Project description:Background : Poxviruses have been shown to be efficient vectors for the production of protective immune responses in host species but also in on host species. Myxoma virus belongs to poxviridae family. In order to evaluate the capacity of (MYXV) as a vaccine vector in ruminants, we invistigated its interactions with bone marrow-derived dendritic cells (BM-DC). DCs are the most potent antigen-presenting cells and play a crucial role during the priming and reactivation of antigen-specific immune responses . Following infection by a pathogen, the functional changes of DC are essential since priming and polarization of the immune response depend on these changes . Therefore a better understanding of the modifications in gene expression of proinflammatory cytokines, chemokines and stimulatory molecules expression may prove useful in predicting whether or not a vaccine vector will be effective. Moreover, it may help identifying modifications for improving its efficacy. Methodology/Principal Findings: We investigated in vitro the interactions between recombinant MYXV expressing GFP protein, SG33-GFP and ovine BM-DCs. To gain a global view of the gene remodelling induced by MYXV, we infected BM-DCs from three sheep with SG33-GFP at different multiplicity of infection (MOI) (0, 1, 3) during different time (0, 3, 8 hours). We measured the expression of 15K probes in BM-DCs after each kind of conditions with ovine Agilent microarrays. Furthermore, a selected number of genes were confirmed by RT-qPCR. MYXV infection induces a strong reprogramming of the cells leading to the expression of pro-inflammatory cytokines and mobilisation of type I IFN pathways, cellular death and features associated with the activation of the adaptive immune response. Conclusion/Significance: Microarray profiling of a poxvirus-DC interaction help to delineate some interesting features of a vector candidate in ovine, and pave the way for additional improvements of a promising platform. Keywords : Myxoma virus ; bone marrow-derived dendritic cells ; transcriptome ; sheep; vaccine 9 samples: 3 time points and 3 replicates of MOI=1

Project description:Dendritic cells (DCs) are antigen sensing and presenting cells that are essential for effective immunity. Existing as a multi-subset population, divided by distinct developmental and functional characteristics1,2, DC subsets play important and unique roles in responses to pathogens, vaccines and cancer therapies, as well as during immune-pathologies. Therefore therapeutic manipulation of the DC compartment is an attractive strategy. However, our incomplete knowledge of the inter-relationship between DC subsets and how they develop from progenitors in the bone marrow (BM) has so far limited the realization of their therapeutic potential. DCs arise from a cascade of progenitors that gradually differentiate in the BM; first, the macrophage DC progenitor (MDP), then common DC progenitor (CDP), and lastly the Pre-DC, which will leave the BM to seed peripheral tissues before differentiating into mature DCs3,4. While the basic outline of this process is known, how subset commitment and development is regulated at the molecular level remains poorly understood. Here we reveal that the Pre-DC population in mice is heterogeneous, containing uncommitted Ly6c+/-Siglec-H+ cells as well as Ly6c+Siglec-H- and Ly6c-Siglec-H- sub-populations that are developmentally fated to become Th2/17-inducing CD11b+ DCs and Th1-inducing CD8α+ DCs, respectively. Using single cell analysis by microfluidic RNA sequencing, we found that DC subset imprinting occurred at the mRNA level from the CDP stage, revealing that subset fate is defined in the BM and not in peripheral tissues. Single cell transcriptome analysis allowed identification of the molecular checkpoints between progenitor stages and revealed new regulators of DC-poiesis, shedding light on the role of cell cycle control and specific transcription factors in DC lineage development. These data advance our knowledge of the steady-state regulation of DC populations and open promising new avenues for investigation of the therapeutic potential of DC subset-specific targeting in vivo to improve vaccine-based and immunotherapeutic strategies.

Dataset Information

Gene expression profiling of dendritic cells derived from bone marrow (BM-DC) in response to myxoma virus infection.

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