Mulberry Leaf Polyphenol Extract and Rutin Induces Autophagy Regulated by p53 in Human Hepatoma HepG2 Cells

The edible leaves of the mulberry (Morus alba L.) plant are used worldwide. They contain abundant polyphenolic compounds with strong anticancer properties. We previously revealed that apoptosis was mediated in p53-negative Hep3B cells, and mulberry leaf polyphenol extract (MLPE) induced autophagy in p53-transfected Hep3B cells. However, how this autophagy is induced by p53 in human hepatoma HepG2 (p53 wild type) cells remains unclear. In the current study, MLPE induced autophagy, as demonstrated by enhanced acidic vesicular organelle staining, by upregulating beclin-1, increasing LC3-II conversion, and phosphorylating AMPK. In HepG2 cells, these processes were associated with p53. Western blot also revealed phosphatidylinositol-3 kinase (PI3K), p-AKT, and fatty acid synthase (FASN) suppression in MLPE-treated cells. Moreover, treatment with the p53 inhibitor pifithrin-α (PFT-α) inhibited autophagy and increased apoptotic response in MLPE-treated HepG2 cells. PFT-α treatment also reversed MLPE-induced PI3K, p-AKT, and FASN suppression. Thus, co-treatment with MLPE and PFT-α significantly increased caspase-3, caspase-8, and cytochrome c release, indicating that p53 deficiency caused the apoptosis. In addition, rutin, a bioactive polyphenol in MLPE, may affect autophagy in HepG2 cells. This study demonstrates that MLPE is a potential anticancer agent targeting autophagy and apoptosis in cells with p53 status. Moreover, this work provides insight into the mechanism of p53 action in MLPE-induced cytotoxicity in hepatocellular carcinoma.

Novel Health Supplement for Recovery of Alcohol Induced Adverse Effects and Associated Disorders: HepG2 Cells Study and Safety Dose Profiling on Rats

The present study objective was to design and develop novel health-supplement formula from plant extracts and was to evaluate the formula for high episodic alcohol toxicities, and associated disorders against alcohol intoxicated and oxidative damaged Human Hepatoma HepG2 cell line.

Gold Nanorods are Selective Cytotoxic Agents

Background: Gold nanorods (GNRs) are very promising agents that have multiple applications in medicine and biology. However, the cytotoxic effects of GNRs have not been fully explored. Objective: Therefore, the main objective of this study was to determine the selective cytotoxic effect of GNRs towards several human tumor cell lines. Methods: To address this issue, three sizes of GNRs (10-nm, 25-nm, and 50-nm) were tested against two human tumor cell lines, namely, human hepatoma HepG2 and human prostate PC3 cancer cells. As GNRs are usually stored in soft tissues inside living bodies, we also tested the effect of GNRs on murine splenocyte viability. To determine if the GNRs displayed selectivity cytotoxicity towards cancer cells, active GNRs with the size showing the least cytotoxicity to splenocytes were then tested against a panel of 11 human tumor cell lines and two human non-tumor cell lines. Results: Our results showed that the most cytotoxic size of GNRs is 10-nm, followed by the 25-nm GNRs, while the 50-nm GNRs did not show a significant effect. In addition, the 25-nm GNRs were the least cytotoxic to splenocytes when tested for 24 and 48 h. These GNRs showed a selective cytotoxic effect to prostate cancer PC3 cells with median inhibitory concentration (IC50) = 8.3 + 0.37 µM, myeloblastic leukemia HL60 cells (IC50 = 19.7 + 0.89 µM), cervical cancer HeLa cells (IC50 = 24.6 + 0.37 µM), renal adenocarcinoma 786.0 cells (IC50 = 27.34 + 0.6 µM), and hepatoma HepG2 cells (IC50 = 27.79 + 0.03 µM) when compared to the effect on the non-tumor human cells; skin fibroblast BJ cell line (IC50 = 40.13 + 0.7 µM) or epithelial breast MCF10A cells (IC50 = 33.2 + 0.89 µM). A high selectivity indices (SI) were observed in GNRs-treated PC3 and HL60 cells with values ranging from 1.69 to 4.83, whereas moderate SIs were observed in GNRs-treated HeLa, 786.0, and HepG2 cells with values ranging from 1.19 to 1.63. Other cells did not show a similar selective effect, including human laryngeal HEp2 cells, colon HCT116, metastatic renal adenocarcinoma ACHN cells, and human breast cancer cells (MCF7, MDA-MB-231, and MDA-MB-468 cells). The effect of GNRs was confirmed using the colony formation assay and the effect was found to be cell cycle specific. Finally, it was shown that laser treatment can potentiate the cytotoxic effect of the 25-nm GNRs. Conclusion: GNRs are selective cytotoxic agents and they have the potential to act as candidate anticancer agents.

Sesquiterpenoids From the Antarctic Fungus Pseudogymnoascus sp. HSX2#-11

The fungal strains Pseudogymnoascus are a kind of psychrophilic pathogenic fungi that are ubiquitously distributed in Antarctica, while the studies of their secondary metabolites are infrequent. Systematic research of the metabolites of the fungus Pseudogymnoascus sp. HSX2#-11 led to the isolation of six new tremulane sesquiterpenoids pseudotremulanes A–F (1–6), combined with one known analog 11,12-epoxy-12β-hydroxy-1-tremulen-5-one (7), and five known steroids (8–12). The absolute configurations of the new compounds (1–6) were elucidated by their ECD spectra and ECD calculations. Compounds 1–7 were proved to be isomeride structures with the same chemical formula. Compounds 1/2, 3/4, 1/4, and 2/3 were identified as four pairs of epimerides at the locations of C-3, C-3, C-9, and C-9, respectively. Compounds 8 and 9 exhibited cytotoxic activities against human breast cancer (MDA-MB-231), colorectal cancer (HCT116), and hepatoma (HepG2) cell lines. Compounds 9 and 10 also showed antibacterial activities against marine fouling bacteria Aeromonas salmonicida. This is the first time to find terpenoids and steroids in the fungal genus Pseudogymnoascus.

Quantitative spectrofluorometric assay detecting nuclear condensation and fragmentation in intact cells

At present, nuclear condensation and fragmentation have been estimated also using Hoechst probes in fluorescence microscopy and flow cytometry. However, none of the methods used the Hoechst probes for quantitative spectrofluorometric assessment. Therefore, the aim of the present study was to develop a spectrofluorometric assay for detection of nuclear condensation and fragmentation in the intact cells. We used human hepatoma HepG2 and renal HK-2 cells cultured in 96-well plates treated with potent apoptotic inducers (i.e. cisplatin, staurosporine, camptothecin) for 6–48 h. Afterwards, the cells were incubated with Hoechst 33258 (2 µg/mL) and the increase of fluorescence after binding of the dye to DNA was measured. The developed spectrofluorometric assay was capable to detect nuclear changes caused by all tested apoptotic inducers. Then, we compared the outcomes of the spectrofluorometric assay with other methods detecting cell impairment and apoptosis (i.e. WST-1 and glutathione tests, TUNEL, DNA ladder, caspase activity, PARP-1 and JNKs expressions). We found that our developed spectrofluorometric assay provided results of the same sensitivity as the TUNEL assay but with the advantages of being fast processing, low-cost and a high throughput. Because nuclear condensation and fragmentation can be typical markers of cell death, especially in apoptosis, we suppose that the spectrofluorometric assay could become a routinely used method for characterizing cell death processes.

Artepillin C Inhibits Cell Proliferation by Cell Cycle Arrest at G0/G1 Phase Accompanied by Up-Regulation of p27Kip1 in Human Hepatoma HepG2 Cells

Artepillin C, 3, 5-diprenyl-4-hydroxycinnamic acid, is one of the bioactive constituents in Brazilian propolis. In the present study, the anticarcinogenic activity of this compound was investigated in human hepatoma HepG2 cells. Artepillin C inhibited the cell proliferation in a dose- and time-dependent manner accompanied by G0/G1 phase arrest in the cell cycle. This compound caused a decrease in the phosphorylation levels of the retinoblastoma protein at Ser780 and Ser807/811 and a decrease in the kinase activity of the cyclinD and CDK4 complex without any change in these protein levels. Artepillin C increased the protein level of p27Kip1, known as a CDK inhibitor. This up-regulation was regulated by both the transcriptional and post-transcriptional levels, i.e., the treatment increased the mRNA of p27Kip1 and decreased the proteosome activity. Thus, artepillin C induces cell cycle arrest at G0/G1 phase accompanied by up-regulation of p27Kip1, resulting in the inhibition of cell proliferation in HepG2 cells. This study suggested that artepillin C will be a promising anti-cancer agent against hepatoma cancer.