Bursatella leachii purple ink secretion extract exerts cytotoxic properties against human hepatocarcinoma cell line (HepG2): In vitro and in silico studies

Liver cancer is the third leading cause of cancer death worldwide. Marine mollusc-derived extracts have gained attention as new potential natural-based anticancer agents to overcome the side effects caused by conventional chemotherapeutic drugs during cancer therapy. We evaluated the cytotoxic effects of a crude extract from the purple-ink released by the sea hare named Bursatella leachii (B. leachii) against human hepatocarcinoma cell line (HepG2) and explored the underlying mechanisms causing the programmed cell death (i.e., apoptosis). Expression of cleaved-caspase-8 and cleaved-caspase-3, key cysteine-aspartic proteases involved in the initiation and completion of the apoptosis process, appeared after HepG2 cell exposure to B. leachii extract. Gene expression levels of pro-apoptotic BAX, tumour suppressor TP53 and Cyclin D1 were increased after treatment with B. leachii. Using liquid chromatography-mass spectrometry, the main biomolecules in the B. leachii extract were identified as hectochlorin, malyngamide X, malyngamide S, bursatellin, and lyngbyatoxin A. Applying in silico approaches, the high scores predicted bioactivities for the five compounds were protease and kinase inhibitors. The ADME and cytochrome profiles for the compounds were also predicted. Altogether, the cytotoxic B. leachii extract presents high pro-apoptotic potentials, suggesting it as a promising safe natural product-based drug for the treatment of liver cancer.

Determination of Perflourooctanoic Acid Toxicity in a Human Hepatocarcinoma Cell Line

Background. Perfluorooctanoic acid (PFOA) is used in different industrial and commercial products. Research shows the presence of PFOA in home dusts, tap and surface water, and in biological samples. The International Agency for Research on Cancer (IARC) has classified PFOA as a possible carcinogen for humans. The liver is thought to be a target organ of PFOA accumulation and toxicity. Objective. Some studies have found toxic effects on the liver and related mechanisms; however, more studies are needed to better understand PFOA - induced hepatotoxicity. Methods. In the present study, a human hepatocarcinoma cell line was exposed to PFOA for 24 hours and cell viability, apoptosis, the oxidative system and immune response were evaluated. Results. While apoptosis was the main cell death pathway at low concentration (86.5%), the necrotic cell fraction increased with higher concentrations (46.7%). Significant changes in the reactive oxygen species (5.3-folds) glutathione (GSH) (1.7-folds) and catalase (CAT) (1.4-folds) levels were observed, as well as changes to interleukin-6 (≤1.8-fold) and interleukin-8 levels (35–40%). Conclusions. In light of the data, PFOA is potentially hepatotoxic through the investigated pathways. The results represent a background for future in vivo mechanistic studies. Competing Interests. The authors declare no competing financial interests.

Bioactivity Profiles of Cytoprotective Short-Chain Quinones

Short-chain quinones (SCQs) have been investigated as potential therapeutic candidates against mitochondrial dysfunction, which was largely thought to be associated with the reversible redox characteristics of their active quinone core. We recently reported a library of SCQs, some of which showed potent cytoprotective activity against the mitochondrial complex I inhibitor rotenone in the human hepatocarcinoma cell line HepG2. To better characterize the cytoprotection of SCQs at a molecular level, a bioactivity profile for 103 SCQs with different compound chemistries was generated that included metabolism related markers, redox activity, expression of cytoprotective proteins and oxidative damage. Of all the tested endpoints, a positive correlation with cytoprotection by SCQs in the presence of rotenone was only observed for the NAD(P)H:quinone oxidoreductase 1 (NQO1)-dependent reduction of SCQs, which also correlated with an acute rescue of ATP levels. The results of this study suggest an unexpected mode of action for SCQs that appears to involve a modification of NQO1-dependent signaling rather than a protective effect by the reduced quinone itself. This finding presents a new selection strategy to identify and develop the most promising compounds towards their clinical use.

Behavioral Changes in Stem-Cell Potency by HepG2-Exhausted Medium

Wharton jelly mesenchymal stem cells (WJ-MSCs) are able to differentiate into different cell lineages upon stimulation. This ability is closely related to the perfect balance between the pluripotency-related genes, which control stem-cell proliferation, and genes able to orchestrate the appearance of a specific phenotype. Here we studied the expression of stemness-related genes, epigenetic regulators (DNMT1, SIRT1), miRNAs (miR-145, miR-148, and miR-185) related to stemness, exosomes, the cell-cycle regulators p21 (WAF1/CIP1) and p53, and the senescence-associated genes (p16, p19, and hTERT). Cells were cultured in the presence or absence of the human hepatocarcinoma cell line HepG2-exhausted medium, to evaluate changes in stemness, differentiation capability, and senescence sensibility. Our results showed the overexpression of SIRT1 and reduced levels of p21 mRNA. Moreover, we observed a downregulation of DNMT1, and a simultaneous overexpression of Oct-4 and c-Myc. These findings suggest that WJ-MSCs are more likely to retain a stem phenotype and sometimes to switch to a highly undifferentiable proliferative-like behavior if treated with medium exhausted by human HepG2 cell lines.

Comparative In Vitro Toxicology of Novel Cytoprotective Short-Chain Naphthoquinones

Short-chain quinones (SCQs) have been identified as potential drug candidates against mitochondrial dysfunction, which largely depends on the reversible redox characteristics of the active quinone core. We recently identified 11 naphthoquinone derivatives, 1–11, from a library of SCQs that demonstrated enhanced cytoprotection and improved metabolic stability compared to the clinically used benzoquinone idebenone. Since the toxicity properties of our promising SCQs were unknown, this study developed multiplex methods and generated detailed toxicity profiles from 11 endpoint measurements using the human hepatocarcinoma cell line HepG2. Overall, the toxicity profiles were largely comparable across different assays, with simple standard assays showing increased sensitivity compared to commercial toxicity assays. Within the 11 naphthoquinones tested, the L-phenylalanine derivative 4 consistently demonstrated the lowest toxicity across all assays. The results of this study not only provide useful information about the toxicity features of SCQs but will also enable the progression of the most promising drug candidates towards their clinical use.

Chemical Constituents and Their Activities From the Twigs of Euscaphis konishii Hayata

A new ellagic acid derivative 3,3′-di- O-methylellagic acid 4 ′-α-l-arabinopyranoside (1), with 9 known compounds identified as 3,3′-di- O-methylellagic acid (2), 3,3′-di- O-methylellagic acid 4′-α-d-arabinofuranoside (3), 3,3′-di- O-methylellagic acid 4′ -β-d-glucopyranoside (4), 3,3′-di- O-methylellagic acid 4 ′-β-d-xylopyranoglucoside (5), 3,3′,4-tri- O-methylellagic acid 4′-β-d-glucopyranoside (6), tormentic acid (7), ursolic acid (8), euscaphic acid (9), and betulinic acid (10), was isolated from the twigs of Euscaphis konishii Hayata. Compounds 1, 3, and 5 to 7 were isolated from this plant for the first time, and compounds 1 and 5 were obtained from the plant genus for the first time. The structure of the new compound was confirmed by HRESIMS, NMR, and compared with data from the literature . The cytotoxicities of 10 isolated compounds were tested, with compounds 1 to 6 showing moderately inhibited activity against the Human Hepatocarcinoma cell line (HepG2 cells) with an IC50 value ranging from 69.7 to 181.8 μM.

Mytoxin B and Myrothecine A Induce Apoptosis in Human Hepatocarcinoma Cell Line SMMC-7721 via PI3K/Akt Signaling Pathway

Trichothecene macrolides comprise a class of valuable leading compounds in developing anticancer drugs, however, there are few reports concerning their anticancer mechanisms, especially the anticancer mechanism of the 10,13-cyclotrichothecane derivatives that are found mainly in symbiotic fungi. In vitro anticancer activity of two trichothecene macrolides mytoxin B and myrothecine A against the human hepatocarcinoma cell line SMMC-7721 was investigated in the present study. MTT assay showed that mytoxin B and myrothecine A inhibited the proliferation of SMMC-7721 cells in dose- and time-dependent manners. Annexin V-FITC/PI dual staining assay revealed that mytoxin B and myrothecine A both could induce SMMC-7721 cells apoptosis in a dose-dependent manner. The decreased expression level of anti-apoptotic protein Bcl-2 and the increased expression level of pro-apoptotic protein Bax were observed apparently in Western blot analysis. The reduced ratio of Bcl-2/Bax further confirmed the apoptosis-inducing effect of mytoxin B and myrothecine A on SMMC-7721 cells. Moreover, the expression levels of caspases-3, -8, and -9, and cleaved caspases-3, -8, and -9 were all upregulated in both mytoxin B and myrothecine A-treated cells in Western blot analysis, which indicated that both compounds might induce SMMC-7721 cells apoptosis through not only the death receptor pathway but also the mitochondrial pathway. Finally, mytoxin B and myrothecine A were found to reduce the activity of PI3K/Akt signaling pathway that was similar to the effect of LY294002 (a potent and specific PI3K inhibitor), suggesting that both mytoxin B and myrothecine A might induce SMMC-7721 cells apoptosis via PI3K/Akt pathway.

Annexin A2 Enhances the Progression of Colorectal Cancer and Hepatocarcinoma via Cytoskeleton Structural Rearrangements

Annexin A2 (ANXA2) is reported to be associated with cancer development. To investigate the roles ANXA2 plays during the development of cancer, the RNAi method was used to inhibit the ANXA2 expression in caco2 (human colorectal cancer cell line) and SMMC7721 (human hepatocarcinoma cell line) cells. The results showed that when the expression of ANXA2 was efficiently inhibited, the growth and motility of both cell lines were significantly decreased, and the development of the motility relevant microstructures, such as pseudopodia, filopodia, and the polymerization of microfilaments and microtubules were obviously inhibited. The cancer cell apoptosis was enhanced without obvious significance. The possible regulating pathway in the process was also predicted and discussed. Our results suggested that ANXA2 plays important roles in maintaining the malignancy of colorectal and hepatic cancer by enhancing the cell proliferation, motility, and development of the motility associated microstructures of cancer cells based on a possible complicated signal pathway.

Inhibitory and Inductive Effects of Opuntia ficus indica Extract and Its Flavonoid Constituents on Cytochrome P450s and UDP-Glucuronosyltransferases

Opuntia ficus indica (OFI) is grown abundantly in arid areas and its fruits are regarded as an important food and nutrient source owing to the presence of flavonoids, minerals, and proteins. The previous report that OFI exerts phytoestrogenic activity makes it plausible for OFI-containing supplements to be used as alternative estrogen replacement therapy. In the case of polypharmacy with the consumption of OFI-containing botanicals in post- or peri-menopausal women, it is critical to determine the potential drug-OFI interaction due to the modulation of drug metabolism. In the present study, the modulating effects on the hepatic drug metabolizing enzymes (DMEs) by OFI and its flavonoid constituents (kaempferol, quercetin, isorhamnetin, and their glycosidic forms) were investigated using the liver microsomal fractions prepared from ovariectomized (OVX) rats, human liver microsomes, and human hepatocarcinoma cell line (HepG2). As a result, the oral administration of extracts of OFI (OFIE) in OVX rats induced hepatic CYP2B1, CYP3A1, and UGT2B1. OFIE, hydrolyzed (hdl) OFIE, and several flavonols induced the transcriptional activities of both CYP2B6 and CYP3A4 genes in HepG2 cells. Finally, OFIE did not inhibit activities of cytochrome P450 (CYPs) or uridine diphosphate (UDP)-glucuronosyltransferases (UGTs), whereas hdl OFIE or flavonol treatment inhibited CYP1A2 and CYP3A1/3A4 in rat and human liver microsomes. Our data demonstrate that OFIE may induce or inhibit certain types of DMEs and indicate that drug-OFI interaction may occur when the substrate or inhibitor drugs of specific CYPs or UGTs are taken concomitantly with OFI-containing products.