Supplementary Material for: Assessment of micronuclei frequency in the peripheral blood of adult and pediatric patients receiving fractionated total body irradiation

The cytokinesis-block micronucleus (CBMN) assay is an established method for assessing chromosome damage in human peripheral blood lymphocytes resulting from exposure to genotoxic agents such as ionizing radiation. The objective of this study is to measure cytogenetic DNA damage and hematology parameters in vivo based on MN frequency in peripheral blood lymphocytes (PBLs) from adult and pediatric leukemia patients undergoing hematopoietic stem cell transplantation preceded by total body irradiation (TBI) as part of the conditioning regimen. CBMN assay cultures were prepared from fresh blood samples collected before and at 4- and 24- h after the start of TBI, corresponding to doses of 1.25 Gy and 3.75 Gy, respectively. For both age groups, there was a significant increase in MN yields with increasing dose (p < 0.05) and dose-dependent decrease in the nuclear division index (NDI; p < 0.0001). In the pre-radiotherapy samples, there was a significantly higher NDI measured in the pediatric cohort compared to the adult due to an increase in the percentage of tri- and quadra-nucleated cells scored. Complete blood counts with differential recorded before and after TBI at the 24 h time point, showed a rapid increase in neutrophil (p = 0.0001) and decrease in lymphocyte (p = 0.0006) counts, resulting in a highly elevated neutrophil-to-lymphocyte (NRL) ratio of 14.45 ± 1.85 after 3.75 Gy TBI (pre-exposure = 4.62 ± 0.49), indicating a strong systemic inflammatory response. Correlation of the hematological cell subset counts with cytogenetic damage, indicated that only the lymphocyte subset survival fraction (after-TBI compared with before-TBI) showed a negative correlation with increasing MN frequency from 0 to 1.25 Gy (r = -0.931; p = 0.007). Further, the data presented here indicate that the combination of CBMN assay endpoints (MN frequency and NDI values) and hematology parameters could be used to assess cytogenetic damage and early hematopoietic injury in the peripheral blood of leukemia patients, 24 h after TBI exposure

胞质分裂阻滞微核（cytokinesis-block micronucleus, CBMN）试验是一种成熟的方法，用于评估人类外周血淋巴细胞因暴露于电离辐射等遗传毒性剂而产生的染色体损伤。本研究的目标是基于接受造血干细胞移植的成人与儿童白血病患者外周血淋巴细胞（peripheral blood lymphocytes, PBLs）中的微核频率，在体内评估细胞遗传学DNA损伤与血液学参数；此类患者在预处理方案中采用了全身照射（total body irradiation, TBI）作为预处理环节。CBMN试验的培养物取自全身照射开始前、照射后4小时与24小时采集的新鲜血液样本，对应照射剂量分别为1.25 Gy与3.75 Gy。两个年龄组的微核产率均随剂量升高呈现显著升高（p < 0.05），而核分裂指数（nuclear division index, NDI）则呈剂量依赖性降低（p < 0.0001）。在放疗前样本中，儿童队列的NDI检测值显著高于成人组，这归因于计数的三核与四核细胞百分比升高。在全身照射前与照射后24小时采集的全血细胞计数及分类检测结果显示，中性粒细胞计数快速升高（p = 0.0001），淋巴细胞计数则出现下降（p = 0.0006），最终在接受3.75 Gy TBI后，中性粒细胞与淋巴细胞比值（NRL）为14.45 ± 1.85，显著高于照射前的4.62 ± 0.49，提示存在强烈的全身性炎症反应。血液细胞亚群计数与细胞遗传学损伤的相关性分析表明，仅淋巴细胞亚群的存活分数（照射后与照射前的比值）与0至1.25 Gy剂量下微核频率的升高呈负相关（r = -0.931; p = 0.007）。此外，本研究数据表明，结合CBMN试验的终点指标（微核频率与NDI值）与血液学参数，可用于评估白血病患者在接受全身照射后24小时外周血中的细胞遗传学损伤与早期造血损伤。