Cyclooxygenase 2 cox 2

Lipopolysaccharide (LPS, Escherichia coli O55:B5) was purchased from Calbiochem. Antibodies for inducible nitric oxide synthase (iNOS) and β-actin were purchased from BD Transduction Lab and Novus Biologicals, respectively. Cyclooxygenase-2 (COX-2) and glial fibrillary acidic protein (GFAP) were purchased from Abcam. Total STAT3 and phospho-STAT3 (p-STAT3) were purchased from Cell Signaling Technology. ELISA kits for TNFα and IL-6 were purchased from Invitrogen. 12-(3-adamantan-1-yl-ureido)-dodecanoic acid (AUDA) and N-acetyl-S-farnesyl-L-cysteine (AFC) were purchased from Sigma-Aldrich. Poly-ornithine was purchased from BD Biosciences. Papain and DNase I were purchased from Worthington Biochemical. Fetal bovine serum was purchased from HyClone and heat-inactivated. Culture media and penicillin/streptomycin were purchased from Gibco, and other common chemicals were from Sigma-Aldrich, unless stated otherwise.

To investigate the signaling pathway related to LTE treatment, the right lung was homogenized. Western blot was performed as previously described [14 (link)]. The primary antibodies were used as follows: p-p65 (dilution 1:1000, Cell signaling, Danvers, MA, USA), matrix-metalloproteinase 9 (MMP-9; dilution 1:1000, Cell signaling), cyclooxygenase2 (COX2; dilution 1:1000, Abcam, Cambridge, UK), hemooxygenase-1 (HO-1; dilution 1:1000, cell signaling), and β-actin (dilution 1:1000, cell signaling). The secondary antibody (Thermo Fisher Scientific) was diluted 1:3000 and used. The density of each protein band was measured using ChemiDoc (Bio-Rad, Hercules, CA, USA).

Lipopolysaccharide (LPS), were obtained from Sigma chemical (St. Louis, MO). Penicillin–streptomycin (PS), Fetal bovine serum (FBS), Dulbecco modified Eagle medium (DMEM) were brought from Invitrogen (Carlsbad, CA, USA). sulfanilamide and 0.1% N-1-napthylethylenediamine dihydrochloride were purchased from sigma chemicals. Prostaglandin E2 (PGE2), Interleukin-6 (IL-6), tumor necrosis factor alpha (TNF-α) and interleukin (IL-1β) elisa kit was purchased from R&D Systems (Minneapolis, MN, USA). Primary antibodies against various proteins such as inducible nitric oxide synthase (iNOS), cyclooxygenase 2 (COX-2), and α-tubulin were obtained from abcam, Santa Cruz technology and cell signaling technology. Secondary antibodies against rabbit, goat and mouse were obtained from sigma chemicals. Murine microglial cell BV2 were used to evaluate the effect of compound against LPS activated microglia. BV2 cell we obtained as a gift sample by Dr. E Choi from Korea University, Seoul South Korea. This cell was maintained with high glucose DMEM supplemented with10% heat-inactivated FBS and 1% of mixture of penicillin–streptomycin (penicillin (1 × 10⁵ U/L), and streptomycin (100 mg/L)). Cultured cells were maintained by keeping the in a humidified incubator with 5% CO₂ at 37 °C.