Stimulation of tissue-type plasminogen activator gene expression by sodium butyrate and trichostatin A in human endothelial cells involves histone acetylation.
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ABSTRACT:
SUBMITTER: Arts J
PROVIDER: S-EPMC1135869 | biostudies-other | 1995 Aug
REPOSITORIES: biostudies-other
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Project description:AimsStimulated release of tissue-type plasminogen activator (t-PA) is pivotal for an intravascular fibrinolytic response and protects the circulation from occluding thrombosis. Hence, an impaired t-PA production is associated with increased risk for atherothrombotic events. A pharmacological means to stimulate the production of this enzyme may thus be desirable. We investigated if the anti-epileptic drug valproic acid (VPA) is capable of enhancing t-PA expression in vitro in vascular endothelial cells, and further examined if its histone deacetylase (HDAC)-inhibitory activity is of importance for regulating t-PA expression.Methods and resultsHuman endothelial cells were exposed to valproic acid and t-PA mRNA and protein levels were quantified. Potential changes in histone acetylation status globally and at the t-PA promoter were examined by western blot and chromatin immunoprecipitation. Valproic acid dose-dependently stimulated t-PA mRNA and protein expression in endothelial cells reaching a 2-4-fold increase at clinically relevant concentrations and 10-fold increase at maximal concentrations. Transcription profiling analysis revealed that t-PA is selectively targeted by this agent. Augmented histone acetylation was detected at the t-PA transcription start site, and an attenuated VPA-response was observed with siRNA knock of HDAC3, HDAC5 and HDAC7.ConclusionsValproic acid induces t-PA expression in cultured endothelial cells, and this is associated with increased histone acetylation at the t-PA promoter. Given the apparent potency of valproic acid in stimulating t-PA expression in vitro this substance may be a candidate for pharmacological modulation of endogenous fibrinolysis in man.
| S-EPMC3282745 | biostudies-literature
Project description:Incubation of cultured human endothelial cells with 5 mM-dibutyryl cyclic AMP led to an approx. 2-fold increase in tissue-type plasminogen-activator (t-PA) production over a 24 h incubation period. The stimulating effect of dibutyryl cyclic AMP could be explained by the slow liberation of butyrate, as the effect could be reproduced by addition of free butyrate to the medium, but not by addition of 8-bromo cyclic AMP or forskolin, agents known to raise intracellular cyclic AMP levels. With butyrate, an accelerated accumulation of t-PA antigen in the conditioned medium (CM) was observed after a lag period of about 6 h. Increasing amounts of butyrate caused an increasingly stimulatory effect, reaching a plateau at 5 mM-butyrate. The relative enhancement of t-PA production in the presence of 5 mM-butyrate varied among different endothelial cell cultures from 6- to 25-fold in 24 h CM. Such an increase in t-PA production was observed with both arterial and venous endothelial cells. The butyrate-induced increases in t-PA production were accompanied by increased t-PA mRNA levels. Analysis of radiolabelled CM and cell extracts by SDS/polyacrylamide-gel electrophoresis indicated that the potent action of butyrate is probably restricted to a small number of proteins. The accumulation of plasminogen activator inhibitor type 1 (PAI-1) in CM from butyrate-treated cells varied only moderately. In our study of the relationship between structure and stimulatory activity, we found that a straight-chain C4 monocarboxylate structure with a methyl group at one end and a carboxy moiety at the other seems to be required for the optimal induction of t-PA in cultured endothelial cells.
| S-EPMC1148455 | biostudies-other
Project description:Establishing an effective method to improve stem cell differentiation is crucial in stem cell transplantation. Here we aimed to explore whether and how sodium butyrate (NaB) induces rat bone marrow mesenchymal stem cells (MSCs) to differentiate into bladder smooth muscle cells (SMCs). We found that NaB significantly suppressed MSC proliferation and promoted MSCs differentiation into SMCs, as evidenced by the enhanced expression of SMC specific genes in the MSCs. Co-culturing the MSCs with SMCs in a transwell system promoted the differentiation of MSCs into SMCs. NaB again promoted MSC differentiation in this system. Furthermore, NaB enhanced the acetylation of SMC gene-associated H3K9 and H4, and decreased the expression of HDAC2 and down-regulated the recruitment of HDAC2 to the promoter regions of SMC specific genes. Finally, we found that NaB significantly promoted MSC depolarization and increased the intracellular calcium level of MSCs upon carbachol stimulation. These results demonstrated that NaB effectively promotes MSC differentiation into SMCs, possibly by the marked inhibition of HDAC2 expression and disassociation of HDAC2 recruitment to SMC specific genes in MSCs, which further induces high levels of H3K9ace and H4ace and the enhanced expression of target genes, and this strategy could potentially be applied in clinical tissue engineering and cell transplantation.
| S-EPMC4280132 | biostudies-literature
Project description:BACKGROUND:In practical production, dairy cows are frequently exposed to bacterial endotoxin (lipopolysaccharide, LPS) when they are subjected to high-concentrate diets, poor hygienic environments, as well as mastitis and metritis. Histone acetylation is an important epigenetic control of DNA transcription and a higher histone acetylation is associated with facilitated transcription. LPS might reduce histone acetylation in the mammary epithelial cells, resulting in lower transcription and mRNA expression of lactation-related genes. This study was conducted to investigate the effect of LPS on histone acetylation in bovine mammary epithelial cells and the efficacy of sodium butyrate (SB) in suppressing the endotoxin-induced adverse effect. Firstly, the bovine mammary epithelial cell line MAC-T cells were treated for 48?h with LPS at different doses of 0, 1, 10, 100, and 1000 endotoxin units (EU)/mL (1 EU?=?0.1?ng), and the acetylation levels of histones H3 and H4 as well as the histone deacetylase (HDAC) activity were measured. Secondly, the MAC-T cells were treated for 48?h as follows: control, LPS (100 EU/mL), and LPS (100 EU/mL) plus SB (10?mmol/L), and the acetylation levels of histones H3 and H4 as well as milk gene mRNA expressions were determined. RESULTS:The results showed that HDAC activity increased linearly with increasing LPS doses (P?<?0.01). The histone H3 acetylation levels were significantly reduced by LPS, while the histone H4 acetylation levels were not affected by LPS (P?>?0.05). Sodium butyrate, an inhibitor of HDAC, effectively suppressed the endotoxin-induced decline of histone H3 acetylation (P?<?0.05). As a result, SB significantly enhanced the mRNA expression of lactation-related genes (P?<?0.05). CONCLUSIONS:The results suggest one of the adverse effects of LPS on the lactation of bovine mammary gland epithelial cells was due to decreasing histone H3 acetylation through increasing HDAC activity, whereas the endotoxin-induced adverse effects were effectively suppressed by SB.
| S-EPMC6664593 | biostudies-literature
Project description:Plasminogen activator inhibitor-1 (PAI-1) paradoxically enhances tumor progression and angiogenesis; however, the mechanism supporting this role is not known. Here we provide evidence that PAI-1 is essential to protect endothelial cells (ECs) from FasL-mediated apoptosis. In the absence of host-derived PAI-1, human neuroblastoma cells implanted in PAI-1-deficient mice form smaller and poorly vascularized tumors containing an increased number of apoptotic ECs. We observed that knockdown of PAI-1 in ECs enhances cell-associated plasmin activity and increases spontaneous apoptosis in vitro. We further demonstrate that plasmin cleaves FasL at Arg144-Lys145, releasing a soluble proapoptotic FasL fragment from the surface of ECs. The data provide a mechanism explaining the proangiogenic activity of PAI-1.
| S-EPMC2630529 | biostudies-literature
Project description:UnlabelledThe expression of the tissue plasminogen activator (t-PA) gene appears to be under epigenetic control and can be affected by histone deacetylation inhibition. The study aimed to test if histone deacetalyase inhibitor treatment lead to increased t-PA release or reduced exhaustion in t-PA release in response to stimulation, as well as change in plasminogen activator inhibitor-1 (PAI-1) in subjects with coronary disease. In this clinical study, 16 post-myocardial infarction subjects, the perfused forearm model was used with isoprenaline provocation during 20 minutes, to stimulate local t-PA release. Each subject was measured twice on the same day (repeated stimuli sequences) as well as on two different occasions, without treatment and after four weeks of treatment with valproic acid (500 mg, twice daily). Net forearm release for t-PA in response to isoprenaline at minutes 1.5, 3, 6, 9, 12, 15 and 18 was measured, allowing assessment of cumulative t-PA release. There was a reduction in the exhaustion of cumulative t-PA release during repeated and prolonged stimulation with valproic acid treatment compared to non-treatment. Plasma PAI-1 antigen was decreased following treatment compared to non-treatment (18.4 ± 10.0 vs. 11.0 ± 7.1 nanograms/ml respectively, mean with 95% confidence interval). These findings demonstrate that histone deacetylation inhibition increases the capacity for endogenous t-PA release in subjects with vascular disease. Furthermore, the fibrinolytic balance is favored with suppressed PAI-1 levels. More studies are needed to establish the clinical relevance of these findings.Trial registrationEU Clinical Trials Register 2012-004950-27.
| S-EPMC4373842 | biostudies-literature
Project description:Widespread changes in gene expression drive tumorigenesis, yet our knowledge of how aberrant epigenomic and transcriptome profiles arise in cancer cells is poorly understood. Here, we demonstrate that metabolic transformation plays an important role. Butyrate is the primary energy source of normal colonocytes and is metabolized to acetyl-CoA, which was shown to be important not only for energetics but also for HAT activity. Due to the Warburg effect, cancerous colonocytes rely on glucose as their primary energy source, so butyrate accumulated and functioned as an HDAC inhibitor. Although both mechanisms increased histone acetylation, different target genes were upregulated. Consequently, butyrate stimulated the proliferation of normal colonocytes and cancerous colonocytes when the Warburg effect was prevented from occurring, whereas it inhibited the proliferation of cancerous colonocytes undergoing the Warburg effect. These findings link a common metabolite to epigenetic mechanisms that are differentially utilized by normal and cancerous cells because of their inherent metabolic differences.
| S-EPMC3513569 | biostudies-literature
Project description:Eukaryotic gene expression requires the coordination of multiple factors to overcome the repressive nature of chromatin. However, the mechanistic details of this coordination are not well understood. The SAGA family of transcriptional coactivators interacts with DNA-binding activators to establish regions of hyperacetylation. We have previously shown that, contrary to the prevailing model in which activator protein increases SAGA affinity for nucleosome substrate, the Gal4-VP16 activator model system augments the rate of acetylation turnover for the SAGA complex from budding yeast. To better understand how this stimulation occurs, we have identified necessary components using both kinetics assays and binding interactions studies. We find that Gal4-VP16-mediated stimulation requires activator binding to DNA flanking the nucleosome, as it cannot be reproduced in trans by activator protein alone or by exogenous DNA containing the activator binding site in combination with the activator protein. Further, activator-mediated stimulation requires subunits outside of the histone acetylation (HAT) module, with the Tra1 subunit being responsible for the majority of the stimulation. Interestingly, for the HAT module alone, nucleosome acetylation is inhibited by activator proteins due to non-specific binding of the activator to the nucleosomes. This inhibition is not observed for the yeast ADA complex, a small complex comprised mostly of the HAT module, suggesting that subunits outside of the HAT module in both it and SAGA can overcome non-specific activator binding to nucleosomes. However, this activity appears distinct from activator-mediated stimulation, as ADA complex acetylation is not stimulated by Gal4-VP16. Graphical abstract Image 1 Highlights • Stimulation of nucleosome acetylation by SAGA requires activator in cis• Tra1 mediates the majority of activator stimulation• The core HAT complex of SAGA is inhibited by activator due to non-specific binding• The related ADA complex is neither stimulated nor inhibited by activator
| S-EPMC7786029 | biostudies-literature
Project description:Therapeutic agents are urgently needed for treating metastatic castration-refractory prostate cancer (mCRPC) that is unresponsive to androgen deprivation and chemotherapy. Our screening assays demonstrated that chemotherapy-resistant prostate cancer (PCa) cells are more sensitive to HDAC inhibitors than paired sensitive PCa cells, as demonstrated by cell proliferation and apoptosis in vitro and in vivo. Kinetic study revealed that TSA-induced apoptosis was significantly dependent on enhanced transcription and protein synthesis in an early stage, which subsequently caused ER stress and apoptosis. ChIP analysis indicated that TSA increased H4K16 acetylation, promoting ER stress gene transcription. The changes in Ac-H4K16, ATF3 and ATF4 were also validated in TSA-treated animals. Further study revealed the higher enzyme activity of HDACs and an increase in acetylated proteins in resistant cells. The higher nucleocytoplasmic acetyl-CoA in resistant cells was responsible for elevated acetylation status of protein and a more vigorous growth state. These results strongly support the pre-clinical application of HDAC inhibitors for treating chemotherapy-resistant mCRPC.
| S-EPMC5824406 | biostudies-literature
Project description:The experiments described in this paper were designed to examine the specific binding of tissue plasminogen activator (tPA) to cultured human aortic endothelial (HAE) cells. When 125I-labelled tPA was incubated with the cells at 4 degrees C, binding was found to plateau within 90 min after incubations were begun. Binding was saturable and the bound enzyme dissociated from the sites with a half-time of approx. 48 min. Scatchard analyses were performed using tPA molecules isolated from human melanoma and colon cells as well as from C127 and Chinese hamster ovary cells that had been transfected with the human tPA gene. These enzymes showed very similar binding characteristics in spite of the fact that they differ substantially in the types of sugars which comprise their side chains. Neither the chainedness of the molecules (one-chain or two-chain) nor the sites at which they are glycosylated (type I or type II) appear to affect their ability to interact with binding sites. The tPA molecules were found to have an average equilibrium dissociation constant of (1.15 +/- 0.10) x 10(-9) M and HAE cells appeared to have a single, homogeneous population of independent binding sites present at a concentration of (1.57 +/- 0.13) x 10(6) sites per cell. Lowering the pH of the binding buffer from 7.4 to 6.5 resulted in a reversible increase in specific binding of between 2-fold and 7-fold depending upon the particular preparation of cells. Preincubation of tPA with plasminogen activator inhibitor 1 (PAI-1) was found to have little effect on binding, suggesting that tPA interacts at sites distinct from surface-bound PAI-1. No evidence for either internalization or degradation of tPA was observed in assays run at 37 degrees C. This suggests that, like urokinase, tPA remains on cell surfaces for an extended period of time.
| S-EPMC1131601 | biostudies-other