Silybin-Induced Apoptosis Occurs in Parallel to the Increase of Ceramides Synthesis and miRNAs Secretion in Human Hepatocarcinoma Cells.

Marcello Dallio,Daniela Vanacore,Stefania Lama,Pasquale Ferranti,Carmelina Loguercio,Paola Stiuso,Pasquale Sperlongano,Silvia Zappavigna,Michele Caraglia,Alessandro Federico,Aniello Russo,Nicoletta Potenza

doi:10.3390/ijms20092190

Abstract

Silybin is a flavonolignan extracted from Silybum marianum (milk thistle) with hepatoprotective, antioxidant, and anti-inflammatory activity. Several studies have shown that silybin is highly effective to prevent and treat different types of cancer and that its antitumor mechanisms involve the arrest of the cell cycle and/or apoptosis. An MTT assay was performed to study cell viability, lipid peroxidation, extracellular NO production, and scavenger enzyme activity were studied by Thiobarbituric Acid-Reactive Species (TBARS) assay, NO assay, and MnSOD assay, respectively. Cell cycle and apoptosis analysis were performed by FACS. miRNA profiling were evaluated by real time PCR. In this study, we demonstrated that Silybin induced growth inhibition blocking the Hepg2 cells in G1 phase of cell cycle and activating the process of programmed cell death. Moreover, the antiproliferative effects of silybin were paralleled by a strong increase of the number of ceramides involved in the modulation of miRNA secretion. In particular, after treatment with silybin, miR223-3p and miR16-5p were upregulated, while miR-92-3p was downregulated (p < 0.05). In conclusion, our results suggest that silybin-Induced apoptosis occurs in parallel to the increase of ceramides synthesis and miRNAs secretion in HepG2 cells.

Highlights

Silybin, a polyphenolic flavonoid, is the main component of silymarin with antioxidant and free radical scavenger activity, used as dietary supplement in the treatment of alcoholic liver disease [1]
We have recently reported that a chronic treatment with Realsil—a food supplement in which silybin is conjugated with phosphatidylcholineand vitamin E —significantly decreases liver damage plasma marker levels (AST, ALT, and γGT) and improves liver histology in ~50% of patients with NAFLD and NASH [2]
We evaluated the cytotoxic effects of different concentrations of silybin (Figure 1) on HepG2 cell growth; growth inhibition was time- and dose-dependent

Summary

Introduction

Silybin (sil), a polyphenolic flavonoid, is the main component of silymarin with antioxidant and free radical scavenger activity, used as dietary supplement in the treatment of alcoholic liver disease [1]. We have recently reported that a chronic treatment with Realsil—a food supplement in which silybin is conjugated with phosphatidylcholineand vitamin E —significantly decreases liver damage plasma marker levels (AST, ALT, and γGT) and improves liver histology in ~50% of patients with NAFLD and NASH [2]. We demonstrated that Realsil treatment restored the normal serum lipidomic profile in NAFDL patients by decreasing the risk to develop NASH-related hepatocellular carcinoma (HCC) [3,4,5,6]. Hepatocellular carcinoma (HCC) is the most common form of hepatic cancer with a poor prognosis in the advanced stage. The main treatment options for patients with HCC include tumor ablation, surgery, and transplantation, but these are not effective in patients with an advanced stage of disease [9,10]

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