Emodin regulates histone acetylation to promote pyroptosis and apoptosis of HpG2 hepatoma cells

doi:10.11958/20230775

Abstract

Abstract:

Objective To study whether emodin, a natural product, can affect the level of histone acetylation in HpG2 hepatocellular carcinoma cells (HCC), and then accelerate the occurrence of pyroptosis and apoptosis in hepatocellular carcinoma cells, so as to provide a new target for the treatment of liver cancer. Methods CCK-8 method was used to detect the effect of different concentrations of emodin on the viability of HpG2 cells. Bioinformatics was used to analyze histone acetylation-related genes in patients with liver cancer in TCGA database. The correlation between the candidate gene lysine acetyltransferase 2A (KAT2A) and the apoptosis pathway was verified. qPCR method was used to detect the mRNA level of KAT2A in HepG2 cells and L02 cells. The effects of emodin on histone acetyltransferase (HAT) and histone deacetyltransferase (HDAC), interleukin 1β (IL-1β) and interleukin 18 (IL-18) in HpG2 cells were detected by ELISA. The effect of emodin on the apoptosis of liver cancer cells was detected by flow cytometry. The expression level of cell apoptosis, pyroptosis-associated protein B lymphocytoma-2 (Bcl-2), Bcl-2-related X protein (Bax), NOD-like receptor thermal protein domain-related protein 3 (NLRP3), Caspase-1, Gasdermin family member DN terminal (GSDMD-N) and KAT2A were detected by Western blot assay. Results Emodin could reduce the activity of HpG2 cells, and the confidence interval of IC₅₀ 95% was 58.12-66.52 μmol/L. Compared with normal liver tissue, the expression of histone acetylation related gene mRNA was increased in HCC tissue, and the change of KAT2A was the highest [log₂(Fold Change)=2.010, P<0.01]. In HCC tissue, the expression of KAT2A mRNA was negatively correlated with apoptosis pathway (r_s=-0.230, P<0.01). Compared with L02 cells, the expression of KAT2A mRNA in HepG2 was higher (P<0.05). Compared with the control group, expression levels of HAT and HDAC decreased in the 60 μmol/L emodin intervention group, expression levels of IL-18 and IL-1β increased, the apoptosis rate increased, expression levels of KAT2A and BAX decreased, and expression levels of Bcl-2, NLRP3, GSDMD-N and Caspase-1 increased (P<0.05). Conclusion Emodin could inhibit the viability of hepatoma cells, accelerate apoptosis and pyroptosis, and its mechanism may be related to the regulation of KAT2A expression.

Key words: Emodin, carcinoma, hepatocellular, acetylation, acetyltransferases, histones, pyroptosis, apoptosis, KAT2A

CLC Number:

R285，R735.7

Figures/Tables 10

Fig.1 Emodin affected IC50 of hepatocellular carcinoma cells

Fig.2 Volcano map of histone acetylation-related differential genes between hepatocellular carcinoma tissue and healthy liver tissue

Fig.3 Comparison of mRNA expression of KAT2A between hepatocellular carcinoma tissue and normal tissue

Fig.4 Spearman analysis of the correlation between KAT2A mRNA and the apoptotic pathway

Tab.1 Comparison of HAT, HDAC, IL-18 and IL-1β levels between the control group and the 60 μmol/L emodin intervention group （n=5，OD450，$\bar{x}±s$）

组别	HAT	HDAC	IL-18	IL-1β
对照组	1.37±0.20	1.29±0.12	0.14±0.01	0.18±0.04
大黄素干预组	0.69±0.13	0.89±0.07	0.31±0.05	0.40±0.05
t	6.043^＊＊	6.386^＊＊	7.077^＊＊	6.753^＊＊

Fig.5 Flow cytometry detection of apoptosis

Fig.6 Western blot detection of KAT2A expression

Fig.7 Western blot detection of apoptosis-associated proteins

Fig.8 Western blot detection of pyroptosis related proteins

Tab.2 Comparison of KAT2A, Bcl-2, BAX, NLRP3, GSDMD-N and Caspase-1 protein expression levels between the control group and the 60 μmol/L emodin intervention group （n=3，$\bar{x}±s$）

组别	KAT2A	Bcl-2	BAX
对照组	0.91±0.26	0.26±0.06	1.04±0.04
大黄素干预组	0.35±0.12	1.08±0.17	0.65±0.12
t	3.426^＊	7.909^＊＊	5.211^＊＊
组别	NLRP3	GSDMD-N	Caspase-1
对照组	0.57±0.02	0.58±0.13	0.52±0.07
大黄素干预组	0.97±0.16	1.03±0.18	0.98±0.04
t	4.392＊	3.632＊	10.507＊＊

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