Expression Profile of Drug and Nutrient Absorption Related Genes in Madin-Darby Canine Kidney (MDCK) Cells Grown under Differentiation Conditions

Abstract

The expression levels of genes involved in drug and nutrient absorption were evaluated in the Madin-Darby Canine Kidney (MDCK) in vitro drug absorption model. MDCK cells were grown on plastic surfaces (for 3 days) or on Transwell® membranes (for 3, 5, 7, and 9 days). The expression profile of genes including ABC transporters, SLC transporters, and cytochrome P450 (CYP) enzymes was determined using the Affymetrix® Canine GeneChip®. Expression of genes whose probe sets passed a stringent confirmation process was examined. Expression of a few transporter (MDR1, PEPT1 and PEPT2) genes in MDCK cells was confirmed by RT-PCR. The overall gene expression profile was strongly influenced by the type of support the cells were grown on. After 3 days of growth, expression of 28% of the genes was statistically different (1.5-fold cutoff, p < 0.05) between the cells grown on plastic and Transwell® membranes. When cells were differentiated on Transwell® membranes, large changes in gene expression profile were observed during the early stages, which then stabilized after 5–7 days. Only a small number of genes encoding drug absorption related SLC, ABC, and CYP were detected in MDCK cells, and most of them exhibited low hybridization signals. Results from this study provide valuable reference information on endogenous gene expression in MDCK cells that could assist in design of drug-transporter and/or drug-enzyme interaction studies, and help interpret the contributions of various transporters and metabolic enzymes in studies with MDCK cells.

Targeting the blood-brain barrier with solid and vesicular nanoparticles decorated with ligands of solute carriers

“…In the epithelial cell lines, the most dominant efflux transporters were MRP2 and MRP3, while in the EPA model MRP1 and MRP5. The data on epithelial cells are in concordance with the findings of Hayeshi et al (2008) and Quan et al (2012).…”

“…Moreover, a correlation was found between normalized mRNA rank and normalized protein abundance rank in Caco-2 cells for selected SLC genes (Ölander et al, 2016). The renal MDCK cell line expressed GLUT1 at high level, as it was already published (Quan et al, 2012), but not the other two GLUT transporters.…”

“…In addition to Pgp, MRP1 was demonstrated at protein level by both Western blot and immunohistochemistry in our EPA model . The canine kidney cells did not express BCRP, as it was demonstrated in an independent study (Quan et al, 2012), which should be taken into account for drug efflux studies. In the epithelial cell lines, the most dominant efflux transporters were MRP2 and MRP3, while in the EPA model MRP1 and MRP5.…”

Influence of P-Glycoprotein Inhibition or Deficiency at the Blood–Brain Barrier on 18F-2-Fluoro-2-Deoxy-d-glucose (18F-FDG) Brain Kinetics

“…We may nonetheless hypothesize that this result may be biased by cell metabolism shift due to prolonged incubation in a Dglucose-free medium, although cell viability was >95% after 120-min incubation. MDCKII cells express GLUT1 and GLUT8 (45). We used a high concentration of D-glucose to unveil the potential transport of 18 F-FDG by P-gp, apart from the GLUT activity.…”

Perspective on the Application of Microphysiological Systems to Drug Transporter Studies

“…Ideally, MPS models would use high-quality primary cells, a source afflicted by high costs and variability from both donors and suppliers. Although prototypical cell lines (e.g., MDCK and Caco-2 cells) can guarantee better reproducibility, they can also have complete transport systems intrinsically abrogated, exemplified by the lack of expression of BCRP and OATs in MDCK cells (Maubon et al, 2007;Quan et al, 2012). Induced pluripotent stem cells (iPSCs) have the potential to fill the gap between primary cells and cell lines, providing an abundant and stable supply of high-quality cells across a range of organ types with native functionalities.…”