Diabetic LDL inhibits cell-cycle progression via STAT5B and p21waf
Maria Felice Brizzi, … , Gianfranco Pagano, Luigi Pegoraro
Maria Felice Brizzi, … , Gianfranco Pagano, Luigi Pegoraro
Published January 1, 2002
Citation Information: J Clin Invest. 2002;109(1):111-119. https://doi.org/10.1172/JCI13617.
View: Text | PDF
Article Article has an altmetric score of 4

Diabetic LDL inhibits cell-cycle progression via STAT5B and p21waf

Abstract

Modified LDL is a major cause of injury to the endothelium in diabetes. In the present study, we analyzed the effects on endothelial cells of LDL recovered from type 2 diabetic patients (dm-LDL) or from nondiabetic subjects (n-LDL). Treatment of human umbilical vein endothelial cells with dm-LDL, but not n-LDL, led to the accumulation of cells in G1. To dissect the molecular mechanisms of this effect, we analyzed the expression and function of the cyclin-dependent kinase inhibitor p21waf, a cell cycle regulator known to be a target of the signal transducers and activators of transcription (STATs). dm-LDL led to transient STAT5 phosphorylation and the formation of a STAT5-containing complex and activated p21waf expression at the transcriptional level. Expression of the dominant-negative form of STAT5B, but not of STAT5A, significantly decreased both p21waf expression and the fraction of cells in G1. Finally, immunofluorescence analysis demonstrated that activated STAT5 is expressed in newly formed intraplaque vessels and in endothelial cells lining the luminal side of the plaque. Similarly, p21waf immunoreactivity was found in the neointimal vasculature. Our results suggest a role of STAT5B as a regulator of gene expression in diabetes-associated vascular disease.

Authors

Maria Felice Brizzi, Patrizia Dentelli, Marzia Pavan, Arturo Rosso, Roberto Gambino, Maria Grazia De Cesaris, Giovanni Garbarino, Giovanni Camussi, Gianfranco Pagano, Luigi Pegoraro

×

Figure 5

Options: View larger image (or click on image) Download as PowerPoint
STAT5 phosphorylation and p21SIE2 binding activity are triggered by dm-L...
STAT5 phosphorylation and p21SIE2 binding activity are triggered by dm-LDL but not ox-LDL. (a) Kinetics of STAT5 phosphorylation. Starved HUVECs were stimulated with dm-LDL or n-LDL for the indicated times. After undergoing SDS-PAGE, lysed proteins were electrophoretically transferred to a nitrocellulose filter, immunoblotted, and photographed. (b) ox-LDL does not activate STAT5. HUVECs processed as above were stimulated with dm-LDL (15 minutes) or ox-LDL for the indicated times. Proteins from lysed HUVECs were processed as above. The filters were immunoblotted with anti–phospho-STAT5 antibody and reprobed with anti-STAT5 antiserum. Four individual experiments were performed with similar results. Unphosphorylated and phosphorylated STAT5 are indicated. (c) p21SIE2 complex formation. Nuclear extracts were prepared from HUVECs stimulated for 30 minutes with n-LDL or dm-LDL. Competition was performed by addition of a 50-fold excess of unlabeled p21SIE2 probe (competitor). The DNA-protein complexes were resolved by nondenaturing PAGE. (d) The p21SIE2-binding complex is antigenically related to STAT5 but not to STAT3. Nuclear extracts from dm-LDL–treated HUVECs were preincubated with an anti-STAT5 or anti-STAT3 antibody as indicated, and the DNA-protein complexes were resolved. Arrows indicate the p21SIE2 complex (lower arrow) and the supershifted species (upper arrow).