Fig. 2

IRE1α silencing by siRNA rescues DHT-induced dysfunction of GCs. GCs from naïve rats were treated with DHT, or were transfected with IRE1α siRNA followed by treatment with DHT. A The mRNA levels of IRE1α in GCs were analyzed by qRT-PCR. B The protein levels of IRE1α in GCs were analyzed by western blot. C Levels of IRE1α in GCs (Alexa Fluor 488) were analyzed by immunofluorescence staining using an Alexa Fluor 488-conjugated antibody (60 ×). D-F The expression of IRE1α, TXNIP, NLRP3 (D), NLRP3 inflammasome activation factors (GSDMD, ASC) (E) and fibrosis factors (β-catenin, TGF-β) (F) in GCs were assessed by qRT-PCR. G-H The protein levels of ASC (G) and α-SMA (H) in GCs were analyzed by immunofluorescence staining (60 ×). I ROS in GCs were assessed by the DCF-DA probe using confocal microscopy (90 ×). Data are shown as the mean ± SD. *p ≤ 0.05, **p ≤ 0.01. IRE1α, inositol-requiring enzyme 1α; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; Ctrl, control; DHT, dihydrotestosterone; DAPI, 4’,6-diamidino-2-phenylindole; NC, normal contrast; si, siRNA; TXNIP, thioredoxin-interacting protein; NLRP3, NOD-like receptor family pyrin domain containing 3; GSDMD, gasdermin D; ASC, apoptosis-associated speck-like protein containing a CARD; TGF-β, transforming growth factor-beta; DCF-DA, dichlorofluorescein diacetate; ROS, reactive oxygen species