Guglielmetti C, Mazza M, Pagano M, Carrella S, Sciuto S, Nodari S, Pezzolato M, Richelmi G B, Baioni E, Caramelli M, Acutis P L, Bozzetta E
a Istituto Zooprofilattico Sperimentale del Piemonte, Liguria e Valle d'Aosta , Turin , Italy.
Food Addit Contam Part A Chem Anal Control Expo Risk Assess. 2014;31(5):833-8. doi: 10.1080/19440049.2014.900191. Epub 2014 Apr 10.
Corticosteroids have become the most widespread illegal growth promoters in veal calves and beef cattle. Testing for corticosteroids relies on either direct detection of compounds or their metabolites or indirect detection to identify changes in biological pathways. We used a comparative proteomic approach, based on two-dimensional electrophoresis (2DE), to identify plasma protein markers after short-term dexamethasone administration in veal calves. Twenty-three male Friesian veal calves were treated experimentally with dexamethasone sodium phosphate: 10 received low-dose administration of the drug (0.4 mg day⁻¹ per os) for 20 consecutive days (treatment group); 10 received the drug at therapeutic dosage (2-4 mg kg⁻¹ i.m.) for 3 consecutive days (comparison group). Three animals were not treated (control group). Plasma samples were collected from each animal at six time points (T1-T6; treatment and control group) and at four time points (T1-T4; comparison group) and stored at -80°C until analysis. Plasma proteins were quantified and analysed in triplicate by 2DE. The images were analysed with Bionumerics® software. Comparison of 2DE maps obtained from blood samples at T1 (before treatment) and at T6 (final sampling) showed a significant disappearance (p < 0.001) of two protein spots at T6 in the treatment group. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis and immunoblotting identified these isoforms as serum paraoxonase/arylesterase 1 precursor (PON1). Synthesised in the liver and released into the blood, PON1 has an important role in lipid metabolism. The absence of variation of this protein in the comparison group suggests that the marker has good specificity for detecting illicit corticosteroid treatment.
皮质类固醇已成为犊牛和肉牛中最普遍使用的非法生长促进剂。皮质类固醇的检测依赖于对化合物或其代谢物的直接检测,或通过间接检测来识别生物途径中的变化。我们采用基于二维电泳(2DE)的比较蛋白质组学方法,来鉴定犊牛短期给予地塞米松后血浆中的蛋白质标志物。23头雄性弗里斯兰犊牛接受了地塞米松磷酸钠的实验性治疗:10头连续20天接受低剂量药物口服给药(每天0.4毫克)(治疗组);10头连续3天接受治疗剂量的药物肌肉注射(每千克体重2 - 4毫克)(比较组)。3头动物未接受治疗(对照组)。在六个时间点(T1 - T6;治疗组和对照组)从每头动物采集血浆样本,在四个时间点(T1 - T4;比较组)采集样本,并储存在 - 80°C直至分析。血浆蛋白质经定量后通过2DE进行三次重复分析。图像用Bionumerics®软件进行分析。对T1(治疗前)和T6(最终采样)血样获得的2DE图谱进行比较,结果显示治疗组在T6时有两个蛋白质斑点显著消失(p < 0.001)。液相色谱 - 串联质谱(LC - MS/MS)分析和免疫印迹确定这些异构体为血清对氧磷酶/芳基酯酶1前体(PON1)。PON1在肝脏中合成并释放到血液中,在脂质代谢中起重要作用。比较组中该蛋白质没有变化,这表明该标志物对检测非法皮质类固醇治疗具有良好的特异性。