OBJECTIVE:To study the inhibitory effect of Ganoderma lucidum, the extract of chloroform, the extract of ethyl acetate and the remains after two-time extraction on BEL-7402 and MGC-803 cells and their protective effects on HL-7702 cells pre-and post-exposed to cisplatin (DDP) and various doses of 60Co gamma irradiation.

METHOD:The antitumor activity and protective effects on damaged HL-7702 cells induced by radiotherapy and chemotherapy of ganoderma lucidum were determined by MTT technique.

RESULT:The anticancer activity of the extract of chloroform Ganoderma lucidum was the best: at the concentration of 0.125 mg x mL(-1), the inhibitory rate was over 50%. To the HL-7702 cells damaged by DDP, four kinds of extracts didn't exert restoring effect, but the pretreatment with the extract of chloroform reduced the damaged degree significantly. To the 60Co gamma irradiated HL-7702 cells, only the extract of chloroform exerted restoring effect to some extent when exposed to middle or high dose of irradiation. The pre-administration of four kinds of extracts reduced the damaged degree by radiation.

CONCLUSION:The extract of chloroform exerts notable antitumor effects on cancer cells and protective effects on damaged normal cells induced by radiotherapy and chemotherapy.

Hepatocellular carcinoma is a cancer of high mortality; therefore, the effective therapy on this cancer is an imperative issue. Recently, anticancer agent combined with natural products has been demonstrated to increase apoptosis of various cancer cells effectively. Accordingly, we investigated the apoptotic effect and possible mechanism of the ethanol extract from Taiwanofungus salmoneus (=Antrodia salmonea) mycelium (TsE) alone or in combination with cisplatin in SK-Hep-1 cells. In this study, the proliferation of SK-Hep-1 cells could be inhibited at various concentrations of TsE for 24 h whereas TsE combined with cisplatin would inhibit the cell proliferation more notably. Moreover, the DNA damage and the interruption of cell cycle of SK-Hep-1 cells would be effectively raised after incubation with TsE combined with cisplatin for 24 h. The apoptosis of cells was dramatically induced, and the expression of caspases 3, 8, and 9, apoptosis-related protein, were significantly upregulated. Therefore, we proposed that the TsE combined with cisplatin inhibited cell proliferation by elevating sub-G1 phase, inducing DNA damage, activating caspases 3, 8, and 9 activities, and triggering cells apoptosis. These results reveal that TsE could be a potential adjuvant chemotherapeutic agent.

Hepatocarcinoma cells (TLT) were incubated in the presence of ascorbate and menadione, either alone or in combination. Cell death was only observed when such compounds were added simultaneously, most probably due to hydrogen peroxide (H2O2) generated by ascorbate-driven menadione redox cycling. TLT cells were particularly sensitive to such an oxidative stress due to its poor antioxidant status. DNA strand breaks were induced by this association but this process did not correspond to oligosomal DNA fragmentation (a hallmark of cell death by apoptosis). Neither caspase-3-like DEVDase activity, nor processing of procaspase-3 and cleavage of poly(ADP-ribose) polymerase (PARP) were observed in the presence of ascorbate and menadione. Cell death induced by such an association was actively dependent on protein phosphorylation since it was totally prevented by preincubating cells with sodium orthovanadate, a tyrosine phosphatase inhibitor. Finally, while H2O2, when administered as a bolus, strongly enhances a constitutive basal NF-kappaB activity in TLT cells, their incubation in the presence of ascorbate and menadione results in a total abolition of such a constitutive activity.

This study investigates the anticancer properties of cannabisin B, purified from hempseed hull, in HepG2 human hepatoblastoma cells. The results indicate that cannabisin B significantly inhibited cell proliferation by inducing autophagic cell death rather than typical apoptosis. Cell viability transiently increased upon the addition of a low concentration of cannabisin B but decreased upon the addition of high concentrations. Cannabisin B-induced changes in cell viability were completely inhibited by pre-treatment with 3-methyladenine (3-MA), indicating that the induction of autophagy by cannabisin B caused cell death. Additionally, cannabisin B induced S phase cell cycle arrest in a dose-dependent manner. Moreover, cannabisin B was found to inhibit survival signaling by blocking the activation of AKT and down-stream targets of the mammalian target of rapamycin (mTOR). These findings suggest that cannabisin B possesses considerable antiproliferative activity and that it may be utilised as a promising chemopreventive agent against hepatoblastoma disease.

INTRODUCTION: Homeopathy is a popular form of complementary and alternative medicine and is used to treat for certain liver ailments. AIM: To analyze the efficacy of homeopathic Chelidonium majus (Chel) 30C and 200C in amelioration of experimentally induced hepato-toxicity in rats. METHODS: Rats were randomized into six sub-groups: negative control; negative control+EtOH; positive control; positive control+EtOH group; Chel 30; Chel 200. Rats were sacrificed at day 30, 60, 90 and 120; various toxicity biomarkers and pathological parameters were evaluated. Gelatin zymography for determination of metalloproteinases activity and Western blot of p53 and Bcl-2 proteins were also employed. All analyses were observer blind. RESULTS: Chronic feeding of p-dimethyl amino azo benzene (p-DAB) and phenobarbital (PB) elevated the levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), gamma glutamyl transferase (GGT), lactate dehydrogenase (LDH), triglyceride, cholesterol, creatinine and bilirubin and lowered the levels of glutathione (GSH), glucose-6-phosphate dehydrogenase (G-6-PD), catalase and HDL-cholesterol. There were statistically significant modulations of these parameters in the treated animals, compared to positive controls. In both treated groups, there was downregulation of metalloproteinases, p53 and Bcl-2 proteins compared to over-expression in the positive control groups. CONCLUSION: Both the potencies of Chel exhibited anti-tumor and anti-oxidative stress potential against artificially induced hepatic tumors and hepato-toxicity in rats. More studies are warranted.

The antiproliferative effect and apoptosis-inducing action of 5-fluorouracil (5-FU) in combination with vitamin C were tested in vitro against the chemosensitive mouse lymphoma, the chemoresistant HEp-2 and a human lung fibroblast cell line. Vitamin C itself had no antiproliferative effect on the fibroblasts, but increased the anticancer effect of 5-FU dose-dependently. In the case of the chemoresistant cell line, only a high concentration of vitamin C increased the cytotoxicity of 5-FU. A combination of 5-FU and vitamin C exerted a significantly enhanced apoptotic effect on the mouse lymphoma cell line, whereas for the HEp-2 cell line this effect was less marked and was achieved only at a high concentration of vitamin C. These findings suggest that the administration of a high dose of vitamin C in combination with 5-FU chemotherapy enhances the chemoresponsiveness of cancer cells and serves as a potential sensitizer, especially in chemo-resistant cell lines. One of the mechanisms by which vitamin C potentiates cytostatics could be apoptosis induction.

BACKGROUND:Mushrooms have been valued for their nutritive content and as traditional medicines; several important medicinal properties of mushrooms have been recognized worldwide.

OBJECTIVE:The purpose of this study was to elucidate the cell growth inhibitory potential of four edible mushrooms; Coprinus comatus (O.F. Mull.) Pers. (Agaricaceae), Tricholoma fracticum (Britzelm.) Kreisel (Tricholomataceae), Rhizopogon luteolus Fr. and Nordholm (Rhizopogonaceae), Lentinus tigrinus (Bull.) Fr. (Polyporaceae) on hepatocellular carcinoma (HepG2) cells in conjunction with their antioxidant and antibacterial capacities.

MATERIALS AND METHODS:Five different extracts of edible mushrooms were obtained using water, methanol, acetone, n-hexane and chloroform as solvent systems for cytotoxic, antioxidant and antibacterial properties.

RESULTS:C. comatus showed substantial in vitro cytotoxic activity against HepG2 cell lines with all extracts especially with chloroform 50% inhibition (IC50 value of 0.086 mg/ml) and acetone (IC50 value of 0.420 mg/ml). Chloroform extract of C. comatus had maximum amount ofβ-carotene (25.94 μg/mg), total phenolic content (76.32 μg/mg) and lycopene (12.00 μg/mg), and n-hexane extract of L. tigrinus had maximum amount of flavonoid (3.67 μg/mg). While chloroform extract of C. comatus showed the highest 2, 2-diphenyl-1-picrylhydrazyl (DPPH) capturing activity (1.579mg/ml), the best result for metal chelating activity was obtained from methanolic extract (0.842 mg/ml). Moreover, all tested mushrooms demonstrated antibacterial activity and n-hexane extract of L. tigrinus and acetone extracts of T. fracticum were the most active against tested microorganism.

CONCLUSION:These results indicate that different extracts of investigated mushroom have considerable cytotoxic, antioxidant and antibacterial properties and may be utilized as a promising source of therapeutics.

Recent studies have demonstrated that vegetable rich diets have protective effects on the occurrence and prognosis of various cancers. In addition to dietary intakes, ascorbic acid and β-carotene are also taken as supplements. The aim of this study was to assess effects of ascorbic acid, β-carotene and their combinations on human hepatocellular carcinoma cell line HepG2. Ascorbic acid and β-carotene were applied to cells as plasma peak concentrations (70 and 8 μM, respectively) and their half concentrations (35 and 4 μM, respectively) for 24 and 48 h. Genotoxic and cytotoxic effects of ascorbic acid and β-carotene were evaluated by alkali single cell gel electrophoresis (SCGE), acridine orange/ethidium bromide staining patterns of cells (apoptosis and necrosis) and lipid peroxidation (thiobarbituric acid reactive substances, TBARS). Results of the SCGE demonstrated that both ascorbic acid and β-carotene caused DNA damage on HepG2 which were also concordant to increased apoptosis and necrosis of cells. Increased TBARS values also demonstrated increased lipid peroxidation in these cells. Results of the present study demonstrates that when dietary intakes of ascorbic acid and β-carotene and their relevant achievable plasma level concentrations were considered, both ascorbic acid and β-carotene induce genotoxic and cytotoxic damage on HepG2 together with increased oxidative damage in contrast to their protective effect on healthy cells. This may be correlated to oxidative status and balance of ROS in hepatocellular carcinoma cells.

OBJECTIVE: To observe the effects of Ganoderma lucidum Spores (GLS) on proliferation and growth cycle in the human hepatoma cell line (HepG2 cells), and study its possible mechanism of activities. METHODS: The growth inhibition of GLS on HepG2 cells was detected by MTT assay. The DNA contents and the distribution of cell cycle were analyzed by flow cytometry. RESULTS: The results of MTT assays showed that GLS could inhibit the HepG2 cells growth at a dose and time-dependent manner directly; the inhibition rate of GLS (2500 microg/ml) on HepG2 cells after 72 h was a maximum up 51.4%. The results of flow cytometry experiments showed that GLS (3 mg/ml) could reduce the G2 phase of HepG2 cells and a clear apoptosis peak would be observed when GLS was 6 mg/ml. CONCLUSION: GLS has a direct inhibitory effect on tumor cell proliferation and its growth cycle, it can reduce the G2 phase; and high doses of GLS can also make tumor cells apoptosised.

The present paper assesses the modifying potential of black pepper on the hepatic biotransformation system in mice. The modulatory effect was assessed on glutathione S-transferase (GST), cytochrome b5 (cyt. b5), cytochrome P-450 (cyt. P-450), acid-soluble sulfhydryl (-SH) content and malondialdehyde (MDA) level. Swiss albino mice of either sex (eight weeks old) were fed on a diet containing 0.5%, 1% and 2% black pepper (w/w) for 10 and 20 days. The findings revealed a significant and dose-dependent increase in GST and -SH content in the experimental groups except the one maintained on 0.5% black pepper diet for 10 days. Elevated levels of cyt. b5 and cyt. P-450 were also statistically significant and dose-dependent. The level of MDA was lowered in the group fed on 2% black pepper diet for 20 days. Being a potential inducer of detoxication system, the possible chemopreventive role of black pepper in chemical carcinogenesis is suggested.

Hepatocellular carcinoma (HCC) has a poor outcome. Most HCCs develop in the context of liver fibrosis and cirrhosis caused by chronic inflammation. Short-term fasting approaches enhance the activity of chemotherapy in preclinical cancer models, other than HCC. Multi-tyrosine kinase inhibitor Sorafenib is the mainstay of treatment in HCC. However, its benefit is frequently short-lived. Whether fasting can alleviate liver fibrosis and whether combining fasting with Sorafenib is beneficial remains unknown. A 24 hr fasting (2% serum, 0.1% glucose)-induced changes on human hepatic stellate cells (HSC) LX-2 proliferation/viability/cell cycle were assessed by MTT and flow cytometry. Expression of lypolysaccharide (LPS)-induced activation markers (vimentin, αSMA) was evaluated by qPCR and immunoblotting. Liver fibrosis and inflammation were evaluated in a mouse model of steatohepatitis exposed to cycles of fasting, by histological and biochemical analyses. A 24 hr fasting-induced changes were also analyzed on the proliferation/viability/glucose uptake of human HCC cells exposed to Sorafenib. An expression panel of genes involved in survival, inflammation, and metabolism was examined by qPCR in HCC cells exposed to fasting and/or Sorafenib. Fasting decreased the proliferation and the activation of HSC. Repeated cycles of short term starvation were safe in mice but did not improve fibrosis. Fasting synergized with Sorafenib in hampering HCC cell growth and glucose uptake. Finally, fasting normalized the expression levels of genes which are commonly altered by Sorafenib in HCC cells. Fasting or fasting-mimicking diet diets should be evaluated in preclinical studies as a mean to potentiate the activity of Sorafenib in clinical use.

Liver resection is the treatment of choice for resectable hepatic metastases; however, most patients have unresectable disease when diagnosed. Hepatic cryotherapy has been advocated to treat unresectable tumours in the liver although its precise role is still being evaluated. This article discusses mechanisms of action, technical considerations, current indications and the early results of cryotherapy in treating metastatic liver disease.

It has been demonstrated that the Hericium erinaceus (HE) mushroom, which primarily consists of polysaccharides, possesses anti-tumor activities. However, the mechanisms by which HE inhibits human hepatocellular carcinoma growth remain unknown. Our study demonstrates that HE acts as an enhancer to sensitize doxorubicin (Dox)-mediated apoptotic signaling, and this sensitization can be achieved by reducing c-FLIP expression via JNK activation and enhancing intracellular Dox accumulation via the inhibition of NF-κB activity. These findings suggest that HE in combination with Dox serves as an effective tool for treating drug-resistant human hepatocellular carcinoma.

The study was undertaken to examine whether Carcinosin-200 (Car-200) could provide additional ameliorative effect, if used intermittently with Natrum sulphuricum-30 (Nat Sulph-30) against hepatocarcinogenesis induced by chronic feeding of p-dimethylaminoazobenzene (p-DAB) and phenobarbital (PB) in mice (Mus mnusculus). Mice were randomly divided into seven sub-groups: (i) normal untreated; (ii) normal + succussed alcohol; (iii) p-DAB (0.06%) + PB (0.05%); (iv) p-DAB + PB + succussed alcohol, (v) p-DAB + PB + Nat Sulph-30, (vi) p-DAB + PB + Car-200, and (vii) p-DAB + PB + Nat Sulph-30 + Car-200. They were sacrificed at 30, 60, 90 and 120 days for assessment of genotoxicity through cytogenetical end-points like chromosome aberrations, micronuclei, mitotic index and sperm head anomaly and cytotoxicity through assay of widely accepted biomarkers and pathophysiological parameters. Additionally, electron microscopic studies and gelatin zymography for matrix metalloproteinases (MMPs) were conducted in liver at 90 and 120 days. Results showed that administration of Nat Sulph-30 alone and in combination with Car-200 reduced the liver tumors with positive ultrastructural changes and in MMPs expression, genotoxic parameters, lipid peroxidation, gamma-glutamyl transferase, lactate dehydrogenase, blood glucose, bilirubin, creatinine, urea and increased GSH, glucose-6-phosphate dehydrogenasc, superoxide dismutase, catalase, glutathione reductase activities and hemoglobin, cholesterol, and albumin levels. Thus, intermittent use of Car-200 along with Nat Sulph-30 yielded additional benefit against genotoxicity, cytotoxicity, hepatotoxicity and oxidative stress induced by the carcinogens during hepatocarcinogenesis.

UNLABELLED:Long-term survival of patients with hepatocellular carcinoma (HCC), a common cancer worldwide, remains poor, due to metastasis and recurrence.

AIM:To investigate the effect of a novel nutrient mixture (NM) containing ascorbic acid, lysine, proline, and green tea extract on human HCC cell line Sk-Hep-1 In vivo and In vitro.

METHODS:After one week of isolation, 5-6 week old male athymic nude mice were inoculated with 3 x 10(6) SK-Hep-1 cells subcutaneously and randomly divided into two groups; group A was fed a regular diet and group B a regular diet supplemented with 0.5% NM. Four weeks later, the mice were sacrificed and their tumors were excised, weighed and processed for histology. We also tested the effect of NM In vitro on SK-Hep-1 cells, measuring cell proliferation by MTT assay, invasion through Matrigel, apoptosis by green caspase detection kit, MMP secretion by zymography, and morphology by H&E staining.

RESULTS:NM inhibited tumor weight and burden of SK-Hep-1 xenografts by 42% and 33% respectively. In vitro , NM exhibited 33% toxicity over the control at 500 and 1,000 microg/ml concentration. Zymography demonstrated MMP-2 and MMP-9 secretion which was inhibited by NM in a dose dependent fashion, with virtual total inhibition at 1000 microg/ml. Invasion through Matrigel was inhibited at 100, 500 and 1,000 microg/ml by 53%, 83% and 100% respectively. NM induced slight apoptosis at 100 microg/ml, and profound apoptosis at 500 microg/ml and 1000 microg/ml concentration.

CONCLUSIONS:These results suggest that NM has therapeutic potential in treatment of HCC.

The present study was conducted to determine whether low intensity-pulsed ultrasound (LIPUS) could induce apoptosis of human hepatocellular carcinoma cells, SMMC-7721, and to define the mechanism of ultrasound-induced apoptosis, in vitro. MTT assay was used to measure cell proliferation. Apoptosis was investigated by multiple methods such as flow cytometry, DNA fragmentation, Ca(2+) mobilizations, pro- and anti-apoptotic protein expression, and light as well as ultramicroscopic morphology. The results provide evidence that LIPUS induced a dose-dependent effect on cell viability and apoptosis of SMMC-7721 cells. Specifically, exposure of cells to>0.5W/cm(2) intensity significantly increased cell apoptosis, caused shifts in cell cycle phase, and induced structural changes. Ultrasound significantly increased intracellular Ca(2+) concentrations and modulated expression of caspase-3, Bcl-2 and Bax. The findings suggest that this novel technology can be used to induce SMMC-7721 apoptosis via the Ca(2+)/mitochondrial pathway and could potentially be of clinical use for the treatment of hepatocellular carcinoma (SMMC-7721 cell line) and other cancers.

BACKGROUND/AIM:Curcumin (CUM) is a promising agent in complementary oncology. The present study analyzed the photoactive properties of curcumin on pediatric epithelial liver tumor cell lines.

MATERIALS AND METHODS:Hepatoblastoma cell lines (HuH6, HepT1) and hepatocellular carcinoma cell lines (HepG2, HC-AFW1) were treated with curcumin and exposed to blue light (phototherapy, 480 nm, 300 W). Cell viability (MTT tests), cellular oxidative stress (production of reactive oxygen species (ROS)) and cellular uptake/degradation of curcumin were analyzed.

RESULTS:Significant loss of viability resulted from 24-48 h incubation with curcumin. With photodynamic therapy (PDT), even short time incubation (1 h) with curcumin resulted in significantly lower half maximal inhibitory concentration (IC50) (p<0.001, two-way ANOVA). Significant ROS production was observed with PDT and curcumin.

CONCLUSION:Phototherapy strongly enhances the anticancer properties of curcumin in pediatric solid liver tumors in vitro.

BACKGROUND/AIM:Curcumin (CUM) is a promising agent in complementary oncology. The present study analyzed the photoactive properties of curcumin on pediatric epithelial liver tumor cell lines.

MATERIALS AND METHODS:Hepatoblastoma cell lines (HuH6, HepT1) and hepatocellular carcinoma cell lines (HepG2, HC-AFW1) were treated with curcumin and exposed to blue light (phototherapy, 480 nm, 300 W). Cell viability (MTT tests), cellular oxidative stress (production of reactive oxygen species (ROS)) and cellular uptake/degradation of curcumin were analyzed.

RESULTS:Significant loss of viability resulted from 24-48 h incubation with curcumin. With photodynamic therapy (PDT), even short time incubation (1 h) with curcumin resulted in significantly lower half maximal inhibitory concentration (IC50) (p<0.001, two-way ANOVA). Significant ROS production was observed with PDT and curcumin.

CONCLUSION:Phototherapy strongly enhances the anticancer properties of curcumin in pediatric solid liver tumors in vitro.

polysaccharide (GLP) is a well-known traditional Chinese medicine, known for its anti-cancer and immunomodulatory properties. The present study aims to investigate whether GLP has a therapeutic effect on hepatocellular carcinoma (HCC) cells exposed to radiation. Immunofluorescence was used to detect the nuclei, the protein expression was measured by western blot analysis and flow cytometry was used to detect the rate of cell apoptosis. GLP treatment was demonstrated to enhance radiation-induced growth inhibition and apoptotic death of HCC cells. At a molecular level, GLP suppressed the activities of DNA repair-associated proteins including ataxia-telangiectasia mutated (ATM) and DNA dependent-protein kinase (DNA-PK) in liver cancer cells under radiation conditions. Furthermore, the addition of an Akt inhibitor elevated the activities of DNA-PK and ATM and attenuated the GLP-induced HepG2 cell injury under the radiation condition. In conclusion, the present study demonstrates that GLP enhances the radiosensitivity of HCC cells via the regulation of Akt signaling pathways, implying a potential therapeutic effect of GLP as a radiation sensitizer in HCC treatment.