Label-retaining liver cancer cells are relatively resistant to sorafenib.

OBJECTIVE: The standard therapy for advanced hepatocellular carcinoma (HCC) is sorafenib, with most patients experiencing disease progression within 6 months. Label-retaining cancer cells (LRCC) represent a novel subpopulation of cancer stem cells (CSC). The objective was to test whether LRCC are resistant to sorafenib. METHODS: We tested human HCC derived LRCC and non-LRCC before and after treatment with sorafenib. RESULTS: LRCC derived from human HCC are relatively resistant to sorafenib. The proportion of LRCC in HCC cell lines is increased after sorafenib while the general population of cancer cells undergoes growth suppression. We show that LRCC demonstrate improved viability and toxicity profiles, and reduced apoptosis, over non-LRCC. We show that after treatment with sorafenib, LRCC upregulate the CSC marker aldehyde dehydrogenase 1 family, wingless-type MMTV-integration-site family, cell survival and proliferation genes, and downregulate apoptosis, cell cycle arrest, cell adhesion and stem cells differentiation genes. This phenomenon was accompanied by non-uniform activation of specific isoforms of the sorafenib target proteins extracellular-signal-regulated kinases and v-akt-murine-thymoma-viral-oncogene homologue (AKT) in LRCC but not in non-LRCC. A molecular pathway map for sorafenib treated LRCC is proposed. CONCLUSIONS: Our results suggest that HCC derived LRCC are relatively resistant to sorafenib. Since LRCC can generate tumours with as few as 10 cells, our data suggest a potential role for these cells in disease recurrence. Further investigation of this phenomenon might provide novel insights into cancer biology, cancer recurrence and drug resistance with important implications for the development of novel cancer therapies based on targeting LRCC.

Chemopreventive activity of sesquiterpene lactones (SLs) from yacon against TPA-induced Raji cells deformation.

Yacon is a medicinal plant used as a traditional medicine by the natives in South America. In Japan, it becomes popular as a health food. Sesquiterpene Lactones (SLs) from yacon leaves were investigated and the active SLs such as enhydrin, uvedalin and sonchifolin, bearing alpha-methylene-gamma-lactone and epoxides as the active functional groups, were identified by 1H-6000 MHz-NMR. Chemopreventive and cytotoxic activities were determined using different primary screening methods. In this study, all tested SLs strongly inhibited TPA-induced deformed of Raji cells. The IC50 values of yacon SLs from anti-deforming assay were 0.04-0.4 microM. Interestingly, yacon SLs showed more potential of chemo preventive activity than both curcumin and parthenolide. However, the cytotoxicity on Raji cells was observed at high concentration of yacon SLs. The degree of anti-deformation was ranked in order: enhydrin >uvedalin >sonchifolin >parthenolide >curcumin. As according to structure-activity relationship, the high activities of enhydrin, uvedalin and sonchifolin may be due to the 2-methyl-2-butenoate and its epoxide moiety.

Garlic constituent diallyl trisulfide induced apoptosis in MCF7 human breast cancer cells.

Identification of agents that are nontoxic but can delay onset and/or progression of breast cancer, which is the main leading cause of cancer-related deaths among women, is highly desirable. Garlic-derived organosulfur compounds (OSCs) have highly effective antitumor effects, but the mechanism has yet to be investigated. The aim of the present study was undertaken to examine the effect of diallyl trisulfide (DATS), a promising cancer chemopreventive constituent of garlic, on growth of two cell lines respectively, MCF-7 human breast cancer cells and nontumorigenic MCF-12a mammary epithelial cells. The effects of DATS were examined by MTT assay, clonogenic survival assay, ELISA based apoptotic assay, TUNEL assay, immunofluoresence staining, flow Cytometry, RT-PCR and western blot analysis. Garlic constituent diallyl trisulfide (DATS) suppresses viability of cultured MCF-7 and MCF-12a cells respectively by decreasing the percent of cells in G(2)/M and inducing apoptotic cell death. DATS-induced apoptosis was markedly elevated in MCF-7 cells compared with MCF-12a cells and this was correlated with elevated levels of cyclin B1. The results from semi-quantitative and real-time RT-PCR indicated that DATS-enhanced the expression levels of FAS and cyclin D1, but in contrast, downregulated the expression levels of Akt and Bcl-2. Furthermore, the DATS-induced apoptosis was correlated with induction of pro-apoptotic Bax protein and p53 protein expression was upregulated and translocation to nucleus in MCF-7 cells. Together, the results of the present study show, for the first time, that DATS administration might offer a novel strategy for the treatment of human breast cancer.

Ex vivo expansion of human hematopoietic stem cells by a small-molecule agonist of c-MPL.

OBJECTIVE: The signaling by thrombopoietin (TPO) via its receptor, c-MPL, plays a crucial role in the maintenance of hematopoietic stem cells (HSCs). Small-molecule c-MPL agonists have recently been shown to be beneficial in the treatment of thrombocytopenia. However, their effects on HSCs have not yet been explored. In this study, we evaluated the effects of NR-101, a novel small-molecule c-MPL agonist, on the ex vivo expansion of human cord blood (hCB) HSCs. MATERIALS AND METHODS: hCB CD34(+) or CD34(+)CD38(-) hematopoietic stem and progenitor cells were cultured for 7 days in the presence of thrombopoietin (TPO) or NR-101, and then subjected to flow cytometric analyses, colony-forming cell assays, and severe combined immunodeficiency-repopulating cell assays. RESULTS: During a 7-day culture of CD34(+) or CD34(+)CD38(-) hematopoietic stem and progenitor cells, NR-101 efficiently increased their numbers, with a greater than twofold increase compared to TPO, although its effect on megakaryocytopoiesis was comparable to that of TPO. Correspondingly, severe combined immunodeficiency-repopulating cells were increased 2.9-fold during a 7-day culture with NR-101 compared to freshly isolated CD34(+) cells, and 2.3-fold compared to that with TPO. Of note, NR-101 persistently activated signal transducer and activator of transcription (STAT) 5 but not signal transducer and activator of transcription 3. Furthermore, NR-101 induced a long-term accumulation of hypoxia-inducible factor-1alpha protein and enhanced activation of its downstream target genes. CONCLUSION: This is the first time that a small-molecule c-MPL agonist has been demonstrated to promote net expansion of HSCs. NR-101 is more efficient in ex vivo expansion of HSCs than TPO. NR-101 could be a useful tool for the therapeutic manipulation of human HSCs.

Lipido-sterolic extract of Serenoa repens (LSESr, Permixon) treatment affects human prostate cancer cell membrane organization.

The molecular mechanism by which the lipido-sterolic extract of Serenoa repens (LSESr, Permixon) affects prostate cells remains to be fully elucidated. In androgen-independent PC3 prostate cancer cells, the LSESr-induced effects on proliferation and apoptosis were evaluated by counting cells and using a FACScan cytofluorimeter. PC3 cells were stained with JC-1 dye to detect mitochondrial membrane potential. Cell membrane lipid composition was evaluated by thin layer chromatography and gas chromatographic analysis. Akt phosphorylation was analyzed by Western blotting and cellular ultrastructure through electron microscopy. LSESr (12.5 and 25 microg/ml) administration exerted a biphasic action by both inhibiting proliferation and stimulating apoptosis. After 1 h, it caused a marked reduction in the mitochondrial potential, decreased cholesterol content and modified phospholipid composition. A decrease in phosphatidylinositol-4,5-bisphosphate (PIP2) level was coupled with reduced Akt phosphorylation. After 24 h, all of these effects were restored to pre-treatment conditions; however, the saturated (SFA)/unsaturated fatty acid (UFA) ratio increased, mainly due to a significant decrease in omega 6 content. The reduction in cholesterol content could be responsible for both membrane raft disruption and redistribution of signaling complexes, allowing for a decrease of PIP2 levels, reduction of Akt phosphorylation and apoptosis induction. The decrease in omega 6 content appears to be responsible for the prolonged and more consistent increase in the apoptosis rate and inhibition of proliferation observed after 2-3 days of LSESr treatment. In conclusion, LSESr administration results in complex changes in cell membrane organization and fluidity of prostate cancer cells that have progressed to hormone-independent status.

Effect of ellagic acid on proliferation, cell adhesion and apoptosis in SH-SY5Y human neuroblastoma cells.

Ellagic acid, a polyphenolic compound found in berries, fruits and nuts, has been shown to possess growth-inhibiting and apoptosis promoting activities in cancer cell lines in vitro. The objective of this study was to investigate the effect of ellagic acid in human neuroblastoma SH-SY5Y cells. In cultures of SH-SY5Y cells incubated with ellagic acid, time- and concentration-dependent inhibitory effects on cell number were demonstrated. Ellagic acid induced cell detachment, decreased cell viability and induced apoptosis as measured by DNA strand breaks. Ellagic acid-induced alterations in cell cycle were also observed. Simultaneous treatment with all-trans retinoic acid did not rescue the cells from ellagic acid effects. Furthermore, the results suggested that pre-treatment with all-trans retinoic acid to induce differentiation and cell cycle arrest did not rescue the cells from ellagic acid-induced cell death.

[New perspectives in the use of human hepatocytes in the preclinical drug development process]. / Nouvelles perspectives d'utilisation des hépatocytes humains au cours du développement préclinique des médicaments.

Human and animal hepatocytes are now widely used for drug metabolism and interaction studies in the drug development process. However, their phenotypic instability and the absence of cell division in primary culture limit their interest for toxicity studies. Hepatoma cell lines are also used but they express very low levels, if any, of cytochromes P450 (CYP) that are essential for metabolism of a number of drugs and other chemicals. A new human hepatoma cell line, named HepaRG, possesses both the metabolic capacity of human hepatocytes in primary culture and the indefinite proliferation potential of hepatoma cell lines. After two weeks of confluence HepaRG cells express the main CYP, phase 2 enzymes, plasma membrane transporters as well as the nuclear receptors such as pregnane X receptor (PXR) and constitutive androstane receptor (CAR). They can be maintained functionally relatively stable for several days or even a few weeks before being seeded and proliferating again, making them suitable for chronic toxicity and mutagenesis/carcinogenesis studies. Another in vitro model system is represented by extrahepatic stem cells but experimental culture conditions allowing their differentiation into mature hepatocytes have not been defined yet.

Caspase-independent death of human osteosarcoma cells by flavonoids is driven by p53-mediated mitochondrial stress and nuclear translocation of AIF and endonuclease G.

Flavonoids have antioxidant and antitumor promoting effects. Rhus verniciflua Stokes (RVS) is a flavonoid-rich herbal medicine that has long been used in Korea as both a food additive and antitumor agent. It was previous reported that a purified flavonoid fraction prepared from RVS, herein named RCMF (the RVS chloroform-methanol fraction), inhibited the proliferation and induced apoptosis in human osteosarcoma (HOS) cells. This study examined the mechanisms involved in the RCMF-mediated apoptosis in HOS cells. RCMF was shown to be capable of inducing apoptosis in HOS cells by inducing p53 in the cells resulting in the decrease in Bcl-2 level, activation of Bax, and cytoplasmic release of cytochrome c, which led to the translocation of apoptosis-inducing factor (AIF) and endonuclease G (EndoG) into the nucleus. However, the RCMF-induced apoptosis was suppressed by transfecting the cells with antisense p53 oligonucleotides but not by treating them with a MAPK or caspase inhibitor. This suppression occurred through the regulation of Bcl-2 members as well as by preventing the nuclear translocation of the mitochondrial apoptogenic factors. Overall, it appears that p53-mediated mitochondrial stress and the nuclear translocation of AIF and EndoG are mainly required for the apoptosis induced by RCMF.

DEP-1 protein tyrosine phosphatase inhibits proliferation and migration of colon carcinoma cells and is upregulated by protective nutrients.

The transmembrane protein-tyrosine phosphatase (PTP) DEP-1 (density-enhanced phosphatase) is a candidate tumor suppressor in the colon epithelium. We have explored the function of DEP-1 in colon epithelial cells by inducible re-expression in a DEP-1-deficient human colon cancer cell line. Density-enhanced phosphatase-1 re-expression led to profound inhibition of cell proliferation and cell migration, and was associated with cytoskeletal rearrangements. These effects were dependent on the PTP activity of DEP-1 as they were not observed with cells expressing the catalytically inactive DEP-1 C1239S variant. shRNA-mediated suppression of DEP-1 in a colon epithelial cell line with high endogenous DEP-1 levels enhanced proliferation, further supporting the antiproliferative function of DEP-1. Nutrients, which are considered to be chemoprotective with respect to colon cancer development, including butyrate, green tea and apple polyphenols, had the capacity to elevate transcription of endogenous DEP-1 mRNA and expression of DEP-1 protein. Upregulation of DEP-1 expression, and in turn inhibition of cell growth and migration may present a previously unrecognized mechanism of chemoprevention by nutrients.

Tea pigments induce cell-cycle arrest and apoptosis in HepG2 cells.

AIM: To investigate the molecular mechanisms by which tea pigments exert preventive effects on liver carcinogenesis. METHODS: HepG2 cells were seeded at a density of 5X10(5)/well in six-well culture dishes and incubated overnight. The cells then were treated with various concentrations of tea pigments over 3 d, harvested by trypsinization, and counted using a hemocytometer. Flow cytometric analysis was performed by a flow cytometer after propidium iodide labeling. Bcl-2 and p21(WAF1) proteins were determined by Western blotting. In addition, DNA laddering assay was performed on treated and untreated cultured HepG2 cells. RESULTS: Tea pigments inhibited the growth of HepG2 cells in a dose-dependent manner. Flow-cytometric analysis showed that tea pigments arrested cell cycle progression at G1 phase. DNA laddering was used to investigate apoptotic cell death, and the result showed that 100 mg/L of tea pigments caused typical DNA laddering. Our study also showed that tea pigments induced upregulation of p21(WAF1) protein and downregulation of Bcl-2 protein. CONCLUSION: Tea pigments induce cell-cycle arrest and apoptosis. Tea pigments may be used as an ideal chemopreventive agent.

Effects of Pinus massoniana bark extract on cell proliferation and apoptosis of human hepatoma BEL-7402 cells.

AIM: To study the effects of Pinus massoniana bark extract (PMBE) on cell proliferation and apoptosis of human hepatoma BEL-7402 cells and to elucidate its molecular mechanism. METHODS: BEL-7402 cells were incubated with various concentrations (20-200 microg/mL) of PMBE for different periods of time. After 48 h, cell proliferation was determined by 3-(4,5-dimethyl-thiazolyl-2)-2,5-diphenyl tetrazolium bromide (MTT) assay. Apoptosis was evaluated by morphological observation, agarose gel electrophoresis, and flow cytometry analysis. Possible molecular mechanisms were primarily explored through immunohistochemical staining. RESULTS: PMBE (20-200 microg/mL) significantly suppressed BEL-7402 cell proliferation in a time- and dose-dependent manner. After treatment of BEL-7402 cells with 160 microg/mL PMBE for 24, 48, or 72 h, a typical apoptotic "DNA ladder" was observed using agarose gel electrophoresis. Nuclear condensation and boundary aggregation or split, apoptotic bodies were seen by fluorescence and electron microscopy. Sub-G1 curves were displayed by flow cytometry analysis. PMBE decreased the expression levels of Bcl-2 protein in a time-dependent manner after treatment of cells with 160 microg/mL PMBE. CONCLUSION: PMBE suppresses proliferation of BEL-7402 cells in a time- and dose-dependent manner and induces cell apoptosis by possibly downregulating the expression of the bcl-2 gene.

Cooking oil fumes improve lung adenocarcinoma cell survival through c-IAP2 induction.

Cooking oil fumes (COF) exposure was demonstrated to be associated with lung cancer development in Taiwanese nonsmoking women. Previous studies identified Cox-2 overexpression and oxidative DNA damage in lung adenocarcinoma cells after exposure to COF. Involvement of COF in lung tumorigenesis may be associated with cell survival, as well as proliferation of lung adenocarcinoma. To test this hypothesis, A549, a lung adenocarcinoma cell line, was used, and MTT assay data showed that the cell viability of A549 was significantly increased in a concentration-dependent manner by COF treatment for 48 h. Flow cytometery results indicated that the proportion of A549 cell at S-phase was markedly increased after exposure of COF. To elucidate whether the anti-apoptotic c-IAP2 (IAP2) was involved in COF-improved cell survival, IAP2 protein levels was determined by Western blot, and the results showed it was significantly induced by COF in a concentration-dependent manner. Moreover, the suppression of BAY, a nuclear factor (NF)-kappaB binding inhibitor, or the COF-induced IAP2 protein levels indicated that NF-kappaB activation by COF may partly be involved in IAP2 induction. These results showed that the positive impact of COF on cell survival and proliferation of A549 lung tumor cells may be through an induction of IAP2 overexpression.

Growth inhibition and apoptosis-induced effect on human cancer cells of toosendanin, a triterpenoid derivative from chinese traditional medicine.

Toosendanin, a triterpenoid derivative isolated from the barks of Melia toosendan Sieb et Zucc, has been used as an anthelmintic vermifuge against ascaris for more than fifty years in China. In the present study, we investigated the growth inhibition and apoptosis-induced effect of toosendanin on human cancer cells. The result showed that toosendanin significantly suppressed the proliferation of tested human cancer cell lines. The IC(50) values were less than 1.7 x 10(-7) M and U937 was the most sensitive cell line with a IC(50) of 5.4 x 10(-9) M. Flow cytometric analysis revealed that treatment of U937 cells with toosendanin resulted in a dose- and time-dependent accumulation of cells in the S phase with a concomitant decrease in cells processing to G(0)/G(1) phase. The growth inhibition of U937 cells after exposure to toosendanin was subsequently associated with the induction of apoptosis, as evidence by the typical condensed and fragmented nuclei, DNA fragmentation, and exposure of phosphatidylserine on the outer leaflet of plasma membrane. All these results indicated that toosendanin could serve as a potential candidate for anticancer drug.

Improvement in biocompatibility of ZrO2-Al2O3 nano-composite by addition of HA.

The biocompatibility of zirconia-alumina (ZA) nano-composites in load-bearing applications such as dental/orthopedic implants was significantly enhanced by the addition of bioactive HA. The ZA matrix was composed of nano-composite powder obtained from the Pechini process and had higher flexural strength than conventionally mixed zirconia-alumina composite. Because the ZA nano-composite powder effectively decreased the contact area between HA and zirconia for their reaction during the sintering process, the HA-added ZA nano-composites contained biphasic calcium phosphates (BCP) of HA/TCP and had higher flexural strength than conventionally mixed ZA-HA composite. From the in vitro test with osteoblastic cell-lines, the proliferation and the differentiation (as expressed by the alkaline phosphatase activity) of the cellular response on the HA-added ZA nano-composites gradually increased as the amount of HA added increased. From the mechanical and biological evaluations of the HA-added ZA nano-composites, 30HA (30 vol% HA + 70 vol% ZA) was found to be the optimal composition for load-bearing biological applications.

Inhibition of substrate-tumor cell adhesion under the effect of gnidilatimonoein purified from Daphne mucronata.

Alteration of the cell surface glycoproteins of cancerous cells correlate with malignancy potential. To evaluate the mediation of membrane glycoproteins in wehi-164 cancerous cells, under the effect of D. mucronata crude extract and one of its purified active components, gnidilatimonoein, their attachment to fibronectin-coated wells were investigated. The plant extract, 27 microl/ml (equivalent to 0.54 mg/of plant leaves powder per ml of culture medium), as well as gnidilatimonoein (0.94 microM), were capable of quenching by 58% and 64%, respectively, the attachment of wehi-164 cells to fibronectin-coated wells (4 microg/ml). In addition to alteration of cell adhesive properties, the morphology of the treated cells were significantly changed upon treatment with the non-toxic dose of the plant extract or gnidilatimonoein. While the untreated cells have polygonal shapes, the treated cells had spherical shapes.

Transfection of human tumour cells with Mre11 siRNA and the increase in radiation sensitivity and the reduction in heat-induced radiosensitization.

Double-strand DNA breaks (DSBs) are potentially lethal DNA lesions induced by ionizing radiation. In eukaryotes, DSBs can be repaired by homologous recombination (HR) or non-homologous end-joining (NHEJ). DNA repair protein Mre11 participates in both the NHEJ and HR DNA repair pathways. Hyperthermia has been used clinically as a radiosensitizer. However, the mechanisms by which radiosensitization is induced by hyperthermia, especially moderate hyperthermia (41 degrees C) are not fully understood. Previous studies suggest that 41 degrees C reduces the nuclear Mre11 protein level in a manner that correlates with heat-induced changes in radiation sensitivity. Therefore, siRNA technology was used in the present study to reduce Mre11 gene expression to determine if reduced Mre11 protein levels induced radiosensitization and if such radiosensitization is similar to that induced by moderate hyperthermia. The results show that (1) the cellular level of the Mre11 protein was reduced about 60 +/- 18% by a 24-h treatment with siRNA. Results from the Mre11 protein turnover assay showed a half-life of 11.6 +/- 0.5 h for the Mre11 protein, which is consistent with reduction in protein level in 24 h after Mre11 siRNA treatment assuming a delay of 4-8 h to reduce RNA levels. After 48 h in siRNA, cellular Mre11 protein levels increased to approximately pretreatment levels. NSY cells were sensitized to ionizing radiation after 24 h of treatment with Mre11 siRNA. Two hours at 41 degrees C did not increase the radiation sensitivity of cells with a reduced Mre11 protein level following a 24-h siRNA treatment. These data support the conclusion that the DSB repair protein, Mre11, appears to be a target for radiosensitization by moderate hyperthermia.

Anti-neoplastic efficacy of Haimiding on gastric carcinoma and its mechanisms.

AIM: To study the anti-neoplastic effect of Haimiding and its mechanisms of action. METHODS: Experiments using MTT and colony formation were carried out to study the in vitro anti-neoplastic action of Haimiding, its in vivo anti-neoplastic action was studied by observing its effect on the weight of tumors in FC mice and S(180), H(22) tumor bearing mice, as well as their life spans. The effect of Haimiding on cell apoptosis and different stages of cell cycles in human gastric carcinoma cells were studied by flow cytometry. Its effect on [Ca(2+)](i) of human gastric carcinoma cells and the source of Ca(2+) during the change of [Ca(2+)](i) were observed by confocal laser scanning technique. RESULTS: Haimiding showed a definite cytotoxicity to 8 human tumor cell lines, which was most prominent against BGC-823, E(ca-109) and HCT-8 tumor cells. It also exhibited an obvious inhibition on colony formation of the above tumor cell lines, which was most prominent in E(ca-109) tumor cells. It showed obvious inhibition on the growth of tumor in FC mice and S(180) bearing mice as well as prolonged the life span of H(22) bearing mice. It was able to induce apoptosis and elevate intracellular [Ca(2+)](i) concentration of tumor cells. The source of Ca(2+) came from both extracellular Ca(2+) influx and intracellular Ca(2+) release. CONCLUSION: Haimiding is composed of a TCM preparation and 5-flurouracil. Its anti-neoplastic potency is highly enhanced by synergism as compared with either one of its components. Its mechanisms of anti-neoplastic action can be attributed to its action to initiate apoptosis of tumor cells by opening the membrane calcium channel and inducing intracellular Ca(2+) release to elevate [Ca(2+)](i) of the tumor cells.

Effects of Ginkgo biloba extract on cell proliferation and cytotoxicity in human hepatocellular carcinoma cells.

AIM: To study the effect of Ginkgo biloba extract (EGb 761) containing 22-27% flavonoids (ginkgo-flavone glycosides) and 5-7% terpenoids (ginkgolides and bilobalides) on cell proliferation and cytotoxicity in human hepatocellular carcinoma (HCC) cells. METHODS: Human HCC cell lines (HepG2 and Hep3B) were incubated with various concentrations (0-1 000 mg/L) of EGb 761 solution. After 24 h incubation, cell proliferation and cytotoxicity were determined by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay and lactate dehydrogenase (LDH) release, respectively. After 48 h incubation, the expression of proliferating cell nuclear antigen (PCNA) and p53 protein was measured by Western blotting. RESULTS: The results showed that EGb 761 (50-1 000 mg/L) significantly suppressed cell proliferation and increased LDH release (P<0.05) in HepG2 and Hep3B cells compared with the control group. The cell proliferation of HepG2 and Hep3B cells treated with EGb 761 (1 000 mg/L) was 45% and 39% of the control group (P<0.05), respectively. LDH release of HepG2 cells without and with EGb 761 (1 000 mg/L) treatment was 6.7% and 37.7%, respectively, and that of Hep3B cells without and with EGb 761 (1 000 mg/L) treatment was 7.2% and 40.3%, respectively. The expression of PCNA and p53 protein in HepG2 cells treated with EGb 761 (1 000 mg/L) was 85% and 174% of the control group, respectively. CONCLUSION: Ginkgo biloba extract significantly can suppress proliferation and increase cytotoxicity in HepG2 and Hep3B cells. Additionally, Ginkgo biloba extract can decrease PCNA and increase p53 expression in HepG2 cells.

Extracts from the roots of Lindera strychifolia induces apoptosis in lung cancer cells and prolongs survival of tumor-bearing mice.

Lindera strychifolia, a scandent shrub Lauraceous medicinal plant, has been used in Chinese traditional medicine as a palliative and an anti-spasmodic. It also shows cytotoxic effects against several tumor cell lines and inhibits marcromolecule biosynthesis. This study investigated the anti-tumor effects of L. strychifolia extract against lung cancer cells using in vitro and in vivo models. Two human lung cancer cell lines A549 (adenocarcinoma) and SBC-3 (small cell carcinoma), and a non-tumor cell line 3T3-L1 (mice fibroblasts) were subjected to L. strychifolia extract treatment. On lung cancer cells, L. strychifolia induced cell growth inhibition in a dose-dependent manner. Conversely, the extract did not show any significant cytotoxic effect on 3T3-L1 cells. Therefore, the extract is specific for tumor cells. Tumor cells treated with L. strychifolia extract showed typical morphological appearance of apoptosis including nuclei fragmentation and cell condensation. The in vivo effects of L. strychifolia extract were investigated in C57BL/6 mice transplanted with Lewis lung cancer (LL-2) cells, and in BALB/c nude mice transplanted with A549 or SBC-3 human lung cancer cells. Oral administration of L. strychifolia extract prolonged survival time and inhibited tumor growth in a dose-dependent manner by inducing apoptosis in the LL-2 cell mice model. Furthermore, in A549 or SBC-3 cell nude mice models, oral administration of L. strychifolia extract also significantly inhibited tumor growth at the 5.0 mg/ml concentration. These findings suggested that the components of L. strychifolia have anticancer activity and may contribute to clinical applications in the prevention and treatment of lung cancer.

Tracking the elusive antiestrogenic effect of melatonin: a new methodological approach.

OBJECTIVES: Detection of the antiestrogenic effect of melatonin on various breast cancer cell lines and its dependence of the differential expression of estrogen receptors (ERalpha and ERbeta) and melatonin receptors (mt1 and RZRalpha). SETTING AND DESIGN: Dose-response curves of estradiol were determined in 6 different breast cancer cell lines using a colorimetric proliferation assay in the absence or presence of various melatonin concentrations. METHODS: In order to detect the minor growth inhibitory effect of melatonin, a simple yet novel approach was employed: instead of incubating cells at single estradiol-concentrations at increasing melatonin levels, breast cancer cells were grown in microwell-plates for 4 days at increasing concentrations of estradiol (10(-12)M - 10(-10)M) in the absence or presence of melatonin (10(-9)M - 10(-8)M). Cell number was determined using Alamar blue and colorimetry. RT-PCR was performed for the expression of ERalpha, ERbeta, RZRalpha and mt1. RESULTS: Melatonin at concentrations of 10(-9)M and 5 x 10(-9)M shifted the dose-response curves of estradiol to higher concentrations. Responsiveness to melatonin depended on expression of ERalpha but not on ERbeta. mRNA of ERbeta was not detectable in the breast cancer cell lines used. Only small amounts of mt1 transcripts were detectable in MCF-7 cells of one source. In MCF-7 cells transfected with the mt1 gene and in an ovarian cancer cell line mt1 was expressed at significant levels. RZRalpha was expressed in all tested cell lines at different amounts. CONCLUSION: The growth of all ERalpha-positive breast cancer cell lines can be inhibited by melatonin. The effect in most cell lines is weak yet clearly reproducible. RZRalpha clearly contributes to the growth inhibitory effect of melatonin.