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.