scholarly journals Synergistic induction of plasminogen activator inhibitor type-1 in HEP G2 cells by thrombin and transforming growth factor-beta

Blood ◽  
1992 ◽  
Vol 79 (1) ◽  
pp. 75-81 ◽  
Author(s):  
WE Hopkins ◽  
S Fujii ◽  
BE Sobel
Keyword(s):  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Plasminogen Activator Inhibitor ◽  
Hep G2 ◽  
Hep G2 Cells ◽  
G2 Cells ◽  
Inhibitor Type ◽  
Activator Inhibitor Type ◽  
Pai 1

Abstract Plasminogen activator inhibitor type-1 (PAI-1) is a physiologic modulator of the fibrinolytic system. Its activity in plasma increases in diverse thrombotic states. The large synthetic capacity of the liver make it a source of potentially large amounts of PAI-1. Because thrombin activity increases in association with thrombotic disorders and because specific binding sites for thrombin have been identified on hepatocytes, we characterized the effect of thrombin on hepatocyte PAI- 1 production. Incubation of Hep G2 cells with human alpha-thrombin resulted in a dose- and time-dependent increase in the concentration of PAI-1 in conditioned media. This effect was inhibited completely by hirudin and by antithrombin III. Steady-state levels of both the 3.2-kb and 2.2-kb forms of PAI-1 mRNA increased after stimulation of the cells with thrombin, indicating that thrombin influences PAI-1 expression in Hep G2 cells at the pretranslational level. Incubation of Hep G2 cells with alpha-thrombin and either platelet lysates or purified transforming growth factor-beta (TGF-beta), both previously shown to augment hepatocyte PAI-1 expression, resulted in a synergistic increase in the concentration of PAI-1 in conditioned media. PAI-1 mRNA appeared to be synergistically increased as well. Thus, thrombin increases expression of both PAI-1 protein and mRNA in Hep G2 cells and exerts synergistic effects with TGF-beta. These results underscore the potential importance of inhibition of thrombin under conditions in which thrombolysis is induced pharmacologically.

scholarly journals Synergistic induction of plasminogen activator inhibitor type-1 in HEP G2 cells by thrombin and transforming growth factor-beta

Blood ◽  
1992 ◽  
Vol 79 (1) ◽  
pp. 75-81
Author(s):  
WE Hopkins ◽  
S Fujii ◽  
BE Sobel
Keyword(s):  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Plasminogen Activator Inhibitor ◽  
Hep G2 ◽  
Hep G2 Cells ◽  
G2 Cells ◽  
Inhibitor Type ◽  
Activator Inhibitor Type ◽  
Pai 1

Plasminogen activator inhibitor type-1 (PAI-1) is a physiologic modulator of the fibrinolytic system. Its activity in plasma increases in diverse thrombotic states. The large synthetic capacity of the liver make it a source of potentially large amounts of PAI-1. Because thrombin activity increases in association with thrombotic disorders and because specific binding sites for thrombin have been identified on hepatocytes, we characterized the effect of thrombin on hepatocyte PAI- 1 production. Incubation of Hep G2 cells with human alpha-thrombin resulted in a dose- and time-dependent increase in the concentration of PAI-1 in conditioned media. This effect was inhibited completely by hirudin and by antithrombin III. Steady-state levels of both the 3.2-kb and 2.2-kb forms of PAI-1 mRNA increased after stimulation of the cells with thrombin, indicating that thrombin influences PAI-1 expression in Hep G2 cells at the pretranslational level. Incubation of Hep G2 cells with alpha-thrombin and either platelet lysates or purified transforming growth factor-beta (TGF-beta), both previously shown to augment hepatocyte PAI-1 expression, resulted in a synergistic increase in the concentration of PAI-1 in conditioned media. PAI-1 mRNA appeared to be synergistically increased as well. Thus, thrombin increases expression of both PAI-1 protein and mRNA in Hep G2 cells and exerts synergistic effects with TGF-beta. These results underscore the potential importance of inhibition of thrombin under conditions in which thrombolysis is induced pharmacologically.

Mediators of Induction of Augmented Expression of Plasminogen Activator Inhibitor Type-1 in Hep G2 Cells by Platelets

1991 ◽  
Vol 66 (02) ◽  
pp. 239-245 ◽  
Author(s):  
William E Hopkins ◽  
Donald R Westerhausen ◽  
Satoshi Fujii ◽  
Joseph J Billadello ◽  
Burton E Sobel
Keyword(s):  
Growth Factors ◽  
Growth Factor ◽  
Transforming Growth Factor ◽  
Plasminogen Activator Inhibitor ◽  
Hep G2 ◽  
Hep G2 Cells ◽  
G2 Cells ◽  
Inhibitor Type ◽  
Activator Inhibitor Type ◽  
Pai 1

SummaryPlasminogen activator inhibitor type-1 (PAI-1) is a physiologic modulator of the fibrinolytic system. We have shown previously that PAI-1 biosynthesis in cultured cells depends on several factors in serum. Because platelets are richly endowed with specific growth factors and because the release reaction is an integral part of thrombosis, the present study was performed to determine whether platelets augment PAI-1 production and if so, to define mediators responsible. Hep G2 cells were used to determine whether platelet lysates increased PAI-1 synthesis in a dose and time-dependent manner. In cells labeled metabolically with 35S-methionine for 6 h, an increase in labeled PAI-1 was elicited indicative of de novo synthesis as well as increased secretion of PAI-1 mediated by platelet lysates. Steady state levels of both the 3.2 and 2.2 kb forms of PAI-1 mRNA increased after 2 h and peaked in 3-5 h in a dose-dependent fashion as well. Incubation of Hep G2 cells with collagen activated platelets resulted in a similar induction of PAI-1 mRNA. The increase in PAI-1 mRNA occurred with exposure of the cells to platelet lysates for intervals as brief as 15 min and was not inhibited by cycloheximide indicating its independence of new protein synthesis. In order to identify the factors in platelets responsible for the induction of PAI-1 synthesis in the Hep G2 cell model system, neutralizing antibodies were used to inhibit specific platelet associated growth factors. Antibodies to transforming growth factor-β (TGF-β) and to the epidermal growth factor (EGF)/transforming growth factor alpha (TGF-α) receptor inhibited the platelet lysate-mediated increase in PAI-1 protein by 77%. The results indicate that physiologic concentrations of platelet derived TGF-β and EGF/TGF-α increase PAI-1 expression in Hep G2 cells and suggest that the augmentation of PAI-1 synthesis induced by platelets is mediated by specific platelet associated growth factors that may contribute to observed increases in circulating PAI-1 levels in patients with disorders manifested by thrombosis.

The Production of Heparin Cofactor li Is Not Regulated by Inflammatory Cytokines in Human Hepatoma Cells: Comparison with Plasminogen Activator Inhibitor Type-1

1996 ◽  
Vol 75 (02) ◽  
pp. 298-302 ◽  
Author(s):  
Chisato Kolke ◽  
Yumiko Hayakawa ◽  
Kenji Niiya ◽  
Nobuo Sakuragawa ◽  
Hideo Sasaki
Keyword(s):  
Plasminogen Activator Inhibitor ◽  
Mrna Levels ◽  
Hep G2 ◽  
Hep G2 Cells ◽  
Tnf Α ◽  
G2 Cells ◽  
Inhibitor Type ◽  
Activator Inhibitor Type ◽  
Activator Inhibitor

SummaryUsing the Northern blot technique, we screened 6 human hepatoma cell lines to investigate the regulation mechanism of heparin cofactor II (HCII) biosynthesis. We found that HuH-7 and Hep G2 cells constitutively expressed the HC II gene. In conditioned medium, HuH-7 cells constantly produced HC II that was functionally active and formed a complex with thrombin in the presence of dermatan sulfate. HC II is thought be an acute phase reactant, and, therefore, we examined the effects of the major inflammatory cytokines, IL-6, IL-1β, and TNF-α, on the regulation of HC II production in HuH-7 and Hep G2 cells. In HuH-7 cells, the antigen and mRNA levels of plasminogen activator inhibitor type-1 (PAI-1), an acute phase protein produced by hepa-tocytes, were increased in response to stimulation with either IL-6 or IL-1 (3 or both, but HC II antigen and mRNA levels were not changed by the same stimulation. Even when Hep G2 cells were treated with a combination of three cytokines, IL-6, IL-1β, and TNF-α, HC II antigen and mRNA levels were not changed; however, PAI-1 antigen and mRNA levels were clearly increased. These results suggest that the production of HC II in hepatoma cells is not regulated by the major inflammatory mediators, IL-6, IL-iβ, and TNF-α.

The Risk of Dialysis Access Thrombosis Is Related to Polymorphisms in the Transforming Growth Factor-β1 and Plasminogen Activator Inhibitor-Type 1 Genes.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1631-1631
Author(s):  
Alejandro Lazo-Langner ◽  
Greg A. Knoll ◽  
Philip S. Wells ◽  
Rachel M. Pilkey ◽  
Nancy Carson ◽  
...  
Keyword(s):  
Transforming Growth Factor ◽  
Plasminogen Activator Inhibitor ◽  
Transforming Growth Factor Β1 ◽  
Inhibitor Type ◽  
Activator Inhibitor Type ◽  
The Relationship ◽  
Activator Inhibitor ◽  
Tgf Β1 ◽  
Pai 1

Abstract Background. Transforming growth factor-β1 (TGF-β1) is involved in cell growth and differentiation and it plays an important role in the genesis of fibrosis through the stimulation of neointima proliferation and accumulation of components of the extracellular matrix and it has been suggested that polymorphisms (polym) in its gene contribute to determining the patency of the vascular access (VA) in patients (pts) on hemodialysis (HD) by contributing to both atherogenesis and VA thrombosis. On the other hand it has been demonstrated that polym in the gene encoding the plasminogen activator inhibitor type-1 (PAI-1) are a risk factor for ischemic heart disease and possibly stroke although their role in other vascular territories is unknown. It is also known that TGF-β1 is an important up-regulator of the PAI-1 gene. Methods. We conducted a case-control study to determine the relationship between TGF-β1 polym of the signal sequence (869 T>C; 915 G>C) and VA thrombosis in 416 HD pts. (107 with VA thrombosis, 309 controls had no thrombosis). We also explored for possible interactions with the 4G/5G polym of the PAI-1 gene. TGF-β1 and PAI-1 polym were amplified using PCR and genotyped using an ABI PRISM 3100 Genetic Analyzer. TGF-β1 producing haplotypes (haplo) were defined as low, intermediate or high as previously reported. All pts were also tested for thrombophilia. Statistical analysis was done using univariate and multivariate logistic regression adjusted for thrombophilia, age, access type, etc. Results. Frequencies for low, intermediate and high TGF-β1 producing haplo were 9.3, 26.2 and 64.5% in cases and 2.6, 22.3 and 75.1% in controls. Odds of thrombosis for TGF-β1 haplotypes Haplotype Crude OR (95% CI) p Adjusted OR (95% CI) p High producing haplotype is reference category Low 5.11 (1.93, 13.5) 0.001 7.31 (2.15, 24.88) 0.001 Intermediate 1.30 (0.74, 2.29) 0.36 1.39 (0.70, 2.75) 0.35 Figure Figure Frequencies for 5G/5G, 4G/5G and 4G/4G PAI-1 polym. were 21.5, 56.1 and 22.4% in cases and 25.6, 50.2 and 24.3% in controls respectively. When we explored the interaction between both gene polym we found a highly significant result for the interaction between the low TGF-β1 producers and the 4G/4G PAI-1 polym (adjusted OR 19.3; 95% CI 2.82, 132.40; p=0.003). Conclusions. Our results show that intermediate and high producing TGF-β1 haplo have a protective effect against VA thrombosis in HD patients that is not modified by PAI-1 polym and also suggest that the interaction between low TGF-β1 producing haplo and the 4G/4G PAI-1 polym might be an important contributor to thrombosis of the VA in HD pts. Further studies are ongoing to determine the relationship between TGF-β1 haplo, TGF-β1 level, and PAI-1 polym with thrombosis in this and other populations as well as to clarify the mechanisms underlying this apparently paradoxical effect.

The risk of dialysis access thrombosis is related to the transforming growth factor–β1 production haplotype and is modified by polymorphisms in the plasminogen activator inhibitor–type 1 gene

Blood ◽  
2006 ◽  
Vol 108 (13) ◽  
pp. 4052-4058 ◽  
Author(s):  
Alejandro Lazo-Langner ◽  
Greg A. Knoll ◽  
Philip S. Wells ◽  
Nancy Carson ◽  
Marc A. Rodger
Keyword(s):  
Growth Factor ◽  
Transforming Growth Factor ◽  
Plasminogen Activator Inhibitor ◽  
Transforming Growth Factor Β1 ◽  
Inhibitor Type ◽  
Activator Inhibitor Type ◽  
Activator Inhibitor ◽  
Tgf Β1 ◽  
Pai 1

Abstract Transforming growth factor–β1 (TGF-β1) and plasminogen activator inhibitor–type 1 (PAI-1) might play a role in the development of fibrosis and stenosis of hemodialysis vascular accesses. We studied polymorphisms in the TGFβ1 (869T>C; 915G>C), and PAI-1 (4G/5G) genes in 416 hemodialysis patients (107 access thrombosis cases, 309 controls), to determine if they are related to vascular access thrombosis. Three TGF-β1 production haplotypes (low, intermediate, and high) were defined according to the combination of polymorphisms found. The adjusted odds ratio (OR) and 95% confidence interval (CI) for access thrombosis in low TGF-β1 producers was 7.31 (2.15-24.88; P = .001). The interaction between low TGF-β1 production haplotype and the 4G/4G PAI-1 genotype was strongly associated with access thrombosis (adjusted OR 19.3; 95% CI 2.82-132.40; P = .003). Mean access thrombosis–free survival times in years (95% CI) were 14.65 (12.05-17.25), 11.96 (8.67-15.25), and 4.94 (3.06-6.83) in high, intermediate, and low TGF-β1 producers, respectively (P = .044). Analysis of the synergy index and the case-only cross-product supported the presence of an interaction. We concluded that low TGF-β1 production haplotype is a risk factor for hemodialysis access thrombosis and that in the presence of the 4G/4G PAI-1 genotype there is an additional increase in risk.

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