Nci h28

All MPM cell lines were maintained in a humidified atmosphere containing 5% CO₂ in appropriate media supplemented with 10% Fetal Bovine Serum and penicillin streptomycin (500 U/mL). Cell culture reagents were purchased from Lonza (Walkersville, MD, USA). The following MPM cell lines were used in the study: LP9, Met5A, NCI-H2596, MMP, MMB, NCI-H2052, NCI-H28, Ju77, One58, RS-5, DM-3, ACC-MESO-1, ACC-MESO-4, Y-MESO-8D, Y-MESO-9, Y-MESO-12, Y-MESO-14, REN, NCI-H226, and MSTO-211H. ACC-MESO-1, ACC-MESO-4, Y-MESO-9, and Y-MESO-12 were generously provided by Yoshitaka Sekido, (Aichi Cancer Center Research Institute, Japan). NCI-H2052, One-58, and Ju77 cells were provided by Duncan Stewart (University of Leicester, UK). LP-9, MMB, and MMP were a generous gift from Warren Thomas (Royal College of Surgeons in Ireland, Dublin, Ireland). The REN and NCI-H226 cell lines were provided by Dean Fennell (Queen's University, Belfast, Northern Ireland). NCI-H28, and the immortalized non-tumorigenic mesothelial cell line, Met-5A were purchased from the ATCC (LGC Promochem, Teddington, UK). STR profiling of the NCI-H226 was conducted by Source Bioscience (Nottingham, UK) to confirm that the cell line had the correct genotype.

The human primary peritoneal mesothelioma cells (MESO-II and STO) were derived from samples of patients who underwent surgery and provided by Dr. Zaffaroni (Molecular Pharmacology Unit, Fondazione IRCCS Istituto Nazionale dei Tumori, Milano, Italy), as described previously [8 (link)]. The use of the patient samples to generate cell lines was approved by the Institutional Review Board of the Fondazione IRCCS Istituto Nazionale dei Tumori (INT). Written informed consent was obtained from all patients to donate the leftover tissue to INT after diagnostic and clinical procedures. The pleural MM (NCI-H2052, NCI-H28) cell lines and human dermal microvascular endothelial cells (HMEC-1; CRL-3243) were obtained from ATCC (Manassas, VA, USA). MESO-II and STO were cultured in Dulbecco’s Modified Eagle’s Medium F12 with 1 g/L glucose (Gibco, Thermo Fisher Scientific, Waltham, MA, USA). H28 and H2052 were cultured in RPMI 1640 medium with L-glutamine (Merck, Germany) supplemented with 10% Fetal Bovine Serum (FBS; Gibco, Thermo Fisher Scientific, Waltham, MA, USA) and 1% penicillin/streptomycin (Merck, Germany), at 37 °C, 5% CO₂. HMEC-1 cells were cultured as described earlier [34 (link)].

Rat MM cell lines (EM 1–5 and SM 1–5) were established from the ascites of mesothelioma-bearing rats models conducted in our previous studies [26 (link)]. Regarding 8 human MM cell lines, four cell lines, ACC-MESO-4, Y-MESO-8A, Y-MESO-9 and Y-MESO-25 were established in our laboratory (Y.S.) [52 (link), 53 (link)]; NCI-H28 and MSTO-211H were purchased from ATCC (USA), and NCI-H290 and NCI-2052 were kindly provided from Dr. Adi F. Gazdar. Rat peritoneal mesothelial cells (RPMCs) were cultured from the omentum of Wistar rats, and full-length HPV16E6 and E7 were transfected for immortalization as previously described [54 (link)]. RPMC and rat/human mesothelioma cell lines were all maintained and used for experiments (except for conditioned medium preparation) in 1640 medium supplemented with 10% FBS and 1% antibiotic-antimycotic solution in a 5% CO₂ incubator at 37°C. A line of immortalized mesothelial cells, MeT-5A, was purchased from ATCC and cultured in M199, according to the instructions. To prepare conditioned medium, subconfluent cultures of 1 × 10⁶ cells were washed twice with PBS and incubated with serum-free medium for 24 h. Conditioned medium was collected and filtered through a 0.22 μm filter, the same amount of which was loaded for SDS-PAGE as described [26 (link)].