Dmem with 4.5 g l glucose and l glutamine

PBMCs were isolated from healthy human donor buffy coats (provided by the Australian Red Cross Blood Service) by density gradient centrifugation with Ficoll-Paque PLUS (GE Healthcare) and resuspended in complete RPMI medium (RPMI 1640 with L-glutamine [Lonza] supplemented with 10% human serum [Sigma-Aldrich]). CD56⁺ lymphocytes were isolated from PBMCs by MACS positive selection using CD56 MicroBeads, following manufacturer’s protocol (Miltenyi Biotech), and subsequently resuspended in complete RPMI medium. For isolation of NK cells by FACS sorting, CD56⁺-selected lymphocytes were stained with fluorochrome-conjugated antibodies in FACS buffer, and sorted to >95% purity using a FACSAria IIu (BD Biosciences).
The ARPE-19 epithelial cell line, human foreskin fibroblasts (HFFs), and the Vero cell line (all ATCC) were maintained in complete DMEM medium (DMEM with 4.5 g/L glucose and L-glutamine [Lonza] supplemented with 10% foetal calf serum [FCS; Sigma-Aldrich] and penicillin streptomycin [Thermo Fisher Scientific]). K562 cells (ATCC) were maintained in complete RPMI/FCS medium (RPMI 1640 with L-glutamine [Lonza] supplemented with 10% FCS and penicillin streptomycin).

Primary human myoblasts from an unaffected control and a patient with FSHD were isolated from muscle biopsies, purified and established as described previously [65 (link),66 (link)]. The myoblasts were grown in 35 mm collagen-coated dishes (Ywaki, Tokyo, Japan) in DMEM with 4.5 g/L glucose and l-glutamine (Lonza, Verviers, Belgium) as well as gentamycin (50 µg/mL, Sigma-Aldrich, Gillingham, UK), 10% fetal bovine serum (Invitrogen/Thermo-Fisher Scientific, Waltham, MA, USA ), and 1% Ultroser G (Pall BioSepra, Cergy-St-Christophe, France) at 37 °C under 5% CO₂. Confluent myoblast cultures were differentiated by switching the medium to DMEM/gentamicin (50 µg/mL) with 2% FBS.
The immortalized myoblasts were grown and differentiated as described [67 (link)].