Km culture medium

The HepG2 (ATCC-HB-8065) and Calu-1 cells were routinely cultured using EMEN, and supplemented with 10% FBS and 1% penicillin/streptomycin (Sigma-Aldrich, St. Louis, MI, USA) as previously described [45 (link),51 (link)]. Additionally, 5 × 10⁵ primary HEKn cells were thawed in KM culture medium (ScienCell Research Laboratories, Carlsbad, CA, USA) supplemented with KGS 100× (ScienCell Research Laboratories, Carlsbad, CA, USA) and 1% penicillin/streptomycin solution (ScienCell Research Laboratories, Carlsbad, CA, USA). After cells reached ~80–90% confluency, they were washed with DPBS and trypsinized to detach from the culture flask. Trypsin-EDTA 0.025% (2 mL) was added to the monolayer and incubated for 3–5 min, and once detached, 2 mL of TNS was added to the cell suspension and centrifuged at 1200 rpm for 3 min. The supernatant was discarded, and the pellet was resuspended in growth medium and replaced in a 1:4 ratio. Passages 2–4 were used for the experiments. All cell lines were grown at 37 °C in 5% CO₂.

The human hepatoma HepG2 cell line (ATCC-HB-8065) was cultured in EMEN medium (Quality Biological, Gaithersburg, MD, USA) supplemented with 10% FBS (Biowest, Riverside, MO, USA) and 1% penicillin/streptomycin (Sigma-Aldrich, St. Louis, MI, USA). The cells were maintained in a humidified atmosphere with 5% CO₂ at 37 °C [36 (link)]. Primary HEKn cells were cultured in KM culture medium (ScienCell Research Laboratories, Carlsbad, CA, USA) supplemented with KGS 100× (ScienCell Research Laboratories, Carlsbad, CA, USA) and 1% penicillin/streptomycin solution (ScienCell Research Laboratories, Carlsbad, CA, USA). All experiments were conducted using cells in the logarithmic growth phase.