Multidrug resistance-associated protein 4 regulates cAMP-dependent signaling pathways and controls human and rat SMC proliferation
Yassine Sassi, Larissa Lipskaia, Grégoire Vandecasteele, Viacheslav O. Nikolaev, Stéphane N. Hatem, Fleur Cohen Aubart, Frans G. Russel, Nathalie Mougenot, Cédric Vrignaud, Philippe Lechat, Anne-Marie Lompré, Jean-Sébastien Hulot
Yassine Sassi, Larissa Lipskaia, Grégoire Vandecasteele, Viacheslav O. Nikolaev, Stéphane N. Hatem, Fleur Cohen Aubart, Frans G. Russel, Nathalie Mougenot, Cédric Vrignaud, Philippe Lechat, Anne-Marie Lompré, Jean-Sébastien Hulot
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Research Article Vascular biology

Multidrug resistance-associated protein 4 regulates cAMP-dependent signaling pathways and controls human and rat SMC proliferation

Abstract

The second messengers cAMP and cGMP can be degraded by specific members of the phosphodiesterase superfamily or by active efflux transporters, namely the multidrug resistance-associated proteins (MRPs) MRP4 and MRP5. To determine the role of MRP4 and MRP5 in cell signaling, we studied arterial SMCs, in which the effects of cyclic nucleotide levels on SMC proliferation have been well established. We found that MRP4, but not MRP5, was upregulated during proliferation of isolated human coronary artery SMCs and following injury of rat carotid arteries in vivo. MRP4 inhibition significantly increased intracellular cAMP and cGMP levels and was sufficient to block proliferation and to prevent neointimal growth in injured rat carotid arteries. The antiproliferative effect of MRP4 inhibition was related to PKA/CREB pathway activation. Here we provide what we believe to be the first evidence that MRP4 acts as an independent endogenous regulator of intracellular cyclic nucleotide levels and as a mediator of cAMP-dependent signal transduction to the nucleus. We also identify MRP4 inhibition as a potentially new way of preventing abnormal VSMC proliferation.

Authors

Yassine Sassi, Larissa Lipskaia, Grégoire Vandecasteele, Viacheslav O. Nikolaev, Stéphane N. Hatem, Fleur Cohen Aubart, Frans G. Russel, Nathalie Mougenot, Cédric Vrignaud, Philippe Lechat, Anne-Marie Lompré, Jean-Sébastien Hulot

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Figure 4

Use of Ad-shMRP4 in vivo.

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Use of Ad-shMRP4 in vivo. (A) Map of the Ad-shMRP4 vector and the MRP4 s...
(A) Map of the Ad-shMRP4 vector and the MRP4 shRNA sequence. (B) Quantitative real-time PCR and (C) western blot of rat SMCs transfected for 72 h with Ad-shMRP4 or Ad-shLuc (n = 3, ***P < 0.001). Viruses were used at an MOI of 30. (D) Representative hematoxylin and eosin–stained sections of non-injured and injured carotid arteries infected with Ad-shLuc or Ad-shMRP4 14 days after surgery. Scale bar: 50 μm. (E) Average intima/media thickness ratios for the above 3 groups (*P < 0.05, ***P < 0.001 compared with Ad-shLuc). ad, adventitia (n = 5 for non-injured carotid, n = 4 for Ad-shLuc, and n = 6 for Ad-shMRP4). (F) PCR on carotid arteries. DNA from rat carotids was extracted and adenovirus DNA was detected by PCR.