Griffith School of Science and Environment, Building 51, Griffith University, Gold Coast Campus, QLD 4222, Australia; The Australian Rivers Institute, Building 51, Griffith University, Gold Coast Campus, QLD 4222, Australia.
Griffith School of Science and Environment, Building 51, Griffith University, Gold Coast Campus, QLD 4222, Australia; The Australian Rivers Institute, Building 51, Griffith University, Gold Coast Campus, QLD 4222, Australia.
Sci Total Environ. 2021 Jan 10;751:141680. doi: 10.1016/j.scitotenv.2020.141680. Epub 2020 Aug 12.
Non-targeted protein expression at the cellular level can provide insights into mechanistic effects of contaminants in wildlife, and hence new and potentially more accurate biomarkers of exposure and effect. However, this technique has been relatively unexplored in the realm of in vitro biomarker discovery in threatened wildlife, despite the vulnerability of this group of animals to adverse sublethal effects of contaminant exposure. Here we examined the usefulness of non-targeted protein expression for biomarker discovery in green sea turtles (Chelonia mydas) by investigating differences in the response of primary cells from five different tissue types that were exposed to three contaminants known to accumulate in this species. Cells derived from C. mydas skin, liver, kidney, ovary and small intestine were exposed to 100 μg/L of either polychlorinated biphenyl 153 (PCB153), perfluorononanoic acid (PFNA) or phenanthrene for 24 h. The global protein expression was then quantitatively evaluated using sequential window acquisition of all theoretical mass spectra (SWATH-MS). Comparison of the global protein profiles revealed that, while a majority of proteins were mutually expressed in controls of all tissue types (~90%), the response to exposure in terms of protein expression strength was significantly different between tissue types. Furthermore, a comparison to known markers of chemical exposure in sea turtles from the literature indicated that in vitro response can reflect known in vivo responses. In particular, markers such as heat shock protein (HSP) 60, glutathione S-transferases (GSTs) and superoxide dismutases (SODs), cytochrome P450 and catalase were dysregulated in response to exposure. Furthermore, potential new markers of exposure were discovered such as annexin, an important protein in cell signalling processes. While this methodology proved promising further studies are required to confirm the accuracy of in vitro protein expression as a tool for biomarker discovery in wildlife.
非靶向蛋白质表达可深入了解污染物对野生动物的机制影响，从而为暴露和效应提供新的、潜在更准确的生物标志物。然而，尽管这组动物容易受到污染物暴露的亚致死效应的不利影响，但这种技术在濒危野生动物体外生物标志物发现领域的应用相对较少。在这里，我们通过研究五种不同组织类型的原代细胞对三种已知在该物种中积累的污染物的反应差异，检验了非靶向蛋白质表达在绿海龟（Chelonia mydas）生物标志物发现中的有用性。从绿海龟皮肤、肝脏、肾脏、卵巢和小肠中提取细胞，将其暴露于浓度为 100μg/L 的多氯联苯 153（PCB153）、全氟壬酸（PFNA）或菲中 24 小时。然后使用序贯窗口采集所有理论质谱（SWATH-MS）定量评估全局蛋白质表达。比较全局蛋白质谱图表明，虽然大多数蛋白质在所有组织类型的对照中都相互表达（约 90%），但暴露对蛋白质表达强度的反应在组织类型之间存在显著差异。此外，与文献中来自海龟的化学暴露已知标志物的比较表明，体外反应可以反映体内已知反应。特别是热休克蛋白 (HSP) 60、谷胱甘肽 S-转移酶 (GST) 和超氧化物歧化酶 (SOD)、细胞色素 P450 和过氧化氢酶等标志物在暴露后出现失调。此外，还发现了一些潜在的暴露新标志物，如膜联蛋白，它是细胞信号转导过程中的一种重要蛋白质。虽然该方法证明有前景，但需要进一步的研究来确认体外蛋白质表达作为野生动物生物标志物发现工具的准确性。
