Fig. 8. Dvl2 is crucial to regulate NLRP3-driven inflammatory response in MSC-mediated immune regulation.

BMMs were isolated from SMO^FL/FL and SMO^M-KO mice and transfected with the CRISPR/Cas9-Dvl2 KO, CRISPR-Dvl2 activation, CRISPR/Cas9-NRX KO or control vector, and then co-cultured with MSCs followed by LPS stimulation. (A) Western blots analysis and relative density ratio of Dvl2, NRX, β-catenin, XBP1, NEK7, NLRP3, ASC, and cleaved caspase-1 in LPS-stimulated macrophages after transfection of CRISPR/Cas9-Dvl2 KO or control vector. (B) Immunofluorescence staining for NLRP3 expression in macrophages (n=3-4 samples/group). DAPI was used to visualize nuclei. Scale bars, 100μm and 40μm. (C) Western blots analysis and relative density ratio of Dvl2, NRX, β-catenin, XBP1, NEK7, NLRP3, ASC, and cleaved caspase-1 in LPS-stimulated macrophages after transfection of CRISPR-Dvl2 activation or control vector. (D) Immunofluorescence staining for NLRP3 expression in macrophages (n=3-4 samples/group). DAPI was used to visualize nuclei. Scale bars, 100μm and 40μm. (E) Western blots analysis and relative density ratio of NRX, β-catenin, XBP1, NEK7, NLRP3, ASC, and cleaved caspase-1 in LPS-stimulated macrophages after transfection of CRISPR/Cas9-NRX KO or control vector. (F) The schematic figure depicts putative molecular mechanisms by which CD47-mediated Hedgehog/SMO/Gli1 signaling regulates NEK7/NLRP3 activation in MSC-mediated immune regulation. All Western blots represent three experiments, and the data represent the mean±SD. *p<0.05, **p<0.01, ***p<0.001.