Differential effects of mitogen-activated protein kinase pathway inhibitors on P-glycoprotein activation

Hirokazu Wakuda,Kazumasa Shinozuka,Kana Maruyama,Shino Miyauchi,Shizuo Yamada,Keizo Umegaki,Satomi Kagota,Kazuki Nakamura

doi:10.5599/admet.3.1.142

Abstract

The aim of this study was to evaluate the effects of the mitogen-activated protein kinase (MAPK) pathway inhibitors SB203580, CMPD-1, SB239063, SP600125, and FR180204 on the activity of P-glycoprotein (P-gp) and to assess whether the MAPK pathway affects P-gp directly. Changes in the fluorescence of residual rhodamine 123, a marker of P-gp activity, in the apical region of Caco-2 cells were measured in the presence of MAPK pathway inhibitors using time-lapse confocal laser scanning microscopy at 0, 10, 20, 30, and 60 min. Significant differences were observed between the fluorescence levels of control cells and cells treated with SB203580 for 20, 30, or 60 min. However, no significant change was observed in the residual rhodamine 123 fluorescence of cells treated with CMPD-1, SB239063, SP600125, or FR180204. Among the p38-MAPK pathway inhibitors investigated, CMPD-1 and SB239063 showed no detectable effect on the activity of P-gp. Further, JNK 1, 2, 3-MAPK pathway (SP600125) and ERK1/2 pathway (FR180204) inhibitors did not affect P-gp activity. However, SB203580 enhanced the transfer of rhodamine 123 across the apical cell membrane. Thus, SB203580 activated P-gp, although not through the p38-MAPK pathway. Importantly, the MAPK pathway did not affect P-gp activity shortly after treatment.

Highlights

P-glycoprotein (P-gp) is the most thoroughly studied member of the adenosine triphosphate-binding cassette transporter superfamily and is expressed throughout the intestinal epithelium, hepatocytes, renal tubular cells, and the blood-brain barrier [1]
We evaluated the effects of mitogen-activated protein kinase (MAPK) pathway inhibitors (SB203580, competitive p38α and β inhibitor; CMPD-1, non-competitive p38α inhibitor; SB239063, competitive p38α and β specific inhibitor; SP600125, competitive Jun Nterminal kinase (JNK) 1, 2, 3 inhibitor; FR180204, competitive ERK1/2 inhibitor) on the Mitogen-activated protein kinase pathway inhibitors activity of P-gp using time-lapse confocal laser scanning microscopy
Hanks’ Balanced Salt Solution (HBSS) with or without MAPK pathway inhibitors was added to the cells (SB203580, 10 μM; CMPD-1, 10 μM; SB239063, 100 nM; SP600125, 100 nM; FR180204, 1 μM), and cells were observed by time-lapse confocal laser scanning microscopy (Fig. 2)

Summary

Introduction

P-glycoprotein (P-gp) is the most thoroughly studied member of the adenosine triphosphate-binding cassette transporter superfamily and is expressed throughout the intestinal epithelium, hepatocytes, renal tubular cells, and the blood-brain barrier [1]. Fujishiro et al reported that activation of the p38-MAPK pathway is not necessary for insulin-induced glucose uptake but that it regulates glucose transporter expression [7]. Nagelin et al reported that murine 12/15-lipoxygenase regulates the expression and function of the ATP-binding cassette transporter G1 through p38- and JNK2-dependent pathways [8].

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Conclusion