Fig. 3.

CX3CR1 deficiency promotes the PI3K-Akt-mTORC1 mediated metabolic signaling pathway. Splenic B cells from WT and CX3CR1 KO mice were labeled with biotinylated AF594-mB-F (ab′)₂-anti-Ig(M + G) without (−) or with streptavidin (sAg) at 4 °C, then washed and activated for varying times. After fixation and permeabilization, cells were stained for pSHIP and analyzed using confocal microscopy (a). The Pearson’s correlation coefficients between BCR and pSHIP were determined using the ZEN 2.3 (blue edition) software (b). Western blot analysis the expression of pSHIP and SHIP (c), pPI3K, PI3K, pFoxO1, FoxO1, pAkt, Akt, pS6, S6, pmTOR, mTOR, pNF-κB, NF-κB, pIKKα/β, IKKα/β and HIF-1α in splenic B cells with total proteins as the loading control (e). Calcium assay of CX3CR1 KO splenic B cells was analysed by flow cytometry (g). OCR of CX3CR1 KO mice was measured by Seahorse XF 24 (l). Shown are representative blots and the ratio of phosphorylated/total proteins from three independent experiments (d, f, h–o). *p < 0.05 **p < 0.01 ****p < 0.0001