Modulation of thrombomodulin-dependent activation of human protein C through differential expression of endothelial Smads

Protein C is a plasma protease that in its active form plays a central role in the regulation of vascular function by modulating thrombosis, inflammation, and apoptosis. A central player in this pathway is the cytokine-regulated receptor thrombomodulin (TM), which functions as a co-factor for the th...

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Vydané v:The Journal of biological chemistry Ročník 277; číslo 51; s. 49815
Hlavní autori: Sandusky, George, Berg, David T, Richardson, Mark A, Myers, Laura, Grinnell, Brian W
Médium: Journal Article
Jazyk:English
Vydavateľské údaje: United States 20.12.2002
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ISSN:0021-9258
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Abstract Protein C is a plasma protease that in its active form plays a central role in the regulation of vascular function by modulating thrombosis, inflammation, and apoptosis. A central player in this pathway is the cytokine-regulated receptor thrombomodulin (TM), which functions as a co-factor for the thrombin-dependent generation of activated protein C. We have found that tumor necrosis factor-beta (TGF-beta)-dependent suppression of TM on endothelial cells is differentially regulated by endothelial Smad6s and Smad7. Overexpression of Smad6s resulted in activation of a TGF-beta reporter alone and enhanced TGF-beta response. Moreover, Smad6s overexpression suppressed TM and subsequently reduced activated protein C generation. Antisense inhibition of Smad6s expression enhanced the TM-dependent activation of protein C, whereas blocking the inhibitory Smad7 by antisense resulted in reduced TM-dependent activation of protein C. The effect of Smad6s appeared to be due, at least in part, to up-regulation of TGF-beta itself. Immunohistochemistry studies in normal versus atherosclerotic vessels showed that TM levels were suppressed in the endothelium over plaque. Consistent with the in vitro data, we found differential expression of Smad6s and Smad7 in normal versus atherosclerotic vessels, with Smad6s expression low in normal vessels but elevated in atherosclerotic vessels. In contrast, the opposite expression pattern was observed for Smad7. Overall, our results suggest that the relative balance of these intracellular Smads modulate the balance of endothelial function with regard to protein C activation.
AbstractList Protein C is a plasma protease that in its active form plays a central role in the regulation of vascular function by modulating thrombosis, inflammation, and apoptosis. A central player in this pathway is the cytokine-regulated receptor thrombomodulin (TM), which functions as a co-factor for the thrombin-dependent generation of activated protein C. We have found that tumor necrosis factor-beta (TGF-beta)-dependent suppression of TM on endothelial cells is differentially regulated by endothelial Smad6s and Smad7. Overexpression of Smad6s resulted in activation of a TGF-beta reporter alone and enhanced TGF-beta response. Moreover, Smad6s overexpression suppressed TM and subsequently reduced activated protein C generation. Antisense inhibition of Smad6s expression enhanced the TM-dependent activation of protein C, whereas blocking the inhibitory Smad7 by antisense resulted in reduced TM-dependent activation of protein C. The effect of Smad6s appeared to be due, at least in part, to up-regulation of TGF-beta itself. Immunohistochemistry studies in normal versus atherosclerotic vessels showed that TM levels were suppressed in the endothelium over plaque. Consistent with the in vitro data, we found differential expression of Smad6s and Smad7 in normal versus atherosclerotic vessels, with Smad6s expression low in normal vessels but elevated in atherosclerotic vessels. In contrast, the opposite expression pattern was observed for Smad7. Overall, our results suggest that the relative balance of these intracellular Smads modulate the balance of endothelial function with regard to protein C activation.Protein C is a plasma protease that in its active form plays a central role in the regulation of vascular function by modulating thrombosis, inflammation, and apoptosis. A central player in this pathway is the cytokine-regulated receptor thrombomodulin (TM), which functions as a co-factor for the thrombin-dependent generation of activated protein C. We have found that tumor necrosis factor-beta (TGF-beta)-dependent suppression of TM on endothelial cells is differentially regulated by endothelial Smad6s and Smad7. Overexpression of Smad6s resulted in activation of a TGF-beta reporter alone and enhanced TGF-beta response. Moreover, Smad6s overexpression suppressed TM and subsequently reduced activated protein C generation. Antisense inhibition of Smad6s expression enhanced the TM-dependent activation of protein C, whereas blocking the inhibitory Smad7 by antisense resulted in reduced TM-dependent activation of protein C. The effect of Smad6s appeared to be due, at least in part, to up-regulation of TGF-beta itself. Immunohistochemistry studies in normal versus atherosclerotic vessels showed that TM levels were suppressed in the endothelium over plaque. Consistent with the in vitro data, we found differential expression of Smad6s and Smad7 in normal versus atherosclerotic vessels, with Smad6s expression low in normal vessels but elevated in atherosclerotic vessels. In contrast, the opposite expression pattern was observed for Smad7. Overall, our results suggest that the relative balance of these intracellular Smads modulate the balance of endothelial function with regard to protein C activation.
Protein C is a plasma protease that in its active form plays a central role in the regulation of vascular function by modulating thrombosis, inflammation, and apoptosis. A central player in this pathway is the cytokine-regulated receptor thrombomodulin (TM), which functions as a co-factor for the thrombin-dependent generation of activated protein C. We have found that tumor necrosis factor-beta (TGF-beta)-dependent suppression of TM on endothelial cells is differentially regulated by endothelial Smad6s and Smad7. Overexpression of Smad6s resulted in activation of a TGF-beta reporter alone and enhanced TGF-beta response. Moreover, Smad6s overexpression suppressed TM and subsequently reduced activated protein C generation. Antisense inhibition of Smad6s expression enhanced the TM-dependent activation of protein C, whereas blocking the inhibitory Smad7 by antisense resulted in reduced TM-dependent activation of protein C. The effect of Smad6s appeared to be due, at least in part, to up-regulation of TGF-beta itself. Immunohistochemistry studies in normal versus atherosclerotic vessels showed that TM levels were suppressed in the endothelium over plaque. Consistent with the in vitro data, we found differential expression of Smad6s and Smad7 in normal versus atherosclerotic vessels, with Smad6s expression low in normal vessels but elevated in atherosclerotic vessels. In contrast, the opposite expression pattern was observed for Smad7. Overall, our results suggest that the relative balance of these intracellular Smads modulate the balance of endothelial function with regard to protein C activation.
Author Myers, Laura
Grinnell, Brian W
Sandusky, George
Berg, David T
Richardson, Mark A
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Snippet Protein C is a plasma protease that in its active form plays a central role in the regulation of vascular function by modulating thrombosis, inflammation, and...
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StartPage 49815
SubjectTerms Animals
Arteriosclerosis - pathology
Cell Survival
Chloramphenicol O-Acetyltransferase - metabolism
DNA-Binding Proteins - metabolism
Dose-Response Relationship, Drug
Down-Regulation
Endothelium, Vascular - cytology
Endothelium, Vascular - metabolism
Enzyme-Linked Immunosorbent Assay
Genes, Reporter
Humans
Immunohistochemistry
Lymphotoxin-alpha - metabolism
Oligonucleotides, Antisense - pharmacology
Protein C - metabolism
Signal Transduction
Smad6 Protein
Smad7 Protein
Thrombomodulin - metabolism
Time Factors
Trans-Activators - metabolism
Transcriptional Activation
Transfection
Transforming Growth Factor beta - metabolism
Up-Regulation
Xenopus
Xenopus Proteins
Title Modulation of thrombomodulin-dependent activation of human protein C through differential expression of endothelial Smads
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