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Oxidized phospholipids in oxidized low-density lipoprotein down-regulate thrombomodulin transcription in vascular

Hidemi Ishii1, Tsuyoshi Tezuka, Hiroyuki Ishikawa

  • 1Department of Public Health, Showa Pharmaceutical University, Machida, Tokyo, Japan. h-ishii@ac.shoyaku.ac.jp

Blood
|February 11, 2003
PubMed

Insights

Oxidized phospholipids in oxidized low-density lipoprotein (ox-LDL) reduce thrombomodulin (TM) gene transcription in endothelial cells. This occurs by inhibiting key protein binding to the TM promoter, decreasing TM expression.

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Endothelial Cell Biology

Background:

  • Thrombomodulin (TM) is an anticoagulant glycoprotein crucial for vascular health.
  • Oxidized low-density lipoprotein (ox-LDL) is implicated in atherosclerosis and endothelial dysfunction.
  • The precise mechanisms by which ox-LDL affects TM expression are not fully understood.

Purpose of the Study:

  • To investigate the molecular mechanisms underlying TM gene down-regulation by ox-LDL in human umbilical vein endothelial cells (HUVECs).
  • To identify the specific components of ox-LDL responsible for TM down-regulation.
  • To elucidate the role of transcription factors and promoter elements in this process.

Main Methods:

  • Cultured HUVECs were treated with lipid extracts from ox-LDL and specific oxidized phospholipids like oxidized 1-palmitoyl-2-arachidonyl-sn-glycero-3-phosphocholine (ox-PAPC).
  • TM mRNA levels and stability were assessed.
  • Nuclear protein binding activity to TM promoter sequences (DR4, Sp1) was measured using electrophoretic mobility shift assays.
  • Reporter gene assays were performed with TM promoter constructs.
  • Expression levels of key transcription factors (RARbeta, RXRalpha, Sp1, Sp3) were analyzed.

Main Results:

  • Phospholipid components of ox-LDL, particularly ox-PAPC, significantly reduced TM mRNA levels in HUVECs.
  • TM mRNA stability was not affected, indicating transcriptional regulation.
  • Ox-LDL and ox-PAPC treatment decreased the binding of nuclear proteins to the DR4 and Sp1 elements in the TM promoter.
  • This decrease correlated with reduced expression of RARbeta, RXRalpha, Sp1, and Sp3.
  • Reporter assays confirmed the importance of DR4 and Sp1 binding sites for TM promoter activity.

Conclusions:

  • Oxidized phospholipids within ox-LDL inhibit TM gene transcription in HUVECs.
  • This inhibition is mediated by reduced binding of RARbeta-RXRalpha heterodimers and Sp proteins (Sp1, Sp3) to the TM promoter.
  • Reduced expression of these transcription factors contributes to the down-regulation of TM expression, potentially impacting endothelial anticoagulant function.

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