MM患者为0.2，p无显著性）。对于H4K20me3，健康受试者的中位数为0.022，CRC患者为0.052，MM患者为0.056。总之，我们的研究结果表明cNUCs与组蛋白甲基化标记之间存在显著相关性。
