P nfκb p50

Cells were lysed with buffer containing 1% NP-40 and proteinase inhibitor cocktail (Thermo Scientific, Rockford, IL). Protein concentrations were determined by Bradford assay (Bio-Rad) and equalized before loading. Cellular protein from each sample was applied to 8% to 12% SDS-PAGE gels and probed with specific antibodies including EEA1, Rab7, RIP1 (Cell Signaling Technology, Danvers, MA), AnxA2, p-IκBα, p-IKKα, p-ERK, p-p38, p-JNK, p-NFκB p65, p-NFκB p50, TNFα, IL-6, IL-1β, and β-actin (Santa Cruz Biotechnology). Blots were developed with horseradish peroxidase-conjugated secondary antibodies (Santa Cruz Biotechnology) and chemiluminescent substrate on Fuji X-ray films52 (link).

Tetrahydrocurcumin (THC, C₂₁H₂₄O₆) was dissolved in DMSO and stored (Cat#sc-391609, Santa Cruz, CA, USA). P. a. LPS was prepared with PBS and stored (Cat#L9143, Sigma-Aldrich). STAT3 inhibitor AG490 was purchased from Cayman Chemical. JAK inhibitor WP1066 was purchased from Calbiochem. Cell Counting Kit-8 (CCK-8) was purchased from Dojindo Molecular Technologies, Inc. (Rockville, MD, USA). Antibodies against p-JAK2, p-STAT1, p-NFκB p50, and HO-1 inhibitor tin protoporphyrin IX (Snpp) were purchased from Santa Cruz Biotechnology Inc. Antibodies against p-STAT3, iNOS, COX2, and HO-1 were purchased from Abcam. Antibody against p-Nrf2 was purchased from Invitrogen. Antibody against glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was purchased from Taiclone.