Fig 1: Blocking miR-21 induction rescues IL-12 expression. (A) C57Bl/6 mice were infected with recombinant wild type or centrin deleted Leishmania parasites secreting CD4+ T cell epitope 2W (EAWGALANWAVDSA) fused to L. donovani 3′ Nucleotidase/Nuclease protein lacking the N' terminus membrane anchoring domain. The mice were administered with LNA chemistry based anti-miR-21 oligonucleotides on days 0, 2, and 4 following parasite infection. (B) Expression of miR-21 5p in the splenocytes 7 days post-infection with LdWT or LdCen−/− parasites in mice treated with anti-miR-21 oligonucleotides. (C) Expression of IL-12 in the splenocytes 7 days post-infection with LdWT or LdCen−/− parasites in mice treated with anti-miR21 oligonucleotides. (D) Epitope-specific activated CD4+ T cells expressing IL-2R and IL7-R markers were identified using flow cytometric analysis 7 days post infection. Gating scheme to identify the activated CD4+ T cells expressing surface markers IL-2R, and IL7R from the spleen and lymph nodes of LdWT2W infected and or LNA-miR-21 treated mice is shown. To enrich 2W epitope specific CD4+ T cell populations, 2W tetramers were used. Dump contained markers for B cells (B220), macrophages (F4/80), DCs, and monocytes (CD11c, CD11b). (E) Proliferation of 2W epitope specific CD4+ T cell population enriched from spleen and lymph nodes in presence or absence of anti-miR-21 oligonucleotides using tetramers is shown. (F) Expression of IL-2R on the activated CD4+ 2W+ T cell population in LdWT2W or LdCen−/−2W infected mice indicating the proliferation is shown. (G) Expression of early CD4+ T cell central memory marker IL-7R on the activated CD4+ 2W+ T cell population in LdWT2W or LdCen−/−2W infected mice is shown. (H). Splenic parasite burden following treatment with LNA-miR-21 in LdWT or LdCen−/− infected mice is shown. Statistical significance between CD4+ T cell proliferation and expression of memory markers in LdWT and LdCen−/− infections 7 days post infection is shown (t-test; *p < 0.05, **p < 0.01, ***p < 0.001).