Identification of Potential mRNA Biomarkers in Milk Small Extracellular Vesicles of Enzootic Bovine Leukosis Cattle.
Identification of Potential mRNA Biomarkers in Milk Small Extracellular Vesicles of Enzootic Bovine Leukosis Cattle.
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DOI：
10.3390/v14051022
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发表时间：
2022-05-11
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地方性牛白血病（EBL）是由牛白血病病毒（BLV）引起的一种疾病;只有一小部分BLV感染的牛发展EBL并出现B细胞淋巴肉瘤。没有针对BLV的疫苗、EBL的治疗或用于预测EBL发作的可能性的方法，因此使得EBL控制困难。在此，为了探索牛奶中EBL的生物标志物，我们通过微阵列分析检测了来自四头BLV未感染和四头EBL牛的牛奶中小细胞外囊泡（sEV）的mRNA谱。结果显示，14种mRNA以显著更高的量包封，因此选择这些mRNA作为生物标志物候选物。这些mRNA的引物进行了设计，和9个引物组可用于定量实时PCR。使用来自7头未感染牛和10头EBL牛的新采集乳汁的sEV，评价9种mRNA作为EBL生物标志物的可用性。EBL牛的乳汁sEV中8种mRNA（TMEM 156、SRGN、CXCL 8、DEFB 4A、FABP 5、LAPTM 5、LGALS 1和Vim）的量显著高于未感染牛。因此，我们的研究结果表明，牛奶sEV中的这八种mRNA可以作为潜在的EBL生物标志物与组合使用，虽然单一的mRNA使用是不够的。因此，可以通过监测这些mRNA在牛奶中的数量波动来识别处于EBL发作风险的牛，然后再发展EBL。
Enzootic bovine leukosis (EBL) is a disease caused by bovine leukemia virus (BLV); only a small percentage of BLV-infected cattle develop EBL and present with B-cell lymphosarcoma. There is no vaccine against BLV, treatment for EBL, or method for predicting the possibility of EBL onset, thus making EBL control difficult. Herein, to explore biomarkers for EBL in milk, we examined the mRNA profiles of small extracellular vesicles (sEVs) in milk from four BLV-uninfected and four EBL cattle by microarray analysis. It was revealed that 14 mRNAs were encapsulated in significantly higher quantities, and these mRNAs were therefore selected as biomarker candidates. Primers for these mRNAs were designed, and nine primer sets were available for quantitative real-time PCR. Nine mRNAs were evaluated for their availability as biomarkers for EBL using sEVs from newly-collected milk of 7 uninfected and 10 EBL cattle. The quantities of eight mRNAs (TMEM156, SRGN, CXCL8, DEFB4A, FABP5, LAPTM5, LGALS1, and VIM) were significantly higher in milk sEVs of EBL cattle than in those of uninfected cattle. Therefore, our findings indicate that these eight mRNAs in milk sEVs can be used as potential EBL biomarkers with combination use, although single mRNA use is not enough. Consequently, cattle at risk of EBL onset can be identified by monitoring the fluctuation in quantities of these mRNAs in milk before they develop EBL.