猪繁殖与呼吸综合征病毒感染对非肌肉肌球蛋白Ⅱ型重链A表达的影响

The effect of PRRSV infection on the expression of NMHC Ⅱ-A

为研究猪繁殖与呼吸综合征病毒(PRRSV)感染对非肌肉肌球蛋白Ⅱ型重链A(NMHCⅡ-A)表达及组织定位的影响,本研究选取5月龄的PRRSV阴性健康长白猪和莱芜黑猪各12头,分别设感染组和对照组,滴鼻感染PRRSV后分别于3 d、7 d、14 d、21 d、28 d迫杀,分别对器脏进行病理组织学观察,免疫组织化学(IHC)和荧光定量PCR(q PCR)检测。组织病理结果显示,PRRSV对商品长白猪产生较强的致病性,感染猪表现明显的免疫抑制,典型的间质性肺炎和各器官炎症;而莱芜黑猪病轻微,仅呈轻微的间质性肺炎,部分淋巴结出现坏死性炎症。IHC检测结果显示,正常组织细胞中均有NMHCⅡ-A表达,但在肺、心肌、脾、扁桃体、软骨中表达较高,其主要定位于细胞浆中,两个品种间无显著差异;PRRSV感染后,NMHCⅡ-A表达趋势增强,且出现于细胞核中,推测PRRSV感染增强了NMHCⅡ-A核浆穿梭功能,两个品种间无显著差异。RT-q PCR检测结果显示,PRRSV在组织中的病毒载量与NMHCⅡ-A的转录水平呈正相关,不同组织间波动规律相似,但在不同品种间波动程度差异明显。综上所述,PRRSV感染对猪体内NMHCⅡ-A的表达产生了明显的影响,但品种间差异不明显。本实验为NMHCII-A作为PRRSV抑制物的候选蛋白提供了实验依据,并为今后PRRSV的防控奠定了基础。

To reveal the effect of PRRSV infection on NMHCII-A expression, 12 five-month-old Laiwu black pigs and Landraces pigs （PRRSV-negative） were divided into two groups at random. Experimental groups were inoculated by nasal instillation of PRRSV isolate TA-12. One pig was killed in each group at 3 d, 7 d, 14 d, 21 d, and all others were killed at 28 dpi in order to collect the main organs. Histopathology, quantitative RT-PCR and immunohistochemistry （IHC） were used in this study. Histopathology observation showed the pathogenicity caused by PRRSV in Landraces pigs was more serious than that in Laiwu black pigs, showing significant interstitial pneumonia in lung, lymphocytes infiltration in main organs, and significant immunosuppression. IHC results showed that NMHCII-A was ubiquitously expressed in normal tissues, and a slight higher expression present in plasma of cells in heart, lung, spleen, tonsil and cartilage. After PRRSV infection, the expressions of NMHCII-A were up-regulated in nuclear which indicated that PRRSV infection enhanced NMHCII-A nucleocytoplasmic shuttling function. Furthermore, no significant differences were observed between the two groups. RT-qPCR results showed that there was a positive correlation between PRRSV load and NMHCII-A mRNA level in different tissues, comparing with the different fluctuation of PRRSV load and NMHCII-A mRNA level in the two groups. The experiment proved that PRRSV infection in pigs had a significant effect on the expression of NMHCII-A, but the difference between varieties was not obvious. This experiment provids an experimental basis for NMHCII-A as a candidate protein for PRRSV inhibitor and layed a foundation for the prevention and control of PRRSV in the future.