Dimethylsulphoxide affects the organisation of microfilaments in the mouse oocyte

The effect of dimethylsulphoxide (DMSO) on microfilament organisation has been studied in the mouse oocyte after staining with (NBD)-phallacidin. The cortical actin meshwork was disrupted by exposure of oocytes to 1.5 M DMSO at 37 degrees C, and this disruption was associated with changes in the cell surface, especially microvilli length and distribution, as observed by scanning electron microscopy. The irregular distribution of actin filaments observed also appears to lead to an irregular expansion of the cell after DMSO removal. However, when exposure to DMSO was combined with cooling, the effects on the microfilament system were much reduced. The reversibility of DMSO action is considered and the potential implications of microfilament disruption on the viability and functions of the oocyte discussed.     

The TX;Y test for the detection of nondisjunction and chromosome breakage in Drosophila melanogaster. II. Results of female exposures

The TX; Y test is a short-term assay for the detection of sex-chromosome nondisjunction and chromosome breakage in Drosophila melanogaster. It has been used in previous work following the exposure of males. In this work, females are exposed. When females are the exposed parent, only chromosome gain can be detected. Positive results for the induction of aneuploidy were obtained following exposures of females to X-rays, 10 degrees C cold shock, and colchicine. No increase in aneuploidy was obtained following exposures of females to DMSO and trifluralin. Comparison with similar work in males reveals no consistent pattern concerning the more appropriate sex to use for aneuploidy testing in Drosophila, as colchicine was found to be positive in females only and DMSO and trifluralin were effective in males only. Further work is necessary to validate the TX; Y test and to understand the relative efficacy of female and male exposures to aneuploidy inducing agents in Drosophila.     

Molecular study of the diffusional process of DMSO in double lipid bilayers

As a way to quantify the diffusion process of molecular compounds through biological membranes, we investigated in this study the dynamics of DMSO through an 1,2-Dipalmitoyl-sn-Glycero-3-Phosphocholine (DPPC) bilayer system. To properly account for the diffusion of DMSO due to a concentration gradient, a double DPPC bilayer was setup for our simulations. In such configuration, the aqueous phases can be explicitly associated with the extra and intracellular domains of the membrane, which is seldom the case in studies of single lipid bilayer due to the periodicity imposed by the simulations. DMSO molecules were initially contained in one of the aqueous phases (extracellular region) at a concentration of 5 wt.%. Molecular dynamics simulation was performed in this system for 95 ns at 350 K and 1 bar. The simulations showed that although many DMSO molecules penetrated the lipid bilayer, only about 10% of them crossed the bilayer to reach the other aqueous phase corresponding to the intracellular region of the membrane. The simulation time considered was insufficient to reach equilibrium of the DMSO concentration between the aqueous phases. However, the simulations provided sufficient information to estimate parameters to apply Fick's Law to model the diffusion process of the system. Using this model, we predicted that for the time considered in our simulation, the concentration of DMSO in the intracellular domain should have been about half of the actual value obtained. The model also predicted that equilibrium of the DMSO concentration in the system would be reached after about 2000 ns, approximately 20 times longer than the performed simulation.     

Molecular study of the diffusional process of DMSO in double lipid bilayers

As a way to quantify the diffusion process of molecular compounds through biological membranes, we investigated in this study the dynamics of DMSO through an 1,2-Dipalmitoyl-sn-Glycero-3-Phosphocholine (DPPC) bilayer system. To properly account for the diffusion of DMSO due to a concentration gradient, a double DPPC bilayer was setup for our simulations. In such configuration, the aqueous phases can be explicitly associated with the extra and intracellular domains of the membrane, which is seldom the case in studies of single lipid bilayer due to the periodicity imposed by the simulations. DMSO molecules were initially contained in one of the aqueous phases (extracellular region) at a concentration of 5 wt.%. Molecular dynamics simulation was performed in this system for 95 ns at 350 K and 1 bar. The simulations showed that although many DMSO molecules penetrated the lipid bilayer, only about 10% of them crossed the bilayer to reach the other aqueous phase corresponding to the intracellular region of the membrane. The simulation time considered was insufficient to reach equilibrium of the DMSO concentration between the aqueous phases. However, the simulations provided sufficient information to estimate parameters to apply Fick's Law to model the diffusion process of the system. Using this model, we predicted that for the time considered in our simulation, the concentration of DMSO in the intracellular domain should have been about half of the actual value obtained. The model also predicted that equilibrium of the DMSO concentration in the system would be reached after about 2000 ns, approximately 20 times longer than the performed simulation.     

Early Early Man in North America and where to look for Him: Geomorphic Contexts

Abstract

There has long been a controversy concerning Man’s antiquity in the New World. Our oldest unquestioned sites are those of the Clovis Complex which date to 11,000-12,000 B.P. In this paper, an attempt is made to set up a testing program that might lead to the discovery of Pre-Clovis (Early Early Man) sites within the contiquous 48 states, should they exist. It is believed that the derived concepts also have application in other parts of the world. Rigorous standards are given for the “best site situation” necessary to solve the problem for everyone’s satisfaction. The methodology involves outlining a list of possible archaeo-geomorphic contexts in which to seek ancient sites, with the idea of determining if any of them are more likely to produce the best site situation than others. Nine archaeo-geomorphic contexts are discussed and evaluated. These include surface sites and the following subsurface sites: underwater, coastal, volcanic, eolian, slope, cave, alluvial, and depression or basin sites. It is concluded, by the process of elimination, that the latter seem to be our best bet to meet the aforementioned rigorous standards. A testing program is suggested on the High Plains where many basins occur, and where some of our better early sites have been found. How we should test is discussed in terms of locating basins, choosing basins to test, and testing procedures. Subsidiary benefits are seen resulting from this program even if the problem of Man’s antiquity should not be determined by this method of research.     

Induction of the tyrosine phosphorylation of a 66 kd soluble protein by DMSO in human peripheral blood T lymphocytes

Previous studies from our laboratory have demonstrated that a 66 KD cytoplasmic protein (TPP 66) is newly phosphorylated on tyrosine when human peripheral blood T lymphocytes are incubated with a variety of agents that activate these cells or augment activation by known mitogens. Since DMSO has been shown to activate tyrosine specific protein kinases we have examined the role of this agent on the phosphorylation of TPP 66. 5 to 40% DMSO induced the phosphorylation of TPP 66 with the maximal increase in phosphorylation seen at 20%. Concentrations greater than 40% were inhibitory. Phosphorylation of TPP 66 in DMSO treated cells could be detected as early as 2 min following the addition of DMSO, with increased [32P]O4 incorporation over the next 60 min. The phosphorylation was on tyrosine residues demonstrated by base hydrolysis of TPP 66 extracted from the gels followed by single dimension high voltage electrophoresis. Since DMSO augments activation of T lymphocytes by lectins, this data provides further support for a critical role for the tyrosine phosphorylation of TPP 66 in the mediation or modulation of T lymphocyte activation.     

Dimethyl Sulfoxide (DMSO) Exacerbates Cisplatin-induced Sensory Hair Cell Death in Zebrafish (Danio rerio)

Inner ear sensory hair cells die following exposure to aminoglycoside antibiotics or chemotherapeutics like cisplatin, leading to permanent auditory and/or balance deficits in humans. Zebrafish (Danio rerio) are used to study drug-induced sensory hair cell death since their hair cells are similar in structure and function to those found in humans. We developed a cisplatin dose-response curve using a transgenic line of zebrafish that expresses membrane-targeted green fluorescent protein under the control of the Brn3c promoter/enhancer. Recently, several small molecule screens have been conducted using zebrafish to identify potential pharmacological agents that could be used to protect sensory hair cells in the presence of ototoxic drugs. Dimethyl sulfoxide (DMSO) is typically used as a solvent for many pharmacological agents in sensory hair cell cytotoxicity assays. Serendipitously, we found that DMSO potentiated the effects of cisplatin and killed more sensory hair cells than treatment with cisplatin alone. Yet, DMSO alone did not kill hair cells. We did not observe the synergistic effects of DMSO with the ototoxic aminoglycoside antibiotic neomycin. Cisplatin treatment with other commonly used organic solvents (i.e. ethanol, methanol, and polyethylene glycol 400) also did not result in increased cell death compared to cisplatin treatment alone. Thus, caution should be exercised when interpreting data generated from small molecule screens since many compounds are dissolved in DMSO.     

Development of a new rapid measurement technique for fish embryo membrane permeability studies using impedance spectroscopy

Information on fish embryo membrane permeability is vital in their cryopreservation. Whilst conventional volumetric measurement based assessment methods have been widely used in fish embryo membrane permeability studies, they are lengthy and reduce the capacity for multi-embryo measurement during an experimental run. A new rapid 'real-time' measurement technique is required to determine membrane permeability during cryoprotectant treatment. In this study, zebrafish (Danio rerio) embryo membrane permeability to cryoprotectants was investigated using impedance spectroscopy. An embryo holding cell, capable of holding up to 10 zebrafish embryos was built incorporating the original system electrods for measuring the impedance spectra. The holding cell was tested with deionised water and a series of KCl solutions with known conductance values to confirm the performance of the modified system. Untreated intact embryos were then tested to optimise the loading capacity and sensitivity of the system. To study the impedance changes of zebrafish embryos during cryoprotectant exposure, three, six or nine embryos at 50% epiboly stage were loaded into the holding cell in egg water, which was then removed and replaced by 0.5, 1.0, 2.0 or 3M methanol or dimethyl sulfoxide (DMSO). The impedance changes of the loaded embryos in different cryoprotectant solutions were monitored over 30 min at 22 degrees C, immediately following embryo exposure to cryoprotectants, at the frequency range of 10-10(6)Hz. The impedance changes of the embryos in egg water were used as controls. Results from this study showed that the optimum embryo loading level was six embryos per cell for each experimental run. The optimum frequency was identified at 10(3.14) or 1,380 Hz which provided good sensitivity and reproducibility. Significant impedance changes were detected after embryos were exposed to different concentrations of cryoprotectants. The results agreed well with those obtained from conventional volumetric based studies.     

NEW BIOLOGICAL DETECTION SYSTEM FOR WEAK ELF MAGNETIC FIELDS AND TESTING OF THE PARAMETRIC RESONANCE MODEL (Lednev 1991)

A new detection system for weak, extremely low frequency (ELF) magnetic fields based on bioluminescence of the autotrophic dinoflagellate Gonyaulax sp. is presented. Due to its sensitivity to external factors, this biological system has already been thoroughly investigated and used for various biological research. In our study, the system was first tested for its capacity to respond to weak ELF magnetic fields (50 Hz, 11.5 mT; 50 Hz, 1.2 mT). After its sensitivity was established, the system was used to test the parametric resonance model proposed by Lednev in 1991. Our results seem to be consistent with the model predictions, especially when monitoring real-time effects of the exposure.     

Long- and Short-Term Health Effects of Pesticide Exposure: A Cohort Study from China

Pesticides are extensively used by farmers in China. However, the effects of pesticides on farmers’ health have not yet been systematically studied. This study evaluated the effects of pesticides exposure on hematological and neurological indicators over 3 years and 10 days respectively. A cohort of 246 farmers was randomly selected from 3 provinces (Guangdong, Jiangxi, and Hebei) in China. Two rounds of health investigations, including blood tests and neurological examinations, were conducted by medical doctors before and after the crop season in 2012. The data on pesticide use in 2009–2011 were collected retrospectively via face-to-face interviews and the 2012 data were collected from personal records maintained by participants prospectively. Ordinary least square (OLS), Probit, and fixed effect models were used to evaluate the relationship between pesticides exposure frequency and the health indicators. Long-term pesticide exposure was found to be associated with increased abnormality of nerve conductions, especially in sensory nerves. It also affected a wide spectrum of health indicators based on blood tests and decreased the tibial nerve compound muscle action potential amplitudes. Short-term health effects included alterations in complete blood count, hepatic and renal functions, and nerve conduction velocities and amplitudes. However, these effects could not be detected after 3 days following pesticide exposure. Overall, our results demonstrate that pesticide exposure adversely affects blood cells, the liver, and the peripheral nervous system. Future studies are needed to elucidate the specific effects of each pesticide and the mechanisms of these effects.     

Dimethyl Sulfoxide Damages Mitochondrial Integrity and Membrane Potential in Cultured Astrocytes

Dimethyl sulfoxide (DMSO) is a polar organic solvent that is used to dissolve neuroprotective or neurotoxic agents in neuroscience research. However, DMSO itself also has pharmacological and pathological effects on the nervous system. Astrocytes play a central role in maintaining brain homeostasis, but the effect and mechanism of DMSO on astrocytes has not been studied. The present study showed that exposure of astrocyte cultures to 1% DMSO for 24 h did not significantly affect cell survival, but decreased cell viability and glial glutamate transporter expression, and caused mitochondrial swelling, membrane potential impairment and reactive oxygen species production, and subsequent cytochrome c release and caspase-3 activation. DMSO at concentrations of 5% significantly inhibited cell variability and promoted apoptosis of astrocytes, accompanied with more severe mitochondrial damage. These results suggest that mitochondrial impairment is a primary event in DMSO-induced astrocyte toxicity. The potential cytotoxic effects on astrocytes need to be carefully considered during investigating neuroprotective or neurotoxic effects of hydrophobic agents dissolved by DMSO.     

Study on fish embryo responses to the treatment of cryoprotective chemicals using impedance spectroscopy

Investigations using electrical impedance spectroscopy to measure the responses of fish embryos to the cryoprotective chemicals, methanol and dimethyl sulphoxide (DMSO), were carried out. Zebrafish (Danio rerio) embryos were used as a model to study the newly proposed technique. The normalised permittivity and conductivity changes of the embryos were measured continuously over a 20-min period in a customised embryo-holding chamber. The normalised permittivity and conductivity spectra were obtained during embryo exposure to different concentrations of methanol (1.0, 2.0 and 3.0 M) and DMSO (0.5, 1.0 and 2.0 M) solutions. The results showed significant permittivity and conductivity changes after embryo exposure to methanol and DMSO at the optimum embryo loading level (six embryos). Embryos in different concentrations of methanol and DMSO also resulted in quantitative responses shown in the normalised permittivity and conductivity spectra. The results demonstrated that fish embryo membrane permeability to cryoprotective chemicals could be monitored in real-time. The measurement of permittivity at a lower frequency range (10–10³ Hz) and conductivity at a higher frequency range (10⁴–10⁶ Hz) during fish embryo exposure to cryoprotective chemicals using impedance spectroscopy can be used as a new tool for the fast screening of most effective cryoprotective chemicals. The results from the present study also demonstrated the possibility of quantifying the level of cryoprotective chemicals penetrating the fish embryos.     

The influence of lipid‐extraction and long‐term DMSO preservation on carbon (δ13C) and nitrogen (δ15N) isotope values in cetacean skin

Stable isotope analysis (SIA) has rapidly become a useful tool to study the ecology of wild animal populations, especially for elusive, wide‐ranging predators like marine mammals. The development of projectile biopsy techniques resulted in the collection of thousands of cetacean tissue samples that were archived in a dimethyl sulfoxide (DMSO) solution for long‐term, multidecadal preservation. Here we examine the influence of DMSO preservation on carbon (δ¹³C) and nitrogen (δ¹⁵N) values by comparing a set of paired delphinid skin samples stored frozen without preservative and in DMSO for up to 22 yr. Treatment of paired frozen and DMSO‐preserved skin in a 2:1 chloroform:methanol solution yielded similar δ¹³C and δ¹⁵N values, revealing that DMSO and lipid contamination have similar isotopic effects on skin, and that these effects can be removed using routine lipid‐extraction methods. Further, amino acid concentrations in DMSO‐preserved and frozen skin tissue were similar, providing independent evidence of minimal protein alteration due to preservation. Access to a rich archive of skin samples preserved in DMSO will expand our ability to examine temporal and spatial variability in the isotope values of cetaceans, which will aid our understanding of how their ecology has been influenced by historical changes in environmental conditions.     

Pesticides and cancer: insights into toxicoproteomic-based findings

Humans are often exposed to a variety of pollutants that contribute to an individual's risk for diseases including cancer. Animal, cell cultures and epidemiological lines of evidence demonstrate that exposure to various environmental pollutants including pesticides are associated with increasing frequency of cancers. Organophosphates, organochlorines, carbamates, pyrethroids, the major groups of pesticides, have been reported to be carcinogenic in various models. However, the results of these studies are still controversial, nevertheless, their mechanism of action is clear. Therefore, new strategies in toxicological research are needed for efficient screening for adverse effects of pesticides on complex living systems. Biomarkers can be employed to identify causal associations and to make better quantitative and qualitative estimates of those associations at relevant levels of exposure. This will enable us to deepen our understanding of mechanism behind their carcinogenic potential. Deciphering the associations between pesticide exposure and cancer, following toxicoproteomics application, will be useful in the development of potential predictive biomarkers of pesticide induced carcinogenicity. Therefore, the thrust of this article was to review the risk of cancer due to pesticide exposure and significant toxicoproteomic-based studies conducted so far, to identify the novel molecules as possible biomarkers for cancer following pesticide exposure.     

Structural analysis of Canavalia maritima and Canavalia gladiata lectins complexed with different dimannosides: new insights into the understanding of the structure-biological activity relationship in legume lectins

Plant lectins, especially those purified from species of the Leguminosae family, represent the best studied group of carbohydrate-binding proteins. The legume lectins from Diocleinae subtribe are highly similar proteins that present significant differences in the potency/efficacy of their biological activities. The structural studies of the interactions between lectins and sugars may clarify the origin of the distinct biological activities observed in this high similar class of proteins. In this way, this work presents a crystallographic study of the ConM and CGL (agglutinins from Canavalia maritima and Canavalia gladiata, respectively) in the following complexes: ConM/CGL:Man(alpha1-2)Man(alpha1-O)Me, ConM/CGL:Man(alpha1-3)Man(alpha1-O)Me and ConM/CGL:Man(alpha1-4)Man(alpha1-O)Me, which crystallized in different conditions and space group from the native proteins. The structures were solved by molecular replacement, presenting satisfactory values for R(factor) and R(free). Comparisons between ConM, CGL and ConA (Canavalia ensiformis lectin) binding mode with the dimannosides in subject, presented different interactions patterns, which may account for a structural explanation of the distincts biological properties observed in the lectins of Diocleinae subtribe.     

Increase in recombination rate in Arabidopsis thaliana plants sharing gaseous environment with X-ray and UVC-irradiated plants depends on production of radicals

X-ray and UVC are the two physical agents that damage DNA directly, with both agents capable of inducing double-strand breaks. Some of our recent work has demonstrated that local exposure to UVC results in a systemic increase in recombination frequency, suggesting that information about exposure can be passed from damaged to non-damaged tissue. Indeed, we recently showed that plants sharing the same enclosed environment with UVC-irradiated plants exhibit similar increase in homologous recombination frequency as irradiated plants. Here, we further tested whether yet another DNA-damaging agent, X-ray, is capable of increasing recombination rate (RR) in neighboring plants grown in a Petri dish. To test this, we grew plants exposed to X-ray or UVC irradiation in an enclosed environment next to non-exposed plants. We found that both X-ray and UVC-irradiated plants and neighboring plants exhibited comparable increases in the levels of strand breaks and the RR. We further showed that pre-exposure of plants to radical scavenger DMSO substantially alleviates the radiation-induced increase in RR and prevents formation of bystander signal. Our results suggest that the increase in RR in bystander plants can also be triggered by X-ray and that radicals may play some role in initiation or maintenance of this signal.     

Low-dose ionizing radiation and chromosome translocations: a review of the major considerations for human biological dosimetry

Chromosome translocations are a molecular signature of ionizing radiation exposure. Translocations persist significantly longer after exposure than other types of chromosome exchanges such as dicentrics. This persistence makes translocations the preferred aberration type for performing radiation dosimetry under conditions of protracted exposure or when exposure assessments are temporally delayed. Low doses of radiation are inherently difficult to quantify because the frequency of induced events is low and the background level of translocations among unexposed subjects can show considerable variability. Analyses of translocation frequencies can be confounded by several factors, including age of the subject, lifestyle choices such as cigarette smoking, the presence of clones of abnormal cells, and possibly genotypic variability among subjects. No significant effects of gender or race have been observed, but racial differences have not been completely ruled out. Translocation analyses may be complicated by the presence of different types of exchanges, i.e., reciprocal or non-reciprocal, and because translocations sometimes occur as a component of complex exchanges that include other forms of chromosome rearrangements. Rates of radiation exposure, ranging from acute to chronic, are known to influence the accumulation of translocations and may also affect their persistence. The influences on translocation frequencies of low-dose radiation hypersensitivity as well as the bystander effect and the adaptive response remain poorly characterized. Thus, quantifying the relationship between radiation dose and the frequency of translocations in any given subject requires attention to multiple issues. Part of the solution to understanding the in vivo dose-response relationship is to have accurate estimates of the baseline levels of translocations in healthy unexposed subjects, and some work in this area has been accomplished. Long-term cytogenetic follow-up of exposed subjects is needed to characterize translocation persistence, which is especially relevant for risk analyses. More work also needs to be done in the area of quantifying the role of known confounders. Characterizing the role of genotype will be especially important. Improvements in the ability to use translocation frequencies for low-dose biological dosimetry will require scoring very large numbers of cells per subject, which may be accomplished by developing a rapid automated image analysis system. This work would enhance our comprehension of the effects of low-dose radiation exposure and could lead to significant improvements in understanding the relationship between chromosome damage and human health.     

Effects of dimethylsulfoxide (DMSO) on the digestive-lysosomal system in Paramecium caudatum

The effects of DMSO (dimethylsulfoxide) on cell growth and on the digestive-lysosomal system of axenically grown Paramecium caudatum were studied. A general protocol of exposing cells to different concentrations of DMSO at the beginning of each of the four processes in the digestive cycle enabled us to analyze the effect of DMSO at each step. Vacuole formation and the beginning of a digestive cycle were initiated by adding latex beads to the cells. Maximum cell densities at stationary phase of growth were found to be inversely proportional to DMSO between 0.5 and 1.75%, and the duration of the generation time was exponentially proportional. At 2% DMSO cellular division was completely blocked, and above 2% it was cytotoxic. P. caudatum survived for 8 h in 4% DMSO and died instantaneously in 10%. This inhibitory effect on growth was reversible, though this reversibility might depend on the duration and level of DMSO exposure. DMSO exerted a dose- and time-dependent inhibitory effect on the rate of DV formation but had little effect on the acidification-condensation and the lysosome fusion-digestion processes. The size of the DV formed was also reduced, and this effect was dose-but not time-dependent; vacuole size reduction occurred immediately with DMSO exposure, and no further reduction was observed during exposures of up to 24 h. DMSO at 3 and 4% inhibited vacuole defecation, but the cells could overcome this inhibition when exposed to DMSO for longer periods.(ABSTRACT TRUNCATED AT 250 WORDS)     

Aerosol Exposure to Rift Valley Fever Virus Causes Earlier and More Severe Neuropathology in the Murine Model,which Has Important Implications for Therapeutic Development

Rift Valley fever virus (RVFV) is an important mosquito-borne veterinary and human pathogen that can cause severe disease including acute-onset hepatitis, delayed-onset encephalitis, retinitis and blindness, or a hemorrhagic syndrome. Currently, no licensed vaccine or therapeutics exist to treat this potentially deadly disease. Detailed studies describing the pathogenesis of RVFV following aerosol exposure have not been completed and candidate therapeutics have not been evaluated following an aerosol exposure. These studies are important because while mosquito transmission is the primary means for human infection, it can also be transmitted by aerosol or through mucosal contact. Therefore, we directly compared the pathogenesis of RVFV following aerosol exposure to a subcutaneous (SC) exposure in the murine model by analyzing survival, clinical observations, blood chemistry, hematology, immunohistochemistry, and virus titration of tissues. Additionally, we evaluated the effectiveness of the nucleoside analog ribavirin administered prophylactically to treat mice exposed by aerosol and SC. The route of exposure did not significantly affect the survival, chemistry or hematology results of the mice. Acute hepatitis occurred despite the route of exposure. However, the development of neuropathology occurred much earlier and was more severe in mice exposed by aerosol compared to SC exposed mice. Mice treated with ribavirin and exposed SC were partially protected, whereas treated mice exposed by aerosol were not protected. Early and aggressive viral invasion of brain tissues following aerosol exposure likely played an important role in ribavirin''s failure to prevent mortality among these animals. Our results highlight the need for more candidate antivirals to treat RVFV infection, especially in the case of a potential aerosol exposure. Additionally, our study provides an account of the key pathogenetic differences in RVF disease following two potential exposure routes and provides important insights into the development and evaluation of potential vaccines and therapeutics to treat RVFV infection.