Characterization of the renal tubular transport of creatinine by activity-based protein profiling and transport kinetics

Identification and characterization of an endogenous biomarker of the renal vectorial transport (OCT2‐MATE1)

“…Blood from patients with chronic kidney disease (serum creatinine level of 443–706 μM) was used as a substrate source for obtaining endogenous substrates of OCT2. To further evaluate the effect of human serum powder on OCT2 activity, d3‐creatinine (creatinine, an endogenous substrate of OCT2 [Ciarimboli et al., 2012; Ma et al., 2023]) uptake in OCT2‐overexpressing cells was performed. The results showed that serum powder could inhibit OCT2‐mediated d3‐creatinine uptake, but the uptake of d3‐creatinine in HEK293T‐OCT2 still markedly higher than that in HEK293T‐MOCK cells (Figure S2).…”

“…There is relatively high expression of these transporters in the kidney, which is the likely major site of renal thiamine reuptake (Dutta et al., 1999; Larkin et al., 2012). Although OCT2 mediates the secretion of creatinine in renal tubules, it is mainly filtered through the glomerulus under normal kidney function, and renal creatinine clearance rate is considered the GFR in the clinic setting (Li et al., 2019; Ma et al., 2023). Therefore, the ratio of renal creatinine clearance rate to renal substrate clearance rate is used as a measure of whether there is renal tubular secretion or reabsorption of substrates.…”

“…HEK293T cell lines stably transfected with human OCT2 and MATE1 were constructed and evaluated as described previously (Ma et al., 2023). Blood from patients with chronic kidney disease (serum creatinine level of 443–706 μM) was collected in accordance with the ethical standards of the Ethics Committee of the First Hospital of Lanzhou University.…”

“…Specificity validation: HEK293T cell lines stably transfected with human high abundance drug transporters OAT1, OAT2, OAT3, MRP4, organic anion transporting polypeptide 4C1 (OATP4C1), P-gp, peptide transporter 2 (PEPT2), urate transporter 1 (URAT1), and OAT4 have been successfully constructed in our laboratory (Ma et al, 2023). HEK293T-MOCK and transporter-overexpressing cells were seeded in 12-well plates (5 � 10 5 cells/mL, 1 mL) and cultured for 24 h, after which the medium was replaced with buffer containing Transport kinetics experiments: HEK293T-MOCK and HEK293T-OCT2 or -MATE1 cells were seeded in 12-well plates (5 � 10 5 cells/ mL, 1 mL) and cultured for 24 h, after which the medium was replaced with buffer containing 100 μM of differential metabolites.…”

“…Blood from patients with chronic kidney disease (serum creatinine level of 443-706 μM) was used as a substrate source for obtaining endogenous substrates of OCT2. To further evaluate the effect of human serum powder on OCT2 activity, d3-creatinine (creatinine, an endogenous substrate of OCT2 [Ciarimboli et al, 2012;Ma et al, 2023]) uptake in OCT2-overexpressing cells was performed.…”