Rbl 2h3

Human follicle dermal papilla (HFDP) cells were purchased from PromoCell (cat# C-12072; Heidelberg, Germany) and cultured in follicular dermal papilla cell growth medium (C-26051; PromoCell) according to the manufacturer’s protocol at 37°C in a humidified atmosphere of 5% CO₂. The rat basophilic leukemia mast cell line RBL-2H3 cells were purchased from American Type Culture Collection (ATCC, VA, USA). RBL-2H3 cells were cultured in DMEM supplemented with 10% heat-inactivated fetal bovine serum (FBS, WELGENE) and 1% penicillin and streptomycin at 37°C in a humidified atmosphere of 5% CO₂. DHT and Compound 48/80 were purchased from Sigma-Aldrich (St. Louis, MO, USA). To measure the mitochondrial activity of HFDP cells, cells were cultured in 96-well plates and treated with WEML. After 24 h, 20 μL MTS solution (Promega, Madison, WI, USA) was added and cells were incubated at 37°C for 1 h (Kim et al. 2020 (link)). Absorbance was measured at 490 nm using a microplate reader (ABS Plus; Molecular Devices, San Jose, CA, USA). Mitochondrial activity was measured using the following equation:
$\mathrm{M}\mathrm{i}\mathrm{t}\mathrm{o}\mathrm{c}\mathrm{h}\mathrm{o}\mathrm{n}\mathrm{d}\mathrm{r}\mathrm{i}\mathrm{a}\mathrm{l}\mathrm{a}\mathrm{c}\mathrm{t}\mathrm{i}\mathrm{v}\mathrm{i}\mathrm{t}\mathrm{y}\left(\text{\%}\right)=({\displaystyle \frac{A_{\mathrm{e}\mathrm{x}\mathrm{p}\mathrm{e}\mathrm{r}\mathrm{i}\mathrm{m}\mathrm{e}\mathrm{n}\mathrm{t}\mathrm{a}\mathrm{l}\mathrm{g}\mathrm{r}\mathrm{o}\mathrm{u}\mathrm{p}}-A_{\mathrm{s}\mathrm{a}\mathrm{m}\mathrm{p}\mathrm{l}\mathrm{e}\mathrm{b}\mathrm{l}\mathrm{a}\mathrm{n}\mathrm{k}}}{A_{\mathrm{c}\mathrm{o}\mathrm{n}\mathrm{t}\mathrm{r}\mathrm{o}\mathrm{l}\mathrm{g}\mathrm{r}\mathrm{o}\mathrm{u}\mathrm{p}}})\times 100}$ where A is the absorbance for the given wavelength.

Human pulmonary bronchial epithelial BEAS-2B, human epithelial cervix carcinoma HeLa, murine macrophage RAW 264.7, and rat basophilic leukemia mast RBL-2H3 cells were purchased from American Type Culture Collection (ATCC, VA, USA). BEAS-2B cells were maintained in bronchial epithelial basal medium (BEBM; Lonza, Basel, Switzerland). HeLa, RAW 264.7, and RBL-2H3 cells were maintained in DMEM supplemented with 10% heat-inactivated fetal bovine serum (Welgene, Gyeongsangbuk-do, Korea) and 1% penicillin and streptomycin (Welgene) at 37 °C in a humidified atmosphere of 5% CO₂. Melittin and its derived peptides (Figure 1A) were synthesized by Peptron, Inc. (Daejeon, Korea).
Figure 1.

In vitro antioxidant activities of melittin and melittin-derived peptides. (A) Location of peptides in melittin protein. (B) ABTS radical scavenging activity of resveratrol (Res.), melittin, and melittin-derived peptides (P1–P4). Results are expressed as means ± SD (n = 3). *p < 0.05, ***p < 0.001.

The rat basophilic leukemia cell line RBL-2H3 was supplied from the Korean Cell Line Bank (Seoul, South Korea). RBL-2H3 cells were cultured in Minimum Essential Medium (MEM) (Welgene, South Korea) supplemented with 10% FBS (Gibco, Rockville, MD, United States) and 1% penicillin-streptomycin under an atmosphere of 5% CO₂ in a humidified 37°C incubator. RAW264.7 macrophage cells, IgEL b4 cells and 293 T cells were purchased from American Type Culture Collection (Manassas, VA, United States) and were cultured in Dulbecco Modified Eagle’s medium (DMEM) with 10% FBS and 1% penicillin-streptomycin. Calcium ionophore A23187 and phorbol 12-myristate 13-acetate (PMA) were purchased from Sigma-Aldrich (St. Louis, MO, United States). Quercetin, 1-Chloro-2,4-dinitrobenzene (DNCB) and dexamethasone were purchased from Sigma-Aldrich (St. Louis, MO, United States). Albumin from bovine serum, 2,4-dinitrophenylated (DNP-BSA) was purchased from Invitrogen by Thermo Fisher Scientific (Eugene, OR, United States). rIL-4 and mTNF-α were purchased from R&D system Inc (Minneapolis, MN, United States). mIL-6 was purchased from Biolegend (San Diego, CA, United States).
Stock solutions of ZCO were diluted using the mixture solution of DMSO and ethanol (1:1) to a concentration of 20% and filter-sterilized, and then diluted with phosphate buffered saline (PBS) for the working concentration.