Multiparametric comparative analysis of all pre- and postinfection related immune cell and clinical traits

Multiparametric analysis of 233 MA/My x C57L intercross and backcross mice segregating MHC class I Dk. Before murine (M)CMV infection, all mice were weighed and bled. Blood leukocytes were stained with fluorochrome-conjugated mAbs for CD3, CD19, and NK receptors and then analyzed by flow cytometry. NK cell subset light-scattering features, absolute number, frequency, and receptor display intensity were recorded as individual pre-infection traits. Afterward, the mice were infected with MCMV. Spleen and liver tissues were collected 3.5 d later for analysis of postinfection traits, including genomic genotypes and viral load. Spleen leukocytes from infected mice were analyzed as for pre-infection traits. The change in trait values were calculated from the pre- (blood) and post- (spleen) infection trait values. In total, 110 distinct traits were assessed. Multiparametric Pearson correlations were performed in the R (versions 2.15.0 and 2.15.2) computing environment using an automated correlation analysis followed with a strict Bonferroni correction for multiple tests. P-values were corrected for multiple comparisons using both the Benjamini-Hochberg false discovery rate and the Bonferroni Correction.

本研究对233只MA/My与C57L的互交及回交小鼠开展多参数分析，这些小鼠均分离携带主要组织相容性复合体I类Dk（MHC class I Dk）。在鼠巨细胞病毒（murine (M)CMV）感染前，对所有小鼠进行称重并采集血液样本。采用荧光素标记的单克隆抗体（monoclonal antibodies, mAbs）对血液白细胞进行染色，靶标涵盖CD3、CD19以及自然杀伤细胞（natural killer, NK）受体，随后通过流式细胞术开展分析。将NK细胞亚群的光散射特征、绝对计数、细胞占比以及受体表达强度作为感染前的单项表型进行记录。随后，对所有小鼠接种MCMV。感染后3.5天采集脾脏与肝脏组织，用于分析感染后表型，包括基因组基因型与病毒载量。对感染小鼠的脾脏白细胞按照感染前表型的检测流程开展分析。根据感染前（血液样本来源）与感染后（脾脏样本来源）的表型数值，计算得到各表型的变化量。本次研究共评估了110项独立表型。在R语言（版本2.15.0与2.15.2）计算环境中，通过自动化相关分析开展多参数Pearson相关分析，并针对多重检验实施严格的Bonferroni校正。针对多重比较得到的P值，同时采用Benjamini-Hochberg错误发现率（false discovery rate, FDR）与Bonferroni校正两种方法进行校正。