Il 1β

Dulbecco’s modified Eagle’s minimum essential medium (DMEM), fetal bovine serum (FBS), penicillin, and streptomycin were purchased from Life Technologies Inc. (Grand Island, NY, USA). The antibodies for iNOS, COX-2, p65, PARP, β-actin, normal rabbit IgG and normal mouse IgG_2b were obtained from Santa Cruz Biotechnology Inc. (Santa cruz, CA, USA). The enzyme immunoassay (EIA) kits for PGE₂, IL-6, IL-1β, and TNF-α were purchased from R&D Systems (Minneapolis, MN, USA). TOPscript™ RT Dry MIX was purchased from Enzynomics Co. Ltd. (Daejeon, Korea). iNOS, COX-2, TNF-α, IL-6, IL-1β, and β-actin oligonucleotide primers were purchased from Bioneer (Seoul, Korea). MTT, l-N⁶-(1-iminoethyl)lysine (l-NIL), NS-398, Escherichia coli LPS and all other chemicals were obtained from Sigma Chemical Co. (St. Louis, MO, USA). HPLC-grade acetonitrile was purchased from Fisher Scientific Korea (Seoul, Korea) and HPLC-grade formic acid was purchased from Wako (Osaka, Japan).

Roswell Park Memorial Institute medium (RPMI-1640) for culturing macrophage, penicillin–streptomycin, fetal bovine serum, and phosphate-buffered saline were acquired from Welgene (Gyeongsan-si, Korea). Oligo-dT and primer sequences, including COX-2, iNOS, IL-1β, IL-6, and TNF-α, were obtained from Bioneer (Daejeon, Republic of Korea). Other reagents were purchased from Invitrogen, Sigma-Aldrich, and Thermo Fisher.

For this study, roxatidine (≥98% purity, Fig. 1A), primary antibodies for p65, p–IκBα, IκBα, p-ERK, p-p38, p-JNK, ERK, p38, JNK, PARP, α-tubulin, caspase-1, MKK3/6, MAPK-activated protein kinase 2 (MK2), and β-actin and peroxidase-conjugated secondary antibodies were purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA, USA). p-IKK, IKK, p-Akt, Akt, p-MKK3/6, and p-MK2 primary antibodies were obtained from Cell Signaling Technology (Danvers, MA) and M5/114.15.2 (MHC class II) antibody was purchased from eBioscience (eBioscience, Affimatrix Phorbol 12-myristate 13-acetate (PMA), calciumionophore A23187 (Calcymycin; C29H37N3O6), 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyl tertazolium bromide (MTT), fluorescein isothiocyanate (FITC) isomer-I, 2,4-dinitrochlorobenzene (DNCB) and all other chemicals were purchased from Sigma Chemical Co. (St. Louis, MO). Iscove’s modified Dulbecco’s medium (IMDM) fetal bovine serum (FBS), penicillin, and streptomycin were obtained from Life Technologies Inc. (Grand Island, NY). The enzyme immunoassay (EIA) kits for TNF-α, IL-6, and IL-1β were obtained from R&D Systems (Minneapolis, MN, USA). SYBR Premix Ex Taq was purchased from Takara (Shuzo, Shiga, Japan). TNF-α, IL-6, IL-1β, and GAPDH oligonucleotide primers were purchased from Bioneer (Daejeon, Chungbuk, South Korea). SB203580 was purchased from Sigma Chemical Co. (St. Louis, MO).

RPMI, penicillin, and streptomycin were purchased from Hyclone (Logan, UT, USA). Bovine serum albumin and LPS were purchased from Sigma (St. Louis, MO, USA). Antibodies against iNOS, COX-2, p-IKK, IKK, IκBα, p-p65 (NF-κB), p65 (NF-κB), β-actin, p-ERK, ERK, p-JNK, JNK, p-p38, and p38 were purchased from Cell Signaling Technology, Inc. (Danvers, MA, USA). Oligonucleotide primers for TNF-α, IL-6, IL-1β, and GAPDH were purchased from Bioneer Corp. (Daejeon, Korea).

Total RNA was isolated from HaCaT cells using TRIzol (Life Technologies, Carlsbad, CA, USA) according to the manufacturer’s protocol and a previously described method with a slight modification [17 (link)]. cDNA was synthesized from 1 µg RNA by a reverse transcriptional reaction. The qPCR amplification was performed with SYBR-Green/ROX qPCR Master Mix (2×) (BioFact Co., Daejeon, Republic of Korea). The PCR primers for IL-6, TNF-α, IL-1β, iNOS, PLTP, APOE, ABCA1, and GAPDH were obtained from Bioneer Corp. Primer sequences are listed in Table 1.