Ab70598

The processed OSCC cells and the ground tumors were increased with the lysates including RIPA (Beyotime, China) and protease inhibitors (Beyotime, China). The extracted proteins were quantified through BCA method, mixed with appropriate loading, and heated at 100℃ for denaturation. Then same amount (50 μg) of proteins were added to 10% SDS-PAGE, separated by electrophoresis at constant pressure, and transferred to PVDF membrane (Millipore). Next, the membrane with protein was sealed with 5% skim milk for 2 h, exposed to primary antibodies, including ASC (Abcam, ab283684, 1:1000), IL-1β (Abcam, ab254360, 1:1000), IL-18 (Abcam, ab207324, 1:1000), Pro-IL-18 (Proteintech, 10,663–1-AP, 1:1000), NLRP3 (Abcam, ab263899, 1:1000), IL-6 (Abcam, ab9324, 1 µg/ml), Sox4 (Abcam, ab70598, 1:500), JAK2 (Abcam, ab108596, 1:1000), pJAK2 (Abcam, ab32101, 1:1000), STAT3 (Abcam, ab68153, 1:1000), pSTAT3 (Abcam, ab267373, 1:1000) at 4℃ overnight, and secondary antibodies (Abcam) for 1 h. Finally, western blotting was developed after processing with ECL kit (Thermo scientific), and the brightness of each strip can be controlled by adjusting the exposure time.

The BCA kit (Beyotime Biotechnology, Shanghai, China) was utilised to measure the concentration of protein was quantified by using. After the electrophoresis, the PVDF membranes were used. We incubated the membranes with different specific primary antibodies at 4°C overnight after blocking with 5% non‐fat milk at a shaker for 2 h. Another day, the corresponding secondary antibodies were used at room temperature for 2 h. Finally, the relative levels were calculated with a Bio‐Imaging System. The product numbers of antibodies were listed: MAPK14 (ab170099, Abcam, 1:1000), MKK6 (ab33866, Abcam, 1:1000), FOXC1 (ab226219, Abcam, 1:2000), p‐MAPK14 phosphorylate T180 + Y182 (ab195049, Abcam, 1:1000), SOX4 (ab70598, Abcam, 1:1000), SOX13 (ab96776, Abcam, 1:1000), MMP10 (ab261733, Abcam, 1:1000), U2AF2(ab37530, Abcam, 1:250), Tubulin (ab6046, Abcam, 1:500) was used as a internal control. H3 (#4499, CST, 1:2000) was used as a nucleus internal control. The experiment was performed in triplicate.

RIPA buffer (Sigma, USA) was used to isolate protein from rat lung tissue samples, and there proteins were then separated by 10% SDS-PAGE, transferred to PVDF membranes (Millipore, MA, USA), and these were blocked for 2 h with 5% non-fat milk, followed by probing overnight with anti-GAPDH (ab8245, Abcam, UK), anti-SDC4 (ab74139, Abcam), anti-SOX4 (ab70598, Abcam), anti-OCT4 (ab200834, Abcam), and anti-Nanog (ab109250, Abcam), all diluted 1:1000. Following three washes, blots were probed with HRP-anti-rabbit IgG (1:2000, ab6721, Abcam) for 2 h, after which bands were detected using an enhanced chemiluminescence system (ECL, ThermoFisher, USA), with the Image Lab™ Software (Bio-Rad, USA) being used to analyze the data.