Trichostatin A inhibits the activation of Hepatic stellate cells by Increasing C/EBP-α Acetylation in vivo and in vitro.

Reversal of activated hepatic stellate cells (HSCs) to a quiescent state and apoptosis of activated HSCs are key elements in the reversion of hepatic fibrosis. CCAAT/enhancer binding protein α (C/EBP-α) has been shown to inhibit HSC activation and promote its apoptosis. This study aims to investigate how C/EBP-α acetylation affects the fate of activated HSCs. Effects of a histone deacetylation inhibitor trichostatin A (TSA) on HSC activation were evaluated in a mouse model of liver fibrosis caused by carbon tetrachloride (CCl4) intoxication. TSA was found to ameliorate CCl4-induced hepatic fibrosis and improve liver function through increasing the protein level and enhancing C/EBP-α acetylation in the mouse liver. C/EBP-α acetylation was determined in HSC lines in the presence or absence of TSA, and the lysine residue K276 was identified as a main acetylation site in C/EBP-α protein. C/EBP-α acetylation increased its stability and protein level, and inhibited HSC activation. The present study demonstrated that C/EBP-α acetylation increases the protein level by inhibiting its ubiquitination-mediated degradation, and may be involved in the fate of activated HSCs. Use of TSA may confer an option in minimizing hepatic fibrosis by suppressing HSC activation, a key process in the initiation and progression of hepatic fibrosis.

TSA increases C/EBP­α expression by increasing its lysine acetylation in hepatic stellate cells.

CCAAT enhancer binding protein­α (C/EBP­α) is a transcription factor expressed only in certain tissues, including the liver. It has been previously demonstrated that C/EBP­α may induce apoptosis in hepatic stellate cells (HSCs), raising the question of whether acetylation of C/EBP­α is associated with HSCs, and the potential associated mechanism. A total of three histone deacetylase inhibitors (HDACIs), including trichostatin A (TSA), suberoylanilide hydroxamic acid and nicotinamide, were selected to determine whether acetylation affects C/EBP­α expression. A Cell Counting Kit­8 assay was used to determine the rate of proliferation inhibition following treatment with varying doses of the three HDACIs in HSC­T6 and BRL­3A cells. Western blot analysis was used to examine Caspase­3, ­8, ­9, and ­12 levels in HSC­T6 cells treated with adenoviral­C/EBP­α and/or TSA. Following treatment with TSA, a combination of reverse transcription­quantitative polymerase chain reaction and western blot analyses was used to determine the inherent C/EBP­α mRNA and protein levels in HSC­T6 cells at 0, 1, 2, 4, 8, 12, 24, 36 and 48 h. Nuclear and cytoplasmic proteins were extracted to examine C/EBP­α distribution. Co­immunoprecipitation analysis was used to examine the lysine acetylation of C/EBP­α. It was observed that TSA inhibited the proliferation of HSC­T6 cells to a greater extent compared with BRL­3A cells, following treatment with the three HDACIs. TSA induced apoptosis in HSC­T6 cells and enhanced the expression of C/EBP­α. Following treatment of HSC­T6 cells with TSA, inherent C/EBP­α expression increased in a time­dependent manner, and its lysine acetylation simultaneously increased. Therefore, the results of the present study suggested that TSA may increase C/EBP­α expression by increasing its lysine acetylation in HSCs.