CNTFRα在ALV自然重组FJ15HT0株感染宿主过程中的表达

Host CNTFRα Expression During Infection by the Natural Recombinant FJ15HT0 Strain of the ALV

本团队之前分离到一株天然重组禽白血病病毒(ALV)毒株FJ15HT0,经RNA-seq分析发现该毒株感染会引起鸡胸腺组织中睫状神经营养因子受体α(CNTFRα)基因表达量上调。为进一步探究CNTFRα在ALV FJ15HT0株感染过程中的表达及作用,本研究在构建ALV先天感染模型的基础上,通过RT-qPCR和免疫组织化学染色(IHC)探究不同时间点CNTFRα在感染鸡肝脏和胸腺中的表达量变化;以RT-qPCR和Western Blot检验病毒的复制水平对DF-1细胞CNTFRα在转录水平和蛋白表达水平的影响。结果表明,感染组与空白组相比,鸡胚出壳率明显降低,鸡只体重极显著降低(P<0.01);RT-qPCR和IHC结果显示同一时间点感染组胸腺和肝脏组织中CNTFRα转录水平及蛋白表达量均极显著高于空白组(P<0.01),胸腺组织中CNTFRα基因相对表达量与病毒gp85基因相对表达量呈极显著正相关关系(r=0.633,P<0.01);14 d、21 d感染组鸡只血清CNTF含量显著高于空白组(P<0.05)。RT-qPCR和Western Blot检测结果表明84 h感染组DF-1细胞CNTFRα表达量随病毒复制水平增加而显著提高(P<0.05),CNTFRα基因与病毒gp85基因相对表达量呈极显著正相关关系(r=0.73,P<0.01),CNTFRα蛋白与p27蛋白的表达量呈极显著正相关关系(r=0.973,P<0.01)。研究表明CNTFRα可能参与ALV FJ15HT0株致病过程。

Previously,we isolated the natural recombinant strain FJ15 HT0 of the avian leukemia virus(ALV),and RNA-sequencing showed that infection with this strain could upregulate expression of the ciliary neurotrophic receptor(CNTFRα)gene in the thymus gland(TG)of chickens.We wished to further explore the expression and role of CNTFRαin infection by the ALV FJ15 HT0 strain.Hence,we measured CNTFRαexpression in the liver and TG of infected chickens at different time points by real-time reverse transcription-quantitative polymerase chain reaction(RT-qPCR)and immunohistochemical(IHC)staining to construct a model of congenital infection by the ALV.RT-qPCR and western blotting were performed to detect the effect of virus replication on the transcriptional and protein expression of CNTFRαin infected DF-1 cells.We discovered that the shelling rate of chicken embryos in the infected group was significantly lower than that in the control group,and that the bodyweight of chickens in the infected group was significantly lower than that in the control group(P<0.01).RT-qPCR and IHC showed that the transcriptional level and protein expression of CNTFRαin the TG and liver of the infection group were significantly higher than those of the control group at the same time point(P<0.01).Relative expression of CNTFRαin the TG was significantly positively correlated with that of viral gp85(r=0.633,P<0.01),and the serum CNTF content of infected birds was significantly higher than that of chickens in the control group(P<0.05)at 14 d and 21 d.RT-qPCR and western blotting showed that CNTFRαexpression in DF-1 cells increased significantly with an increase in virus replication at 84 h in the infection group(P<0.05).There was a significant positive correlation between the relative expression of CNTFRαand viral gp85(r=0.73,P<0.01),CNTFRαprotein,and p27 protein(r=0.973,P<0.01).Our study showed that CNTFRαmay be involved in the pathogenic process of the ALV FJ15 HT0 strain.