Molecular characterization of hypothalamus-infiltrating T cells following Pandemrix® vaccination of Orex-HA mice through bulk RNA sequencing of T cell subsets

To identify the T cell subsets and molecular pathways potentially involved in the killing of orexinergic neurons in Pandemrix®-vaccinated Orex-HA mice, we cell-sorted activated (CD44high CD62Llow) transferred HA-specific (CD45.1+) and endogenous (CD45.1-) CD4 and CD8 T cells from dissected hypothalami, spleen and cervical lymph nodes at the peak of hypothalamic infiltration (day 14 post-vaccination). RNA sequencing on these sorted T cell populations allowed us to compare the gene expression profile of a given T cell subset across different tissue sites and of different T cell subsets in a given tissue. Orex-HA mice were immunized intramuscularly with Pandemrix® vaccine 1 day after the adoptive transfer of naïve (CD45.1+CD62L+CD25-) HA-specific CD4+ and CD8+ T cells. At day 14 post-vaccination, endogenous (CD45.1-) and transferred HA-specific (CD45.1+) populations of CD44highCD62LlowCD4+ and CD44highCD62LlowCD8+ T cells were isolated from spleens, cervical lymph nodes and hypothalami by FACS sorting in 4 independent experiments involving 20 to 30 mice each. 4 experiments X 3 tissues per experiment X 4 T cell populations (CD4 and CD8 both with endogenous & transferred) = 48 samples (1 samples did not sequence)…. Total of 47 samples available.