Hepatocyte nuclear factor-1alpha (HNF1a) is one of the key transcription factors of the HNF family, which plays a critical role in hepatocyte differentiation. Substantial evidence has suggested that down-regulation of HNF1a may contribute to the development of hepatocellular carcinoma (HCC). Herein, human cancer cells and tumor-associated fibroblasts (TAFs) were isolated from human HCC tissues, respectively. A recombinant adenovirus carrying the HNF1a gene (AdHNF1a) was constructed to determine its effect on HCC in vitro and in vivo. Our results demonstrated that HCC cells and HCC tissues revealed reduced expression of HNF1a. Forced reexpression of HNF1a significantly suppressed the proliferation of HCC cells and TAFs and inhibited the clonogenic growth of hepatoma cells in vitro. In parallel, HNF1a overexpression reestablished the expression of certain liver-specific genes and microRNA 192 and 194 levels, with a resultant increase in p21 levels and induction of G 2 /M arrest. Additionally, AdHNF1a inhibited the expression of cluster of differentiation 133 and epithelial cell adhesion molecule and the signal pathways of the mammalian target of rapamycin and transforming growth factor beta/Smads. Furthermore, HNF1a abolished the tumorigenicity of hepatoma cells in vivo. Most interestingly, intratumoral injection of AdHNF1a significantly inhibited the growth of subcutaneous HCC xenografts in nude mice. Systemic delivery of AdHNF1a could eradicate the orthotopic liver HCC nodules in nonobese diabetic/severe combined immunodeficiency mice. Conclusion: These results suggest that the potent inhibitive effect of HNF1a on HCC is attained by inducing the differentiation of hepatoma cells into mature hepatocytes and G 2 /M arrest. HNF1a might represent a novel, promising therapeutic agent for human HCC treatment. Our findings also encourage the evaluation of differentiation therapy for tumors of organs other than liver using their corresponding differentiation-determining transcription factor.
The expression of cartilage-derived retinoic acid-sensitive protein (CD-RAP) is initiated at the beginning of chondrogenesis and continues throughout the cartilage development. In chondrocytes, CD-RAP is down-regulated by retinoic acid. To understand the molecular mechanism underlying this regulation and the cell-specific expression, the deletion constructs of the mouse CD-RAP promoter were transfected into chondrocytes and a melanoma cell line. The results revealed a domain that demonstrated high levels of expression specifically in chondrocytes. In this functional domain, we show that a cis-acting element, 5-GCCTGAGGC-3, binds to the trans-acting factor protein AP-2. Mutation of the AP-2 site on the CD-RAP promoter led to decreased transcription in C5.18 chondrocytes, indicating that this site may act as an activator of transcription. In contrast, increased concentration of AP-2, stimulated by retinoic acid, led to decreased transcription of the CD-RAP promoter, an effect that was abolished by mutation of the AP-2 binding site. The effect of AP-2 was further examined by co-transfection of C5.18 and HepG2 cells with the CD-RAP promoter constructs and an AP-2 expression plasmid. In a dose-dependent manner, cotransfection with AP-2 elevated and then decreased CD-RAP promoter activity. Taken together, these results suggest that AP-2 is involved in the biphasic regulation of CD-RAP transcription.
Background : Liver cancer stem cells (LCSCs) are responsible for the initiation, progression and chemoresistance of liver cancer. However, no agent targeting LCSC is available in the clinic to date. Here, we investigated the effects of targeting protein arginine methyltransferase 5 (PRMT5), an epigenetic regulator, on LCSCs and HCC using a novel PRMT5 inhibitor DW14800. Methods : Tumor spheroid formation culture was used to enrich LCSCs and assess their self-renewal capability. Human alpha-1-antitrypsin (A1AT) ELISA, acetylated low-density lipoprotein (ac-LDL) uptake, periodic acid-Schiff (PAS) reactions and senescence associated β-galactosidase (SA-β-gal) activity assays were performed to examine the differentiation status of HCC cells. The effects of DW14800 on HCC malignancy were assessed in HCC cell lines and on an HCC xenograft model in mice. Chromatin immunoprecipitation was applied to clarify the transcriptional regulation of HNF4α by PRMT5-mediated Histone H4 arginine-3 symmetrical dimethylation (H4R3me2s). Results : Quantitative real-time PCR revealed that the expression of PRMT5 was upregulated in LCSCs. DW14800 specifically decreased the symmetrical dimethylation of arginine residues in HCC cells. Treatment of DW14800 suppressed the self-renewal capacity of LCSCs while re-establishing hepatocyte-specific characteristics in HCC cells. DW14800 displayed antitumor effects in HCC cells in vitro and in xenograft HCC in vivo . Importantly, ChIP assay showed that PRMT5 and H4R3me2s bound to the promoter region of HNF4α gene, and DW14800 increased the expression of HNF4α via reducing the H4R3me2s levels and enhancing the transcription of HNF4α. Conclusions : Our data revealed the significance of targeting PRMT5 activity in LCSC elimination and HCC differentiation, and proposed that DW14800 may represent a promising therapeutic agent for HCC in the clinic.
The liver is known as an organ with high proliferation potential. Clarifying the cellular origin and deepening the understanding of liver regeneration mechanisms will help provide new directions for the treatment of liver disease. With the development and application of lineage tracing technology, the specific distribution and dynamic changes of hepatocyte subpopulations in homeostasis and liver injury have been illustrated. Self‐replication of hepatocytes is responsible for the maintenance of liver function and mass under homeostasis. The compensatory proliferation of remaining hepatocytes is the main mechanism of liver regeneration following acute and chronic liver injury. Transdifferentiation between hepatocytes and cholangiocytes has been recognized upon severe chronic liver injury. Wnt/β‐catenin, Hippo/YAP and Notch signalling play essential roles in the maintenance of homeostatic liver and hepatocyte‐to‐cholangiocyte conversion under liver injury. In this review, we summarized the recent studies on cell origin of newly generated hepatocytes and the underlying mechanisms of liver regeneration in homeostasis and liver injury.
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