Fbs gold

The Dulbecco’s modified Eagle’s medium (DMEM), containing glucose at 4.5 mg/mL with GlutaMax^TM, trypsin-EDTA, penicillin, streptomycin and fetal bovine serum Gold (FBS Gold) were provided by Gibco (San Diego, CA, USA). Passive lysis buffer, ReliaPrep RNA Cell Miniprep System and luminescent Caspase-Glo 9 Assay were provided by Promega (Madison, WI, USA), and BCA Protein Assay Kit by Thermo Scientific (Rockford, IL, USA). Annexin V Apoptosis Detection Kit I, JC-1 MitoScreen Kit, APO-Direct Kit were product of BD Pharmingen^TM (San Diego, CA, USA). Sigma-Fast BCIP/NBT reagent, acridine orange, fumed silica dioxide amorphous powder 7 nm, silica dioxide spherical, porous nanopowder 5–15 nm and silica dioxide nanopowder 10–20 nm, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and dichlorodihydrofluorescein diacetate were provided by Sigma (St. Louis, MO, USA). Polyclonal (rabbit) anti-human LC3 antibody and alkaline phosphatase-labeled anti-rabbit immunoglobulin G were provided by Cell Signaling Technology (Boston, MA, USA). High Capacity RNA-to-cDNA Kit was purchased from Applied Biosystems (Foster City, CA, USA).

Media and reagents for culture cells: L-15 Medium (1×) + GlutaMAX^TM-I, trypsin-EDTA, RPMI Medium 1640 (1×) + GlutaMAX^TM-I, DPBS, penicillin, streptomycin and fetal bovine serum Gold (FBS Gold) were provided by Gibco (San Diego, CA, USA). Kits for apoptosis detection: FITC Annexin V apoptosis detection Kit I was obtained from BD Pharmingen^TM (San Diego, CA, USA); FAM-FLICA Caspase-8 and FAM-FLICA Caspase-9 Assay were provided by ImmunoChemistry Technologies (Davis, CA, USA). Chemical treatment of cells: reduced graphene oxide (rGO) was obtained from Sigma-Aldrich (St. Louis, MO, USA), while the proteasome inhibitor (MG-132) was provided by Selleckchem (Cologne, Germany). Kit for cytotoxicity analysis: the LDH-Cytotoxicity Assay kit was provided by BioVision (St. Louis, CA, USA). The compounds for oxidative stress analysis: SDS, TCA, TBA, Folin–Ciocalteu reagent and 2′,7′-dichlorodihydrofluorescein diacetate (H₂DCFDA) were provided by Sigma-Aldrich (St. Louis, MO, USA), and GSH/GSSG-Glo^TM Assay kit by Promega (Madison, WI, USA). The compounds for ultrastructure analysis: glutaraldehyde, paraformaldehyde, cacodylate buffer, osmium tetroxide, ethanol and glycid ether were provided by Sigma-Aldrich (St. Louis, MO, USA).

Dulbecco’s Modified Eagle Medium (DMEM) with GlutaMax, trypsin–EDTA, penicillin, and streptomycin, fetal bovine serum Gold (FBS Gold) were provided by Gibco (Paisley, UK), and Annexin V Apoptosis Detection Kit I by BD Pharmingen (San Diego, CA, USA). Dichlorodihydrofluorescein diacetate (DCFHDA), fumed silica powder 7 nm, silica dioxide nanopowder 5–15 nm, and silica dioxide nanopowder 10–20 nm were purchased from Sigma (St Louis, MO, USA).

Human alveolar epithelial A549 cells (ATCC, CCL-185™) were cultured in DMEM/F12 (Gibco) medium supplemented with 10% FBS Gold (PAA)_. The cells were seeded into 6-well plates at a density of 2 × 10⁵ cells/well and incubated overnight at 37 °C in a humidified atmosphere at 5% CO₂. Prior to the treatment, the cells were starved for 24 h in DMEM/F12 media containing 0.1% FBS and then treated for 72 h with either vehicle control (0.1% BSA in 4 mM HCl) or recombinant human TGF-β1 (2 ng/mL) (R&D Systems, USA). Thereafter, cells were lysed with Qiazol (Qiagen, Hilden, Germany) and the lysates were stored at −20 °C until RNA isolation. All stimulations were carried out in triplicate and repeated independently three times.

Dulbecco’s modified Eagle’s medium (DMEM) containing glucose at 4.5 mg/ml (25 mM) with Glutamax, penicillin, streptomycin and trypsin–EDTA were provided by Invitrogen (San Diego, USA), passive lysis buffer by Promega (Madison, USA), FBS Gold by Gibco (San Diego, USA), BCA Protein Assay Kit by Thermo Scientific (Rockford, USA), PE Annexin V Apoptosis Detection Kit I by BD Pharmingen™ (CA, USA), Senescence Detection Kit by bioVision (CA, USA), Sigma-Fast BCIP/NBT reagent by Sigma (St Louis, MO, USA), monoclonal (mouse) anti-human ORP150 antibody by IBL (Gunma, Japan), monoclonal (mouse) anti-HIF-1α antibody by BD Biosciences (CA, USA), and alkaline phosphatase-labelled anti-mouse immunoglobulin G by Rockland (PA, USA), precision plus protein standards by Bio-Rad Laboratories (USA).

The Dulbecco’s modified Eagle’s medium (DMEM), containing glucose at 4.5 mg/mL with GlutaMax^TM, DPBS, trypsin-EDTA, medium RPMI, penicillin, streptomycin and fetal bovine serum Gold (FBS Gold) were provided by Gibco (San Diego, CA, USA). FITC Annexin V apoptosis detection Kit I was provided by BD Pharmingen^TM (San Diego, CA, USA). Reduced graphene oxide (rGO), paraformaldehyde, 3,8-Diamino-5-[3-(diethylmethylammonio)propyl]-6-phenylphenanthridinium diiodide-PI, 2′,7′-dichlorofluorescin diacetate-DCFH-DA,4′,6′-di-amidino-2-phenylindole-DAPI, and crystal violet by Sigma-Aldrich (St. Louis, MO, USA). SDS, TCA, TBA, Folin-Ciocalteu reagent were provided by Sigma-Aldrich (St. Louis, MO, USA). GSH/GSSG-Glo^TM Assay kit was provided by Promega (Madison). LDH-Cytotoxicity Assay kit (C) was provided by BioVision (CA, Milpitas, USA). Methanol, NaOH, and formaldehyde were provided by POCH (Gliwice, Poland).

Media for cells cultivation: L-15 Medium (1×) + GlutaMAX^TM-I, trypsin-EDTA, RPMI Medium 1640 (1×) + GlutaMAX^TM-I, DPBS, penicillin, streptomycin, and fetal bovine serum Gold (FBS Gold) were provided by Gibco (San Diego, CA, USA).
Agents for apoptosis detection: JC-1 MitoScreen Kit, PE Active Caspase-3 Apoptosis Kit, and FITC Annexin V apoptosis detection Kit I were obtained from BD Pharmingen^TM (San Diego, CA, USA); FAM-FLICA Caspase-9 Assay was provided by ImmunoChemistry Technologies (Davis, CA, USA). Chemical treatment of cells: silica nanoparticles 5–15 nm (SiNPs) were obtained from Sigma-Aldrich (St. Louis, MO, USA).
Chemical compounds for oxidative stress analysis: SDS, TCA, TBA, Folin-Ciocalteu reagent, N-acetylcysteine, and 2’,7’-dichlorodihydrofluorescein diacetate (H₂DCFDA) were provided by Sigma-Aldrich (St. Louis, MO, USA) and GSH/GSSG-Glo^TM Assay kit by Promega (Madison, WI, USA).