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. 2017 Jan 12;7:40418. doi: 10.1038/srep40418

Figure 8. Both NOD1 and NOD2 are involved in bacteria-stimulated activation of the NF-κB pathway in BLP-tolerised macrophages.

Figure 8

(A) Murine BMMs were transfected with the NOD1 specific siRNA (siNOD1), NOD2 specific siRNA (siNOD2), or their scrambled siRNA (scrRNA). Expression of NOD1 and NOD2 protein was assessed by Western blot analysis 24 hrs after transfection. (B,C) BLP-tolerised BMMs transfected with siNOD1/NOD2 or scrRNA were stimulated with S. typhimurium for the indicated time periods. Confocal images were taken after cells were stained with the anti-p65 Ab and Alexa Flour 594-conjugated secondary Ab (B), and nuclear translocation of p65 was quantitatively analysed and expressed as mean fluorescence intensity (MFI) ratio (C). Results shown represent one experiment from a total of three separate experiments. Cell nuclei were stained with DAPI. Scale bar = 10 μm. (D–G) BLP-tolerised BMMs transfected with siNOD1, siNOD2, or their scrRNA were stimulated with S. typhimurium (S. typhi) for the indicated time periods. Expression of Acp5 and Rab10 mRNA (D,E) was assessed by quantitative real-time RT-PCR. Expression of Acp5 and Rab10 protein (F,G) was assessed by Western blot analysis. Results shown represent one experiment from a total of three separate experiments. (H) Intracellular killing of the ingested S. typhimurium by BLP-tolerised BMMs transfected with either siNOD1/NOD2 or scrRNA. All data are mean ± SD from four to six independent experiments in duplicate or triplicate. *p < 0.05, **p < 0.01 compared with BLP-tolerised BMMs transfected with scrRNA.