Porcine reproductive and respiratory syndrome virus (PRRSV) has disrupted the global swine industry since the 1980s. PRRSV-host interactions are largely still unknown but may involve host ISG15 protein. In this study, we developed a monoclonal antibody (Mab-3D5E6) specific for swine ISG15 (sISG15) by immunizing mice with recombinant sISG15. A sandwich enzyme-linked immunosorbent assay (ELISA) incorporating this sISG15-specific Mab was developed to detect sISG15 and provided a lower limit of sISG15 detection of 200 pg/mL. ELISA results demonstrated that infection of porcine alveolar macrophages (PAMs) with low-virulence or attenuated PRRSV vaccine strains induced intracellular ISG15 expression that was independent of type I IFN production, while PAMs infection with a PRRSV vaccine strain promoted extracellular ISG15 secretion from infected PAMs. Conversely, the addition of recombinant sISG15 to PAMs mimicked natural extracellular ISG15 effects whereby sISG15 functioned as a cytokine by activating PAMs. Once activated, PAMs could inhibit PRRSV replication and resist infection with PRRSV vaccine strain TJM. In summary, a sandwich ELISA incorporating homemade anti-ISG15 Mab detected ISG15 secretion induced by PAMs infection with a PRRSV vaccine strain. Recombinant ISG15 added to cells exhibited cytokine-like activity that stimulated PAMs to assume an anti-viral state that enabled them to inhibit PRRSV replication and resist viral infection.

中文翻译：

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PRRSV疫苗株诱导的细胞外ISG15分泌物可刺激猪肺泡巨噬细胞针对PRRSV的抗病毒反应。

自1980年代以来，猪繁殖与呼吸综合症病毒（PRRSV）扰乱了全球养猪业。PRRSV与宿主的相互作用在很大程度上仍然未知，但可能涉及宿主ISG15蛋白。在这项研究中，我们通过用重组sISG15免疫小鼠，开发了对猪ISG15（sISG15）有特异性的单克隆抗体（Mab-3D5E6）。已开发出一种结合了sISG15特异性Mab的夹心酶联免疫吸附测定（ELISA）来检测sISG15，并且提供的sISG15检测下限为200 pg / mL。ELISA结果表明，用低毒力或减毒PRRSV疫苗株感染猪肺泡巨噬细胞（PAM）会诱导细胞内ISG15表达，而这种ISG15表达不依赖于I型IFN的产生，而用PRRSV疫苗株感染的PAM会促进感染PAM的细胞外ISG15分泌。相反，向PAM中添加重组sISG15可以模仿天然的细胞外ISG15效应，其中sISG15通过激活PAM发挥细胞因子的作用。一旦激活，PAMs可以抑制PRRSV复制并抵抗PRRSV疫苗株TJM的感染。总之，包含自制抗ISG15 Mab的夹心ELISA检测到由PRRSV疫苗株感染的PAM诱导的ISG15分泌。添加到细胞中的重组ISG15表现出类似细胞因子的活性，可刺激PAM呈现抗病毒状态，从而使其能够抑制PRRSV复制并抵抗病毒感染。PAM可以抑制PRRSV复制并抵抗PRRSV疫苗株TJM的感染。总之，包含自制抗ISG15 Mab的三明治ELISA检测到由PRRSV疫苗株感染的PAM诱导的ISG15分泌。添加到细胞中的重组ISG15表现出类似细胞因子的活性，可刺激PAM呈现抗病毒状态，从而使其能够抑制PRRSV复制并抵抗病毒感染。PAM可以抑制PRRSV复制并抵抗PRRSV疫苗株TJM的感染。总之，包含自制抗ISG15 Mab的三明治ELISA检测到由PRRSV疫苗株感染的PAM诱导的ISG15分泌。添加到细胞中的重组ISG15表现出类似细胞因子的活性，可刺激PAM呈现抗病毒状态，从而使其能够抑制PRRSV复制并抵抗病毒感染。