Alba Andrew C, Strauch Trista A, Keisler Duane H, Wells Kevin D, Kesler Dylan C
Fisheries and Wildlife Sciences Department, University of Missouri Columbia, 1105 E Rollins St., Columbia, MO 65211, United States; Disney's Animal Kingdom, 1200 North Savannah Circle East, Bay Lake, FL 32830, United States.
Fisheries and Wildlife Sciences Department, University of Missouri Columbia, 1105 E Rollins St., Columbia, MO 65211, United States; Division of Animal Sciences, University of Missouri, 160 Animal Science Research Center, Columbia, MO 65211, United States.
Gen Comp Endocrinol. 2019 Jan 1;270:35-40. doi: 10.1016/j.ygcen.2018.10.002. Epub 2018 Oct 3.
Stress in animals is a concern in conservation breeding programs and livestock production facilities. The biological stress response is mediated by the release of glucocorticoids, which can suppress reproduction, growth, and immunity if recurrently activated. Feathers can be used to extract and monitor concentrations of corticosterone, a primary glucocorticoid in birds. However, current techniques for extracting feather corticosterone present challenges, including difficulty assessing extraction efficiency or hormone recovery, inconsistent extraction across feather lengths or pieces, and several uncertainties regarding the mechanisms of hormone deposition into feathers. To overcome such challenges and to provide tools useful for facilitating conservation breeding and livestock production, we developed and validated an alternative procedure for extracting feather glucocorticoids. We first developed a protocol to enzymatically digest the protein matrix of feathers using a keratinase, such that non-protein analytes could be isolated by organic extraction. We then developed an extraction protocol and evaluated techniques by measuring extraction efficiency and by testing parallelism and hormone recovery (accuracy) using radioimmunoassay. Our results demonstrated high and consistent extraction efficiency, as well as high accuracy and reliable parallelism to a standard curve upon measurement of corticosterone concentrations from extracts. By dissolving feather material into solution prior to extraction, we were able to replicate hormone deposition into the feather matrix and ensure consistent extraction across feathers. This work provides additional support for the validity and practicality of extracting glucocorticoids from feathers. Our extraction protocol is likely to extend to other applications as well, including the isolation of numerous non-protein analytes from various keratinized tissues.
动物应激是保护育种计划和畜牧生产设施中关注的问题。生物应激反应由糖皮质激素的释放介导，如果反复激活，糖皮质激素会抑制繁殖、生长和免疫。羽毛可用于提取和监测鸟类主要糖皮质激素皮质酮的浓度。然而，目前提取羽毛皮质酮的技术存在挑战，包括难以评估提取效率或激素回收率、不同羽毛长度或片段的提取不一致，以及关于激素沉积到羽毛中的机制存在若干不确定性。为了克服这些挑战并提供有助于促进保护育种和畜牧生产的工具，我们开发并验证了一种提取羽毛糖皮质激素的替代方法。我们首先制定了一个方案，使用角蛋白酶酶解羽毛的蛋白质基质，以便通过有机萃取分离非蛋白质分析物。然后我们制定了提取方案，并通过测量提取效率以及使用放射免疫测定法测试平行性和激素回收率（准确性）来评估技术。我们的结果表明，在测量提取物中的皮质酮浓度时，提取效率高且一致，准确性高，与标准曲线的平行性可靠。通过在提取前将羽毛材料溶解到溶液中，我们能够复制激素沉积到羽毛基质中，并确保不同羽毛的提取一致。这项工作为从羽毛中提取糖皮质激素的有效性和实用性提供了额外支持。我们的提取方案也可能扩展到其他应用，包括从各种角质化组织中分离多种非蛋白质分析物。
