Plants from Northwestern Anatolia Display Selective Cytotoxicity and Induce Mitotic Catastrophe: A Study on Anticancer and Genotoxic Activities.

Anatolia is rich in floristic diversity with a high rate of endemism. Eight plant species from northwestern Anatolia were evaluated for their anti-growth properties in two malignant (MCF-7 and MDA-MB-231) and a non-malignant (MCF-10A) breast cell lines. The two most active extracts, Achillea multifida (AME) and Astragalus sibthorpianus (ASE), induced apoptotic cell death in all cell lines. The major phenolic compounds in AME were identified as chlorogenic acid, and catechins in ASE. ASE displayed selective cytotoxicity against breast cancer cells, with DNA damage repair in non-malignant cells contributing to its selectivity. Conversely, AME induced DNA damage in a time-dependent manner and displayed a dual dose-dependent biological activity, resulting in mitotic catastrophe and apoptosis at different doses. Most plant species exhibited moderate to strong cytotoxicity, highlighting their medicinal and economic potential and the need for their protection.

Simultaneous inhibition of PFKFB3 and GLS1 selectively kills KRAS-transformed pancreatic cells.

Activating mutations of the oncogenic KRAS in pancreatic ductal adenocarcinoma (PDAC) are associated with an aberrant metabolic phenotype that may be therapeutically exploited. Increased glutamine utilization via glutaminase-1 (GLS1) is one such feature of the activated KRAS signaling that is essential to cell survival and proliferation; however, metabolic plasticity of PDAC cells allow them to adapt to GLS1 inhibition via various mechanisms including activation of glycolysis, suggesting a requirement for combinatorial anti-metabolic approaches to combat PDAC. We investigated whether targeting the glycolytic regulator 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase-3 (PFKFB3) in combination with GLS1 can selectively prevent the growth of KRAS-transformed cells. We show that KRAS-transformation of pancreatic duct cells robustly sensitizes them to the dual targeting of GLS1 and PFKFB3. We also report that this sensitivity is preserved in the PDAC cell line PANC-1 which harbors an activating KRAS mutation. We then demonstrate that GLS1 inhibition reduced fructose-2,6-bisphosphate levels, the product of PFKFB3, whereas PFKFB3 inhibition increased glutamine consumption, and these effects were augmented by the co-inhibition of GLS1 and PFKFB3, suggesting a reciprocal regulation between PFKFB3 and GLS1. In conclusion, this study identifies a novel mutant KRAS-induced metabolic vulnerability that may be targeted via combinatorial inhibition of GLS1 and PFKFB3 to suppress PDAC cell growth.

Lichens exerts an anti-proliferative effect on human breast and lung cancer cells through induction of apoptosis.

Successful cancer treatment still requires new complexes or compounds from natural sources. Therefore, we investigated anti-growth/apoptotic effects of methanol extracts of the lichen species (Xanthoparmelia somloensis (Gleyn.) Hale, Usnea intermedia (A. Massal.) Jatta, Bryoria capillaris (Ach.) Brodo & D. Hawksw and Lobaria pulmonaria (L.) Hoffm.) on human lung (A549, H1299) and breast (MCF-7, MDA-MB-231) cancer cell lines. Anti-growth effects were monitored by the MTT and ATP viability assays. Cell death mode was evaluated by employing the fluorescence staining of nucleus, caspase-cleaved cytokeratin 18 detection, caspase 3/7 activity assay, Anneksin V cytofluorimetric assay and mitochondria membrane potential assay. Among the lichen extracts, Usnea intermedia exhibited strong anti-growth activity in a dose-dependent manner (1.56-100 µg/ml) compared to the others. Usnea intermedia was especially cytotoxic against MDA-MB-231 and H1299 cells (IC50 value for was found 3.0 and 10.2 µg/ml respectively). The cytotoxicity was resulted from apoptosis as proved by the presence of pyknotic nuclei, caspase 3/7 activity, phosphatidylserine translocation and loss of mitochondria membrane potential. In conclusion, Usnea intermedia warrants for further in vivo evaluation as a new alternative in cancer treatment.

Toxicity assessment of Hypericum olympicum subsp. olympicum L. on human lymphocytes and breast cancer cell lines.

There is a limited number of studies about the constituents of Hypericum olympicum subsp. olympicum and its genotoxic and cytotoxic potency. We examined the possible antigenotoxic/genotoxic properties of methanolic extract of H. olympicum subsp. olympicum (HOE) on human lymphocytes by employing sister chromatid exchange, micronucleus and comet assay and analyzed its chemical composition by GCxGC-TOF/MS. The anti-growth activity against MCF-7 and MDA-MB-231 cell lines was assessed by using the ATP viability assay. Cell death mode was investigated with fluorescence staining and ELISA assays. The major components of the flower and trunk were determined as eicosane, heptacosane, 2-propen-1-ol, hexahydrofarnesyl acetone and α-muurolene. HOE caused significant DNA damage at selected doses (250-750 µg/ml) while chromosomal damage was observed at higher concentrations (500 and 750 µg/ml). HOE demonstrated anti-growth activity in a dose-dependent manner between 3.13-100 µg/ml. Pyknotic nuclei were observed at 100 µg/ml concentration of HOE in both cell lines. In conclusion, HOE demonstrated cytotoxic effects in a cell type-dependent manner, however its genotoxic effects were observed at relatively higher doses.

6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase-3 is required for transforming growth factor ß1-enhanced invasion of Panc1 cells in vitro.

Transforming growth factor ß1 (TGFß1) is a well-established inducer of the epithelial-mesenchymal transition (EMT) that is essential for the acquisition of malignant properties, such as invasion, in tumor cells. Although recent studies suggest that the EMT in tumor cells is associated with reprogramming of energy metabolism and TGFß1 has been shown to stimulate glycolysis in multiple primary cell lines, little is known about TGFß1's effect on glycolysis and glycolytic regulators in transformed cells. Given the known regulatory role of 6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase-3 (PFKFB3) in glycolysis and association of glycolytic activity with malignant features such as invasion, we sought to investigate whether TGFß1 regulates PFKFB3 expression and if PFKFB3 is involved in the TGFß1-mediated increase in the invasive ability of the Panc1 cell cline-a well-established model of TGFß1-initiated EMT. Herein we demonstrate that TGFß1 induces PFKFB3 expression and stimulates glycolysis in Panc1 cells. We also show that siRNA silencing of PFKFB3 prevents the stimulation of glycolysis and in vitro invasive ability of Panc1 cells by TGFß1. Furthermore, PFKFB3 silencing suppresses the TGFß1-mediated induction of the Snail protein, suggesting that PFKFB3 is required for the regulation of Snail expression by TGFß1. Taken together, our study identifies PFKFB3 as a key TGFß1 effector protein that mediates TGFß1's effect on Snail expression, invasion, and glycolysis.

Evaluation of genotoxic and apoptotic potential of Hypericum adenotrichum Spach. in vitro.

Hypericum adenotrichum Spach. is an endemic plant from Turkey that is also used in folk medicine. In this study, following analyses of its chemical composition, the genotoxic/antigenotoxic effects of the methanol extract of H. adenotrichum in human lymphocyte culture were investigated using in vitro sister chromatid exchange, micronucleus and comet assays. In addition, the anti-growth effect of the extract was investigated in human breast cancer cell lines (MCF-7 and MDA-MB-231) using MTT and ATP viability assays. The mode of cell death was determined using fluorescence microscopy and biochemical methods. We found that the H. adenotrichum extract demonstrated cytotoxic and genotoxic effects in a cell type-dependent manner. At selected doses (125-500 µg/ml), the H. adenotrichum extract exhibited significant genotoxic activity in human lymphocytes, whereas it showed anti-growth effects on cancer cell lines between 0.2 and 100 µg/ml concentrations. The mode of cell death in cancer cells was shown to be apoptosis due to the presence of pyknotic nuclei, the cleavage of poly-(ADP-ribose) polymerase (PARP) and/or the activation of caspase-3. These results suggest that H. adenotrichum might show both cytotoxic and genotoxic effects depending on the cell type. This should be taken into account in its use for therapeutic purposes.

Genotoxic, cytotoxic, and apoptotic effects of Hypogymnia physodes (L.) Nyl. on breast cancer cells.

The aim of this study is to determine the chemical composition, and evaluate the genotoxic, and anti-growth potency of the methanol extracts of lichen species Hypogymnia physodes (L.) Nyl. (HPE). Anti-growth effect was tested in two different human breast cancer cell lines (MCF-7 and MDA-MB-231) by the MTT and ATP viability assays and apoptosis was assayed by the caspase-cleaved cytokeratin 18 (M30-antigen). Genotoxic activity of HPE was studied using chromosome aberration and micronuclei tests in human lymphocytes culture in vitro. The chemical composition of H. physodes was analyzed by using direct thermal desorption method coupled with comprehensive gas chromatography-time of flight mass spectrometry (GCXGC-TOF/MS). Our results indicate that HPE has an anti-growth effect at relatively lower concentrations, while relatively higher concentrations are required for genotoxic activity. HPE, therefore, seems to represent a therapeutic potential and poses new challenges for medicinal chemistry.

Protective and antigenotoxic effect of Ulva rigida C. Agardh in experimental hypothyroid.

The presence of chromosomal damage in bone marrow cells affected by several diseases such as thyroid, cancer etc., was detected by the micronucleus (MN) assay. The present study was designed to evaluate: i) volatile components of Ulva rigida, ii) effects of hypothyroidism on bone marrow MN frequency, iii) effects of oral administration of Ulva rigida ethanolic extract (URE) on MN frequency produced by hypothyroidism, and iv) thyroid hormone levels in normal and 6-n-Propylthiouracil (PTU)-induced hypothyroid rats. The volatile components of Ulva rigida was studied using a direct thermal desorption (DTD) technique with comprehensive two-dimensional gas chromatography time-of-flight mass spectrometry (GCxGC-TOF/MS). URE administration was of no significant impact on thyroid hormone levels in control group, while PTU administration decreased thyroid hormone levels compared to control group (p < 0.001). Moreover, URE supplementation resulted in a significant decrease in MN frequency in each thyroid group (p < 0.0001). This is the first in vivo study that shows the strong antigenotoxic and protective effect of URE against the genotoxicity produced by hypothyroidism.

Ulva rigida improves carbohydrate metabolism, hyperlipidemia and oxidative stress in streptozotocin-induced diabetic rats.

This study was designed to investigate the effects of Ulva rigida, one of the green algae, on the lipid profile and oxidative-antioxidative systems in streptozotocin-induced diabetic rats. Forty Wistar rats randomly divided into four groups: control (C), control + U. rigida extract (C + URE), diabetes (D) and diabetes + U. rigida extract (D + URE). U. rigida (2%) was administered in drinking water for 5 weeks after the induction of diabetes. U. rigida reduced the blood glucose, serum total cholesterol, triglyceride levels and plasma and tissue malondialdehyde (MDA) levels in the D + URE group. Insulin levels were significantly higher in the D + URE than those of the D group. Serum total cholesterol and tissue MDA levels were reduced in the C + URE group. Whole blood glutathione peroxidase and erythrocyte superoxide dismutase activities were higher in the D and C + URE groups compared with the C group. Paraoxonase and arylesterase activities were lower in the D group while U. rigida increased paraoxonase activities in C + URE and D + URE groups. This is the first study which showed U. rigida has antidiabetic and antihyperlipidemic effects and improves oxidative stress in diabetic rats. We conclude that U. rigida might have a potential use as a protective and/or therapeutic agent in diabetes mellitus.

Palladium (II) Complex Enhances ROS-Dependent Apoptotic Effects via Autophagy Inhibition and Disruption of Multiple Signaling Pathways in Colorectal Cancer Cells.

BACKGROUND: Inhibition of autophagy is reported to be a therapeutically effective strategy in overcoming resistance that is a deadly outcome in cancer. One of the most common reasons for chemo-resistance to treatment is the patients with tumors exhibiting a KRAS mutation, which occurs in approximately 40% of colorectal cancer patients. OBJECTIVE: Hence, we assessed whether a Palladium (Pd)(II) complex is a promising anticancer complex, compared to 5-fluorouracil in KRAS wt HT-29 and KRAS mutant HCT-15 cells. METHODS: HCT-15 and HT-29 cells were used for colorectal cancer and Chloroquine (CQ) was used as an inhibitor of autophagy. In this context, cells were treated with Pd(II) complex and 5-FU in combination with CQ for 48h and cell viability was measured by SRB assay. Cell death mode was examined with M30 and M65 ELISA assays, using annexin V/propidium iodide. Autophagy was determined by Acridine Orange (AO) staining. Furthermore, the expressions of various autophagy and apoptosis-related proteins were evaluated with Western blotting. Luminex assay and the level of Reactive Oxygen Species (ROS) were examined. RESULTS: Cell viability was found to decrease in a dose-dependent manner and CQ enhanced cytotoxic effect in Pd(II) and 5-FU treated cells in colorectal cancer cells. Our data showed that inhibition of autophagic flux significantly increased intrinsic apoptosis through the activation of ROS. We showed that combinatorial treatment with CQ induced apoptosis via the caspase-dependent mitochondrial pathway. Luminex analysis revealed that the combination resulted in a down-regulation of NF-κB/AKT/CREB signaling pathways in both cell lines, however, decreased Erk1/2 protein expression was only observed after treatment with CQ combination in HCT-15 cells. CONCLUSION: We suggest that the inhibition of autophagy along with Pd(II) and 5-FU treatment has a synergistic effect on KRAS-mutant colorectal cancer cells. Autophagy inhibition by CQ promotes apoptosis via blockade of the NF-κB/AKT/CREB and activation of ROS.

αvß3 Integrin Antagonists Enhance Chemotherapy Response in an Orthotopic Pancreatic Cancer Model.

Pancreatic cancer decreases survival time and quality of life because of drug resistance and peripheral neuropathy during conventional treatment. This study was undertaken to investigate whether αvß3 integrin receptor antagonist compounds NDAT and XT199 can suppress the development of cisplatin resistance and cisplatin-induced peripheral neuropathy in an orthotopic pancreatic SUIT2-luc cancer cell mouse model. Anticancer effects of these compounds and their combination with cisplatin were assessed in this tumor mouse model with bioluminescent signaling and histopathology, and a cytokine assay was used to examine expression of inflammatory cytokines IL-1ß, IL-6, IL-10, and TNF-α from plasma samples. To determine the neuroprotective effects of the compounds on cisplatin-induced peripheral neuropathy, behavioral hind-limb posture of the mice was evaluated. The combination therapy of NDAT or XT199 with cisplatin elicited greater inhibition of tumor growth and increased tumor necrosis compared to cisplatin alone. NDAT and XT199 in combination with cisplatin significantly decreased expression of pro-inflammatory cytokines IL-1ß, IL-6, and TNF-α and significantly increased expression of anti-inflammatory cytokine IL-10 in comparison to cisplatin alone. Cisplatin-treated groups showed stocking-glove hind-limb posture, whereas NDAT and XT199 with cisplatin-treated groups displayed normal hind-limb posture. Results clearly suggest that NDAT and XT199 treatment with cisplatin that inactivates NF-κB may contribute to increased antitumor and anti-inflammatory efficacy as well as alleviate cisplatin-mediated loss of motor function in this pancreatic tumor mouse model.

A promising therapeutic combination for metastatic prostate cancer: Chloroquine as autophagy inhibitor and palladium(II) barbiturate complex.

Autophagy is a catabolic process for cells that can provide energy sources and allows cancer cells to evade cell death. Therefore, studies on the combination of autophagy inhibitors with drugs are increasing as a new treatment modality in cancer. Previously, we reported the anti-tumor activity of a Palladium (Pd)(II) complex against different types of cancer in vitro and in vivo. Chloroquine (CQ), the worldwide used anti-malarial drug, has recently been focused as a chemosensitizer in cancer treatment. The aim of this study was to investigate the efficacy of a combined treatment of these agents that work through different mechanisms to provide an effective treatment modality for metastatic prostate cancer that is certainly fatal. Metastatic prostate cancer cell lines (PC-3 and LNCaP) were treated with Pd (II) complex, CQ, and their combination. The combination enhanced apoptosis by increasing phosphatidylserine translocation and pro-apoptotic proteins. Apoptosis was confirmed by the use of apoptosis inhibitor. The formation of acidic vesicular organelles (AVOs) was observed by acridine orange staining in fluorescence microscopy. The Pd (II) complex increased AVOs formation in prostate cancer cells and CQ-pretreatment has potentiated this effect. Importantly, treatment with CQ suppressed the pro-survival function of autophagy, which might have contributed to enhanced cytotoxicity. In addition, PI3K/AKT/mTOR-related protein expressions were altered after the combination of treatments. Our results suggest that combination treatment enhances apoptotic cell death possibly via the inhibition of autophagy, and may therefore be regarded as a novel and better approach for the treatment of metastatic prostate cancer.

Evaluation of chromosome aberrations, sister chromatid exchange and micronuclei in patients with type-1 diabetes mellitus.

Oxidative stress-induced DNA damage seems to play a role in the pathogenesis of type-1 diabetes mellitus and its complications. Several in vitro assays have been used to measure the DNA damage produced by oxidative stress. In the present study, we aimed to investigate the frequency of sister chromatid exchange (SCE), chromosomal aberrations (CA) and micronuclei (MN) in type-1 diabetes mellitus patients compared with healthy controls. SCE, CA and MN tests were carried out with the blood-cell cultures from 35 type-1 diabetic patients and 15 healthy, age- and sex-matched control subjects. The mean age of the type-1 diabetic patients was 31.89 +/- 10.01 years, with a mean duration of the diabetes of 7.8 +/- 6.02 years. The mean level of HbA1c of the type-1 diabetic patients was 8.37+/-1.36%. Only three (8.5%) patients with type-1 diabetes mellitus had an HbA1c level below 7%. Patients with type-1 diabetes mellitus showed a higher frequency of SCE compared with controls (5.44 +/- 1.47 and 2.54 +/- 0.82, respectively, p < 0.001), but there was no significant correlation between the duration of diabetes, HbA1c and SCE. No significant difference was found in CA or MN frequency in type-1 diabetic patients compared with controls. In conclusion, these results suggest that type-1 diabetes mellitus is a condition with genomic instability characterized by an increased level of SCE. Hyperglycemia-induced oxidative stress may be the underlying factor of the increased SCE frequency.

A biomonitoring study of genotoxic risk to workers of transformers and distribution line stations.

A cytogenetic monitoring study was carried out on a group of workers from transformer and distribution line stations in the Bursa province of Turkey, to investigate the genotoxic risk of occupational exposure to extremely low frequency electric (ELF) and magnetic fields (EMF). Cytogenetic analysis, namely chromosomal aberrations (CAs) and micronucleus (MN) tests were performed on a strictly selected group of 55 workers and compared to 17 controls. CA and MN frequencies in electrical workers appeared significantly higher than in controls (p < 0.001, 0.05, respectively). The frequency of CA in exposed groups were significantly enhanced with the years of exposure (p < 0.01). The effect of smoking on the level of CA and MN was not significant in the control and exposure groups. The results of this study demonstrated that a significant induction of cytogenetic damage in peripheral lymphocytes of workers engaged to occupational exposure to ELMF in electric transformer and distribution stations.

Cytotoxic and genotoxic effects of an endemic plant of Turkey Salvia kronenburgii on breast cancer cell lines.

CONTEXT: The natural products derived from plants are the important sources that can be used for breast cancer treatment. Salvia species and their derived products were recommended as potential antitumor substances. AIM: The potential cytotoxic and genotoxic effects of Salvia kronenburgii have been investigated on breast cancer cell lines, MCF-7 and MDA-MB-231. MATERIALS AND METHODS: Determination of chemical compounds of S. kronenburgii was done using a gas chromatography coupled to time-of-flight mass spectrometry system and a dual-stage commercial thermal desorption injector. Growth inhibition of the S. kronenburgii was measured by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and ATP viability assays. The cell death mode was detected by fluorescent dyes. Genotoxic effect of S. kronenburgii was measured by comet assay. RESULTS: S. kronenburgii showed antiproliferative effect in a dose-dependent manner on MCF-7 and MDA-MB-231 cell lines by inducing apoptosis-like cell death. The pyknotic cell nuclei were observed at the cell lines in response to S. kronenburgii. Furthermore, significant increase was shown in genetic damage index and frequencies in the damaged cells. CONCLUSION: S. kronenburgii might be a promising natural source for cancer therapy. Further experiments need to be done in vivo to understand of the anticancer effects of this plant.

In vivo evaluation of the toxic effects of pyrimethamine on spermatogenesis in male mice.

Pyrimethamine is known to have antimalarial activities and used clinically in the therapy of toxoplasmosis and human immunodeficiency virus-associated pneumonia. In this study we aimed to test the effects of pyrimethamine on spermatogenesis in mice. For this aim, animals were given pyrimethamine as a single application and the doses were 5, 10, 20, and 40 mg/kg. For the spermatogenic effects, the sperm shape abnormality, epididymal sperm counts, and testes weights were evaluated at the end of days 7, 14, 21, 28, and 35 after single pyrimethamine injection at the first day. Pyrimethamine increased the frequency of abnormal sperm shape for all studied weeks except the first week and its germ cell stage-specific effects correspond to spermatozoa, spermatids, and spermatocytes. It also decreased the epididymal sperm counts at the end of days 28 and 35, which corresponds to the spermatocyte stage of mouse spermatogenesis.

In vitro antigenotoxicity of Ulva rigida C. Agardh (Chlorophyceae) extract against induction of chromosome aberration, sister chromatid exchange and micronuclei by mutagenic agent MMC.

OBJECTIVE: To determine the in vitro possible clastogenic and cytotoxic activities of Ulva rigida crude extracts (URE), and identify their antigenotoxic and protective effects on chemotherapeutic agent mitomycine-C (MMC). METHODS: Anti-clastogenic and anti-genotoxic activities of Ulva rigida crude extracts (URE) were studied using chromosome aberration (CA), sister chromatid exchange (SCE), and micronuclei (MN) tests in human lymphocytes cultured in vitro. RESULTS: The chromosome aberration, sister chromatid exchange or micronuclei tests showed that URE at concentrations of 10, 20, and 40 microg/mL had no clastogenic activity in human lymphocyte cell culture. Three doses of URE significantly decreased the number of chromosomal aberrations and the frequencies of SCE and MN when compared with the culture treated with MMC (P < 0.0001). CONCLUSION: Although URE itself is not a clastogenic or cytotoxic substance, it possesses strong antigenotoxic, anti-clastogenic, and protective effects on MMC in vitro.

Angelica sylvestris and Delphinium staphisagria Extracts Induces Antiproliferation through Caspase-mediated Apoptosis on Human Cancer Cells

Abstract: Angelica sylvestris and Delphinium staphisagria are medicinal and aromatic herbs with a long history in medicine and food industry. In this study, we have investigated anti-cancer activity of Angelica sylvestris and Delphinium staphisagria extracts on various cell lines of lung (A549), breast (MCF-7), colon (HT-29), and cervix (HeLa) origin. Also, cytotoxicity was tested on human healthy bronchial epithelial (BEAS-2B) cells. In vitro experiments showed that plant extracts suppressed cell growth and proliferation at low concentrations by reducing cell viability on cancer cells in a time and concentration-dependent manner. It was observed that Angelica sylvestris was more effective in HT-29 and HeLa cells and Delphinium staphisagria in A549 and MCF-7 cells by suppressing cell proliferation and increasing cell death. Cell death mode (apoptosis/necrosis) was investigated via fluorescent imaging, caspase-cleaved cytokeratin 18, activated caspase-3, and cleaved-PARP (poly (ADP-ribose) polymerase). In order to evaluate the cell death mode by plant extracts apoptotic markers were investigated by fluorescence staining. Delphinium staphisagria extract (50-200 µg/mL) caused a decrease in cell density in A549 and MCF-7 cells compared to untreated controls. A similar situation was observed in HT-29 and HeLa cell lines when treated with ASE. As a result, Delphinium staphisagria extracts induced apoptosis in A549 and MCF-7, while Angelica sylvestris extracts induced apoptosis in HT-29 and HeLa cancer cells.

Germination responses to GA3 and stratification of threatened Festuca L. species from Eastern Mediterranean.

The seed germination characteristics of three threatened Festuca sp. [F. punctoria Sm., F. cyllenica Boiss. et Heldr. subsp. uluana Markgr.-Dannenb., F. paphlagonica (St.-Yves) Markgr.-Dannenb. subsp. paphlagonica] were investigated. These species are endemic and spread on alpine belt. The study was carried out with wet-cold and dry-cold stratification throughout 15 days, different doses of GA3 (50, 100 and 150 ppm) and hormone-stratification combined treatments, and non-treatment series. We found that the germination rates of three fescue seeds for various treatment series were different. The mean germination percentage of F. cyllenica was higher (80%) than that of F. punctoria and F. paphlagonica which were fairly low (50-60%). Germination rates increased by wet-stratification treatment in F. punctoria and also increased with 100 ppm GA3 application to the seeds of F. paphlagonica. When taken into consideration the germination percentages of all fescue species, the seeds of F. punctoria and F. paphlagonica can be dormant, but the seeds of F. cyllenica are non-dormant.

Genotoxic effects induced by fotemustine and vinorelbine in human lymphocytes.

The aim of this study was to investigate the in vitro genotoxic effects of the anticancer drugs fotemustine and vinorelbine on human lymphocytes and to determine individual and sex-related responses to these drugs. Fotemustine is a DNA-alkylating drug while vinorelbine is a semi-synthetic Vinca alkaloid. The study was carried out with twenty independent healthy donors for each drug. We have tested the ability of these drugs to induce chromosome aberrations (CAs) and sister chromatid exchanges (SCEs) as well as effect on the mitotic index (MI) in cultured human lymphocytes. Fotemustine was shown to induce CAs and SCEs at all concentrations tested (2, 4 and 8 microg/ml) in a dose-dependent manner. Additionally it also decreased the mitotic index in a similar dose-dependent manner. Vinorelbine had no effect on structural CAs, but it significantly increased the numerical CAs at all doses tested (0.5, 1 and 2 microg/ml). Vinorelbine also induced SCE events and increased the MI values. Two-way analyses of variance were used to compare the individual and gender-related susceptibilities to fotemustine and vinorelbine with respect to the CA, SCE and MI values. The results indicated that individuals in fotemustine treatment groups showed different genotoxic responses with respect to CA and SCE induction and additional findings indicated a gender-specific response in this group. Individuals in the vinorelbine test group also exhibited statistically significant numerical CA, SCE and MI responses to vinorelbine. A statistically significant gender-related SCE response to this drug was also evident. This study indicates that these drugs have potentially harmful effects on human health.