Cell and Tissue Research

, Volume 368, Issue 1, pp 145–157 | Cite as

The critical role of ABCG1 and PPARγ/LXRα signaling in TLR4 mediates inflammatory responses and lipid accumulation in vascular smooth muscle cells

  • Xiaojie Cao
  • Lili Zhang
  • Chunhai Chen
  • Qingsong Wang
  • Lu Guo
  • Qinlong Ma
  • Ping Deng
  • Gang Zhu
  • Binghu Li
  • Yan Pi
  • Chunyan Long
  • Lei Zhang
  • Zhengping Yu
  • Zhou Zhou
  • Jingcheng Li
Regular Article

Abstract

Toll-like receptor 4 (TLR4) plays critical roles in vascular inflammation, lipid accumulation and atherosclerosis development. However, the mechanisms underlying these processes are still not well established, especially in vascular smooth muscle cells (VSMCs). ATP-binding cassette transporter G1 (ABCG1) is one of the key genes mediating inflammation and cellular lipid accumulation. The function of TLR4 in regulating the expression of ABCG1 and the underlying molecular mechanisms remain to be elucidated. In this study, we cultured VSMCs from the thoracic aortas of mice and treated the cells with 50 μg/ml oxidized low-density lipoprotein (oxLDL) to activate TLR4 signaling. We observed that activating TLR4 with oxLDL induced inflammatory responses and lipid accumulation in VSMCs. The expression of peroxisome proliferator-activated receptor gamma (PPARγ), liver X receptor alpha (LXRα) and ABCG1 was inhibited by TLR4 activation. However, these effects could be reversed by knocking out TLR4. PPARγ activation by rosiglitazone rescued LXRα and ABCG1 expression and reduced TLR4-induced inflammation and lipid accumulation. Silencing PPARγ expression with a specific small interfering RNA (siRNA) inhibited LXRα and ABCG1 expression and, importantly, enhanced TLR4-induced inflammation and lipid accumulation. In conclusion, ABCG1 expression was down-regulated by TLR4, which induces inflammation and lipid accumulation in VSMCs via PPARγ/LXRα signaling. These findings indicate a novel molecular mechanism underlying TLR4-induced inflammation and lipid accumulation.

Keywords

Vascular smooth muscle cells Toll-like receptor 4 ATP-binding cassette transporter G1 Peroxisome proliferator-activated receptor gamma Lipid accumulation 

Abbreviations

ABCG1

ATP-binding cassette transporter G1

LXRα

Liver X receptor alpha

oxLDL

Oxidized low-density lipoprotein

PPARγ

Peroxisome proliferator-activated receptor gamma

RSG

Rosiglitazone

TLR4

Toll-like receptor 4

VSMCs

Vascular smooth muscle cells

Supplementary material

441_2016_2518_MOESM1_ESM.doc (34 kb)
Supplementary Table S1(DOC 34 kb)
441_2016_2518_Fig7_ESM.gif (671 kb)
Supplementary Figure 1

Identification of mice and primary VSMCs.a PCR identification of TLR4 KO mice. The 390-bp PCR products represent the WT TLR4 genotype, the 140-bp PCR products represent the mutant TLR4 genotype and both the 390-bp and 140-bp PCR products represent the heterozygous TLR4 genotype. b The first generation of cells came from thoracic aorta pieces cultured for 8 days and observed under an ordinary light microscope. Scale bar 200 μm. c Primary VSMC identification. Immunofluorescence images show the expression of α-SMA (green) and SM-22α (red) in VSMCs. Representative pictures of three independent experiments are shown. Scale bar 50 μm. (GIF 670 kb)

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High Resolution (TIF 16481 kb)
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Supplementary Figure 2

OxLDL stimulatedTLR4expression and promoted inflammatory responses, reversed byTLR4deaficiency. a The images show TLR4 mRNA levels, which were normalized to β-actin mRNA levels (n = 3). b Representative immunoblotting and statistics data revealed the TLR4 protein expression in WT VSMCs (n = 3). The mRNA levels of TNF-α, IL-6, MCP-1 and VCAM-1 were detected via real-time PCR (c, n = 4). The data are represented as the fold change relative to the control and are presented as the mean ± SEM. *P < 0.05, **P < 0.01 compared with the untreated control group. (GIF 134 kb)

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High Resolution (TIF 1818 kb)
441_2016_2518_Fig9_ESM.gif (47 kb)
Supplementary Figure 3

PPARγ-siRNA silencing efficiency. a The images show PPARγ mRNA levels, which were normalized to β-actin mRNA levels (n = 3). b Representative immunoblotting and statistics data revealed PPARγ expression in WT VSMCs (n = 3). The data are represented as the fold change relative to the control and are presented as the mean ± SEM. *P < 0.05, **P < 0.01 compared with the untreated control group. (GIF 46 kb)

441_2016_2518_MOESM4_ESM.tif (1.3 mb)
High Resolution (TIF 1310 kb)

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Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  • Xiaojie Cao
    • 1
    • 2
  • Lili Zhang
    • 1
  • Chunhai Chen
    • 3
  • Qingsong Wang
    • 2
  • Lu Guo
    • 1
  • Qinlong Ma
    • 3
  • Ping Deng
    • 3
  • Gang Zhu
    • 3
  • Binghu Li
    • 1
  • Yan Pi
    • 1
  • Chunyan Long
    • 1
  • Lei Zhang
    • 3
  • Zhengping Yu
    • 3
  • Zhou Zhou
    • 3
  • Jingcheng Li
    • 1
  1. 1.Department of Neurology, Institute of Surgery Research, Daping HospitalThird Military Medical UniversityChongqingPeople’s Republic of China
  2. 2.Department of NeurologyChengdu Military General HospitalChengduPeople’s Republic of China
  3. 3.Department of Occupational Health, Faculty of Preventive MedicineThird Military Medical UniversityChongqingPeople’s Republic of China

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