Fig 2: Pathogenic bacteria influence the expression of NLRP3 inflammasome pathway components in the human GEN2.2 pDC cell line. GEN2.2 cells were treated with E. coli (MOI 10) in a time-dependent manner (A,B). The expression of NLRP3, ASC, pro-caspase-1 and pro-IL-1ß was measured at the protein level by Western blotting. Representative blots are shown in (A). Data are shown as mean ± SD from 5 independent experiments in (B). Data were analyzed using one-way ANOVA followed by Bonferroni’s post-hoc test. * p < 0.05, ** p < 0.01 vs. control (ctrl). EC: E. coli.

Fig 3: Western blotting of the NLRP3-ASC-Caspase-1 pathway proteins in the liver (A); the relative expression of NLRP3 inflammasome proteins in the liver (B). Data were presented as the means ± SD (n = 5-7), group differences were determined by ANOVA test followed by Tukey's post hoc test, and *P < 0.05 indicates a difference between the two groups. ASC, apoptosis-associated speck-like protein containing a CARD; Caspase-1, cysteinyl aspartate-specific protease-1; IL, interleukin; MFGM, milk fat globule membrane; NLRP3, nucleotide-binding oligomerization domain leucine-rich repeat and pyrin domain-containing protein 3; SBS, short bowel syndrome.

Fig 4: HAP1-gC1qR knockout cells display loss of OXPHOS and cell proliferation activity. (A) Whole protein extracts were isolated from parental HAP1 cells that express wild-type gC1qR (wt) and HAP1-gC1qR knockout cells (–/–). Western blot analyses were performed using primary antibodies specific for gC1qR (60.11 Ab) or ß-actin. (B) Cellular morphology of parental HAP1 cells and HAP1-gC1qR knockout cells was assessed by light microscopy. (C,D) Oxygen consumption rate (OCR) of HAP1-gC1qR-wt and HAP1-gC1qR-/- cells was determined using the Seahorse XF Cell Mito Stress Test. (E) Graphical presentation of data received from the Seahorse XF Cell Mito Stress Test to visualize cellular metabolic state. (F) Whole protein extracts were isolated from parental HAP1 and -/- cells. Western blot analyses were performed using primary antibodies specific for pp44/42, p44/42, pAMPKa, AMPKa, pAKT, AKT or ß-actin. (G) Wt and –/– cells were cultivated for 72 h and cell viability was assayed by MTS assay. (H) The mRNA expression level of Caspase-1, PYCARD, and NLRP3 in wt and –/– cells were quantified by qPCR. Results are expressed as mean ± SEM and are presented from at least three independent experiments. *p = 0.05, **p = 0.01, ****p = 0.0001.

Fig 5: SIRT1 knockout activates the XBP1/NLRP3 inflammatory pathway(A) mRNA expression of XBP1, NLRP3, IL-1ß, and IL-18 in liver of WT control mice and SIRT1 knockout mice both in response to different treatment. (B) The levels of IL-1ß, TNF-a, and IL-18 in the serum of WT control mice and SIRT1 knockout mice both in response to different treatment as detected by ELISA. (C) The protein expression of XBP1, NLRP3, C-caspase-1, ASC, and NF-?B in liver of WT control mice and SIRT1 knockout mice both in response to different treatment as detected by western blot analysis as well as the corresponding histogram. *p < 0.05, **p < 0.01 versus the sham-operated WT mice or knockout mice; #p < 0.05, ##p < 0.01 versus WT control mice subjected to IR. The data are measurement data, presented as mean ± standard deviation. Data between two groups were compared using an unpaired t test. The experiment was independently repeated three times. n = 10.