Effect of Trichostatin A on Anti HepG2 Liver Carcinoma Cells: Inhibition of HDAC Activity and Activation of Wnt/β-Catenin Signaling

Abstract

Purpose: To investigate the effect of deacetylase inhibitory trichostatin A (TSA) on anti HepG2 liver carcinoma cells and explore the underlying mechanisms. Materials and Methods: HepG2 cells exposed to different concentrations of TSA for 24, 48, or 72h were examined for cell growth inhibition using CCK8, changes in cell cycle distribution with flow cytometry, cell apoptosis with annexin V-FTIC/PI double staining, and cell morphology changes under an inverted microscope. Expression of β-catenin, HDAC1, HDAC3, H3K9, CyclinD1 and Bax proteins was tested by Western blotting. Gene expression for β-catenin, HDAC1and HDAC3 was tested by q-PCR. β-Catenin and H3K9 proteins were also tested by immunofluorescence. Activity of Renilla luciferase (pTCF/LEF-luc) was assessed using the Luciferase Reporter Assay system reagent. The activity of total HDACs was detected with a HDACs colorimetric kit. Results: Exposure to TSA caused significant dose-and time-dependent inhibition of HepG2 cell proliferation (p<0.05) and resulted in increased cell percentages in G0/ G1 and G2/M phases and decrease in the S phase. The apoptotic index in the control group was 6.22±0.25%, which increased to 7.17±0.20% and 18.1±0.42% in the treatment group. Exposure to 250 and 500nmol/L TSA also caused cell morphology changes with numerous floating cells. Expression of β-catenin, H3K9and Bax proteins was significantly increased, expression levels of CyclinD1, HDAC1, HDAC3 were decreased. Expression of β-catenin at the genetic level was significantly increased, with no significant difference in HDAC1and HDAC3 genes. In the cytoplasm, expression of β-catenin fluorescence protein was not obvious changed and in the nucleus, small amounts of green fluorescence were observed. H3K9 fluorescence protein were increased. Expression levels of the transcription factor TCF werealso increased in HepG2 cells following induction by TSA, whikle the activity of total HDACs was decreased. Conclusions: TSA inhibits HDAC activity, promotes histone acetylation, and activates Wnt/β-catenin signaling to inhibit proliferation of HepG2 cell, arrest cell cycling and induce apoptosis.

Both bulk and cancer stem cell subpopulations in triple‐negative breast cancer are susceptible to Wnt, HDAC, and ERα coinhibition

“…This may be associated with an upregulation of Wnt signaling upon exposure to HDAC inhibitors such as VPA and trichostatin A in a patient-dependent manner. Trichostatin A and VPA have been reported to activate Wnt signaling in HepG2 liver carcinoma and Neuro 2A cells [46,47]. This is also demonstrated in breast cancer cell lines where HDAC inhibitors have been found to upregulate b-catenin expression leading to increased Wnt signaling [29].…”

Clinical significance of the phosphorylation of MAPK and protein expression of cyclin D1 in human osteosarcoma tissues

“…Studies have indicated that the overexpression of cyclin D1 may result in a reduction of the G1 phase of the cell cycle (15)(16)(17), leading to progression into the S phase and completing the duplication of DNA. The increase in the protein expression of cyclin D1 is observed in certain primary malignant tumors, including parathyroid adenoma, neck squamous-cell carcinoma, breast cancer, esophageal cancer, and hepatocellular carcinoma (18)(19)(20)(21). As MAPK is expressed as an upstream gene of cyclin D1, the increase in the level of p-MAPK in osteosarcoma was directly proportional to the intensity of cyclin D1-positive staining.…”

Short hairpin RNA silencing of TGF-βRII and FZD-7 synergistically suppresses proliferation and metastasis of hepatocellular carcinoma cells

“…Cyclin D1 forms a complex with cyclin-dependent kinases 4 and 6 and functions as a regulatory subunit of this complex. Cyclin D1 is required to facilitate the transition between G1 and S phases; therefore, it promotes the proliferation of cells and may contribute to tumorigenesis ( 23 , 24 ). In addition, cyclin D1 acts as a downstream effector molecule for Wnt/β-catenin signaling.…”