Figure 6

Knockdown of TXNIP decreases inflammasome activation. (A) LP9/hTERT cells (90% confluent) were transiently transfected with siTXNIP or siControl siRNA for 48 h and knockdown of TXNIP expression was assessed by qRT-PCR. (*p < 0.05 compared to siControl) (B) siControl and siTXNIP transfected cells were exposed to crocidolite asbestos for 48 h and the medium were collected, concentrated and analyzed for the presence of caspase-1 (p20) by Western blot analysis. (C) The transcript levels of ERK 1/2 as well as TXNIP were verified in shERK2 HMESO cells by qRT-PCR in the presence and absence of 5 μM Dox along with priming of the inflammasome (*p < 0.05 compared to shCon alone; †p < 0.05 compared to shCon + Dox; n = 2 per group). (D) Inflammasome activation was assessed by Western blot analysis for the p20 fragment of caspase-1 in the media supernatants after treatment with Dox (*p < 0.05 compared to shCon alone; †p < 0.05 compared to shCon + Dox; n = 2 per group). (E) LP9 cells were pretreated with 40 μM of the caspase-1 inhibitor VI (cas-1 inh) prior to exposure to asbestos for 48 h and cells were counted to determine survival (*p < 0.05 compared to control; †p < 0.05 compared to Croc 75 alone). (F) Western blot analysis for Cas-1 p20 fragment in medium supernatant from LP9 cells pretreated with the caspase-1 inhibitor prior to exposure to asbestos (n = 3 per group).