实时xCELLigence细胞分析系统在药物心脏毒性筛选中的应用

药物的心脏毒性越来越受到关注,大量临床上使用的药物由于心脏毒性而被撤市或监管。因此,药物安全评价是研发过程中非常重要的一步。实时xCELLigence细胞分析系统是一种基于电子阻抗的非标记性、非侵入性、实时动态的细胞分析检测技术,它可以广泛地应用于药物的毒性筛选中。xCELLigence RTCA Cardio系统具有高的时间分辨率,能灵敏地、精确地监测到药物作用于心肌细胞后心肌细胞搏动频率、节律、幅度等指标的改变,并通过Poincaréplots分析其心率变异性。这些指标均与药物的心脏毒性有着密切的关系。因此,该系统具有高的特异性,能在药物研发的早期阶段筛选其心脏毒性。本文对RTCA的基本原理和其在药物心脏毒性筛选中的应用作一综述。

Dynamic monitoring of the cytotoxic effects of protoberberine alkaloids from Rhizoma Coptidis on HepG2 cells using the xCELLigence system

AIM: To investigate the cytotoxic effects of the six protoberberine alkaloids(PAs) from Rhizoma Coptidis on HepG2 cells. METHOD: A systematic screening was conducted to investigate the dynamic response of HepG2 cells to the PAs using the impedance-based xCELLigence system. Cisplatin was selected as the positive control. The real time, concentration-response curves and the 50% inhibitory concentrations(IC50) were acquired to evaluate the anticancer activity of the PAs. RESULTS: All of the six PAs inhibited cell growth and induce death in HepG2 cells in a time- and concentration-dependent manner. The IC50 values of cisplatin, berberine, columbamine, coptisine, epiberberine, jatrorrhizine, and palmatine were 5.13, 42.33, 226.54, 36.90, 302.72, 383.54, and 456.96 μg·mL-1, respectively. The results obtained using the xCELLigence system corresponded well with those of the conventional methods. CONCLUSION: The xCELLigence system is a reliable and efficient tool for real-time screening of the cytotoxic effect of compounds in cell-based in vitro assays. Coptisine and berberine, with methylenedioxy group at C2 and C3 on the phenyl ring showed stronger effect.than the other four PAs. However, compared with cisplatin, the six PAs didn't show obvious cytotoxic effect on HepG2 cells.These results provided some useful data for the evaluation of the anticancer compounds, and the clinical application of traditional Chinese medicine.

Effects of tyrosine kinase inhibitor E7080 and eNOS inhibitor L-NIO on colorectal cancer alone and in combination

Objective：To investigate the effects of E7080 and N5-（1-iminoethyl）-L-ornithine dihydrochloride （L-NIO）on colorectal cancer alone and in combination.Methods：HT29 colorectal cancer cell line from Sap Institute was used.Real-time cell analysis （xCELLigence system） was performed to determine the effects of E7080 and L-NIO on colorectal cell proliferation.While apoptosis was determined with Annexin V staining,and the effect of agents on angiogenesis was determined with chorioallantoic membrane （CAM） model.Results：We found that E7080 has a strong antiproliferative effect with an half maximum inhibition of concentration （IC50） value of 5.60×10-8 mol/L.Also it has been observed that E7080 showed antiangiogenic and apoptotic effects on HT29 colorectal cancer cells.Antiangiogenic scores of E7080 were 1.2,t.0 and 0.6 for 100,10 and 1 nmol/L E7080 concentrations,respectively.Furthermore,apoptosis has been detected in 71％ of HT29 colorectal cancer cells after administration of 100 nmol/L E7080 which may indicate strong apoptotic effect.Meanwhile administration of L-NIO alone did not show any effect,but the combination of E7080 with L-NIO increased the antiproliferative,antiangiogenic and apoptotic effects of E7080.Conclusions：Results of this study indicate that E7080 may be a good choice in treatment of colorectal tumors.Furthermore the increased effects of E7080 when combined with L-NIO raise the possibility to use a lower dose of E7080 and therefore avoid/minimize the side effects observed with E7080.

RTCA法用于医疗器械体外细胞毒性试验的研究

目的了解RTCA法是否适用于医疗器械体外细胞毒性的评价。方法采用L929细胞,对RTCA法的线性范围、精密度和重复性进行考察,并与MTT比色法进行结果比较。结果接种14个浓度的细胞后,2 h时细胞数量和细胞指数的相关性最好。线性范围为98~2.5×10~4个/ml;高、中、低3个质控浓度的相对标准偏差（RSD）均〈15%,日内、日间变异均〈15%,符合样品分析要求;连续接种中浓度的细胞悬液测定的RSD值〈15%,重复性良好;同时采用常规细胞毒性检测法MTT比色法,对8种医疗器械产品进行检测,两种方法结果的相关较高（R=0.994）。结论 RTCA法方便快捷、结果准确可靠,适用于医疗器械体外细胞毒性评价。

外源性化合物心脏毒性体外研究新进展

外源性化合物导致的心脏毒性问题一直是毒理学研究中的一个非常重要的关注点。心脏毒性体外研究可以降低研究成本,缩短实验周期,在心脏毒性研究中起重要作用。传统的心脏毒性体外研究方法常存在通量低、操作复杂、预测准确性不好等缺点。近年来,随着科学技术的不断发展,新的心脏毒性研究技术和方法不断出现,包括实时x CELLigence细胞分析技术、微电极阵列技术、扫描离子电导显微镜技术、新型线粒体膜电位检测技术、in-silico模型及新型心肌损伤生物标志物等。本文主要就以上心脏毒性研究的新技术和方法进行了综述。