Germacranolides from Carpesium divaricatum: Some New Data on Cytotoxic and Anti-Inflammatory Activity.

Carpesium divaricatum Sieb. & Zucc., a traditional medicinal plant used as an inflammation-relieving remedy, is a rich source of terpenoids. At least 40 germacrane-type sesquiterpene lactones, representatives of four different structural groups, were isolated from the plant. Cytotoxicity against cancer cells in vitro is the most frequently described biological activity of the compounds. However, little is known about the selectivity of the cytotoxic effect. The anti-inflammatory activity of the germacranolides is also poorly documented. The objective of the present study was to assess the cytotoxic activity of selected C. divaricatum germacranolides-derivatives of 4,5,8,9-tetrahydroxy-3-oxo-germacran-6,12-olide towards cancer and normal cell lines (including cells of different p53 status). Moreover, to assess the anti-inflammatory effect of the compounds, the release of four proinflammatory cytokines/chemokines (IL-1ß, IL-8, TNF-α and CCL2) by lipopolysaccharide-stimulated human neutrophils was measured by ELISA. The investigated sesquiterpene lactones demonstrated nonselective activity towards prostate cancer (Du145 and PC3) and normal prostate epithelial cells (PNT2) as well as against melanoma cells (A375 and HTB140) and keratinocytes (HaCaT). Cytotoxic activity against osteosarcoma cells was independent of their p53 status. In sub-cytotoxic concentrations (0.5-2.5 µM) the studied compounds significantly decreased cytokine/chemokine release by lipopolysaccharide-stimulated human leukocytes.

Marine Algae of the Genus Gracilaria as Multi Products Source for Different Biotechnological and Medical Applications.

BACKGROUND: Gracilaria has been shown to be an important source of marine bioactive natural biomaterials and compounds. Although there are no enough patents used Gracilaria worldwide, the current study tries to put the Gracilaria on the spot for further important patents in the future. OBJECTIVE: The current study investigates the pharmaceuticals and biochemical activity of Gracilaria because no previous studies have been carried out to examine the biochemical and pharmaceutical activates of Gracilaria from the Suez Canal of Egypt as an excellent source for bioactive compounds. METHODS: Different advanced experimental models and analytical techniques, such as cytotoxicity, total antioxidant capacity, anticancer, and anti-inflammatory profiling were applied. The phytochemical analysis of different constituents was also carried out. RESULTS: The mineral analysis revealed the presence of copper (188.3 ppm) and iron (10.07 ppm) in addition to a remarkable wealth of selenium and sulfur contents giving up to 36% of its dry mass. The elemental analysis showed high contents of sulfur and nitrogen compounds. The GCMS profiling showed varieties of different bioactive compounds, such as fatty acids, different types of carotenoids in addition to pigments, alkaloids, steroids. Many other compounds, such as carbohydrates and amino acids having antioxidant, anti-inflammatory, and antiviral activities, etc. were identified. The cytotoxicity activity of Gracilaria marine extract was very effective against cancerous cell lines and showed high ability as a potent antitumor due to their bioactive constituents. Specialized screening assays using two anticancer experimental models, i.e., PTK and SKH1 revealed 77.88% and 84.50% inhibition anticancer activity; respectively. The anti-inflammatory activities investigated using four different experimental models, i.e., COX1, COX2, IL6, and TNF resulted in 68%, 81.76%, 56.02% and 78.43% inhibition; respectively. Moreover, Gracilaria extracts showed potent anti-Alzheimer with all concentrations. CONCLUSION: Gracilaria proved to be a multi-product source of marine natural products for different biotechnological applications. Our recommendation is to investigate the Gracilaria bioactive secondary metabolites in order to create and innovate in more patents from current important seaweeds (Gracilaria).

Evaluation of cytotoxicity and mutagenicity of the benzodiazepine flunitrazepam in vitro and in vivo

Flunitrazepam (FNZ) is a sedative benzodiazepine prescribed for the short-term treatment of insomnia. However, there are concerns regarding possible carcinogenic or genotoxic effects of this medicine. Thus, the aim of this study was to evaluate the cytotoxic, clastogenic and aneugenic effects of FNZ in hepatoma cells from Rattus norvegicus (HTC) in vitro and in bone marrow cells of Wistar rats in vivo. These effects were examined in vitro following treatment with 0.2, 1.0, 5.0 or 10 μg/mL FNZ using a micronucleus test with a cytokinesis block or in vivo using a chromosomal aberration test following treatment with 7, 15 or 30 μg/mL/kg body weight. The results showed that the benzodiazepine concentrations tested were not cytotoxic, aneugenic or clastogenic. However, considering the adverse effects of using this benzodiazepine, more studies are required.

Flunitrazepam (FNZ) é um sedativo benzodiazepínico prescrito para o tratamento da insônia em curto prazo. Entretanto, existe a preocupação com relação aos possíveis efeitos carcinogênicos ou genotóxicos causados por este fármaco. Então, o objetivo deste estudo foi avaliar os efeitos citotóxicos, clastogênicos e aneugênicos do FNZ em células de hepatoma de Rattus norvegicus (HTC) in vitro e em células de medula óssea de ratos Wistar in vivo. Foram testadas as concentrações de 0,2, 1,0 e 10 μg/mL de FNZ pelo teste do micronúcleo com bloqueio de citocinese in vitro e 7, 15 e 30 μg/mL/kg de peso corpóreo para o teste de aberração cromossômica in vivo. Os resultados mostraram que as concentrações do benzodiazepínico testadas não foram citotóxicas, aneugênicas ou clastogênicas. Entretanto, considerando os efeitos adversos do uso deste benzodiazepínico, mais estudos são necessários.

Mutagenicity and DNA damage of bisphenol A and its structural analogues in HepG2 cells.

Environmental oestrogen bisphenol A (BPA) and its analogues are widespread in our living environment. Because their production and use are increasing, exposure of humans to bisphenols is becoming a significant issue. We evaluated the mutagenic and genotoxic potential of eight BPA structural analogues (BPF, BPAF, BPZ, BPS, DMBPA, DMBPS, BP-1, and BP-2) using the Ames and comet assay, respectively. None of the tested bisphenols showed a mutagenic effect in Salmonella typhimurium strains TA98 and TA100 in either the presence or absence of external S9-mediated metabolic activation (Aroclor 1254-induced male rat liver). Potential genotoxicity of bisphenols was determined in the human hepatoma cell line (HepG2) at non-cytotoxic concentrations (0.1 µmol L(-1) to 10 µmol L(-1)) after 4-hour and 24-hour exposure. In the comet assay, BPA and its analogue BPS induced significant DNA damage only after the 24-hour exposure, while analogues DMBPS, BP-1, and BP-2 induced a transient increase in DNA strand breaks.

Functional and phylogenetic characterization of Vaginolysin, the human-specific cytolysin from Gardnerella vaginalis.

Pore-forming toxins are essential to the virulence of a wide variety of pathogenic bacteria. Gardnerella vaginalis is a bacterial species associated with bacterial vaginosis (BV) and its significant adverse sequelae, including preterm birth and acquisition of human immunodeficiency virus. G. vaginalis makes a protein toxin that generates host immune responses and has been hypothesized to be involved in the pathogenesis of BV. We demonstrate that G. vaginalis produces a toxin (vaginolysin [VLY]) that is a member of the cholesterol-dependent cytolysin (CDC) family, most closely related to intermedilysin from Streptococcus intermedius. Consistent with this predicted relationship, VLY lyses target cells in a species-specific manner, dependent upon the complement regulatory molecule CD59. In addition to causing erythrocyte lysis, VLY activates the conserved epithelial p38 mitogen-activated protein kinase pathway and induces interleukin-8 production by human epithelial cells. Transfection of human CD59 into nonsusceptible cells renders them sensitive to VLY-mediated lysis. In addition, a single amino acid substitution in the VLY undecapeptide [VLY(P480W)] generates a toxoid that does not form pores, and introduction of the analogous proline residue into another CDC, pneumolysin, significantly decreases its cytolytic activity. Further investigation of the mechanism of action of VLY may improve understanding of the functions of the CDC family as well as diagnosis and therapy for BV.

Multiple regression analysis as a tool for the identification of relations between semi-quantitative LC-MS data and cytotoxicity of extracts of the fungus Fusarium avenaceum (syn. F. arthrosporioides).

The cytotoxicity of methanolic extracts from rice cultures of 53 Fusarium avenaceum strains, which had been isolated from different host organisms in Northern Europe, Canada and Australia/New Zealand, was investigated in a rat hepatoma (H4IIE-W), porcine epithelial kidney (PK-15), foetal feline lung fibroblast, dog lymphoblast (D3447), and a human hepatocarcinoma (Hep G2) cell line using the Alamar Bluetrade mark assay. All extracts were screened for known fungal metabolites using high-performance liquid chromatography with photodiode array and mass spectrometric detection, and both known and unknown metabolites were semi-quantified. Known metabolites that were determined in the cultures include acuminatopyrone, 2-amino-14,16-dimethyloctadecan-3-ol (2-AOD-3-ol), antibiotic Y, aurofusarin, chlamydosporol, chlamydospordiol, enniatins, fusarin A and C, and moniliformin. Multiple regression analysis was used in order to relate fungal metabolites to the cytotoxicity of the extracts. Separate linear regression models were constructed for each cell line. Eleven different fungal metabolites were related to the cytotoxicity (P<0.05). Out of these, nine metabolites were siginificantly related to the cytotoxicity in only one of the five models, while two, namely enniatins and 2-AOD-3-ol, were significant contributors in three or four regression models, respectively. This paper describes how multiple regression analysis may be applied for the assignment of bioactivity/toxicity to the constituents of a multi-component mixture.

Interactions between membranes and cytolytic peptides.

The physico-chemical and biological properties of cytolytic peptides derived from diverse living entities have been discussed. The principal sources of these agents are bacteria, higher fungi, cnidarians (coelenterates) and the venoms of snakes, insects and other arthropods. Attention has been directed to instances in which cytolytic peptides obtained from phylogenetically remote as well as from related sources show similarities in nature and/or mode of action (congeneric lysins). The manner in which cytolytic peptides interact with plasma membranes of eukaryotic cells, particularly the membranes of erythrocytes, has been discussed with emphasis on melittin, thiolactivated lysins and staphylococcal alpha-toxin. These and other lytic peptides are characterized in Table III. They can be broadly categorized into: (a) those which alter permeability to allow passage of ions, this process eventuating in colloid osmotic lysis, signs of which are a pre-lytic induction or latent period, pre-lytic leakage of potassium ions, cell swelling and inhibition of lysis by sucrose. Examples of lysins in which this mechanism is involved are staphylococcal alpha-toxin, streptolysin S and aerolysin; (b) phospholipases causing enzymic degradation of bilayer phospholipids as exemplified by phospholipases C of Cl. perfringens and certain other bacteria; (c) channel-forming agents such as helianthin, gramicidin and (probably) staphylococcal delta-toxin in which toxin molecules are thought to embed themselves in the membrane to form oligomeric transmembrane channels.