Antiproliferative and Cytotoxic Activities of Fluorescein-A Diagnostic Angiography Dye.

Fluorescein is a fluorescent dye used as a diagnostic tool in various fields of medicine. Although fluorescein itself possesses low toxicity, after photoactivation, it releases potentially toxic molecules, such as singlet oxygen (1O2) and, as we demonstrate in this work, also carbon monoxide (CO). As both of these molecules can affect physiological processes, the main aim of this study was to explore the potential biological impacts of fluorescein photochemistry. In our in vitro study in a human hepatoblastoma HepG2 cell line, we explored the possible effects on cell viability, cellular energy metabolism, and the cell cycle. We observed markedly lowered cell viability (≈30%, 75-2400 µM) upon irradiation of intracellular fluorescein and proved that this decrease in viability was dependent on the cellular oxygen concentration. We also detected a significantly decreased concentration of Krebs cycle metabolites (lactate and citrate < 30%; 2-hydroxyglutarate and 2-oxoglutarate < 10%) as well as cell cycle arrest (decrease in the G2 phase of 18%). These observations suggest that this photochemical reaction could have important biological consequences and may account for some adverse reactions observed in fluorescein-treated patients. Additionally, the biological activities of both 1O2 and CO might have considerable therapeutic potential, particularly in the treatment of cancer.

Effects of Low Dose Space Radiation Exposures on the Splenic Metabolome.

Future space missions will include a return to the Moon and long duration deep space roundtrip missions to Mars. Leaving the protection that Low Earth Orbit provides will unavoidably expose astronauts to higher cumulative doses of space radiation, in addition to other stressors, e.g., microgravity. Immune regulation is known to be impacted by both radiation and spaceflight and it remains to be seen whether prolonged effects that will be encountered in deep space can have an adverse impact on health. In this study, we investigated the effects in the overall metabolism of three different low dose radiation exposures (γ-rays, 16O, and 56Fe) in spleens from male C57BL/6 mice at 1, 2, and 4 months after exposure. Forty metabolites were identified with significant enrichment in purine metabolism, tricarboxylic acid cycle, fatty acids, acylcarnitines, and amino acids. Early perturbations were more prominent in the γ irradiated samples, while later responses shifted towards more prominent responses in groups with high energy particle irradiations. Regression analysis showed a positive correlation of the abundance of identified fatty acids with time and a negative association with γ-rays, while the degradation pathway of purines was positively associated with time. Taken together, there is a strong suggestion of mitochondrial implication and the possibility of long-term effects on DNA repair and nucleotide pools following radiation exposure.

Ameliorating mitochondrial dysfunction restores carbon ion-induced cognitive deficits via co-activation of NRF2 and PINK1 signaling pathway.

Carbon ion therapy is a promising modality in radiotherapy to treat tumors, however, a potential risk of induction of late normal tissue damage should still be investigated and protected. The aim of the present study was to explore the long-term cognitive deficits provoked by a high-linear energy transfer (high-LET) carbon ions in mice by targeting to hippocampus which plays a crucial role in memory and learning. Our data showed that, one month after 4 Gy carbon ion exposure, carbon ion irradiation conspicuously resulted in the impaired cognitive performance, neurodegeneration and neuronal cell death, as well as the reduced mitochondrial integrity, the disrupted activities of tricarboxylic acid cycle flux and electron transport chain, and the depressed antioxidant defense system, consequently leading to a decline of ATP production and persistent oxidative damage in the hippocampus region. Mechanistically, we demonstrated the disruptions of mitochondrial homeostasis and redox balance typically characterized by the disordered mitochondrial dynamics, mitophagy and glutathione redox couple, which is closely associated with the inhibitions of PINK1 and NRF2 signaling pathway as the key regulators of molecular responses in the context of neurotoxicity and neurodegenerative disorders. Most importantly, we found that administration with melatonin as a mitochondria-targeted antioxidant promoted the PINK1 accumulation on the mitochondrial membrane, and augmented the NRF2 accumulation and translocation. Moreover, melatonin pronouncedly enhanced the molecular interplay between NRF2 and PINK1. Furthermore, in the mouse hippocampal neuronal cells, overexpression of NRF2/PINK1 strikingly protected the hippocampal neurons from carbon ion-elicited toxic insults. Thus, these data suggest that alleviation of the sustained mitochondrial dysfunction and oxidative stress through co-modulation of NRF2 and PINK1 may be in charge of restoration of the cognitive impairments in a mouse model of high-LET carbon ion irradiation.

Gas Chromatography/Mass Spectrometry Metabolomics of Urine and Serum from Nonhuman Primates Exposed to Ionizing Radiation: Impacts on the Tricarboxylic Acid Cycle and Protein Metabolism.

Ionizing radiation (IR) directly damages cells and tissues or indirectly damages them through reactive free radicals that may lead to longer term adverse sequelae such as cancers, persistent inflammation, or possible death. Potential exposures include nuclear reactor accidents, improper disposal of equipment containing radioactive materials or medical errors, and terrorist attacks. Metabolomics (comprehensive analysis of compounds <1 kDa) by mass spectrometry (MS) has been proposed as a tool for high-throughput biodosimetry and rapid assessment of exposed dose and triage needed. While multiple studies have been dedicated to radiation biomarker discovery, many have utilized liquid chromatography (LC) MS platforms that may not detect particular compounds (e.g., small carboxylic acids or isomers) that complementary analytical tools, such as gas chromatography (GC) time-of-flight (TOF) MS, are ideal for. The current study uses global GC-TOF-MS metabolomics to complement previous LC-MS analyses on nonhuman primate biofluids (urine and serum) 7 days after exposure to 2, 4, 6, 7, and 10 Gy IR. Multivariate data analysis was used to visualize differences between control and IR exposed groups. Univariate analysis was used to determine a combined 26 biomarkers in urine and serum that significantly changed after exposure to IR. We found several metabolites involved in tricarboxylic acid cycle function, amino acid metabolism, and host microbiota that were not previously detected by global and targeted LC-MS studies.

Urinary metabolic signatures and early triage of acute radiation exposure in rat model.

After a large-scale radiological accident, early-response biomarkers to assess radiation exposure over a broad dose range are not only the basis of rapid radiation triage, but are also the key to the rational use of limited medical resources and to the improvement of treatment efficiency. Because of its high throughput, rapid assays and minimally invasive sample collection, metabolomics has been applied to research into radiation exposure biomarkers in recent years. Due to the complexity of radiobiological effects, most of the potential biomarkers are both dose-dependent and time-dependent. In reality, it is very difficult to find a single biomarker that is both sensitive and specific in a given radiation exposure scenario. Therefore, a multi-parameters approach for radiation exposure assessment is more realistic in real nuclear accidents. In this study, untargeted metabolomic profiling based on gas chromatography-mass spectrometry (GC-MS) and targeted amino acid profiling based on LC-MS/MS were combined to investigate early urinary metabolite responses within 48 h post-exposure in a rat model. A few of the key early-response metabolites for radiation exposure were identified, which revealed the most relevant metabolic pathways. Furthermore, a panel of potential urinary biomarkers was selected through a multi-criteria approach and applied to early triage following irradiation. Our study suggests that it is feasible to use a multi-parameters approach to triage radiation damage, and the urinary excretion levels of the relevant metabolites provide insights into radiation damage and repair.

Variations of metabolites and proteome in Lonicera japonica Thunb. buds and flowers under UV radiation.

Lonicera japonica Thunb., also known as Jin Yin Hua and Japanese honeysuckle, is used as a herbal medicine in Asian countries. Its flowers have been used in folk medicine in the clinic and in making food or healthy beverages for over 1500years in China. To investigate the molecular processes involved in L. japonica development from buds to flowers exposed to UV radiation, a comparative proteomics analysis was performed. Fifty-four proteins were identified as differentially expressed, including 42 that had increased expression and 12 that had decreased expression. The levels of the proteins related to glycolysis, TCA/organic acid transformation, major carbohydrate metabolism, oxidative pentose phosphate, stress, secondary metabolism, hormone, and mitochondrial electron transport were increased during flower opening process after exposure to UV radiation. Six metabolites in L. japonica buds and flowers were identified and relatively quantified using LC-MS/MS. The antioxidant activity was performed using a 1,1-diphenyl-2-picrylhydrazyl assay, which revealed that L. japonica buds had more activity than the UV irradiated flowers. This suggests that UV-B radiation induces production of endogenous ethylene in L. japonica buds, thus facilitating blossoming of the buds and activating the antioxidant system. Additionally, the higher metabolite contents and antioxidant properties of L. japonica buds indicate that the L. japonica bud stage may be a more optimal time to harvest than the flower stage when using for medicinal properties.

Effects of anthropogenic sound on digging behavior, metabolism, Ca(2+)/Mg(2+) ATPase activity, and metabolism-related gene expression of the bivalve Sinonovacula constricta.

Anthropogenic sound has increased significantly in the past decade. However, only a few studies to date have investigated its effects on marine bivalves, with little known about the underlying physiological and molecular mechanisms. In the present study, the effects of different types, frequencies, and intensities of anthropogenic sounds on the digging behavior of razor clams (Sinonovacula constricta) were investigated. The results showed that variations in sound intensity induced deeper digging. Furthermore, anthropogenic sound exposure led to an alteration in the O:N ratios and the expression of ten metabolism-related genes from the glycolysis, fatty acid biosynthesis, tryptophan metabolism, and Tricarboxylic Acid Cycle (TCA cycle) pathways. Expression of all genes under investigation was induced upon exposure to anthropogenic sound at ~80 dB re 1 µPa and repressed at ~100 dB re 1 µPa sound. In addition, the activity of Ca(2+)/Mg(2+)-ATPase in the feet tissues, which is directly related to muscular contraction and subsequently to digging behavior, was also found to be affected by anthropogenic sound intensity. The findings suggest that sound may be perceived by bivalves as changes in the water particle motion and lead to the subsequent reactions detected in razor clams.

Plant Responses to High Frequency Electromagnetic Fields.

High frequency nonionizing electromagnetic fields (HF-EMF) that are increasingly present in the environment constitute a genuine environmental stimulus able to evoke specific responses in plants that share many similarities with those observed after a stressful treatment. Plants constitute an outstanding model to study such interactions since their architecture (high surface area to volume ratio) optimizes their interaction with the environment. In the present review, after identifying the main exposure devices (transverse and gigahertz electromagnetic cells, wave guide, and mode stirred reverberating chamber) and general physics laws that govern EMF interactions with plants, we illustrate some of the observed responses after exposure to HF-EMF at the cellular, molecular, and whole plant scale. Indeed, numerous metabolic activities (reactive oxygen species metabolism, α- and ß-amylase, Krebs cycle, pentose phosphate pathway, chlorophyll content, terpene emission, etc.) are modified, gene expression altered (calmodulin, calcium-dependent protein kinase, and proteinase inhibitor), and growth reduced (stem elongation and dry weight) after low power (i.e., nonthermal) HF-EMF exposure. These changes occur not only in the tissues directly exposed but also systemically in distant tissues. While the long-term impact of these metabolic changes remains largely unknown, we propose to consider nonionizing HF-EMF radiation as a noninjurious, genuine environmental factor that readily evokes changes in plant metabolism.

CD47 Receptor Globally Regulates Metabolic Pathways That Control Resistance to Ionizing Radiation.

Modulating tissue responses to stress is an important therapeutic objective. Oxidative and genotoxic stresses caused by ionizing radiation are detrimental to healthy tissues but beneficial for treatment of cancer. CD47 is a signaling receptor for thrombospondin-1 and an attractive therapeutic target because blocking CD47 signaling protects normal tissues while sensitizing tumors to ionizing radiation. Here we utilized a metabolomic approach to define molecular mechanisms underlying this radioprotective activity. CD47-deficient cells and cd47-null mice exhibited global advantages in preserving metabolite levels after irradiation. Metabolic pathways required for controlling oxidative stress and mediating DNA repair were enhanced. Some cellular energetics pathways differed basally in CD47-deficient cells, and the global declines in the glycolytic and tricarboxylic acid cycle metabolites characteristic of normal cell and tissue responses to irradiation were prevented in the absence of CD47. Thus, CD47 mediates signaling from the extracellular matrix that coordinately regulates basal metabolism and cytoprotective responses to radiation injury.

[Characteristics of indices of biological oxidation under the therapeutic influence of electromagnetic radiation of millimeter range in a human organism]. / Kharakterystyka pokaznykiv biolohichnoho okysnennia pry diï elektromahnitnoho vyprominiuvannia milimetrovoho diapazony na orhanizm liudyny.

The results of studying the process of biological oxidation of carbohydrates and Krebs cycle enzymes under the therapeutical influence of electromagnetic radiation of mm-range as a response of patient with gastroduodenal pathology are presented. Positive clinical effect was accompanied by multidirectional changes of biochemical parameters and activation of main mechanism of sanogenesis. Correcting action of EMR of m-range on the homeostasis system and process of biological oxidation of carbohydrates and Krebs and the functional state of a human's systems and organs were ascertained.

[The hormonal status and carbohydrate-energy metabolism of rats after the long-term action of low doses of ionizing radiation and heat]. / Gormonal'nyi status i uglevodno-énergeticheskii obmen u krys posle dlitel'nogo deistviia malykh doz ioniziruiushchego izlucheniia i tepla.

Hormonal status (blood content of triiodothyronine, thyroxin, insulin, 11-hydroxycorticosteroids), dehydration in the Krebs cycle, and activity of the first enzyme in the pentose-phosphate cycle, glucose-6-phosphate dehydrogenase, in the brain and myocardium of white rats were studied at different time periods after separate and combined prolonged exposure to radiation in relatively small doses and heat. It was found that combination of ionizing radiation and heat led to hypofunction of the endocrine glands and inhibition of dehydration processes in the Krebs Cycle.

[Functional interrelationships between dehydrogenases and transaminases in various regions of the rat brain as affected by small doses of gamma-radiation]. / Funktsional'nye vzaimootnosheniia mezhdu degidrogenazami i transaminazami v razlichnykh otdelakh golovnogo mozga krys pri deistvii malykh doz gamma-izlucheniia.

Wistar rats were exposed to 10, 20 and 40 sGy of gamma-irradiation. Maximal activity of 2-oxoglutarate- and succinate dehydrogenases, as well as of aspartate-2-oxoglutarate and pyruvate-2-oxoglutarate in the brain tissue was measured within 30 days after the exposure. Dehydrogenase activity was measured in the brain cortex, limbic system and in the cerebellum; transaminase activity was determined in the brain cortex, diencephalic zone and in the hypophysis. It was found that small doses of irradiation resulted in an oscillating increase of dehydrogenase activity with a decrease of the amplitude by the 30th day of the experiment. As the dose of irradiation increases, the periods of normal activity change into phases of inhibition of dehydrogenase action, the period of oscillations diminishes. Transaminase activity, as a rule, changes in a phase in a reversible manner as compared with dehydrogenase activity. These observations are discussed from the point of view of structural relations between these two systems in the intramitochondrial supramolecular structures, the so-called metabolons.

[The effect of helium-neon laser radiation on the energy metabolic indices of the myocardium]. / Vliianie izlucheniia gelii-neonovogo lazera na pokazateli énergeticheskogo metabolizma miokarda.

It was shown in experiments on white rats, that intravenous and direct myocardium helium-neon laser irradiation leads to the some activation of lactate, glucose-6-phosphate, succinate and reduced NAD degydrogenases. During direct myocardium irradiation these changes are in a less degree. It is suggested that helium-neon laser irradiation displays some active influence on the energy metabolism enzymes of the myocardium, and the mechanisms of this action are discussed.

[Energy metabolism during stressful exposures (ionizing radiation), its self-regulation and correction]. / Energeticheskii obmen pri stressovykh vozdeistviiakh (na primere ioniziruiushchei radiatsii), ego samoreguliatsiia i korrektsiia.

Normal hormonal regulation of energy metabolism is mainly realized by glucocorticoids and insulin, their physiological antagonist. Under the effect of different extremal factors (including ionizing radiation) there arises non-specific stress, a syndrome the main component of which is the hyperfunction of glucocorticoids--the intermediate hormonal link in the stress reaction. Stimulation of hypercorticism by administering hydrocortisone to intact animals as well as its stimulation by administering this preparation to irradiated animals causes development and intensification of inhibition and uncoupling of the oxidative phosphorylation as well as disturbance in adenylic nucleotides metabolism. The administered insulin, softening the reaction of hypercorticism and changing the ratio of the hormone levels in favour of insulin, weakens essentially the stress (ray) disturbances in the energy metabolism.