Anti p16ink4a

The following primary antibodies were used: anti‐p53 (Abcam, UK), anti‐p16^INK4a (Abcam, UK), anti‐p16^INK4a (Abcam, UK), anti‐p21^CIP1/WAF1 (Abcam, UK), anti‐GATA4 (Santa‐Cruz, USA), anti‐GATA4 (Abcam, UK), anti‐CCN1 (Abcam, UK), anti‐γ‐H2AX (Abcam, UK), anti‐γ‐H2AX (CST, USA), anti‐GAPDH (Proteintech, China), anti‐β‐actin (Boster, China), anti‐interleukin (IL)‐1α (Proteintech, China), anti–monocyte chemotactic protein‐1 (Abcam, UK), anti–tumor necrosis factor‐α (TNF‐α) (Abcam, UK), and anti‐α‐actin (Proteintech, China). All secondary antibodies were from ZSGB‐BIO Company (China). Reagents wheat germ agglutinin and Alda‐1 were purchased from Sigma‐Aldrich, USA. All of the catalog numbers of the antibodies are listed in Table S1.

After extracting the total protein from tissues and cellular samples using RIPA buffer with protease inhibitors, BCA quantitative measurements of protein concentrations were conducted. The relative expressions of the target proteins were normalized to β-actin. A total of 20 μg protein for each sample was separated by SDS-PAGE and transferred onto PVDF membrane (Cat No.: IEVH00005, Millipore, USA). After incubating with the milk in TBS for blocking, the membrane would be incubated the corresponding primary antibody, including anti-p16^INK4a (Cat No.: ab108349, Abcam, UK), p21^Waf/Cip1 (Cat No.: ab109520, Abcam, UK), SIRT1 (Cat No.: 2310, Cell Signaling, USA), NLRP3 (Cat No.: ab263899, Abcam, UK), apoptosis-associated speck-like protein (ASC, Cat No.: 13833, Cell Signaling, USA), pro-caspase-1 (Cat No.: ab179515, Abcam, UK), caspase 1 (Cat No.: ab207802, Abcam, UK) and β-actin (Cat No.: sc-47724, Santa Cruz, USA), at 4°C overnight. On the second day, the membrane was washed three times with TBST and incubated with the secondary antibody (Cat No.: sc-2357 and sc-2005, Santa Cruz, USA) for 1 h. The density of the bands was quantified using Labworks image acquisition software (UVP, USA) and was quantized using ImageJ software (NIH, USA).

Cells collected by FACS were washed by PBS and lysed in RIPA buffer. Protein content was determined by the Bradford assay (Beyotime Institute of Biotechnology, Haimen, China). 50μg proteins were separated in a 15% SDS-polyacrylamide gel electrophoresis and transferred to PVDF membrane. The membranes were first blocked with 5% (w/v) nonfat dry milk in TBST and then probed with the indicated primary antibodies with gentle shaking at 4°C overnight. After washing 3 times, the membranes were incubated with the HRP-conjugated secondary antibodies for 1 h. The signals were detected using an enhanced chemiluminescence detection kit (Thermo Scientific, Illinois, USA). The anti-E-Cadherin (1: 500 dilution), ani-Vimentin (1: 1000 dilution), anti-IL-8(1: 200 dilution), anti-p16^INK4a (1: 500 dilution) and anti-GAPDH (1: 5000 dilution) antibodies were purchased from Abcam Co., MA, USA (Cat # ab323410, ab8069, ab18672, ab108349 and ab8245).