Fetal bovine serum influences the effect of a ligand on the activity of the SV23 splice variant of human constitutive androstane receptor (1148.1)

Abstract

The naturally occurring SV23 splice variant of human constitutive androstane receptor (hCAR‐SV23) is activated by di‐(2‐ethylhexyl)phthalate (DEHP), which is detected as a contaminant in fetal bovine serum (FBS). We determined the effect of charcoal‐stripped, dextran‐treated FBS (CS‐FBS), dialyzed FBS, and regular FBS on hCAR‐SV23 activity in a cell‐based reporter gene assay. In transfected HepG2 cells cultured in Minimum Essential Medium (MEM) supplemented with 10% FBS, basal hCAR‐SV23 activity varied with the type of FBS (regular > dialyzed > CS). DEHP increased hCAR‐SV23 activity when 10% CS‐FBS, but not regular or dialyzed FBS, was used to supplement MEM. With increasing concentrations (1‐10%) of regular or CS‐FBS, hCAR‐SV23 basal activity increased, whereas the DEHP‐mediated hCAR‐SV23 activity remained constant (regular FBS) or slightly increased (CS‐FBS). Subsequent experiments identified the following optimal conditions to conduct the hCAR‐SV23 activity assay: 1% CS‐FBS in Minimum Essential Medium‐Reduced Serum (MEM‐RS) for plating of cells and for the first 24 h after plating; and then in serum‐free MEM‐RS. Under these conditions, artemisinin, artemether, and arteether increased hCAR‐SV23 activity. In contrast, they decreased it in cells cultured in MEM supplemented with 10% regular FBS. In conclusion, the type and concentration of FBS influence the effect of a ligand on hCAR‐SV23 activity.Grant Funding Source: Canadian Institutes of Health Research and Michael Smith Fdn. for Health Res.