Tài liệu Báo cáo khoa học: Modulation of cyclin D1 and early growth response factor-1 gene

Modulation of cyclin D1 and early growth response factor-1 gene

expression in interleukin-1b-treated rat smooth muscle cells

by n-6 and n-3 polyunsaturated fatty acids

Souad Bousserouel, Michel Raymondjean, Arthur Brouillet, Gilbert Be´re´ ziat and Marise Andre´ ani

UMR 7079 Physiologie et Physiopathologie, Universite´ Pierre et Marie Curie, Paris, France

The proliferation of smooth muscle cells (SMC) is a key

event in the development of atherosclerosis. In addition to

growth factors or cytokines, we have shown previously that

n-3 polyunsaturated fatty acids (PUFAs) act in opposition

to n-6 PUFAs by modulating various steps of the inflam￾matory process. We have investigated the molecular mech￾anisms by which the incorporation of the n-6 PUFA,

arachidonic acid, increases the proliferation of rat SMC

treated with interleukin-1b, while the n-3 PUFAs eicosa￾pentaenoic acid (EPA) and docosahexaenoic acid (DHA),

elicit no mitogenic response. Incorporation of EPA or DHA

into SMC, which are then activated by interleukin-1b to

mimic inflammation, decreases promoter activity of the

cyclin D1 gene and phosphorylation of the retinoblastoma

protein. Together, our data demonstrate that n-3 effects are

dependent on the Ras/Raf-1/extracellular signal regulated

kinase (ERK)/mitogen-activated protein kinase pathway,

and that down-regulation of the cyclin D1 promoter activity

is mediated by the specific binding of the early growth re￾sponse factor-1. Finally, we have shown that the incorpor￾ation of EPA and DHA also increased the concentration of

caveolin-1 and caveolin-3 in caveolae, which correlated with

n-3 PUFA inhibition of SMC proliferation through the

mitogen-activated protein kinase pathway. We provide evi￾dence indicating that, in contrast to n-6 PUFAs, n-3 PUFAs

exert antiproliferative effects on SMC through the mitogen￾activated protein kinase/ERK pathway.

Keywords: caveolae; cyclin D1 gene expression; interleukin￾1b; n-3 PUFA; vascular smooth muscle cells.

Early atherosclerosis lesions have many features of an

inflammatory process, and the proliferation of smooth

muscle cells (SMC) from the medial layer of the vessel to the

intima is a key event in the development of this disorder [1].

As interleukin-1 b (IL-1b) seems to act (probably in

association with other growth factors) as a mitogen in

cultured SMC, its release in large quantities by activated

endothelial cells and macrophages may contribute to

atherosclerotic lesions [2]. Long-chain n-3 and n-6 poly￾unsaturated fatty acids (PUFAs) may also modulate the

mitogenic response. Arachidonic acid (AA, 20:4 n-6)

and its metabolites stimulate growth by activating the

mitogen-activated protein kinase (MAPK) pathway in

many cell types, including SMC [3,4]. There is also good

evidence that growth factors and cytokines stimulate AA

release [5,6].

Eicosapentaenoic acid (EPA, 20:5 n-3) and docosahexa￾enoic acid (DHA, 22:6 n-3) from fish oil lipids, also known

as anti-inflammatory precursors, exert their antiatheroscle￾rotic effects by inhibiting the proliferation of SMC [7]. The

molecular mechanisms underlying these opposing effects of

n-3 and n-6 PUFAs are still not clear. MAPK plays a central

role by transducing extracellular signals, including growth

factors and cytokines [8]. Recent evidence indicates that

phosphatidylinositol 3-kinase (PI3-K) is involved in the

regulation of MAPK in various cell systems, including SMC

[9]. Studies have demonstrated that the classical Ras/

extracellular signal regulated kinase (ERK) pathway regu￾lates G1 progression by directly controlling cyclin D1

production via phosphorylation of various transcription

factors that bind to defined elements within the cyclin D1

promoter [10,11]. Sequential cyclin activation leads to the

progressive phosphorylation of the retinoblatoma protein

(Rb) that is essential for entry into the S-phase.

The transcription factor Egr-1 (early growth response

factor-1) also seems to be involved in the control of cell

proliferation initiated by the MAPK cascade [12,13].

Above-normal concentrations of Egr-1 are found in

atherosclerotic lesions and these are associated with

increased activity of the Egr-1 target genes implicated in

the proliferative and chemotactic responses of SMC to

injury [14]. EPA and DHA also seem to reduce the

Correspondence to M. Raymondjean, UMR 7079 Physiologie et

Physiopathologie, Universite´ Pierre et Marie Curie, Case Courrier

256, Baˆtiment A, 5e`me e´tage, 7 Quai Saint Bernard, 75252 Paris Cedex

5, France. Fax: +33 1 44 27 51 40, Tel.: +33 1 44 27 32 06,

E-mail: [email protected]

Abbreviations: AA, arachidonic acid; COX, cyclooxygenase; DHA,

docosahexaenoic acid; Egr-1, early growth response factor-1; EMSA,

electrophoretic mobility shift assay; EPA, eicosapentaenoic acid;

ERK, extracellular signal regulated kinase; IL-1b, interleukin-1b;

MAPK, mitogen-activated protein kinase; MEK, MAPK kinase;

NF-jB, nuclear factor-jB; PDGF, platelet-derived growth factor;

PI3-K, phosphatidylinositol 3-kinase; PGE2, prostaglandin E2;

PUFA, polyunsaturated fatty acid; Rb, retinoblastoma protein;

SMC, smooth muscle cells.

(Received 6 May 2004, revised 9 September 2004,

accepted 29 September 2004)

Eur. J. Biochem. 271, 4462–4473 (2004)
FEBS 2004 doi:10.1111/j.1432-1033.2004.04385.x