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Abstract Chromatin remodeling enzymes are important "writers'', "readers'' and "erasers'' of the epigenetic code. These proteins are responsible for the placement, recognition, and removal of molecular marks in histone tails that trigger structural and functional changes in chromatin. This is also the case for histone deacetylases (HDACs), i.e., enzymes that remove acetyl groups from histone tails, signaling heterochromatin formation. Chromatin remodeling is necessary for cell differentiation processes in eukaryotes, and fungal pathogenesis in plants includes many adaptations to cause disease. Macrophomina phaseolina (Tassi) Goid. is a nonspe-cific, necrotrophic ascomycete phytopathogen that causes charcoal root disease. M. phaseolina is a frequent and highly destructive pathogen in crops such as common beans (Phaseolus vulgaris L.), particularly under both water and high temperature stresses. Here, we evaluated the effects of the classical HDAC inhibitor trichostatin A (TSA) on M. phaseolina in vitro growth and virulence. During inhibition assays, the growth of M. phaseolina in solid media, as well as the size of the microsclerotia, were reduced (p <0.05), and the colony morphology was remark-ably affected. Under greenhouse experiments, treatment with TSA reduced (p <0.05) fungal virulence in common bean cv. BAT 477. Tests of LIPK, MAC1 and PMK1 gene expression during the interaction of fungi with BAT 477 revealed noticeable deregulation. Our results provide additional evidence about the role of HATs and HDACs in important biological processes of M. phaseolina.
Resumen Las enzimas remodeladoras de la cromatina son «escritores¼, «lectores¼ y «borradores¼ importantes del código epigenético. Estas proteínas son responsables de la localización, el reconocimiento y la remoción de las marcas moleculares sobre las terminaciones de las histonas que desencadenan cambios funcionales y estructurales en la cromatina. Es el caso de las desacetilasas de histonas (HDAC), enzimas que remueven grupos acetilo de las «colas¼ de las histonas, señalizando la formación de heterocromatina. La anterior es una actividad necesaria en los procesos de diferenciación celular de los eucariotas, y se conoce que la patogénesis fúngica en las plantas requiere de adaptaciones diversas para ocasionar enfermedad. Macrophomina phaseolina (Tassi) Goid. es un ascomiceto fitopatógeno, necrótrofo e inespecífico, causante de la pudrición carbonosa. Este es un hongo frecuente y altamente destructivo en cultivos como fríjol común (Phaseolus vulgaris L.), particularmente bajo estrés hídrico y térmico. En este trabajo evaluamos los efectos del inhibidor de HDAC clásicas tricostatina A (TSA) sobre el crecimiento in vitro y la virulencia de M. phaseolina. El TSA redujo el crecimiento de M. phaseolina en medio sólido y el tamano de los microesclerocios (p < 0,05), lo que afectó la morfología colonial. En invernadero, el tratamiento con TSA disminuyó (p<0,05) la gravedad de la infección en la variedad de frijol BAT 477. La expresión de los genes de patogenicidad LIPK, MAC1 y PMK1 durante la interacción del hongo con la planta reveló una desregulación importante. Estos resultados proporcionan evidencia adicional del papel que cumplen las HDAC en la regulación de procesos biológicos fundamentales de M. phaseolina. © 2023 Asociación Argentina de Microbiología. Publicado por Elsevier Espana, S.L.U.
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Krüppel-like transcription factor 2 (KLF2) plays a key regulatory role in endothelial inflammation, thrombosis, angiogenesis and macrophage inflammation and polarization, and up-regulation of KLF2 expression has the potential to prevent and treatment atherosclerosis. In this study, trichostatin C (TSC) was obtained from the secondary metabolites of rice fermentation of Streptomyces sp. CPCC 203909 as a KLF2 up-regulator by using a high throughput screening model based on a KLF2 promoter luciferase reporter assay. TSC significantly inhibited the adhesion of tumor necrosis factor-α (TNFα) induced monocytes (THP-1) to human umbilical vein endothelial cells (HUVECs). Western blot results showed that TSC decreased TNFα induced the protein expression increase of vascular cell adhesion molecule-1 (VCAM-1), and thereby inhibited endothelial inflammation. The results of histone deacetylase (HDAC) overexpression and molecular docking experiments showed that TSC upregulated the expression of KLF2 by inhibiting subtypes of HDAC 4/5/7. In conclusion, this study suggests that TSC up-regulates the expression of KLF2 through inhibiting HDAC 4/5/7 and thus inhibits TNFα induced endothelial inflammation, and it has the potential to prevent and treat atherosclerosis.
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Objective:To investigate the effect of histone deacetylase inhibitor trichostatin A(TSA) on the lipopolysaccharide(LPS)-induced injury and apoptosis of human microvascular endothelial cell(HMEC).Methods:HMECs were used as research cells to establish LPS-induced septic cell model, which were divided into three groups according to different treatments: control group (150 μL of phosphate buffer), LPS group (150 μL of 5 μg/mL LPS), LPS+ TSA group (150 μL of 5 μg/mL LPS and 500 μg/L TSA). After cells of each group were cultured for 24 h and 48 h, the concentration of lactate dehydrogenase(LDH)in the culture supernatant was detected by enzyme-linked immunosorbent assay and the apoptosis rate of HMECs was detected by Annexin V-FTTC/PI staining, then comparison between different groups were made.Results:Compared with the control group, LDH concentration in LPS group increased significantly at 24 h[(4.67±1.27) ng/L vs. (11.57±0.83) ng/L ] and 48 h[(7.93±0.80) ng/L vs. (12.72±0.89) ng/L ]; Compared with LPS group, LDH concentration in LPS + TSA group decreased significantly at 24 h[(6.01±0.29) ng/L ] and 48 h[(5.96±0.27) ng/L ], and the differences were statistically significant ( P<0.05). Compared with the control group, the apoptosis rates of HMEC cells in LPS group were significantly higher at 24 h[(0.92±0.89)% vs. (1.66±0.09)% ] and 48 h[(1.09±0.14)% vs. (5.01±0.16)%]; Compared with LPS group, the apoptosis rate of HMEC cells in LPS + TSA group significantly decreased at 24 h[(1.36±0.01)% ] and 48 h[(4.19±0.23)% ], the differences were statistically significant ( P<0.05). Conclusion:TSA has the protective effect of reducing cell injury and apoptosis in sepsis.
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@#BACKGROUND: Sepsis-induced liver injury is a fatal complication of sepsis. Trichostatin A (TSA) regulates inflammation and autophagy in some human diseases, and forkhead box O3a (FoxO3a) has been shown to regulate autophagy. The present study aims to investigate whether TSA exerts its effects on septic liver injury through the FoxO3a/autophagy signaling pathway. METHODS: A sepsis mouse model was constructed by the cecal ligation and puncture (CLP) method, and AML12 cells were pretreated with lipopolysaccharide (LPS) (1 μg/mL) to establish a sepsis cell model. Forty mice were divided into four groups, namely control group, TSA group, CLP group, and CLP+TSA group, with 10 mice in each group. Cells were divided into control group, TSA group, LPS group, and LPS+TSA group. Hematoxylin-eosin (H&E) staining and biochemical methods were used to evaluate liver tissue injury. Enzyme-linked immunosorbent assay (ELISA) was applied to detect the expression of proinflammatory cytokines, and Western blotting and immunofluorescence were used to measure autophagy-related protein expression. RESULTS: Compared with the CLP group (mice), the proinflammatory cytokines (interleukin-β [IL-β] 2,665.27±324.90 pg/mL to 2,080.26±373.66 pg/mL; interleukin-6 [IL-6] 399.01±60.98 pg/mL to 221.90±46.89 pg/mL) and the hepatocyte injury markers (aspartate transaminase [AST] from 198.18±27.07 U/L to 128.42±20.55 U/L; alanine aminotransferase [ALT] from 634.98±74.10 U/L to 478.60±32.56 U/L) were notably decreased after TSA intervention. Moreover, LC3 II and FoxO3a showed an obvious increase and P62 showed an obvious decrease in the CLP+TSA group. Cell experiment results showed the similar trend. After FoxO3a gene was knocked down in AML12 cells, the promotion of autophagy and the improvement of liver enzyme index and inflammation by TSA were weakened. CONCLUSION: TSA may improve the inflammatory response and liver injury in septic mice through FoxO3a/autophagy.
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Objective:To investigate the mechanism of microRNA-4298 (miR-4298) targeting inhibition of peptidylarginine deiminase 4 (PADI4) expression in U251 cells apoptosis induced by histone deacetylase inhibitor trichostatin A (TSA).Methods:The experiment was divided into TSA group (0.2 μmol/L TSA) and contral group. CCK8 method was used to detect the effect of TSA on the proliferation of U251 cells. Flow cytometry was used to detect the apoptosis level of U251 cells after drug action. Reverse transcription PCR and Western blotting experiments were used to determine the changes of PADI4 gene and protein expression after miR-4298 interference. Luciferase assay was used to determine the effects of miR-4298 on targeted binding and luciferase activity in the 3′UTR region of PADI4. U251 cells were transfected with PADI4 to observe the rescue effect of miR-4298 on apoptosis. Each experiment was divided into 3 groups, NC group, miR-4298 mimic group, TSA+ miR-4298 mimic group and NC group, miR-4298 inhibitor group, TSA+ miR-4298 inhibitor group.Results:U251 cells were treated with 0.2 μmol/L TSA for 4 days, the Absorbancy ( A) values of the control group was 1.168±0.148, which was higher than those of the TSA group (0.737±0.007), with statistically significant difference ( t=4.948, P=0.008). Compared with the control group, after treatment with 0.2 μmol/L TSA for 48 h, U251 cells showed obvious apoptosis (27.62%±3.49% vs. 4.99%±0.13%, t=11.190, P<0.001). Compared with those before the drug treatment, the PADI4 gene expression (0.386±0.020 vs. 0.903±0.021) and protein expression (0.276±0.041 vs. 0.777±0.031) after TSA treatment for 48 h both were decreased, with statistically significant differences ( t=30.400, P<0.001; t=16.770, P<0.001). The expression of miR-4298 during the process of TSA-induced U251 cell apoptosis increased (2.573±0.289 vs. 1.003±0.136; t=8.487, P=0.001). There was a negative correlation between the miR-4298 expression and PADI4 expression ( r=-0.877, P=0.002). Luciferase experiment confirmed that miR-4298 has targeted binding and luciferase inhibitory effects on the 3′UTR region of wild-type PADI4. Compared with NC group (0.920±0.026), the relative expression levels of PADI4 gene of miR-4298 mimic group (0.413±0.049) and TSA+ miR-4298 mimic group (0.213±0.035) were decreased, with statistically significant differences (all P<0.001). The PADI4 protein expression level change was consistent with the gene change trend. Compared with NC group (3.78%±0.68%), the apoptosis levels of miR-4298 mimic group (7.96%±1.10%) and TSA+ miR-4298 mimic group (13.74%±1.26%) were increased, with statistically significant differences ( P=0.005; P<0.001). Compared with NC group (0.183±0.025), the PADI4 gene expression of miR-4298 inhibitor group (0.483±0.032) and TSA+ miR-4298 inhibitor group (0.386±0.025) were increased, with statistically significant differences ( P<0.001; P=0.015). The PADI4 protein expression level change was consistent with the gene change trend. Compared with NC group (4.96%±0.59%), the apoptosis levels of miR-4298 inhibitor group (23.83%±2.20%) and TSA+ miR-4298 inhibitor group (9.55%±1.49%) were increased, with statistically significant differences (all P<0.001). In the rescue experiment, the expression level of PADI4 in miR-4298 mimic group was significantly increased ( P<0.001), while the expression level of PADI4 in miR-4298 mimic+ PADI4 group was relatively reversed ( P=0.002). Conclusion:miR-4298 can participate in the process of U251 cell apoptosis by targeting the expression of PADI4 gene, and miR-4298 may be the target of targeted intervention therapy for glioma.
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Previously, we evaluated the effect of trichostatin A (TSA) on the expression of DNA methyltransferase 1 (DNMT1) in Hepatocellular Carcinoma (HCC). Fragile histidine triad (FHIT) and WW domain-containing oxidoreductase (WWOX) are two of the most common down-regulated genes in many cancers located on chromosome 3p14.2 and 16q23.3-24.1 respectively. The aim of the current study was to assess the effect of TSA on these genes expression, cell growth, and apoptosis in HCC WCH 17 cell. The cells were seeded and treated with TSA at different times. Then, MTT assay, flow cytometry, and qRT-PCR were achieved to determine viability, apoptosis and gene expression respectively. Cell growth was significantly inhibited, 92 to 36% after 24 h, 86 to 28% after 48 h, and 78 to 24% after 72 h. The results of flow cytometry confirmed that TSA increased apoptosis compared to the control group, the apoptosis percentage increased to 12%, 16%, and 18% in comparison to control groups (2%). Significant up-regulation of the genes was observed in all treated groups. We concluded that re-expression of silenced WWOX and FHIT genes could be achieved by TSA resulting in cell growth inhibition and apoptosis induction in WCH 17 cell.
Subject(s)
Apoptosis/physiology , Carcinoma, Hepatocellular/pathology , WW Domain-Containing Oxidoreductase , Growth/physiology , Chromosomes/classification , Flow Cytometry/instrumentation , Neoplasms/classificationABSTRACT
Objective: The main objective of the research work is to develop and validate a rapid UHPLC method for the estimation of assay and its related substances of Trichostatin A (TSA) in pharmaceutical samples. Methods: The UHPLC method developed for chromatographic separation between TSA and its related compounds on Poroshell 120 SB C18(50×4.6) mm; 2.7 µm RRLC column using Agilent RRLC (UHPLC) system with linear gradient elution. Results: The developed UHPLC method has shown excellent chromatographic separation between TSA and its related compounds within 12 min run time, during validation experiments, specificity study revealed that the peak threshold was more than the peak purity and no purity flag was observed. Repeatability, intra, and inter-day precision results were well within the tolerable limits. Limits of detection concentrations were found between 0.075 to 0.077 ppm and the limit of quantitation is between 0.252 to 0.258 ppm for related compounds and TSA. The related substances method recoveries were found between 80 and 120 % and assay method recovery was found between 98.0 to 102.0%. Conclusion: The developed method capability was proven for the assay of TSA and its related compounds in pharmaceutical samples and the method shows eco-friendlier than routine, conventional HPLC methods in terms of analysis time, cost and HPLC effluent waste.
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Objective To investigate the effect of histone deacetylase inhibitor trichostatin A (TSA) on the migration of human esophageal squamous carcinoma cells(ESCC) and the possible mechanism. Methods KYSE-150 cells and EC9706 cells were cultured and Transwell assay was performed to detect the role of TSA alone and combined with protein kinase C (PKC) inhibitor AEB071 on cell migration; the images of morphology after cells treatment with TSA or combination of AEB071 with TSA; Western blotting was conducted to examine the protein level of epithelial-mesenchymal transition (EMT) related signaling molecules. Results TSA promoted the migration of ESCC cells significantly, and PKC inhibitor AEB071 partly inhibited the effect of TSA-promoted ESCC cells migration. Treatment with TSA resulted in the cell morphology transitioned from epithelia oval-like to mesenchymal spindle-like, indicating the EMT. AEB071 partially rescued ESCC cells morphological changes which TSA induced. Western blotting showed that TSA reduced the expression of E-cadherin and augmented the expression of vimentin, β-catenin, Slug and acH3, whereas AEB071 obviously blocked the EMT-related protein level changes which induced by TSA. Conclusion TSA promotes ESCC cells migration via inducing EMT process and the mechanism may be mediated by PKC signaling pathway.
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Objective Trihostatin A (TSA) is a histone deacetylase inhibitor of oxime salts, which has certain anti-tumor activity. This article mainly investigates the molecular mechanism of TSA inhibiting cell proliferation through p-AKT/p-mTOR pathway in gastric cancer cells. Methods Gastric cancer cell line-SGC-7901 were treated with TSA of different concentrations, and the inhibition rate of TSA on the cells was detected by MTT assay. The cells were divided into control group (without any treatment), TSA-treated group (200ng/ml TSA), p-AKT covering group (200 ng/mL TSA+8 μg/mL SC79) and autophagy inhibition group (5 mmol/mL 3-methyladenine+200 ng/mL TSA). The protein expression distribution of Lc3 in control and TSA group were detected by cell immunofluorescence staining. The relative expression levels of p-AKT, p-mTOR and autophagy related proteins Lc3 and P62 in control group, TSA group and p-AKT covering group were detected by Western blot. The proliferation of cells in control group, TSA group, p-AKT covering group and autophagy inhibition group were measured by EdU and cell count assay. Results After 24h of treatment, Lc3 in TSA group formed a large number of granular aggregates in the cytoplasm, and the fluorescence distribution changed from the initial diffuse to dense. The TSA group showed a significant reduction in green fluorescence compared with the control group in the EdU experiment. The expression levels of p-AKT in the control group, TSA group and the autophagy inhibition group were 1.78±0.19, 0.92±0.03 and 1.71±0.19, respectively, and Lc3 were 0.21±0.01, 0.79±0.06 and 0.55±0.10, respectively. Compared with the control group, the relative expression level of p-AKT in the TSA group all decreased, while the expression level of Lc3 increased (P <0.05). p-mTOR in the three groups was 0.80±0.16, 0.45±0.04 and 0.98±0.16, respectively. Compared with the control group, the relative expression level of p-mTOR in the TSA group all decreased (P <0.05). P62 in the three groups was 1.17±0.15, 0.48±0.08 and 0.77±0.10, respectively. Compared with the control group, the relative expression level of P62 in the TSA group all decreased (P<0.05). Compared with the TSA group, p-AKT, p-mTOR and P62 expression in the p-AKT covering group increased (P<0.05), while Lc3 expression decreased (P<0.05). Compared with the control group, the inhibition effect of cell growth curve was the most obvious in the TSA group, while the cell growth curve of p-AKT covering group and autophagy inhibition group showed a partial recovery compared with the TSA group. Conclusion TSA can promote autophagy by inhibiting p-AKt/p-mTOR pathway, thus inhibiting the proliferation of gastric cancer cells.
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Objective To investigate the effect of trichostatin A (TSA) on migration and invasion in human gastric carcinoma SGC-7901 cells and its possible mechanism. Methods SGC-7901 cells were cultured in vitro and treated with TSA (5, 10, 20, 40, 80, 160 nmol/L) for 48 hours, and then the cell viability was detected by cell counting kit 8 (CCK-8) assay. The protocadherin 9 (PCDH9) high-expression SGC-7901 cells were stably established by transfecting with eukaryotic expression vector (pCMV6-PCDH9). Transwell assay was used to determine the abilities of migration and invasion. The mRNA expression level of PCDH9 were measured by RT-PCR. Western blotting was performed to analyze the protein expression of PCDH9, Snail, E-cadherin, matrix metalloproteinase (MMP)-2 and MMP-9. Results TSA remarkably reduced the cell viability of SGC-7901 cells in excess of 80 nmol/L (P<0. 05). However, in a dose-dependent manner, low-level TSA (5-20 nmol/L) suppressed migration and invasion of SGC-7901 cells (P<0. 05), down-regulated the protein levels of Snail, MMP-2 and MMP-9 (P<0. 05), and up-regulated the protein levels of PCDH9 and E-cadherin (P<0. 05). Meanwhile, high expression of PCDH9 also inhibited migration and invasion of SGC-7901 cells (P<0. 05), down-regulated the protein levels of Snail, MMP-2 and MMP-9 (P<0. 05), and up-regulated the protein level of E-cadherin (P<0. 05). Conclusion TSA may inhibit migration and invasion of SGC-7901 cells most likely via up-regulating PCDH9, and then down-regulating the protein levels of Snail, MMP-2 and MMP-9, and up-regulating the protein level of E-cadherin.
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OBJECTIVE@#To explore the analgesic mechanism of intrathecal trichostatin A (TSA) injection in a rat model of neuropathic pain induced by chronic constrictive injury (CCI).@*METHODS@#Male SD rats were randomized into sham operation+ DMSO group (group S), CCI +DMSO group (group C), CCI +10 μg TSA group (group T), and in the latter two groups, rat models of neuropathic pain were established induced by CCI. The rats were given intrathecal injections of 10 μL 5% DMSO or 10 μg TSA (in 5% DMSO) once a day on days 7 to 9 after CCI or sham operation. The rats were euthanized after behavioral tests on day 10, and the lumbar segment of the spinal cord was sampled to determine the expression of histone deacetylase 4 (HDAC4) protein and mRNA and detect the differentially expressed miRNAs using a miRNA chip. MiR-190b-5p and miR-142-3p were selected for validation of the results using RT-qPCR.@*RESULTS@#Compared with those in group S, the rats in group C showed significantly decreased paw withdrawal mechanical threshold (PWMT) from day 3 to day 10 after CCI ( < 0.05); intrathecal injection of TSA significantly reversed the reduction of PWMT following CCI ( < 0.05). Positive HDAC4 expression was detected mainly in the cytoplasm of the neurons in the gray matter of the spinal cord, and was obviously up-regulated after CCI ( < 0.05). Intrathecal injection of TSA significantly suppressed CCI-induced up-regulation of HDAC4 at 10 days after the operation ( < 0.05). Compared with the miRNA profile in group S, miRNA profiling identified 83 differentially expressed miRNAs in group C (fold change ≥2 or ≤0.5, < 0.05); TSA treatment reversed the expressions of 58 of the differentially expressed miRNAs following CCI, including 41 miRNAs that were decreased after CCI but up-regulated following TSA treatment. The results of real-time PCR validated the changes in the expressions of miR-190b-5p and miR-142-3p.@*CONCLUSIONS@#TSA suppresses CCI-induced up-regulation of HDAC4 and reverses differential expressions of miRNAs in the spinal cord of rats, which may contribute to the analgesic effect of TSA on neuropathic pain.
Subject(s)
Animals , Male , Rats , Histone Deacetylases , Hydroxamic Acids , MicroRNAs , Rats, Sprague-Dawley , Spinal Cord , Up-RegulationABSTRACT
Objective To investigate the effects of histone deacetylase inhibitors trichostatin A (TSA) on acute lung injury in septic mice.Methods Septic mice model was induced by cecal ligation and puncture(CLP).Ninty male BALB/c mice of clean grade were randomly(random number) divided into six groups(n=15),namely sham operation group,CLP group,CLP+DMSO group,CLP+TSA 1 mg group,CLP+TSA 5 mg group,and CLP+TSA 10 mg group.TSA(1 mg/kg,5 mg/kg,10 mg/kg) was administrated 12 hours before operation by intraperitoneal injection.And mice in sham group were only treated with laparotomy without CLP,and 24 h later,all survived mice were sacrificed to obtain specimens.ELISA method was employed to detect the concentrations of TNF-α and IL-1β in BALF.The lung wet/dry ratio was calculated.Histopathology changes of lung tissues were observed under light microscope.Lung tissue cell apoptosis was detected by TUNEL method.Caspase-3,Caspase-9 and CytC were assayed by Western blotting.The survival rate of mice in each group was calculated by additional 120 mice.Data were analyzed by SPSS 23.0.Statistical analyses were performed using independent sample t-test to compare between two groups or one-way analysis of variance test to compare among muhiple groups.The survival rate of mice was analyzed by univariate analysis using log-rank test.Results The lung W/D(P=0.021),the concentrations of TNF-α(P=0.000 1)and IL-1β(P=0.000 6)in BALF,puhnonary pathological change(P=0.001 6),lung tissue cell apoptotic index(P=0.000 9),the levels of apoptosis proteins (P＜0.05) in CLP group were higher than those in sham group,while survival rate (P=0.000 1) in CLP groups was lower than that in sham group.Compared with DMSO,the TSA significantly reduced the lung W/D,the levels of TNF-α.IL-1β in BALF,pathologic changes of lung tissue,lung tissue cell apoptotic index and the levels of apoptosis proteins in septic mice(P＜0.05).The increase in survival rate (P=0.007 2) associated with TSA(10 mg/kg)administration.Conclusion TSA exerts protective effects through attenuating pro-inflammatory cytokines and lung tissue cell apoptosis in sepsis induced acute lung injury in mice.
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AIM:To investigate influence of demethylation/acetylation by 5-Aza-2'-deoxycytidine/trichostatin A(5-Aza/TSA)treatment on B-cell specific phenotype of non-Hodgkin lymphoma cells.METHODS:CD19 promoter-driven reporter with NEO cassette was constructed to realize transfection and stable selection of Hodgkin and non -Hodgkin lymphoma cells.The exogenous CD19 promoter activity in both cell line clusters with and without 5-Aza/TSA treatment was detected and compared.The B-cell specific expression profiling in Eμ-myc transgenic mouse model developed lymphoma was isolated and identified.The effects of 5-Aza/TSA treatment on B-cell specific phenotype were analyzed.RESULTS:Epigenetic modification via 5-Aza/TSA repressed B-cell specific phenotype in B-cell-derived non-Hodgkin lymphoma cells. CONCLUSION:Epigenetic modification of pivotal master repressor genes plays an essential role in B -cell phenotype of both human and murine developed B-cell non-Hodgkin lymphoma cells.
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Nasal polyposis is a multi-factorial disease associated with chronic inflammation of the paranasal sinuses. Myofibroblast differentiation and extracellular matrix (ECM) accumulation are involved in the pathogenesis of nasal polyposis. Epigenetics, DNA methylation, and chromatin modifications are critical for generating cellular diversity and for maintaining distinct gene expression profiles. Based on our recent study that evaluated the inhibitory effect of Trichostatin A on myofibroblast differentiation in nasal polyposis, we hypothesized that HDAC inhibition is associated with myofibroblast differentiation and extracellular matrix accumulation in nasal polyposis and suggested that Trischostatin A may be useful as an inhibitor of nasal polyp growth and thus has potential to be used as a novel treatment option for nasal polyposis. In this review, we present general concept of epigenetics and results of recent research that elucidate the role of epigenetics in the pathogenesis of nasal polyps.
Subject(s)
Chromatin , DNA Methylation , Epigenomics , Extracellular Matrix , Fibroblasts , Inflammation , Myofibroblasts , Nasal Polyps , Paranasal Sinuses , TranscriptomeABSTRACT
Objective Lysine-specific demethylase 1 (LSD1) is a flavin adenine dinucleotide-dependent oxidase, which participates in many biological processes , such as cell proliferation and differentiation and gene activation and repression .The aim of this study was to investigate LSD1 acetylation by histone deacetylase inhib -itor trichostatin A ( TSA) and its effect on TSA-induced apoptosis of ovarian cancer cells . Methods LSD1 shRNA was synthesized and implanted into the pLKO-Tet-On lentiviral vector , which was transfected into HO8910 and SKOV3 ovarian cancer cell lines , and then the transfected cells were screened with 1.5μg/mL puromycin for one week until stable clones were established .The cells were treated for 48 hours with methanol (2 mg/mL, control), TSA (200 nmol/L), TCP (100μmol/L), or TSA+TCP.And in the experiment of RNA interfering the LSD1 expression, the cells were also treated for 48 hours with methanol (2 mg/mL, control), TSA (200 nmol/L), Dox (100 ng/mL), or TSA +Dox.The levels of LSD1 acetylation and its substrate histone H3 lysine 4 dimethylation (H3K4me2) were de-tected by immunoprecipitation (IP) and Western blot.The apoptosis of the cells was determined by Annexin Ⅴ/PI staining and flow cytometry, the transcription levels of the Bax and p21 genes detected by real-time quantitative PCR, and the H3K4me2levels in the promoter regions of Bax and p21 measured by chromatin immunoprecipitation ( ChIP ) .Results In comparison with the methanol control, the TSA group showed significantly increased levels of LSD 1 acetylation in the HO8910(1.00±0.29 vs 5.83±0.46, P<0.01) and SKOV3 cells ( 1.00±0.24 vs 5.07±0.35, P<0.01) as well as that of H3K4me2 ( P<0.01);the total apoptosis rates of HO 8910 and SKOV3 cells were remarkably increased in the TSA, TCP, and TSA+TCP groups (P<0.05), even more significantly in the TSA+TCP than in the TSA and TCP groups ( P<0.05) .The mRNA expressions of Bax and p21 in the HO8910 cells were markedly upregulated in the TSA, Dox, and TSA+Dox groups (P<0.05), even more significantly in the latter than in the former two groups (P<0.05).The TSA group exhibited a higher level of H 3K4me2 than the methanol control in the promoters of Bax(2 .92±0.26 vs 0.68±0.19, P<0.01) and p21 (3.07±0.29 vs 0.93±0.17, P<0.01). Conclusion TSA induces the LSD1 acetylation, while suppression of LSD1 expres-sion and activity may enhance the antitumor activity of TSA .
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Objective To investigate the effect of trichostatin A(TSA),a histone deacetylase inhibitor, on s-100-induced autoimmune hepatitis in mice.Methods A total of 26 six-week-old male C57BL/6 mice were randomly divided into control group,model group and TSA group(six in each group),and the rest 8 mice were used to extract the s-100 protein from liver tissue.Mice of model group and TSA group were injected intraperitoneally with s-100 with complete Freund's adjuvant to induce autoimmune hepatitis model.At day 21, TSA group mice were injected intraperitoneally with TSA 2 mg/(kg·d)for 7 days,and 0.9% sodium chloride solution containing 1% dimethyl sulfoxide was injected into the control and model group mice.Alanine transaminase(ALT)and aspartate aminotransferase(AST)in serum were measured and liver histopathology was observed.The protein levels of nuclear factor(NF)-κB and acetylated histone H3 in liver tissue were detected by Western Blot.The hepatic mRNA levels of NF-κB,HDAC3,toll-like receptor 4(TLR4)and TNF-α were measured by real-time PCR.ELISA was used to determine the TNF-α in serum.The results were analyzed with t test.Results The serum levels of ALT in control group,model group and TSA group were(122.00 ± 45.29),(459.33 ± 167.58)and(217.33 ± 49.25)U/L,respectively.The differences between model group and control group or TSA group were significant(t=4.76 and 3.41,respectively,both P<0.05).The serum levels of AST in control group,model group and TSA group were(127.83 ± 18.55),(389.67 ± 87.14)and (249.50 ± 71.72)U/L,respectively.The differences between model group and control group or TSA group were also significant(t= 7.20 and 3.04,respectively,both P< 0.05).The inflammation of the liver histopathology induced by s100 was alleviated by TSA.The relative expressions of NF-κB protein,NF-κB mRNA,TNF-α mRNA,HDAC3 mRNA and TLR4 mRNA in the liver tissue of model group mice were 2.43 ± 0.42,9.51 ± 0.36,10.53 ± 0.74,2.90 ± 0.22,and 4.50 ± 0.73,respectively,which were significantly higher than those of the control group(1.28 ± 0.49,1.28 ± 0.49,1.06 ± 0.14,1.72 ± 0.73,and 1.01 ± 0.31, respectively)(t=4.68,37.14,30.69,4.33 and 10.85,respectively,all P <0.05).In TSA group,the relative expressions of NF-κB protein,NF-κB mRNA,TNF-α mRNA,HDAC3 mRNA and TLR4 mRNA were decreased(1.30 ± 0.36,1.30 ± 0.36,2.38 ± 0.36,2.13 ± 0.32 and 2.40 ± 0.51,respectively),which were statistically lower than those in model group(t=4.58,30.62,24.12,2.81 and 5.81,respectively,all P<0.05).The serum TNF-α levels in control group,model group and TSA group were(122.37 ± 68.12), (1361.44 207.13)and(691.64 ± 162.12)ng/L,respectively.Compared with model group,the differences were statistically significant(t=13.92 and 6.24,respectively,both P<0.05).The relative expression of ac-H3 protein in the model group was 1.10 ± 0.08,which was higher than that in the control group 0.96 ± 0.17(t=2.27,P<0.05).That in TSA group was 1.30 ± 0.04,which was higher than the model group(t=-0.30, P <0.05).Conclusion Histone deacetylase inhibitor TSA alleviates autoimmune hepatitis by enhancing histone acetylation and inhibiting NF-κB and inflammatory factors.
ABSTRACT
Objective Lysine-specific demethylase 1 (LSD1) is a flavin adenine dinucleotide-dependent oxidase, which participates in many biological processes , such as cell proliferation and differentiation and gene activation and repression .The aim of this study was to investigate LSD1 acetylation by histone deacetylase inhib -itor trichostatin A ( TSA) and its effect on TSA-induced apoptosis of ovarian cancer cells . Methods LSD1 shRNA was synthesized and implanted into the pLKO-Tet-On lentiviral vector , which was transfected into HO8910 and SKOV3 ovarian cancer cell lines , and then the transfected cells were screened with 1.5μg/mL puromycin for one week until stable clones were established .The cells were treated for 48 hours with methanol (2 mg/mL, control), TSA (200 nmol/L), TCP (100μmol/L), or TSA+TCP.And in the experiment of RNA interfering the LSD1 expression, the cells were also treated for 48 hours with methanol (2 mg/mL, control), TSA (200 nmol/L), Dox (100 ng/mL), or TSA +Dox.The levels of LSD1 acetylation and its substrate histone H3 lysine 4 dimethylation (H3K4me2) were de-tected by immunoprecipitation (IP) and Western blot.The apoptosis of the cells was determined by Annexin Ⅴ/PI staining and flow cytometry, the transcription levels of the Bax and p21 genes detected by real-time quantitative PCR, and the H3K4me2levels in the promoter regions of Bax and p21 measured by chromatin immunoprecipitation ( ChIP ) .Results In comparison with the methanol control, the TSA group showed significantly increased levels of LSD 1 acetylation in the HO8910(1.00±0.29 vs 5.83±0.46, P<0.01) and SKOV3 cells ( 1.00±0.24 vs 5.07±0.35, P<0.01) as well as that of H3K4me2 ( P<0.01);the total apoptosis rates of HO 8910 and SKOV3 cells were remarkably increased in the TSA, TCP, and TSA+TCP groups (P<0.05), even more significantly in the TSA+TCP than in the TSA and TCP groups ( P<0.05) .The mRNA expressions of Bax and p21 in the HO8910 cells were markedly upregulated in the TSA, Dox, and TSA+Dox groups (P<0.05), even more significantly in the latter than in the former two groups (P<0.05).The TSA group exhibited a higher level of H 3K4me2 than the methanol control in the promoters of Bax(2 .92±0.26 vs 0.68±0.19, P<0.01) and p21 (3.07±0.29 vs 0.93±0.17, P<0.01). Conclusion TSA induces the LSD1 acetylation, while suppression of LSD1 expres-sion and activity may enhance the antitumor activity of TSA .
ABSTRACT
AIM: To investigate the effect of 3-methyladenine (3-MA) combined with trichostatin A (TSA) on triple-negative breast cancer cells and the mechanism involved.METHODS: The viability of MDA-MB-231 cells was detected by MTT assay, the migration ability was determined by scratch assay, and the expression of autophagy-related proteins was detected by Western blot.RESULTS: TSA significantly inhibited the viability of MDA-MB-231 cells in a time-and dose-dependent manner.The results of scratch assay showed that TSA inhibited cell migration ability.Western blot data indicated that TSA resulted in a moderate increase in LC3-Ⅱ expression.Moreover, 3-MA inhibited cell autophagy induced by TSA, and combination of 3-MA and TSA further inhibited the viability of MDA-MB-231 cells.CONCLUSION: Combination of 3-MA and TSA may effectively inhibit the growth of triple-negative breast cancer cells.
ABSTRACT
Objective To investigate the effects of 5-aza-2'-deoxycytidine (5-Aza-dC) alone or combined with trichostatin A(TSA) on cell proliferation, promoter methylation and mRNA expression level of PDX-1 gene in pancreatic β cells induced by high glucose toxicity. Method NIT-1 cells were treated in vitro by high glucose (33.3 mmol/L), then divided into five groups, control group, HG grpup, 5-Aza-dC treatment group, TSA interfere group and 5-Aza-dC + TSA group. Proliferation of NIT-1 cells, insulin secretion, promoter methylation and mRNA expression of PDX-1 gene were detected respectively. Results 5-Aza-dC and TSA alone or in combination could promote cell proliferation and recover insulin secretion in NIT-1 cells , could also reduce PDX-1 gene methylation and enhance expression of PDX-1 mRNA. Compared with single-treatment group , combined group was significantly different (all P < 0.05). Conclusion 5-Aza-dC and TSA could activate the expression of PDX-1 and, then recover insulin secretion in NIT-1 cells induced by high glucose. Combination of them had synergistic effect.