Lipopolysaccharide increases cell surface P-glycoprotein that exhibits diminished activity in intestinal epithelial cells.
Increasingly, it is recognized that commensal microflora regulate epithelial cell processes through the dynamic interaction of pathogen-associated molecular patterns and host pattern recognition receptors such as Toll-like receptor 4 (TLR4). We therefore investigated the effects of bacterial lipopolysaccharide (LPS) on intestinal P-glycoprotein (P-gp) expression and function. Human SW480 (P-gp+/TLR4+) and Caco-2 (P-gp+/TLR4-) cells were treated with medium control or LPS (100 ng/ml) for 24 h prior to study. P-gp function was assessed by measuring the intracellular concentration of rhodamine 123 (Rh123). To confirm P-gp-specific effects, breast cancer resistance protein (BCRP/ABCG2) and multidrug resistance-associated protein 2 (MRP-2/ABCC2) were also analyzed. Treatment of SW480 cells with LPS led to diminished P-gp activity, which could be prevented with polymyxin B (control: 207+/-16 versus LPS: 402+/-22 versus LPS+polymyxin B: 238+/-26 pmoles Rh123/mg protein, p<0.05 control versus LPS). These effects could be blocked by using polymyxin B and were not seen in the P-gp+/TLR4--Caco-2 cell line (control: 771+/-28 versus LPS: 775+/-59 pmoles Rh123/mg protein). Total cellular levels of P-gp did not change in LPS-treated SW480 cells; however, a significant increase in cell surface P-gp was detected. No change in activity, total protein, or apically located MRP-2 was detected following LPS treatment. Sequence analysis confirmed wild-type status of SW480 cells. These data suggest that activation of TLR4 in intestinal epithelial cells leads to an increase in plasma membrane P-gp that demonstrates a diminished capacity to transport substrate.
Mishra
Jayshree
J
Department of Clinical Pharmacy, University of Tennessee College of Pharmacy, 19 South Manassas St., Rm. 262, Memphis, TN 38163, USA.
Zhang
Qiuye
Q
Rosson
Jessica L
JL
Moran
John
J
Dopp
John M
JM
Neudeck
Brien L
BL
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism* / Q000502:physiology; D001483:Base Sequence; D015153:Blotting, Western; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D017931:DNA Primers; D005456:Fluorescent Dyes / Q000378:metabolism; D006801:Humans; D047468:Immunoprecipitation; D007413:Intestinal Mucosa; D008070:Lipopolysaccharides / Q000494:pharmacology*; D018613:Microscopy, Confocal; D000089762:Multidrug Resistance-Associated Protein 2
C000715087:ABCC2 protein, human; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D017931:DNA Primers; D005456:Fluorescent Dyes; D008070:Lipopolysaccharides; D000089762:Multidrug Resistance-Associated Protein 2