Wang Lianghai, Zhang Hexiao, Suo Xiong, Zheng Shijun, Feng Wen-Hai
State key laboratory for Agrobiotechnology, Department of Microbiology and Immunology, College of Biological Science, China Agricultural University, Beijing 100193, China.
Vet Immunol Immunopathol. 2011 Jun 15;141(3-4):209-20. doi: 10.1016/j.vetimm.2011.03.001. Epub 2011 Mar 9.
Porcine reproductive and respiratory syndrome virus (PRRSV) has a restricted tropism mainly for porcine alveolar macrophages (PAMs), but not for peripheral blood monocytes (BMo) in vivo. Previous research showed that only a few BMo became susceptible to PRRSV infection after 1 day culture. Porcine sialoadhesin (PoSn) and CD163 are identified to be the two main PRRSV receptors for binding and internalization. Both receptors are not expressed on BMo, or only expressed at low levels, which may explain why PRRSV cannot infect them. The relationship of BMo differentiation/aging, PRRSV receptor level, and susceptibility to PRRS virus infection has not been thoroughly investigated. In this study, BMo were successfully cultured with pig serum plus L929 cell culture supernatant. Our results showed that both the mRNA and protein expression levels of PoSn were significantly increased after 5-day culture. The mRNA level of CD163 was enhanced more than 20-fold after 1-day culture; CD163-positive BMo increased dramatically from about 2% after 2h- culture to about 50% after 96-h culture. Furthermore, cultured BMo became much more permissive to PRRSV infection, and the percentage of PRRSV-infected BMo was at least the same as PAMs, if not higher, when infected with CH-1a, the first PRRSV strain isolated in China, or HV, a highly virulent strain. Three other PRRSV strains including VR2332, and two classical Chinese isolates could also infect cultured BMo as well. Most importantly, PRRS virus was successfully isolated from 14 of 15 antibody-positive serum samples using cultured BMo. These results suggest that the enhanced susceptibility of cultured BMo to PRRS virus is coordinated with increased CD163 expression, but less related to the delayed (day 5) increased expression of PoSn. Thus, cultured BMo could be an alternative choice for PRRS virus isolation and identification.
猪繁殖与呼吸综合征病毒（PRRSV）具有受限的嗜性，主要针对猪肺泡巨噬细胞（PAM），但在体内不针对外周血单核细胞（BMo）。先前的研究表明，仅少数BMo在培养1天后变得易受PRRSV感染。猪唾液酸粘附素（PoSn）和CD163被确定为PRRSV结合和内化的两个主要受体。这两种受体在BMo上均不表达，或仅低水平表达，这可能解释了PRRSV为何无法感染它们。BMo分化/衰老、PRRSV受体水平与对PRRS病毒感染的易感性之间的关系尚未得到充分研究。在本研究中，BMo成功地用猪血清加L929细胞培养上清液进行培养。我们的结果表明，培养5天后，PoSn的mRNA和蛋白表达水平均显著增加。培养1天后，CD163的mRNA水平提高了20多倍；CD163阳性BMo从培养2小时后的约2%急剧增加到培养96小时后的约50%。此外，培养的BMo对PRRSV感染的易感性大大增加，当用中国分离的第一株PRRSV毒株CH-1a或高毒株HV感染时，PRRSV感染的BMo百分比至少与PAM相同，甚至更高。包括VR2332在内的其他三株PRRSV毒株以及两株中国经典分离株也能感染培养的BMo。最重要的是，使用培养的BMo从15份抗体阳性血清样本中的14份成功分离出PRRS病毒。这些结果表明，培养的BMo对PRRS病毒易感性的增强与CD163表达的增加相关，但与PoSn表达延迟（第5天）增加的相关性较小。因此，培养的BMo可作为PRRS病毒分离和鉴定的替代选择。
