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. 2019 Jul 3;10:1519. doi: 10.3389/fimmu.2019.01519

Figure 6.

Figure 6

rBCG-DisA induced innate immune responses. (A) RAW264.7 macrophages were stimulated with c-di-AMP or infected with indicated bacterial strains at MOI = 10. IFN-β and IL-1β transcripts were analyzed by qRT-PCR. Relative expression of each gene was normalized by expression of GAPDH. (B) RAW264.7 macrophages were treated as indicated in (A), and transcription of autophagy related genes (Atgs) including LC3, Beclin1, Atg5, and Atg7 was determined by qRT-PCR. Relative expression of each gene was normalized by expression of GAPDH. (C) Splenocytes from Naïve, BCG-, and rBCG-DisA-immunized mice were stimulated with BCG proteins for 72 h. Cells were stained with CD4 and TNF-α fluorescently-labeled antibodies and analyzed by flow cytometry. (D) TNF-α transcription in lungs and spleens of Naive, BCG-, and rBCG-DisA immunized mice was measured by qRT-PCR. (E) Eight weeks after Mtb infection, splenocytes from Naïve, BCG-, and rBCG-DisA-immunized mice were stimulated with BCG proteins for 72h. IL-1β, IL-6, and TNF-α production was measured by ELISA. (F) Eight weeks after Mtb infection, histone 3 trimethylation of lysine 4 (H3K4me3) expressions in lung tissues of Naïve, UN, BCG-, and rBCG-DisA- immunized mice were detected by immunohistochemistry (IHC) and observed by microscope (400×). */#P < 0.05, **P < 0.01.