Abstract / Summary
Abstract P‐glycoprotein (P‐gp, encoded by ABCB1 ) and breast cancer resistance protein (BCRP/ ABCG2 ), present in the blood–brain barrier, protect the fetal brain from exposure to neurotoxicants. P‐gp and BCRP are modulated by microRNA‐145‐5p (miR‐145‐5p) in cancer cells. In the present study, we hypothesized that miR‐145‐5p modulates P‐gp and BCRP in human fetal brain endothelial cells (hfBECs) and in an adult BEC line (human cerebral microvascular endothelial cell line; hCMEC/D3). miR‐145‐5p mimic and inhibitor were transfected into hfBECs ( n = 6, mid‐gestation) and hCMEC/D3 cells. P‐gp and BCRP mRNA, protein expression and efflux activity were assessed using qPCR, Western blotting, immunofluorescence and calcein‐AM and chlorin e6 assays. Target specificity was evaluated using luciferase reporter assays. hfBECs exhibited lower ABCB1 and ABCG2 mRNA and protein expression and reduced P‐gp activity, but significantly increased BCRP activity, compared with hCMEC/D3 cells. The miR‐145‐5p mimic reduced luciferase activity of an ABCB1 3′‐untranslated region (3’‐UTR) reporter but had no effect on the ABCG2 3′‐UTR. In hfBECs, miR‐145‐5p overexpression reduced P‐gp and BCRP levels and efflux activity. In contrast, miR‐145‐5p overexpression in hCMEC/D3 cells increased P‐gp activity while reducing BCRP activity. Inhibition of miR‐145‐5p in hfBECs increased P‐gp protein levels and exhibited a trend towards increased P‐gp activity, without altering BCRP levels, although BCRP activity significantly increased. miR‐145‐5p directly modulates the expression and activity of P‐gp and indirectly modulates BCRP in fetal and adult human BECs. These data suggest that miR‐145‐5p plays an important role in the regulation of key drug transporters at the BBB in fetal brain and may modulate BBB function in adulthood. image Key points microRNA‐145‐5p (miR‐145‐5p) directly targets the 3′‐untranslated region (3′‐UTR) of ABCB1 , but not ABCG2 , as shown by luciferase assays. In human fetal brain endothelial cells (hfBECs), miR‐145‐5p mimic reduces P‐glycoprotein (P‐gp) and breast cancer resistance protein (BCRP) levels and transporter activity. Inhibition of miR‐145‐5p in hfBECs increases P‐gp and BCRP activities. In adult human cerebral microvascular endothelial cell line (hCMEC/D3) cells, miR‐145‐5p mimic increases P‐gp activity and decreases BCRP activity, indicating developmental differences in regulation. miR‐145‐5p directly modulates P‐gp and indirectly modulates BCRP, suggesting a key role in blood–brain barrier (BBB) transporter regulation during fetal development and potentially in adulthood.