Figures & data
Figure 1. Acute arsenic treatment increases IL-1α, IL-1β and IL-18 mRNA levels as well as secretion levels in HaCaT cells. (A) HaCaT cells were treated with 0.5, 2 or 10 µM sodium arsenite for 24 h, or treated with 2 µM sodium arsenite for 24, 48 or 72 h. The mRNA levels of IL-1α, IL-1β and IL-18 in the cells were determined by real-time PCR. (B) The protein secretion levels of IL-1α, IL-1β and IL-18 in the media were determined by ELISA. (C) Cell viabilities were determined by cell counting kit-8. The experiments were repeated three times. Data are expressed as the mean ± SEM, *p < 0.05 vs. control.
Figure 2. Arsenic activates AIM2 inflammasome in HaCaT cells. (A) HaCaT cells were treated with sodium arsenite at 2 µM for 24, 48 or 72 h. Expressions of cleaved caspase-1 were detected by Western-blot. (B) Real-time PCR showed the mRNA level of AIM2, NLRP1, NLRP3, NLRC4 and ASC in HaCaT cells after arsenic treatment. (C) The protein level of AIM2 was determined by Western-blot and quantified by densitometry. (D) HaCaT cells were transiently transfected with AIM2-shRNA plasmids. The downregulation of AIM2 was determined by Western-blot. (E) Control or AIM2-knockdown HaCaT cells were treated with arsenic for 24 h. The levels of cleaved caspase-1 were determined by Western-blot. Cytokine secretions in the media of arsenic-treated or control cells were determined by ELISA. The experiments were repeated three times. Data are represented as mean ± SEM of three experiments. *p < 0.05 vs. control.
Figure 2. (Continued).
Figure 3. AIM2 deficiency inhibits arsenic-induced cleavage of pro-casepase-1 and secretion of IL-1β and IL-18 in the skin of AIM2 KO mice. (A) AIM2 deficiency was confirmed by protein gel blot (upper panel) and arsenic-induced cleavage of pro-casepase-1 was inhibited in the skin of AIM2 KO mice. (B) Arsenic-induced secretion of IL-1β and IL-18, but not IL-1α, in the skin of wildtype mice was inhibited in AIM2 KO mice. The experiments were repeated three times. Data are represented as mean ± SEM of three experiments. *p < 0.05 vs. control.
Figure 4. Arsenic activates AIM2 inflammasome through activation PKR. (A) HaCaT cells were treated with sodium arsenite at 2 µM for 24, 48 or 72 h. The levels of p-PKR, PKR, p-eIF2α and eIF2α were determined by Western-blot. (B) HaCaT cells were treated with 2 µM arsenic together with PKR inhibitors C16 or 2-AP for 24 h, or transiently transfected with PKR siRNA and then treated with arsenic for 24 h. The levels of p-PKR, PKR, p-eIF2α, eIF2α, AIM2 and cleaved caspase-1 were determined by Western-blot and quantified by densitometry. (D) Cytokine secretions in the media of arsenic-treated or control cells were determined by ELISA. The experiments were repeated three times. Data are represented as mean ± SEM of three experiments. *p < 0.05 vs. control.
Figure 4. (Continued).
Figure 5. PKR mutation inhibits arsenic-induced activation of AIM2 inflammasome and secretion of IL-1β and IL-18, but not IL-1α, in the skin of C-PKR−/− mice. The C-PKR−/− mice and controlled C57BL/6J mice were treated with 0.25 µM sodium arsenite in drinking water for 8 weeks. The skin tissues were collected. (A) The protein levels of AIM2, cleaved caspase-1, IL-1α, IL-1β and IL-18 were determined by Western-blot and quantified by densitometry. (B) The secretion levels of IL-1α, IL-1β and IL-18 were determined by ELISA assay. The experiments were repeated three times. Data are represented as mean ± SEM of three experiments. *p < 0.05.
