The transcriptome of human PBMCs incubated with or without Indoxyl sulfate-(Trans.fpkm) The pharmacokinetics of tacrolimus in PBMCs and whole blood of the two rat models of CKD-(MS data)

Human PBMCs were isolated from a healthy male Chinese volunteer, then resuspended in RPMI containing 10% FBS under standard cell culture conditions (humidified atmosphere, 5% CO2, and 37 ℃). Indoxyl sulfate (Sigma) was dissolved in RPMI containing 10% FBS and diluted to 0.2 mM in the range of recipients with CKD. Transcriptome analysis of PBMCs was then performed. Tac blood concentration measurement was performed using an HPLC-MS/MS system comprising an Agilent-1260 system and an API4000 triple quadrupole mass spectrometer (Applied Biosystems, Singapore) in positive mode. The analyte was separated on an ACE C18 column (20×2.1 mm, 2 μm) with an XDB pre-column. The mobile phase consisted of Phase A (water containing 10 mM Ammonium acetate with 0.1% Formic acid) and Phase B (Acetonitrile containing 0.1% Formic acid) at 20% (0-0.01 minutes), 40% (0.3 minutes), 60% (0.8 minutes), 80% (1.2 minutes), 95% (1.5 minutes), 98% (1.8 minutes), 95% (4.5 minutes), 60% (4.6 minutes), 40% (4.8 minutes), 20% (5-6 minutes). Multiple reaction monitoring modes had the m/z transitions of 821.8/768.6 (TAC); 931.8/864.7 (sirolimus [IS]). The linear range for the calibration curve in WB was 0.5-150 ng∙mL-1, and the lower limit of quantitation was 0.5 ng∙mL-1 PBMCs TAC concentration measurement was performed using a UPLC-MS/MS system comprising an LC30-AD (Shimadzu, Kyoto, Japan) system and an API5500 triple quadrupole mass spectrometer (Applied Biosystems, Singapore) in positive mode. The analyte was separated on a Waters C18 column (50×2.1 mm, 1.7 μm). The mobile phase consisted of Phase A (water containing 2 mM Ammonium format with 0.1% Formic acid) and Phase B (methanol) at 25% (0-0.5 minutes), 40% (1 minute), 60% (1.2 minutes), 80% (1.5 minutes), 95% (2 minutes), 95% (4.5 minutes), 60% (4.6 minutes), 40% (4.8 minutes), 25% (5-6 minutes). Multiple reaction monitoring modes had the m/z transitions of 821.8/768.6 (TAC); 931.8/864.7 (sirolimus [IS]). The method was validated according to Clinical Laboratory Standard Institute (CLSI) guidelines and showed high sensitivity and specificity over a range of 0.005-2.5 ng∙10-6 PBMCs in PBMC.