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Oxidized LDL activates PAI-1 transcription through autocrine activation of TGF-beta signaling in mesangial cells.
Kidney Int. 2005 May; 67(5):1743-52.KI

Abstract

BACKGROUND

Lipid abnormalities and oxidative stress may be involved in the development of glomerulosclerosis. Plasminogen activator inhibitor-1 (PAI-1) is a component of extracellular matrix (ECM) and target gene of transforming growth factor-beta (TGF-beta). Smad proteins play a key role in TGF-beta signaling, and Smad binding CAGA boxes are present in the PAI-1 promoter. This study examined whether oxidized low-density lipoprotein (Ox-LDL) activates PAI-1 transcription in human mesangial cells, mediated by increased Smad/DNA interactions.

METHODS

Quiescent HMC were incubated with 50 microg/mL of Cu(++)-catalyzed Ox-LDL for 15 minutes to 4 hours, and the effects of Ox-LDL on TGF-beta1 and PAI-1 mRNA expression, PAI-1 promoter activity, and DNA binding activity of Smad proteins were examined.

RESULTS

Ox-LDL induced TGF-beta1 and PAI-1 mRNA expression. Ox-LDL increased the transiently transfected PAI-1 promoter activity as compared with controls to 3.9-fold. Ox-LDL-treated cells increased Smad3 protein levels two times the control levels in the nuclei. Electrophoretic mobility shift assay (EMSA) performed using a CAGA sequence probe and nuclear extracts showed that Ox-LDL increased DNA/protein complexes. When nuclear extracts were preincubated with 100 molar excess of unlabeled CAGA oligonucleotide or SB-431542, an inhibitor of the TGF-beta type I receptor, the formation of complex was prevented. The DNA binding protein was shown to be Smad3 by antibody supershift. Transfection of phosphorothioate CAGA oligonucleotides, which compete with the CAGA-containing PAI-1 promoter for Smad3 binding, inhibited the Ox-LDL-induced PAI-1 mRNA expression. Cotransfection of phosphorothioate CAGA oligonucleotides with PAI-1 reporter vector also blocked the Ox-LDL-induced PAI-1 promoter activity.

CONCLUSION

These results suggest that Ox-LDL activates TGF-beta/Smad signaling to stimulate PAI-1 transcription in human mesangial cells. Thus, progression of glomerular disease may be promoted by PAI-1 up-regulation in human mesangial cells mediated by the Ox-LDL-induced TGF-beta/Smad signaling pathways.

Authors+Show Affiliations

Department of Pathology, Seoul National University College of Medicine, Seoul, Korea.No affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

15840021

Citation

Song, Chi Young, et al. "Oxidized LDL Activates PAI-1 Transcription Through Autocrine Activation of TGF-beta Signaling in Mesangial Cells." Kidney International, vol. 67, no. 5, 2005, pp. 1743-52.
Song CY, Kim BC, Hong HK, et al. Oxidized LDL activates PAI-1 transcription through autocrine activation of TGF-beta signaling in mesangial cells. Kidney Int. 2005;67(5):1743-52.
Song, C. Y., Kim, B. C., Hong, H. K., & Lee, H. S. (2005). Oxidized LDL activates PAI-1 transcription through autocrine activation of TGF-beta signaling in mesangial cells. Kidney International, 67(5), 1743-52.
Song CY, et al. Oxidized LDL Activates PAI-1 Transcription Through Autocrine Activation of TGF-beta Signaling in Mesangial Cells. Kidney Int. 2005;67(5):1743-52. PubMed PMID: 15840021.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Oxidized LDL activates PAI-1 transcription through autocrine activation of TGF-beta signaling in mesangial cells. AU - Song,Chi Young, AU - Kim,Bong Cho, AU - Hong,Hye Kyoung, AU - Lee,Hyun Soon, PY - 2005/4/21/pubmed PY - 2005/8/23/medline PY - 2005/4/21/entrez SP - 1743 EP - 52 JF - Kidney international JO - Kidney Int VL - 67 IS - 5 N2 - BACKGROUND: Lipid abnormalities and oxidative stress may be involved in the development of glomerulosclerosis. Plasminogen activator inhibitor-1 (PAI-1) is a component of extracellular matrix (ECM) and target gene of transforming growth factor-beta (TGF-beta). Smad proteins play a key role in TGF-beta signaling, and Smad binding CAGA boxes are present in the PAI-1 promoter. This study examined whether oxidized low-density lipoprotein (Ox-LDL) activates PAI-1 transcription in human mesangial cells, mediated by increased Smad/DNA interactions. METHODS: Quiescent HMC were incubated with 50 microg/mL of Cu(++)-catalyzed Ox-LDL for 15 minutes to 4 hours, and the effects of Ox-LDL on TGF-beta1 and PAI-1 mRNA expression, PAI-1 promoter activity, and DNA binding activity of Smad proteins were examined. RESULTS: Ox-LDL induced TGF-beta1 and PAI-1 mRNA expression. Ox-LDL increased the transiently transfected PAI-1 promoter activity as compared with controls to 3.9-fold. Ox-LDL-treated cells increased Smad3 protein levels two times the control levels in the nuclei. Electrophoretic mobility shift assay (EMSA) performed using a CAGA sequence probe and nuclear extracts showed that Ox-LDL increased DNA/protein complexes. When nuclear extracts were preincubated with 100 molar excess of unlabeled CAGA oligonucleotide or SB-431542, an inhibitor of the TGF-beta type I receptor, the formation of complex was prevented. The DNA binding protein was shown to be Smad3 by antibody supershift. Transfection of phosphorothioate CAGA oligonucleotides, which compete with the CAGA-containing PAI-1 promoter for Smad3 binding, inhibited the Ox-LDL-induced PAI-1 mRNA expression. Cotransfection of phosphorothioate CAGA oligonucleotides with PAI-1 reporter vector also blocked the Ox-LDL-induced PAI-1 promoter activity. CONCLUSION: These results suggest that Ox-LDL activates TGF-beta/Smad signaling to stimulate PAI-1 transcription in human mesangial cells. Thus, progression of glomerular disease may be promoted by PAI-1 up-regulation in human mesangial cells mediated by the Ox-LDL-induced TGF-beta/Smad signaling pathways. SN - 0085-2538 UR - https://www.unboundmedicine.com/medline/citation/15840021/Oxidized_LDL_activates_PAI_1_transcription_through_autocrine_activation_of_TGF_beta_signaling_in_mesangial_cells_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0085-2538(15)50652-2 DB - PRIME DP - Unbound Medicine ER -