Sodium lauryl sulfate

Ciclopirox olamine (CPO) batch 19D11-B023-0486 was purchased from Fagron (Rotterdam, The Netherlands). Micolamina^® lacquer batch T266 (CPO 8%, isopropyl alcohol, ethyl acetate, polymeth-acryliccopolyethylacrylate and dimethylsulfoxide) was purchased from a local distributor in Recife, Brazil. Isopropyl myristate, polyethylene glycol 400, Tween^® 20 (Polyoxy-ethylene sorbitan monolaurate), Span^® 80 (Sorbitan monooleate), Tween^® 80 (Polyox-yethylene sorbitan monooleate), ethyl alcohol, Kolliphor^® EL (Macrogolglycerol ricinoleate), Poloxamer 407, propylene glycol, isopropyl alcohol, potassium hydroxide, urea, hydroxypropylmethylcellulose (HPMC), sodium chloride, potassium chloride, sodium phosphate, potassium phosphate, sodium azide, oleic acid, BRIJ^®20 (Polyoxy-ethylene 20 oleyl ether), sodium lauryl sulfate (SLS), and HPLC solvents methanol and acetonitrile from JT Baker, Phillipsburg, NJ, USA, were purchased from Casa do laboratório, Pernambuco, Brazil. Sabouraud dextrose agar and mycosel agar (PL1340) were purchased from Sigma-Aldrich (St. Louis, MO, USA) and Plastlabor (Rio de Janeiro, Brazil), respectively.

The 96-well plates were precoated with 100 mg/ml fibronectin at 4 °C overnight and blocked with 1% BSA at 37 °C for 1 h. Next, 2 × 10⁴ Calu-3 cells were inoculated into the 96-well plates and cultured in serum-free DMEM. After 2 h of culture, cells were rinsed with PBS 3 times to gently remove nonadherent cells. Thereafter, attached cells were fixed with 4% paraformaldehyde, and stained with 0.5% crystal violet (Sangon Biotech). The stained crystal violet was dissolved with lauryl sodium sulfate (Amresco, Solon, OH, USA). Absorbance at 570 nm was read with a microplate reader.

The reagents used in this study were as follows: liposomes were purchased from Invitrogen (Carlsbad, CA, USA); glycine, lauryl sodium sulfate, tetramethylethylenediamine, TRIzol, and tris(hydroxymethyl)aminomethane were purchased from Amresco (Solon, OH, USA); acrylic amide was purchased from Merck (Darmstadt, Germany); bovine serum albumin (BSA) was purchased from Roche (Basel, Switzerland); fluorescent protein solutions were purchased from Pierce (Rockford, IL, USA); ammonium peroxydisulfate, dimethyl sulfoxide (DMSO), N, N’-methylenebisacrylamide, and puromycin were purchased from Sigma (St. Louis, MO, USA); trypsin, Dulbecco’s modified Eagle’s medium (DMEM) with high glucose, and fetal bovine serum were purchased from HyClone (Logan, UT, USA); PrimeSTAR DNA polymerase and T4 DNA ligase were purchased from TaKaRa (Tokyo, Japan); GAPDH, P62, CHK1, CHK2, MYTl, WEEl, CDC25, CDC25C, ATM, CDK1, LC3, p68-CHK1, p216-CDC25C, p15-CDK1, p1981-ATM, SQSTM1/P62, pCHK2, pCHK1, CDC, pCDC25C, and MPM2 antibodies were purchased from Santa Cruz Biotechnology (Dallas, TX, USA), Cell Signaling Technology (Danvers, MA, USA), or Millipore/Upstate(NY, USA); and the pLKO.1 plasmid was purchased from Sigma (Darmstadt, Germany).