摘要
The safe use of intraoperative blood salvage （IBS） in cancer surgery remains controversial. Here, we investigated the kil ing effect of cisplatin combined with hyperthermia on human hepatocarcinoma （HepG2） cel s and erythrocytes from IBS in vitro. HepG2 cel s were mixed with concentrated erythrocytes and pretreated with cisplatin （50, 100, and 200μg/ml） alone at 37 °C for 60 min and cisplatin （25, 50, 100, and 200μg/ml） combined with hyperthermia at 42 °C for 60 min. After pretreatment, the cel viability, colony formation and DNA metabolism in HepG2 and the Na＋-K＋-ATPase activity, 2,3-diphosphoglycerate （2,3-DPG） concentration, free hemoglobin （Hb） level, osmotic fragility, membrane phosphatidylserine externalization, and blood gas variables in erythrocytes were determined. Pretreatment with cisplatin （50, 100, and 200μg/ml） combined with hyperthermia （42 °C） for 60 min significantly decreased HepG2 cel viability, and completely inhibited colony formation and DNA metabolism when the HepG2 cel concentration was 5×104 ml？1 in the erythrocyte （P0.05）. In conclusion, pre-treatment with cisplatin （50μg/ml） combined with hyperthermia （42 °C） for 60 min effectively eliminated HepG2 cel s from IBS but did not significantly affect erythrocytes in vitro.
目的:研究离体热处理和顺铂联合热处理对术中回收血红细胞功能的影响及其中混杂的肝肿瘤细胞株(HepG2)的杀灭作用。创新点:(1)采用多种评价指标研究了不同时间离体热处理对术中回收血中混杂的HepG2的杀灭作用及对红细胞的影响,并确定了对红细胞安全且能有效杀灭HepG2的离体热处理时间。(2)从多个角度评价了离体顺铂联合热处理对术中回收血红细胞的影响及对其中混杂的HepG2的杀灭作用,确定了该方案中对红细胞安全且能有效去除HepG2的顺铂浓度。方法:采用3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐(MTT)、5-乙炔基-2'脱氧尿嘧啶核苷(EdU)和平板克隆形成评估HepG2的细胞存活率、DNA复制率和克隆形成能力(图1和3);从红细胞渗透脆性、携氧能力(2,3-二磷酸甘油酸(2,3-DPG)、半饱和氧分压(P50))、能量代谢(Na+-K+-ATPase、pH)、膜完整性(游离血红蛋白(Hb)、血清K+和Na+浓度、细胞膜磷脂酰丝氨酸外翻比例)等角度评估红细胞功能(图2和4;表3)。结论:肝肿瘤术中回收血经离体顺铂联合热处理(42°C,50μg/ml)60min后,能有效清除其中混杂的HepG2,但对红细胞无显著影响,值得体内进一步研究顺铂热处理有效应用于肿瘤手术自体血液回输的安全方案。
基金
Project supported by the Scientific Research from Chinese Ministryof Health-Zhejiang Health Department,China(Nos.WKJ2008-2-021and WKJ2013-2-019)
