Institute of Biological, Environmental and Rural Sciences, Aberystwyth University, United Kingdom.
J Proteome Res. 2010 Oct 1;9(10):4940-51. doi: 10.1021/pr1000785.
Control of Fasciola hepatica infections of livestock in the absence of vaccines depends largely on the chemical triclabendazole (TCBZ) because it is effective against immature and adult parasites. Overdependence on a single drug and improper application is considered a significant factor in increasing global reports of fluke resistant to TCBZ. The mode(s) of action and biological target(s) of TCBZ are not confirmed, delaying detection and the monitoring of early TCBZ resistance. In this study, to further understand liver fluke response to TCBZ, the soluble proteomes of TCBZ-resistant and TCBZ-susceptible isolates of F. hepatica were compared with and without in vitro exposure to the metabolically active form of the parent drug triclabendazole sulphoxide (TCBZ-SO), via two-dimensional gel electrophoresis (2-DE). Gel image analysis revealed proteins displaying altered synthesis patterns and responses both between isolates and under TCBZ-SO exposure. These proteins were identified by mass spectrometry supported by a F. hepatica expressed sequence tag (EST) data set. The TCBZ responding proteins were grouped into three categories; structural proteins, energy metabolism proteins, and "stress" response proteins. This single proteomic investigation supported the reductionist experiments from many laboratories that collectively suggest TCBZ has a range of effects on liver fluke metabolism. Proteomics highlighted differences in the innate proteome profile of different fluke isolates that may influence future therapy and diagnostics design. Two of the TCBZ responding proteins, a glutathione transferase and a fatty acid binding protein, were cloned, produced as recombinants, and both found to bind TCBZ-SO at physiologically relevant concentrations, which may indicate a role in TCBZ metabolism and resistance.
在没有疫苗的情况下，控制家畜的肝片吸虫感染在很大程度上依赖于化学药物三氯苯达唑（TCBZ），因为它对未成熟和成年寄生虫都有效。过度依赖单一药物和不当应用被认为是导致全球报道的吸虫对 TCBZ 产生抗药性的一个重要因素。TCBZ 的作用模式和生物靶标尚未得到证实，这延迟了对 TCBZ 早期抗性的检测和监测。在这项研究中，为了进一步了解肝片吸虫对 TCBZ 的反应，我们比较了 TCBZ 耐药和 TCBZ 敏感的肝片吸虫分离株在体外暴露于母体药物三氯苯达唑砜（TCBZ-SO）的代谢活性形式前后的可溶性蛋白质组，方法是通过二维凝胶电泳（2-DE）。凝胶图像分析显示，在分离株之间以及在 TCBZ-SO 暴露下，蛋白质的合成模式和反应都发生了改变。这些蛋白质通过质谱鉴定得到支持，同时还利用了一个肝片吸虫表达序列标签（EST）数据集。TCBZ 反应蛋白被分为三类；结构蛋白、能量代谢蛋白和“应激”反应蛋白。这项单一的蛋白质组学研究支持了许多实验室的还原实验，这些实验共同表明 TCBZ 对肝片吸虫代谢有多种作用。蛋白质组学强调了不同肝片吸虫分离株固有蛋白质组谱的差异，这可能影响未来的治疗和诊断设计。我们克隆了两种 TCBZ 反应蛋白，一种谷胱甘肽转移酶和一种脂肪酸结合蛋白，并将它们作为重组蛋白进行生产，两者都被发现能够在生理相关浓度下结合 TCBZ-SO，这可能表明它们在 TCBZ 代谢和抗性中发挥作用。
