K. Fay1, N. Klingensmith1, J. Lyons1, C. Chen1, C. Coopersmith2, M. L. Ford3 1Emory University School Of Medicine,Department Of Surgery,Atlanta, GA, USA 2Emory University,Critical Care Center,Atlanta, GA, USA 3Emory University,Transplant Center,Atlanta, GA, USA
Introduction: Previous studies have shown that mice with identical genetic backgrounds obtained from different vendors possess differential immuno-phenotypic characteristics based on environmental factors including the composition of the microbiome. However, immune function in the setting of murine septic peritonitis has been previously assumed to be consistent within mice of the same strain. Here, we aimed to determine whether vendor-specific environmental factors impact survival and immune function following CLP.
Methods: 6 week old B6 mice from Charles River (CR) and Jackson Lab (JAX) underwent cecal ligation and puncture (CLP) one to two days after arrival to prevent acclimation to an environment different from their original vendor. To assess survival, each group underwent CLP and was followed for seven days. To assess immunophenotype, mice were sacrificed at 24 hours after CLP and underwent splenic harvest. Cells were then processed and stained for phenotypic and functional markers to be analyzed via flow cytometry. Data are presented as mean±SEM.
Results: CR B6 mice exhibited worsened lymphopenia in the setting of sepsis compared to JAX B6 mice, and a significantly decreased frequency of CD4+ and CD8+ T cells (36.9±0.8 vs 44.1±2.0, p<0.05 and 25.0±1.8 vs 29.5±0.8, p<0.01). When function of these T cells was examined via cytokine analysis, CR B6 mice possessed significantly reduced frequencies of IL-2- secreting T cells as compared to JAX B6 mice, and this was true for both the CD4+ and CD8+ T cell compartments (8.7±0.4 vs 15.7±2.1, <p0.05 and 5.8±0.3 vs 12.2±1.5, p<0.01, respectively). This immune profile of worsened lymphopenia and decreased frequencies of IL-2-secreting CD4+ T cells observed in CR septic mice has been previously associated with worsened mortality. However, CR B6 mice exhibited profoundly superior survival following CLP as compared to JAX B6 mice (90% vs. 0%, p<0.0001). Further evaluation for possible mechanisms underlying this finding revealed that CR septic mice possessed significantly increased frequencies of IFNγ+ CD4+ cells compared to JAX septic mice (2.4±0.1 vs 1.5±0.2, p<0.01). This was not associated with a difference in the frequencies of dendritic cells between the two groups (21.1±1.6 vs. 25.9±3.7). However, CR septic mice did demonstrate significantly higher frequencies of effector memory CD4+ T cells compared to JAX septic mice (10.6±0.3 vs 7.3±0.4, p<0.01).
Conclusions: Having the same genetic background does not result in an identical immunophenotype following the induction of sepsis in mice obtained from different vendors. Interestingly, despite having some immunologic features previously associated with increased mortality, Charles River B6 mice exhibited markedly better survival compared to Jackson Lab B6 mice. We conclude that the presence of increased IFNγ-secreting CD4+ and effector memory cells could underlie the improved survival observed in Charles River B6 mice. Previous work has demonstrated an effect of the microbiome on antigen specific T cell proliferation and function, which may play a role in the differences with see between Charles River and Jackson Lab mice. Evaluation of differences between the microbiota of the two groups will be an avenue of future exploration.