Chronic Exposure to Environmentally Relevant Concentrations of Microcystin-Leucine Arginine Causes Lung Barrier Damage through PP2A Activity Inhibition and Claudin1 Ubiquitination.

Microcystin-leucine arginine (MC-LR), ubiquitous in water and food, is a threat to public health. In the present study, after C57BL/6J mice were fed with environmental concentrations of MC-LR (0, 1, 30, 60, 90, and 120 µg/L) for 6, 9, and 12 months, it was found that MC-LR could enter into mouse lung tissues and cause microstructural damage, as shown by western blotting and HE staining. Electron microscopy examination showed that MC-LR could damage the lung barrier by disruption of the tight junctions, which was confirmed by the decreased expression of tight junction markers, including Occludin, Claudin1, and ZO-1. In addition, MC-LR also increased the ubiquitination of Claudin1, indicating that MC-LR could disrupt tight junctions by promoting the degradation of Claudin1. Furthermore, MC-LR increased the levels of TNF-α and IL-6 in mouse lung tissues, leading to pneumonia. Importantly, pretreatment with PP2A activator D-erythro-sphingosine (DES) was found to significantly alleviate MC-LR-induced decrease of Occludin and Claudin1 by inhibiting the P-AKT/Snail pathway in vitro. Together, this study revealed that chronic exposure to MC-LR causes lung barrier damage, which involves PP2A activity inhibition and enhancement of Claudin1 ubiquitination. This study broadens the awareness of the toxic effects of MC-LR on the respiratory system, which has deep implications for public health.

Reproductive and developmental safety of nirmatrelvir (PF-07321332), an oral SARS-CoV-2 Mpro inhibitor in animal models.

Nirmatrelvir (PF-07321332; NMV) the antiviral component of PAXLOVID™ is a potent and selective inhibitor of the SARS-CoV-2 main protease (Mpro), which plays a critical role in viral replication. PAXLOVID, comprised of nirmatrelvir and ritonavir (used as a pharmacokinetic enhancer), is an oral therapy currently in development as a therapeutic option for those infected with SARS-CoV-2 to prevent progression to severe disease, hospitalization, and death. PAXLOVID has been shown to be efficacious against hospitalization and death in two Phase 2/3 clinical studies that evaluated non hospitalized patients both with and without high risk factors for progression to severe illness. Given that males and females of reproductive age are included in the intended patient population, we assessed the potential effects of NMV up to the limit dose of 1000 mg/kg/day in ICH guideline embryo-fetal development studies in rats and rabbits, and a fertility and early embryonic development study in rats. There were no effects on male and female fertility or early embryonic development in rats, and no severe manifestations of developmental toxicity in rats or rabbits. The lack of adverse findings reported here in nonclinical species is consistent with the intended therapeutic target of NMV (a virus specific protein not present in mammalian cells), the favorable off-target selectivity profile, and lack of genetic toxicity. The results of these nonclinical studies with NMV along with existing ritonavir safety information indicate that there are no clinically relevant risks associated with PAXLOVID administration during pregnancy and in males and females of reproductive age.

Male reproductive toxicity induced by Microcystin-leucine-arginine (MC-LR).

Microcystin-LR (MC-LR) has been identified to pose an increasing threat to the male reproductive system in vivo and in vitro studies with the objects like mammal animals, amphibians, aquatic organisms, etc. This review demonstrates the latest research advances of the male reproductive toxicity induced by MC-LR in detail, which mainly consists of two aspects, namely pathological injuries to testis and prostate, as well as the endocrine disruption. Apart from the direct toxicity to the male reproductive system, we also underline the transgenerational reproductive toxicity that prenatal exposure may pass on to male offspring. This review also demonstrates the interactive effects between MC-LR and other compounds, including synergistic effects with some toxicants and antagonistic effects with some medicine or chemical modification. In terms of the mechanisms of MC-LR-induced toxicity, we mainly focus on the epigenetic modification and non-coding RNAs (ncRNAs)-related mechanisms which have provided a new perspective.

Analysis of miRNA and mRNA expression in the dysregulation of insulin secretion in MIN6 cells exposed to microcystin-leucine-arginine.

Microcystin -leucine-arginine (MC-LR), produced by freshwater cyanobacteria, is a potential pancreatic ß-cell toxin. In this study, the function of the mouse pancreatic ß-cell line, MIN6, was evaluated after MC-LR exposure, and the underlying molecular mechanisms were explored. Exposure to MC-LR for 24 h was found to inhibit cell viability and impair insulin secretion. Such findings indicate that ß-cell function would be impaired following MC-LR treatment. The microarray results revealed altered miRNA and mRNA expression profiles that might be responsible for the abnormal function of MIN6 cells. Further, miRNA-gene network analysis demonstrated that miR-29b-3p, miR-6967-5p, miR-3473, miR-7061-5p, Xkr4, Tmem178b, Scp2, Ypel2, and Kcnj11 are key miRNAs and genes in the MC-LR-induced MIN6-cell toxicity. The altered expression levels of several miRNAs (e.g., miR-320-5p, miR-770-5p, miR-99a-3p, and miR-375-5p) and genes (e.g., Pklr and Gpd2) involved in insulin secretion or the onset of diabetes were also identified in MIN6 cells after treatment with MC-LR. Collectively, these findings provide evidence of the toxic effects of MC-LR on ß-cells and the underlying molecular mechanisms of its glycometabolism toxicity. MCs may thus possibly play an important role in the development of diabetes mellitus in humans.

Repeated exposure to microcystin-leucine-arginine potentiates excitotoxicity induced by a low dose of kainate.

Microcystin-leucine-arginine (MLCR) is a cyanobacterial toxin, and has been demonstrated to cause neurotoxicity. In addition, MCLR has been identified as an inhibitor of protein phosphatase (PP)1 and PP2A, which are known to regulate the phosphorylation of various molecules related to synaptic excitability. Thus, in the present study, we examined whether MCLR exposure affects seizures induced by a low dose of kainic acid (KA; 0.05 µg, i.c.v.) administration. KA-induced seizure occurrence and seizure score significantly increased after repeated exposure to MCLR (2.5 or 5.0 µg/kg, i.p., once a day for 10 days), but not after acute MCLR exposure (2.5 or 5.0 µg/kg, i.p., 2 h and 30 min prior to KA administration), and hippocampal neuronal loss was consistently facilitated by repeated exposure to MCLR. In addition, repeated MCLR significantly elevated the membrane expression of kainate receptor GluK2 subunits, p-pan-protein kinase C (PKC), and p-extracellular signal-related kinase (ERK) at 1 h after KA. However, KA-induced membrane expression of Ca2+/calmodulin-dependent kinase II (CaMKII) was significantly reduced by repeated MCLR exposure. Consistent with the enhanced seizures and neurodegeneration, MCLR exposure significantly potentiated KA-induced oxidative stress and microglial activation, which was accompanied by increased expression of p-ERK and p-PKCδ in the hippocampus. The combined results suggest that repeated MCLR exposure potentiates KA-induced excitotoxicity in the hippocampus by increasing membrane GluK2 expression and enhancing oxidative stress and neuroinflammation through the modulation of p-CaMKII, p-PKC, and p-ERK.

Effects of Chronic Exposure to Microcystin-LR on the Gut Microbiota of Male Mice.

Increasing evidence indicates that environmental pollutants can change human gut microbiota. Microcystin-leucine arginine (MC-LR), considered a major hazard to mammals, is one of the important contaminants. However, little is known about the long-term influence of MC-LR on gut microbial communities. We aimed to investigate the effect of MC-LR on gut microbiota composition and functions by conducting a chronic exposure of male mice to MC-LR via the oral route. Using 16S rRNA gene sequencing analysis on cecum samples of mice, our results showed that significant changes of species diversity were observed in the gut microbiota of MC-LR-exposed mice. In addition, comparative analysis of the microbial communities showed that the reduction of the Actinobacteria and Saccharibacteria populations was detected in MC-LR-exposed mice. Collectively, our study highlighted the significant effects of MC-LR on the shift of gut microbial communities which could contribute to the development of metabolic syndromes.

Glutathione peroxidase-1 overexpressing transgenic mice are protected from neurotoxicity induced by microcystin-leucine-arginine.

Although it has been well-recognized that microcystin-leucine-arginine (MCLR), the most common form of microcystins, induces neurotoxicity, little is currently known about the underlying mechanism for this neurotoxicity. Here, we found that MCLR (10 ng/µL/mouse, i.c.v.) induces significant neuronal loss in the hippocampus of mice. MCLR-induced neurotoxicity was accompanied by oxidative stress, as shown by a significant increase in the level of 4-hydroxynonenal, protein carbonyl, and reactive oxygen species (ROS). Superoxide dismutase-1 (SOD-1) activity was significantly increased, but glutathione peroxidase (GPx) level was significantly decreased following MCLR insult. In addition, MCLR significantly inhibited GSH/GSSG ratio, and significantly induced NFκB DNA binding activity. Because reduced activity of GPx appeared to be critical for the imbalance between activities of SODs and GPx, we utilized GPx-1 overexpressing transgenic mice to ascertain the role of GPx-1 in this neurotoxicity. Genetic overexpression of GPx-1 or NFκB inhibitor pyrrolidine dithiocarbamate (PDTC) significantly attenuated MCLR-induced hippocampal neuronal loss in mice. However, PDTC did not exert any additive effect on neuroprotection mediated by GPx-1 overexpression, indicating that NFκB is a neurotoxic target of MCLR. Combined, these results suggest that MCLR-induced neurotoxicity requires oxidative stress associated with failure in compensatory induction of GPx, possibly through activation of the transcription factor NFκB.

Polymer-Lipid Microparticles for Pulmonary Delivery.

Toward engineering approaches that are designed to optimize the particle size, morphology, and mucoadhesion behavior of the particulate component of inhaler formulations, this paper presents the preparation, physicochemical characterization, and preliminary in vitro evaluation of multicomponent polymer-lipid systems that are based on "spray-drying engineered" α-lactose monohydrate microparticles. The formulations combine an active (budesonide) with a lung surfactant (dipalmitoylphosphatidylcholine) and with materials that are known for their desirable effects on morphology (polyvinyl alcohol), aerosolization (l-leucine), and mucoadhesion (chitosan). The effect of the composition of formulations on the morphology, distribution, and in vitro mucoadhesion profiles is presented along with "Calu-3 cell monolayers" data that indicate good cytocompatibility and also with simulated-lung-fluid data that are consistent with the therapeutically useful release of budesonide.

A Toxicological Assessment of Creatyl-l-Leucine.

A battery of toxicological studies was conducted to investigate the genotoxicity and repeated-dose oral toxicity of creatyl-l-leucine, a synthetic compound, in rats in accordance with internationally accepted guidelines. There was no evidence of mutagenicity in a bacterial reverse mutation test and in an in vitro mammalian chromosomal aberration test. There was no genotoxic activity observed in an in vivo mammalian micronucleus test at concentrations up to the limit dose of 2,000 mg/kg bw/d. Creatyl-l-leucine did not cause mortality or toxic effects in Hsd.Han Wistar rats in a 90-day repeated-dose oral (gavage) toxicity study at doses of 1,250, 2,500, and 5,000 mg/kg bw/d. The no observed adverse effect level from the 90-day study was determined to be 5,000 mg/kg bw/d, the highest dose tested, for both male and female rats.

From the Cover: Roles of mmu_piR_003399 in Microcystin-Leucine Arginine-Induced Reproductive Toxicity in the Spermatogonial Cells and Testis.

Our previous work has demonstrated that microcystin-leucine arginine (MC-LR) is a potent toxin for the reproductive system of male mice and it exerts cytotoxicity in spermatogonial cells, resulting in the constitutional and functional changes of the mouse testis. The present study was designed to investigate the functions of P-element-induced wimpy (piwi)-interacting RNAs (piRNAs) in MC-LR-induced reproductive toxicity in male mice. We observed an increase in the mmu_piR_003399 level in spermatogonial cells and mouse testicular tissues following treatment with MC-LR. Moreover, our data confirmed that cyclin-dependent kinase 6 (CDK6) was the target gene of mmu_piR_003399. Increases in the concentration of mmu_piR_003399 were correlated with the reduced expression of CDK6 both in vitro and in vivo. mmu_piR_003399 induced cell cycle arrest at the G1-phase, down-regulated sperm counts and sperm motility, and compromised sperm morphology. On the contrary, suppressing the expression of mmu_piR_003399 could substantially attenuate MC-LR-induced pathology in mice including cell cycle arrest, reduced mature sperm counts, sperm viability loss and abnormal sperm morphology. Furthermore, our data supported that mmu_piR_003399 existed in mouse serum and plasma, and its level was increased in MC-LR-treated mice. In conclusion, we demonstrate that mmu_piR_003399 plays a crucial role in regulating MC-LR-induced reproductive toxicity.

Roles of piRNAs in microcystin-leucine-arginine (MC-LR) induced reproductive toxicity in testis on male offspring.

In the present study, we evaluated the toxic effects on the testis of the male offspring of MC-LR exposure during fetal and lactational periods. Pregnant females were distributed into two experimental groups: control group and MC-LR group which were exposed to 0 and 10 µg/L of MC-LR, respectively, through drinking water separately during fetal and lactational periods. At the age of 30 days after birth, the male offspring were euthanized. The body weight, testis index, and histomorphology change were observed and the global changes of piwi-interacting RNA (piRNA) expression were evaluated. The results revealed that MC-LR was found in the testis of male offspring, body weight and testis index decreased significantly, and testicular tissue structure was damaged in the MC-LR group. In addition, the exposure to MC-LR resulted in an altered piRNA expression profile and an increase of the cell apoptosis and a decrease of the cell proliferation in the testis of the male offspring. It was reasonable to speculate that the toxic effects on reproductive system of the male offspring in MC-LR group might be mediated by piRNAs through the regulation of the target genes. As far as we are aware, this is the first report showing that MC-LR could play a role in disorder of proliferative and cell apoptosis in the testis of the male offspring by the maternal transmission effect of toxicity.

Activation of the Neuroprotective Angiotensin-Converting Enzyme 2 in Rat Ischemic Stroke.

The angiotensin-converting enzyme 2/angiotensin-(1-7)/Mas axis represents a promising target for inducing stroke neuroprotection. Here, we explored stroke-induced changes in expression and activity of endogenous angiotensin-converting enzyme 2 and other system components in Sprague-Dawley rats. To evaluate the clinical feasibility of treatments that target this axis and that may act in synergy with stroke-induced changes, we also tested the neuroprotective effects of diminazene aceturate, an angiotensin-converting enzyme 2 activator, administered systemically post stroke. Among rats that underwent experimental endothelin-1-induced ischemic stroke, angiotensin-converting enzyme 2 activity in the cerebral cortex and striatum increased in the 24 hours after stroke. Serum angiotensin-converting enzyme 2 activity was decreased within 4 hours post stroke, but rebounded to reach higher than baseline levels 3 days post stroke. Treatment after stroke with systemically applied diminazene resulted in decreased infarct volume and improved neurological function without apparent increases in cerebral blood flow. Central infusion of A-779, a Mas receptor antagonist, resulted in larger infarct volumes in diminazene-treated rats, and central infusion of the angiotensin-converting enzyme 2 inhibitor MLN-4760 alone worsened neurological function. The dynamic alterations of the protective angiotensin-converting enzyme 2 pathway after stroke suggest that it may be a favorable therapeutic target. Indeed, significant neuroprotection resulted from poststroke angiotensin-converting enzyme 2 activation, likely via Mas signaling in a blood flow-independent manner. Our findings suggest that stroke therapeutics that target the angiotensin-converting enzyme 2/angiotensin-(1-7)/Mas axis may interact cooperatively with endogenous stroke-induced changes, lending promise to their further study as neuroprotective agents.

Recovery: revisão sistemática de um conceito / Recovery: systematic review of a concept

O conceito de recovery tem sido descrito em artigos como um estado de recuperação ou restabelecimento de funções psíquicas, físicas e sociais no funcionamento cotidiano. O objetivo do artigo é analisar concepções terminológicas em diferentes metodologias investigativas e a evolução paradigmática da noção de recovery. Pesquisa bibliográfica sistemática na base Pubmed com as palavras "recovery + schizophrenia", limitada a dois anos retrospectivos e a artigos completos gratuitos. Dezenove artigos foram analisados. A maioria destes busca associações entre dada característica e recovery, poucos são aqueles que discutem a sua concepção de forma que se distinga de termos comuns como "cura" e "reabilitação". Recovery como um estado em que o portador de transtorno mental grave possa sentir-se criador de seus caminhos tende a estar presente em estudos com metodologia qualitativa e em revisões bibliográficas, em que a medida de recovery deixa de relacionar-se à ausência de sintomas e passa a priorizar o quão participativa pode ser a vida de um indivíduo apesar da doença. Alguns estudos quantitativos vislumbram essa diferença conceitual. Em pesquisas qualitativas ocorre expansão na concepção de recovery e nas formas de promovê-lo.

The concept of recovery has been described in papers as a state of psychic, physical and social recuperation of day-to-day functions. The scope of this article is to analyze the concepts of the term in different research methodologies and the paradigmatic evolution of the recovery concept. Systematic bibliographical research was conducted in the Pubmed database using the words "recovery + schizophrenia" limited to freely available full papers published in the previous two years. Nineteen papers were analyzed. The majority of the papers sought associations between characteristic data and recovery; few papers discussed the concept in a way to distinguish it from other words like cure or rehabilitation. Recovery as a state in which people with severe mental illness can feel like the creators of their own itinerary tend to be found in qualitative studies and in bibliographic reviews in which the meaning of recovery is not related to the lack of symptoms and tends to prioritize how participative the life of an individual can be despite the disease. Some quantitative studies detect this conceptual difference. In qualitative research there is an increase in the concept of recovery and in ways of promoting it.

Diagnosis, classification and treatment of gestational trophoblastic neoplasia / Diagnóstico, classificação e tratamento da neoplasia trofoblástica gestacional

Gestational trophoblastic neoplasia (GTN) is the term to describe a set of malignant placental diseases, including invasive mole, choriocarcinoma, placental site trophoblastic tumor and epithelioid trophoblastic tumor. Both invasive mole and choriocarcinoma respond well to chemotherapy, and cure rates are greater than 90%. Since the advent of chemotherapy, low-risk GTN has been treated with a single agent, usually methotrexate or actinomycin D. Cases of high-risk GTN, however, should be treated with multiagent chemotherapy, and the regimen usually selected is EMA-CO, which combines etoposide, methotrexate, actinomycin D, cyclophosphamide and vincristine. This study reviews the literature about GTN to discuss current knowledge about its diagnosis and treatment.

Neoplasia trofoblástica gestacional (NTG) é o termo que descreve o conjunto de anomalias malignas da placenta, incluindo a mola invasora, coriocarcinoma, tumor trofoblástico do sítio placentário e tumor trofoblástico epitelióide. Ambos a mola invasora e o coriocarcinoma respondem bem à quimioterapia, com taxas de cura superiores a 90%. Desde o advento da quimioterapia, NTG de baixo risco tem sido tratada com monoquimioterapia, pelo geral methotrexate ou actinomicina-D. Casos de NTG de alto risco, contudo, devem ser tratados com poliquimioterapia, e o regime usualmente escolhido é o EMA-CO que combina etoposide, methotrexate, actinomicina-D, ciclofosfamida e vincristina. Esse estudo revê a literatura sobre NTG a fim de discutir os conhecimentos atuais sobre seu diagnóstico e tratamento.

Cytotoxicity and pro-inflammatory action of chemo-mechanical caries-removal agents against oral cells.

BACKGROUND: Chemo-mechanical caries removal eliminates the outermost portion of the infected layer, leaving behind healthy dentine surfaces, with scarce dental tissue damage; however, the safety of caries solvents has not been established. The aim of the present study was to investigate the possible cytotoxicity of two popular chemo-mechanical caries removal agents. MATERIALS AND METHODS: The cytotoxicity of Carisolv, Papacarie Duo and control vehicle solution (0.155-20% v/v) against human oral squamous cell carcinoma cells (HCS-2, HSC-3, HSC-4, Ca9-22) human gingival fibroblast (HGF), pulp (HPC) and periodontal ligament fibroblast (HPLF) was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide method. Prostaglandin E2 (PGE2) was quantified by enzyme-linked immunosorbent assay. Changes in fine cell structure were assessed by transmission electron microscopy. RESULTS: Carisolv exhibited neither cytotoxicity nor hormetic growth stimulation. Papacarie Duo significantly reduced the viable cell number within 30 min. HSC-4 exhibited the highest sensitivity, followed by HSC-2>HSC-3>HPLF>Ca9-22>HPC>HGF cells. Interleukin-1ß (3 ng/ml) stimulated HGF, but not HPC cells to produce PGE2 in the culture medium. Papacarie Duo stimulated HGF cells to produce PGE2 in synergistic fashion with interleukin-1ß. CONCLUSION: Carisolv had acceptable biocompatibility with both normal and cancerous oral cells. On the other hand, Papacarie Duo had a rapid but slight cytotoxicity and pro-inflammatory action against oral cells, suggesting the importance of careful application of this agent.

Summary of workshop discussions on establishing upper limits for amino acids with specific attention to available data for the essential amino acids leucine and tryptophan.

The morning of the first day of the 8th Amino Acid Assessment Workshop was organized and co-sponsored by the International Council on Amino Acid Science (ICAAS) and the International Life Sciences Institute Research Foundation and was focused on the International Life Sciences Institute Research Foundation's approach to establishing upper limits of nutrients. The remainder of d 1 and all of d 2 were focused on the safety of leucine and tryptophan, with special emphasis on determining the upper level of the safe range of intake. It was recognized that some toxicological frameworks, mainly the key-events dose response framework, might be applicable to amino acids and provide appropriate assistance to regulators in establishing upper limits for amino acids as a group of nutrients used in dietary supplements. ICAAS-funded projects for determining the upper intake limits for the essential amino acid leucine provided the main pool of leucine data discussed at the workshop. The acute clinical study suggests 500 mg/(kg · d) as a possible upper limit for leucine in healthy humans, but the safety margin needed to widen this limit to the general population has not been determined. For tryptophan, the workshop participants found less ground for consensus. Older efficacy studies suggested that tryptophan at 8-15 g/d was well tolerated, but human research was abruptly terminated in the late 1980s and no new data are available. Animal results obtained in pigs and rodents were discussed and 2 possible strategies for applying those outcomes to humans were described.

Dry powder inhalers of gentamicin and leucine: formulation parameters, aerosol performance and in vitro toxicity on CuFi1 cells.

The high hygroscopicity of gentamicin (G) as raw material hampers the production of respirable particles during aerosol generation and prevents its direct use as powder for inhalation in patients suffering from cystic fibrosis (CF). Therefore, this research aimed to design a new dry powder formulation of G studying dispersibility properties of an aminoacid, L-leucine (leu), and appropriate process conditions. Spray-dried powders were characterized as to water uptake, particle size distribution, morphology and stability, in correlation with process parameters. Aerodynamic properties were analyzed both by Single Stage Glass Impinger and Andersen Cascade Impactor. Moreover, the potential cytotoxicity on bronchial epithelial cells bearing a CFTR F508/F508 mutant genotype (CuFi1) were tested. Results indicated that leu may improve the aerosol performance of G-dried powders. The maximum fine particle fraction (FPF) of about 58.3% was obtained when water/isopropyl alcohol 7:3 system and 15-20% (w/w) of leu were used, compared to a FPF value of 13.4% for neat G-dried powders. The enhancement of aerosol efficiency was credited both to the improvement of the powder flowability, caused by the dispersibility enhancer (aminoacid), and to the modification of the particle surface due to the influence of the organic co-solvent on drying process. No significant degradation of the dry powder was observed up to 6 months of storage. Moreover, particle engineering did not affect either the cell viability or cell proliferation of CuFi1 over a 24 h period.

First-time report of metalloproteinases in fish bile and their potential as bioindicators regarding environmental contamination.

Gallbladder bile from 2 fish species, mullet (Mugil liza) and tilapias (Tilapia rendalli), contain substantial matrix metalloproteinases (MMPs). Extensive purification studies were conducted in order to obtain workable samples for SDS-PAGE and zymography analysis. Proteinase activities were assayed by gelatin substrate zymography. Several protein bands were observed, corresponding to molecular weights of 200, 136, 43, 36, 34, 29, 23 and 14 kDa in mullet bile and 179, 97, 79, 61, 54, 45, 36, 33 and 21 kDa in tilapia bile. Specific inhibitor studies were conducted, in which MMPS were inhibited by EDTA and 1,10 phenanthroline, but not by serine and cysteine protease inhibitors, such as phenylmethylsulfonyl fluoride (PMSF) and transepoxysuccinyl-l-leucylamido-l-guanidino butane (E-64), confirming the proteinase identities as MMPs. Differences in proteinase expression were observed in fish from a contaminated and reference site. Some studies regarding MMPs in different fish tissues exist, however this is the first study conducted in fish bile, and their involvement in detoxification processes and organism protection against the effects of aquatic contaminants may be a possibility.

Development of an early-tier laboratory bioassay for assessing the impact of orally-active insecticidal compounds on larvae of Coccinella septempunctata (Coleoptera: Coccinellidae).

Early-tier studies are the initial step in the environmental risk assessment of genetically engineered plants on nontarget arthropods. They are conducted in the laboratory where surrogate species are exposed to higher concentrations of the arthropod-active compound than those expected to occur in the field. Thus, early-tier tests provide robust data and allow to make general conclusions about the susceptibility of the surrogate to the test substance. We have developed an early-tier test for assessing the toxicity of orally-active insecticidal compounds to larvae of the ladybird beetle Coccinella septempunctata L. (Coleoptera: Coccinellidae). Using potassium arsenate and the protease inhibitor E-64 as model compounds, we validated the bioassay set-up for C. septempunctata. Sucrose solution containing the test compound was offered to larvae for 24 h on the first day of each of its four larval instars. Subsequently, larvae were fed ad libitum with Ephestia kuehniella (Zeller) (Lepidoptera: Pyralidae) eggs. Both compounds negatively affected C. septempunctata larval survival and development, and adult weight, indicating that the bioassay setup was able to detect dietary effects of insecticidal substances on the ladybird. Power analyses revealed that sample sizes of 20 or 45 per treatment are sufficient to detect 50 or 20% differences between the control and treatment groups, respectively, for the various measurement endpoints.

Synthesis, characterization, cytotoxic activities, and DNA-binding studies of ternary copper(II) complexes with new coumarin derivatives.

In this study, two novel complexes [Cu(MCVH)phen(H(2)O)].ClO(4) (1) and [Cu(MCLH)phen(H(2)O)].ClO(4) (2) (here, MCVH(2)=(7-hydroxy-4-methyl-8-coumarinyl) valine, MCLH(2)=(7-hydroxy-4-methyl-8-coumarinyl) leucine) have been synthesized and characterized by elemental analyses, molar conductance, infrared spectra (IR), (1)H-NMR and UV-Vis measurements. The interactions of them with calf thymus DNA (ct DNA) have been investigated by absorption spectroscopy, fluorescence spectroscopy, circular dichroism spectroscopy and viscosity measurements. Experimental results reveal an intercalative interaction with DNA for the complexes, furthermore the binding affinity of 2 is higher than that of 1 according to the calculated binding constant values. In addition, they were evaluated for their cytotoxic activities toward human prostate cancer cell (PC3), human liver cell (L02) and human myeloid leukemia cancer cell (HL-60) by acid phosphoatase assay. Both of them showed significant cytotoxic potency.