Amphotericin

Poly (1,4-butylene succinate) extended with 1,6-diisocyanatohexane (T_m 120 °C) (PBS), 1,1,1,3,3,3-hexafluoroisopropanol (HFIP) and Dulbecco’s phosphate buffered saline (DPBS) were purchased from Aldrich Milan, Italy. This merchant specifies only the T_m for PBS, but Fabbri et al. performed a GPC analysis to evaluate the molecular weight (81.2 kDa) [21 (link)].
Dulbecco’s modified Eagle’s medium (DMEM), fetal bovine serum (FBS), trypsin, l-glutamine, penicillin, streptomycin, and amphotericin were purchased from Euroclone group (Milan, Italy).
Porcine bile was extracted from pigs at Istituto Zooprofilattico della Sicilia “A. Mirri,” Palermo, Italy accordingly with European rules on animal experiments.
Human blood was extracted from volunteers upon informed consent and isolated at the University of Palermo, Palermo, Italy.
NHDF-Ad-Human Dermal Fibroblasts, Adult were obtained from Lonza bioscience and used after 9 doublings. The cell line was grown in a minimum essential medium [Dulbecco’s modified Eagle’s medium (DMEM)] supplemented with 10% (v/v) fetal bovine serum (FBS), 2 mM l-glutamine, 100 um/mL penicillin, 100 μg/mL streptomycin, and 2.5 μg/mL amphotericin B (all reagents were from Euroclone, Milan, Italy) under standard conditions (95% relative humidity, 5% CO₂, 37 °C).

Primary tubular cell cultures were obtained from 44 independent human renal cortex tissue specimens and characterized as previously described [22 (link)–27 (link)]. Normal renal cortex specimens were obtained from adult human kidneys surgically removed because of renal carcinoma, after written patients’ informed consent and accordingly with the Declaration of Helsinki recommendations and with Ethical Committee “Comitato Etico Azienda Ospedaliera San Gerardo” (No. 1532, 17 November 2011). At the first confluence, the cells were detached with trypsin and replated to reach the second confluence at the end of each treatment period. After 24h of serum starvation, the cells were cultured in low glucose Dulbecco’s modified Eagle’s medium (DMEM) (100 mg/dl glucose; control medium), or in high glucose DMEM (450 mg/dl glucose; HG medium), both supplemented with 10% fetal bovine serum (FBS), 1% glutamine, 1% penicillin-streptomycin and 1% amphotericin (Euroclone, Milan, Italy) for up to 7 days. The glucose concentration was regularly checked with Atellica CH Glucose Hexokinase_3 (GluH_3) assay by Atellica Solution Analyzer (Siemens, Munich, Germany), and, when necessary, restored by addition of fresh D-glucose (Sigma-Aldrich, St Louis, MO, USA). Osmolality balance was obtained by addition of D-mannitol (350 mg/dl; Sigma-Aldrich) in control medium.

Rat embryonic cardiomyoblasts (H9c2) (ATCC CRL-1446, Mannassas, VA, USA) were maintained at 37 °C in humidified atmosphere (5% CO₂) and cultured in growth medium composed by Dulbecco’s Modified Eagle Medium (DMEM) (Euroclone, Milano, Italy) supplemented with 10% (v/v) fetal bovine serum (FBS), penicillin (100 U/mL), streptomycin (0.1 mg/mL), amphotericin (0.25 µg/mL) and L-Glutamine (2 mM) (all products from Euroclone, Milano, Italy). Cells were passed after 70–80% confluence was achieved. The differentiation of embryonic cardiomyoblasts into a mature phenotype was induced 24 h after seeding, reducing the percentage of serum (1% FBS) followed by the addition of 1 µM retinoic acid (RA) (Sigma, Milano, Italy). The stimulus (1% FBS + RA) was renewed every 2 days.