Cytotoxicity of anti-tumor herbal Marsdeniae tenacissimae extract on erythrocytes

Abstract

Marsdeniae tenacissimae extract (MTE) has been used as an adjuvant medicine for cancer therapy for a long time. Although massive studies demonstrated its considerable anti-cancer effect, there is no research on its influence on erythrocytes, which are firstly interacted with MTE in the circulation. To investigate the influence of MTE on erythrocytes, we used a flow cytometer to detect the MTE-treated alternations of morphology, calcium concentration, and reactive oxygen species (ROS) level in erythrocytes. We used hemolysis under different osmotic solutions to evaluate the fragility of erythrocytes. Data showed that MTE treatment dose-dependently increased the ratio of erythrocyte fragmentation (P<0.001) and shrinking, and elevated the forward scatter (FSC) value (P<0.001) and calcium accumulation (P<0.001). MTE induced ROS production of erythrocytes under the high glucose condition (P<0.01) and consequently caused a rise in fragility (P<0.05). These results suggest that MTE induces cytotoxicity and aging in erythrocytes in a dose-dependent manner, and presents the possibility of impairment on cancer patients’ circulating erythrocytes when MTE is used as an anti-cancer adjuvant medicine.

中文概要

题目

乌骨藤提取物对人红细胞的毒性研究

目的

评估乌骨藤提取物对红细胞形态及生理功能的影响,并初步探讨其作用机制。

创新点

首次检测乌骨藤提取物对红细胞的毒性作用,并 且初步探索此毒性作用与乌骨藤导致的红细胞 内活性氧(ROS)和钙离子水平的相关性。

方法

不同浓度乌骨藤提取物(0、64、128 和256 μg/ml) 处理红细胞24 h,通过流式细胞术检测前向角散 射(FSC)和侧向角散射(SSC),计算红细胞破 碎率和大小改变,通过显微镜观察形态学的变 化;装载2',7'-二氯荧光黄双乙酸盐(DCFH-DA) 和钙荧光探针Fluo-4 染料,通过流式细胞术分别 检测红细胞ROS 和钙离子荧光强度的变化;通过 观察一系列低渗生理盐水下红细胞的溶胀情况, 评估乌骨藤提取物对红细胞脆性的改变。

结论

乌骨藤提取物剂量依赖性地增加红细胞的破碎率 (图1),导致FSC 增加(图2),同时高浓度乌 骨藤提取物(256 μg/ml)导致红细胞圆盘状结构 消失(图3);128 和256 μg/ml 乌骨藤提取物处 理红细胞导致胞内钙离子显著增加(图4);乌骨 藤提取物在高糖环境中诱导红细胞ROS 累计,并 最终导致红细胞脆性增加(图5)。上述结果显示, 乌骨藤提取物对红细胞有毒性作用,并有可能在 临床抗癌治疗中破坏红细胞引起潜在的毒副作 用(图6)。