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Apolipoprotein E3 Inhibits Rho to Regulate the Mechanosensitive Expression of Cox2.

Hsu BY, Bae YH, Mui KL, Liu SL, Assoian RK - PLoS ONE (2015)

Bottom Line: However, apoE3 also has cholesterol-independent effects that contribute to its anti-atherogenic properties.Both of these cholesterol-independent effects result from an apoE3-mediated induction of cyclooxygenase-2 (Cox2).Nevertheless, how apoE3 regulates Cox2 remains unknown.

View Article: PubMed Central - PubMed

Affiliation: Program in Translational Biomechanics, Institute of Translational Medicine and Therapeutics, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America; Department of Systems Pharmacology and Translational Therapeutics, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.

ABSTRACT
Apolipoprotein E3 (apoE3) is thought to protect against atherosclerosis by enhancing reverse cholesterol transport. However, apoE3 also has cholesterol-independent effects that contribute to its anti-atherogenic properties. These include altering extracellular matrix protein synthesis and inhibiting vascular smooth muscle cell proliferation. Both of these cholesterol-independent effects result from an apoE3-mediated induction of cyclooxygenase-2 (Cox2). Nevertheless, how apoE3 regulates Cox2 remains unknown. Here, we show that apoE3 inhibits the activation of Rho, which reduces the formation of actin stress fibers and focal adhesions and results in cellular softening. Inhibition of Rho-Rho kinase signaling or direct cellular softening recapitulates the effect of apoE3 on Cox2 expression while a constitutively active Rho mutant overrides the apoE3 effect on both intracellular stiffness and Cox2. Thus, our results describe a previously unidentified mechanism by which an atheroprotective apolipoprotein uses Rho to control cellular mechanics and Cox2.

No MeSH data available.


Related in: MedlinePlus

Constitutively active Rho rescues actin stress fibers and paxillin in the presence of apoE3.(A) VSMCs infected with adenoviruses encoding LacZ or RhoV14 were incubated in 10% FBS with or without apoE3 for 24 hr. Cells were co-stained with phalloidin and anti-paxillin. Scale bar = 50 μm. The zooms show magnified views of paxillin staining in the boxed areas. The average phalloidin (B) and paxillin (C) intensity of single cells was analyzed using ImageJ and normalized to control cells. n = 3 independent experiments with at least 8 cells analyzed per experiment. Data information: Graphs show mean + SEM. *p<0.05 or **p<0.01.
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pone.0128974.g002: Constitutively active Rho rescues actin stress fibers and paxillin in the presence of apoE3.(A) VSMCs infected with adenoviruses encoding LacZ or RhoV14 were incubated in 10% FBS with or without apoE3 for 24 hr. Cells were co-stained with phalloidin and anti-paxillin. Scale bar = 50 μm. The zooms show magnified views of paxillin staining in the boxed areas. The average phalloidin (B) and paxillin (C) intensity of single cells was analyzed using ImageJ and normalized to control cells. n = 3 independent experiments with at least 8 cells analyzed per experiment. Data information: Graphs show mean + SEM. *p<0.05 or **p<0.01.

Mentions: The Rho-ROCK signaling pathway regulates acto-myosin-dependent contraction, intracellular tension, and the formation of actin stress fibers and associated focal adhesions [6–8]. Consistent with the inhibitory effect of apoE3 on Rho activity, f-actin stress fiber intensity (Figs 1C and 1D and 2A and 2B; LacZ) was reduced in VSMCs treated with apoE3. Focal adhesion abundance and areas were also reduced as determined by staining for either paxillin (Fig 2A and 2C; LacZ and S1A, S1B and S1C Fig) or vinculin (S1D and S1E Fig). The levels of total paxillin (S1F Fig) and vinculin (S1G Fig) were unchanged in apoE3-treated cells compared to controls. Thus, inhibition of intracellular stiffness by apoE3 affects the distribution rather than total quantity of focal adhesion proteins. These effects closely resembled those seen with dominant negative Rho (RhoN19, S2A Fig) or a selective ROCK inhibitor (Y27632, S2B Fig).


Apolipoprotein E3 Inhibits Rho to Regulate the Mechanosensitive Expression of Cox2.

Hsu BY, Bae YH, Mui KL, Liu SL, Assoian RK - PLoS ONE (2015)

Constitutively active Rho rescues actin stress fibers and paxillin in the presence of apoE3.(A) VSMCs infected with adenoviruses encoding LacZ or RhoV14 were incubated in 10% FBS with or without apoE3 for 24 hr. Cells were co-stained with phalloidin and anti-paxillin. Scale bar = 50 μm. The zooms show magnified views of paxillin staining in the boxed areas. The average phalloidin (B) and paxillin (C) intensity of single cells was analyzed using ImageJ and normalized to control cells. n = 3 independent experiments with at least 8 cells analyzed per experiment. Data information: Graphs show mean + SEM. *p<0.05 or **p<0.01.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4465925&req=5

pone.0128974.g002: Constitutively active Rho rescues actin stress fibers and paxillin in the presence of apoE3.(A) VSMCs infected with adenoviruses encoding LacZ or RhoV14 were incubated in 10% FBS with or without apoE3 for 24 hr. Cells were co-stained with phalloidin and anti-paxillin. Scale bar = 50 μm. The zooms show magnified views of paxillin staining in the boxed areas. The average phalloidin (B) and paxillin (C) intensity of single cells was analyzed using ImageJ and normalized to control cells. n = 3 independent experiments with at least 8 cells analyzed per experiment. Data information: Graphs show mean + SEM. *p<0.05 or **p<0.01.
Mentions: The Rho-ROCK signaling pathway regulates acto-myosin-dependent contraction, intracellular tension, and the formation of actin stress fibers and associated focal adhesions [6–8]. Consistent with the inhibitory effect of apoE3 on Rho activity, f-actin stress fiber intensity (Figs 1C and 1D and 2A and 2B; LacZ) was reduced in VSMCs treated with apoE3. Focal adhesion abundance and areas were also reduced as determined by staining for either paxillin (Fig 2A and 2C; LacZ and S1A, S1B and S1C Fig) or vinculin (S1D and S1E Fig). The levels of total paxillin (S1F Fig) and vinculin (S1G Fig) were unchanged in apoE3-treated cells compared to controls. Thus, inhibition of intracellular stiffness by apoE3 affects the distribution rather than total quantity of focal adhesion proteins. These effects closely resembled those seen with dominant negative Rho (RhoN19, S2A Fig) or a selective ROCK inhibitor (Y27632, S2B Fig).

Bottom Line: However, apoE3 also has cholesterol-independent effects that contribute to its anti-atherogenic properties.Both of these cholesterol-independent effects result from an apoE3-mediated induction of cyclooxygenase-2 (Cox2).Nevertheless, how apoE3 regulates Cox2 remains unknown.

View Article: PubMed Central - PubMed

Affiliation: Program in Translational Biomechanics, Institute of Translational Medicine and Therapeutics, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America; Department of Systems Pharmacology and Translational Therapeutics, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.

ABSTRACT
Apolipoprotein E3 (apoE3) is thought to protect against atherosclerosis by enhancing reverse cholesterol transport. However, apoE3 also has cholesterol-independent effects that contribute to its anti-atherogenic properties. These include altering extracellular matrix protein synthesis and inhibiting vascular smooth muscle cell proliferation. Both of these cholesterol-independent effects result from an apoE3-mediated induction of cyclooxygenase-2 (Cox2). Nevertheless, how apoE3 regulates Cox2 remains unknown. Here, we show that apoE3 inhibits the activation of Rho, which reduces the formation of actin stress fibers and focal adhesions and results in cellular softening. Inhibition of Rho-Rho kinase signaling or direct cellular softening recapitulates the effect of apoE3 on Cox2 expression while a constitutively active Rho mutant overrides the apoE3 effect on both intracellular stiffness and Cox2. Thus, our results describe a previously unidentified mechanism by which an atheroprotective apolipoprotein uses Rho to control cellular mechanics and Cox2.

No MeSH data available.


Related in: MedlinePlus