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Discoid lupus erythematosus (DLE) is a common autoimmune skin disease in dogs. Conventional treatments, such as corticosteroids, can be effective but often have side effects. This case report presents a successful use of cannabinoid therapy (CT) in a dog with DLE resistant to conventional treatment. A 2-year-old mixed-breed dog with a history of DLE presented with worsening lesions despite treatment with corticosteroids and other medications. Liver enzymes levels were elevated, indicating corticosteroid-induced side effects. CT with a CBD-rich full spectrum Cannabis oil was initiated. The dosage was gradually adjusted until the minimum effective dose was found. Within a few weeks of starting CT, the dog showed significant improvement in skin lesions and in liver enzymes levels. After 1 year, the dog remains clinically stable on a low dose of full-spectrum CBD-rich oil. No evidence of DLE recurrence was observed. This case suggests that CT may be a viable alternative or complementary therapy for DLE in dogs, particularly for those experiencing adverse effects from conventional treatments. Further research is warranted to confirm the efficacy and safety of CT for DLE management in dogs.

Ambient air pollutants have been reported to induce oxidative stress based inflammatory responses in humans and experimental animals. However, most of these reports describe the actions of the particulate phase of ambient and exhaust samples. We describe here results of studies investigating the actions of the vapor phase of ambient air samples collected in the midtown area of Los Angeles on human bronchial epithelial BEAS-2B cells using DNA microarray analysis. Among 26 genes whose expression increased fourfold or more, four genes were associated with detoxifying genes regulated by the transcription factor Nrf2. Consistent with these results, the vapor samples activate the Nrf2-ARE pathway, resulting in up-regulation of heme oxygenase-1 (HO-1), glutamate cysteine ligase modifier subunit, and cystine transporter (xCT) mRNA and proteins. No appreciable increases in pro-inflammatory genes were observed. These results suggest that ambient vapor samples activate the Nrf2-ARE pathway but not an inflammatory response. Also, treatment of the vapor samples with glutathione resulted in reduction in the Nrf2 activation and HO-1 induction, suggesting that electrophiles in vapor samples contribute to this Nrf2-dependent antioxidant or adaptive response.

Discoid lupus erythematosus (DLE) is a common autoimmune skin disease in dogs. Conventional treatments, such as corticosteroids, can be effective but often have side effects. This case report presents a successful use of cannabinoid therapy (CT) in a dog with DLE resistant to conventional treatment. A 2-year-old mixed-breed dog with a history of DLE presented with worsening lesions despite treatment with corticosteroids and other medications. Liver enzymes levels were elevated, indicating corticosteroid-induced side effects. CT with a CBD-rich full spectrum Cannabis oil was initiated. The dosage was gradually adjusted until the minimum effective dose was found. Within a few weeks of starting CT, the dog showed significant improvement in skin lesions and in liver enzymes levels. After 1 year, the dog remains clinically stable on a low dose of full-spectrum CBD-rich oil. No evidence of DLE recurrence was observed. This case suggests that CT may be a viable alternative or complementary therapy for DLE in dogs, particularly for those experiencing adverse effects from conventional treatments. Further research is warranted to confirm the efficacy and safety of CT for DLE management in dogs. [ABSTRACT FROM AUTHOR]

Background: Transmissible venereal tumour (TVT), also called Sticker Tumour, is a round cell tumour whose transmission occurs mainly through sexual contact between animals or direct transplantation of the neoplastic cell through licking or through direct contact with the lesioned area. TVT is the most common tumor found in stray dogs and predominantly affects the genital area and, in case of bitches, occurs in the vagina and foreskin, as well as the oral and nasal cavities. Intracavitary TVT is rarely reported. The aim of this paper is to report a case of an old neutered bitch affected by vaginal and intracavitary TVT, treated at the Veterinary School Clinic (CVE) of the Federal University of Santa Catarina (UFSC), Curitibanos campus, Brazil. Case: An approximately 10-year-old mixed breed bitch, weighing 18,5 kg and, neutered 2 years previously at the same institution, was brought by her owner, who reported dyskinesia, constipation, and serosanguinous vaginal discharge for a few weeks. Physical examination showed a row of 4 smooth, round masses with a soft consistency ranging from 1 to 2 cm at the bottom of the vaginal roof. The patient was referred for an ultrasound (US) and a vaginal swab was taken for cytological examination, whose result demonstrated TVT. The result of the US scan showed a heterogeneously shaped structure, with a more hypoechogenic center in relation to the periphery and irregular contours, measuring around 2.63 x 6.10 cm, in the uterine topography caudally to the bladder. Exploratory laparotomy was performed with total removal of the mass, which was surrounding the proximal urethra, and the material was sent for histological and cytological analysis, whose both results indicated TVT. Chemotherapy was then initiated, with 6 intravenous administrations of vincristine (0.5 mg/m2) with an interval of 15 days between each administration. The animal underwent a total of 6 chemotherapy sessions, without complete improvement of the vaginal tumours. A 2nd chemotherapy protocol based on doxorubicin was suggested, which was not accepted by the owner. Three months after the end of the original chemotherapy protocol, the patient was assessed again and it was found that there had been no progression of the vaginal masses on vaginal palpation and no signs of recurrence of the intracavitary mass on ultrasound. Discussion: TVT is a venereal tumor that affects almost 100% of non-neutered animals, males and females. In the case of bitches, the most common sites are the vagina (53%), vulva (13%) and extragenital areas (14%). This animal had no intracavitary mass at the time of her castration 2 years earlier. At the time of diagnosis, she had the extragenital form of the tumor, consisting of an intra-abdominal mass, in addition to the genital masses in the vaginal roof. The chemotherapy drug vincristine is the drug of choice for the treatment of transmissible venereal tumors, due to its high efficacy associated with 4 to 6 sessions. However, in this case, there was no total improvement, so it was classified as a case of proven pharmacological resistance as the chemotherapy was ineffective for the complete regression of the tumor. It was therefore concluded that the occurrence of intracavitary TVTs, although rare, warrants a thorough physical examination combined with diagnostic tools such as ultrasound and histology, even in castrated old patients. [ABSTRACT FROM AUTHOR]

Platinosomosis is a systemic parasitic disease with a hepatic predilection caused by the trematode Platynosomum spp. It occurs mainly in felines through ingestion of the paratenic host, or from a second intermediate host, which would be terrestrial isopods in endemic tropical areas around the world. The pathophysiology includes cases of hepatic lipidosis, cholangitis and cholangiohepatitis, with their severity related to the level of parasite load in the final host. The present work aims to report two cases of feline platinosomosis seen at the Clinical Veterinary School (CVE) of the Federal University of Santa Catarina (UFSC), in Curitibanos, SC. Both cases correspond to two domiciled female felines aged six to eight months with complaints of emesis and hematochezia. Furthermore, one of the animals had diarrhea and the other had pasty feces, both with a history of delayed vaccinations and deworming. Laboratory tests showed thrombocytopenia, changes in ALT, GGT and FA enzymes, abdominal ultrasound demonstrated a small thickening of the gastric wall and fecal parasitology showed a positive result for the presence of the species of Platynosomum fastosum (= P. illiciens) in fecal samples. The present work consists of the report of the first two cases described in the central region of the Santa Catarina highland, with the disease previously described only in the municipality of Araquari, in the north of the state. [ABSTRACT FROM AUTHOR]

This article demonstrates some aspects of semiotic theory applied at the analysis of the graphical user interface elements of a vdeo game. Initially the concepts and terminology used in the context of the analyzed object it will be defined. Then, the method of choice for the analysis exercice, which follows an application path based on the propositions of Santaella (2002), will be presented. The object choosen for this analysis, is the Head-up Display from the video game Dead Space. As closing comments, a strong appeal to the diegetic elements is highlighted, which can contribute to build a solid game environment and generate a more immersive experience.

THIS STUDY AIMED TO PERFORM AN EXPLORATORY ANALYSIS ON HEURISTIC EVALUATION OF VIDEOGAMES GRAPHICAL USER INTERFACES, CORRELATING THE HEURISTIC PRINCIPLES PROPOSED BY FEDEROFF (2002), DESURVIRE, CAPLAN AND TOTH (2004) AND NIELSEN (1994) WITH THE HEADS-UP DISPLAYS DESIGN-RELATED ASPECTS. THEORETIC FUNDAMENTALS WERE GATHERED BY REVISION OF THE LITERATURE IN ORDER TO DEFINE THE APROACHED TERMINOLOGY AND CONCEPTS SUCH AS: USABILIT; PLAYABILITY; GENERAL HEURISTIC PRINCIPLES; AND VIDEOGAME EVALUATION ORIENTED HEURISTIC PRINCIPLES. AS RESULTS OF THIS STUDY, ENTRIES WERE MADE ABOUT THE VIABILITY OF HEURISTIC PRINCIPLES FOR EVALUATION OF VIDEOGAMES AND HUDS.

The seamless transition through stages of pluripotency relies on a balance between transcription factor networks and epigenetic mechanisms. Here, we reveal the crucial role of the transgene activation suppressor (TASOR), a component of the human silencing hub (HUSH) complex, in maintaining cell viability during the transition from naive to primed pluripotency. TASOR loss in naive pluripotent stem cells (PSCs) triggers replication stress, disrupts H3K9me3 heterochromatin, and impairs silencing of LINE-1 (L1) transposable elements, with more severe effects in primed PSCs. Notably, the survival of Tasor knockout PSCs during this transition can be restored by inhibiting caspase or deleting the mitochondrial antiviral signaling protein (MAVS). This suggests that unscheduled L1 expression activates an innate immune response, leading to cell death specifically in cells exiting naive pluripotency. Our findings highlight the importance of epigenetic programs established in naive pluripotency for normal development., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

The zebrafish Danio rerio has become a popular model host to explore disease pathology caused by infectious agents. A main advantage is its transparency at an early age, which enables live imaging of infection dynamics. While multispecies infections are common in patients, the zebrafish model is rarely used to study them, although the model would be ideal for investigating pathogen-pathogen and pathogen-host interactions. This may be due to the absence of an established multispecies infection protocol for a defined organ and the lack of suitable image analysis pipelines for automated image processing. To address these issues, we developed a protocol for establishing and tracking single and multispecies bacterial infections in the inner ear structure (otic vesicle) of the zebrafish by imaging. Subsequently, we generated an image analysis pipeline that involved deep learning for the automated segmentation of the otic vesicle, and scripts for quantifying pathogen frequencies through fluorescence intensity measures. We used Pseudomonas aeruginosa, Acinetobacter baumannii, and Klebsiella pneumoniae, three of the difficult-to-treat ESKAPE pathogens, to show that our infection protocol and image analysis pipeline work both for single pathogens and pairwise pathogen combinations. Thus, our protocols provide a comprehensive toolbox for studying single and multispecies infections in real-time in zebrafish., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Schmitz et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

How to derive principles of community dynamics and stability is a central question in microbial ecology. Bottom-up experiments, in which a small number of bacterial species are mixed, have become popular to address it. However, experimental setups are typically limited because co-culture experiments are labor-intensive and species are difficult to distinguish. Here, we use a four-species bacterial community to show that information from monoculture growth and inhibitory effects induced by secreted compounds can be combined to predict the competitive rank order in the community. Specifically, integrative monoculture growth parameters allow building a preliminary competitive rank order, which is then adjusted using inhibitory effects from supernatant assays. While our procedure worked for two different media, we observed differences in species rank orders between media. We then parameterized computer simulations with our empirical data to show that higher order species interactions largely follow the dynamics predicted from pairwise interactions with one important exception. The impact of inhibitory compounds was reduced in higher order communities because their negative effects were spread across multiple target species. Altogether, we formulated three simple rules of how monoculture growth and supernatant assay data can be combined to establish a competitive species rank order in an experimental four-species community., Competing Interests: The authors declare that they have no conflict of interest., (© The Author(s) 2024. Published by Oxford University Press on behalf of the International Society for Microbial Ecology.)

Embryogenesis necessitates harmonious coordination between embryonic and extraembryonic tissues. Although stem cells of both embryonic and extraembryonic origins have been generated, they are grown in different culture conditions. In this study, utilizing a unified culture condition that activates the FGF, TGF-β, and WNT pathways, we have successfully derived embryonic stem cells (FTW-ESCs), extraembryonic endoderm stem cells (FTW-XENs), and trophoblast stem cells (FTW-TSCs) from the three foundational tissues of mouse and cynomolgus monkey (Macaca fascicularis) blastocysts. This approach facilitates the co-culture of embryonic and extraembryonic stem cells, revealing a growth inhibition effect exerted by extraembryonic endoderm cells on pluripotent cells, partially through extracellular matrix signaling. Additionally, our cross-species analysis identified both shared and unique transcription factors and pathways regulating FTW-XENs. The embryonic and extraembryonic stem cell co-culture strategy offers promising avenues for developing more faithful embryo models and devising more developmentally pertinent differentiation protocols., Competing Interests: Declaration of interests Y.W., L.Y., T.T., and J.W. are inventors on a patent application (applied through the Board of Regents of The University of Texas System, application number 63/488,401) entitled “Methods For the Derivation Culture Of Embryonic and Extra-Embryonic Stem Cells” arising from this work., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Bacterial infections are often polymicrobial, leading to intricate pathogen-pathogen and pathogen-host interactions. There is increasing interest in studying the molecular basis of pathogen interactions and how such mechanisms impact host morbidity. However, much less is known about the ecological dynamics between pathogens and how they affect virulence and host survival. Here we address these open issues by co-infecting larvae of the insect model host Galleria mellonella with one, two, three or four bacterial species, all of which are opportunistic human pathogens. We found that host mortality was always determined by the most virulent species regardless of the number of species and pathogen combinations injected. In certain combinations, the more virulent pathogen simply outgrew the less virulent pathogen. In other combinations, we found evidence for negative interactions between pathogens inside the host, whereby the more virulent pathogen typically won a competition. Taken together, our findings reveal positive associations between a pathogen's growth inside the host, its competitiveness towards other pathogens and its virulence. Beyond being generalizable across species combinations, our findings predict that treatments against polymicrobial infections should first target the most virulent species to reduce host morbidity, a prediction we validated experimentally.

Our understanding of human early development is severely hampered by limited access to embryonic tissues. Due to their close evolutionary relationship with humans, nonhuman primates are often used as surrogates to understand human development but currently suffer from a lack of in vivo datasets, especially from gastrulation to early organogenesis during which the major embryonic cell types are dynamically specified. To fill this gap, we collected six Carnegie stage 8-11 cynomolgus monkey (Macaca fascicularis) embryos and performed in-depth transcriptomic analyses of 56,636 single cells. Our analyses show transcriptomic features of major perigastrulation cell types, which help shed light on morphogenetic events including primitive streak development, somitogenesis, gut tube formation, neural tube patterning and neural crest differentiation in primates. In addition, comparative analyses with mouse embryos and human embryoids uncovered conserved and divergent features of perigastrulation development across species-for example, species-specific dependency on Hippo signalling during presomitic mesoderm differentiation-and provide an initial assessment of relevant stem cell models of human early organogenesis. This comprehensive single-cell transcriptome atlas not only fills the knowledge gap in the nonhuman primate research field but also serves as an invaluable resource for understanding human embryogenesis and developmental disorders., (© 2022. The Author(s).)

Although features of ribosome assembly are shared between species, our understanding of the diversity, complexity, dynamics, and regulation of ribosome production in multicellular organisms remains incomplete. To gain insights into ribosome biogenesis in human cells, we perform a genome-wide loss-of-function screen combined with differential labeling of pre-existing and newly assembled ribosomes. These efforts identify two functionally uncharacterized genes, C1orf109 and SPATA5. We provide evidence that these factors, together with CINP and SPATA5L1, control a late step of human pre-60S maturation in the cytoplasm. Loss of either C1orf109 or SPATA5 impairs global protein synthesis. These results link ribosome assembly with neurodevelopmental disorders associated with recessive SPATA5 mutations. Based on these findings, we propose that the expanded repertoire of ribosome biogenesis factors likely enables multicellular organisms to coordinate multiple steps of ribosome production in response to different developmental and environmental stimuli., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

Dynamic pluripotent stem cell (PSC) states are in vitro adaptations of pluripotency continuum in vivo. Previous studies have generated a number of PSCs with distinct properties. To date, however, no known PSCs have demonstrated dual competency for chimera formation and direct responsiveness to primordial germ cell (PGC) specification, a unique functional feature of formative pluripotency. Here, by modulating fibroblast growth factor (FGF), transforming growth factor β (TGF-β), and WNT pathways, we derived PSCs from mice, horses, and humans (designated as XPSCs) that are permissive for direct PGC-like cell induction in vitro and are capable of contributing to intra- or inter-species chimeras in vivo. XPSCs represent a pluripotency state between naive and primed pluripotency and harbor molecular, cellular, and phenotypic features characteristic of formative pluripotency. XPSCs open new avenues for studying mammalian pluripotency and dissecting the molecular mechanisms governing PGC specification. Our method may be broadly applicable for the derivation of analogous stem cells from other mammalian species., Competing Interests: Declaration of Interests L.Y. and J.W. are inventors on a patent application arising from this work. The other authors declare no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Limited access to embryos has hampered the study of human embryogenesis and disorders that occur during early pregnancy. Human pluripotent stem cells provide an alternative means to study human development in a dish 1-7 . Recent advances in partial embryo models derived from human pluripotent stem cells have enabled human development to be examined at early post-implantation stages 8-14 . However, models of the pre-implantation human blastocyst are lacking. Starting from naive human pluripotent stem cells, here we developed an effective three-dimensional culture strategy with successive lineage differentiation and self-organization to generate blastocyst-like structures in vitro. These structures-which we term 'human blastoids'-resemble human blastocysts in terms of their morphology, size, cell number, and composition and allocation of different cell lineages. Single-cell RNA-sequencing analyses also reveal the transcriptomic similarity of blastoids to blastocysts. Human blastoids are amenable to embryonic and extra-embryonic stem cell derivation and can further develop into peri-implantation embryo-like structures in vitro. Using chemical perturbations, we show that specific isozymes of protein kinase C have a critical function in the formation of the blastoid cavity. Human blastoids provide a readily accessible, scalable, versatile and perturbable alternative to blastocysts for studying early human development, understanding early pregnancy loss and gaining insights into early developmental defects.

We assessed the chemical properties and oxidative stress of particulate matter (PM) emissions from underfired charbroiled meat operations with and without the use of aftertreatment control technologies. Cooking emissions concentrations showed a strong dependence on the control technology utilized, with all emission rates showing decreases with the control technologies compared to the baseline testing. The organic acids profile was dominated by the saturated nonanoic, myristic, palmitic, and stearic acids, and the unsaturated oleic, elaidic, and palmitoleic acids. Cholesterol was also found in relatively high concentrations. Lower and medium-weight polycyclic aromatic hydrocarbons (PAHs) were the dominant species for all cooking experiments. Heavier PAHs were also detected in high concentrations, especially in the particle-phase. For the nitrated PAH emissions (nitro-PAHs), low molecular weight compounds dominated the cooking emissions. Under the present experimental conditions, the heterocyclic aromatic amines (HAAs) showed very low concentrations, which suggests these species are rarely formed in meat cooking PM. The most efficient control technology for reducing the majority of the toxic pollutants was the electrostatic precipitator, which resulted in total emissions reductions on the order of 95%, 79%, 90%, 96%, 90%, and 94%, respectively, for particle-phase PAHs, gas-phase PAHs, particle-phase nitro-PAHs, gas-phase nitro-PAHs, particle-phase HAAs, and gas-phase HAAs compared to the baseline testing. Our experiment showed that cooking aerosol contained higher levels of prooxidants in the particle-phase and the corresponding vapors contained higher levels of electrophiles. Overall, the use of control technologies reduced the redox and electrophilic activities of cooking PM., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The emissions and the potential health effects of particulate matter (PM) were assessed from two heavy-duty trucks with and without emission control aftertreatment systems when operating on CARB ultra-low sulfur diesel (ULSD) and three different biodiesel blends. The CARB ULSD was blended with soy-based biodiesel, animal fat biodiesel, and waste cooking oil biodiesel at 50vol%. Testing was conducted over the EPA Urban Dynamometer Driving Schedule (UDDS) in triplicate for both trucks. The aftertreatment controls effectively decreased PM mass and number emissions, as well as the polycyclic aromatic hydrocarbons (PAHs) compared to the uncontrolled truck. Emissions of nitrogen oxides (NO x ) exhibited increases with the biodiesel blends, showing some feedstock dependency for the controlled truck. The oxidative potential of the emitted PM, measured by means of the dithiothreitol (DTT) assay, showed reductions with the use of biodiesel blends relative to CARB ULSD for the uncontrolled truck. Overall, the cellular responses to the particles from each fuel were reflective of the chemical content, i.e., particles from CARB ULSD were the most reactive and exhibited the highest cellular responses., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Unlabelled: The potential adverse health effects of PM2.5 (particulate matter with an aerodynamic diameter<2.5 μm) and vapor samples from three communities that neighbor railyards, Commerce (CM), Long Beach (LB), and San Bernardino (SB), were assessed by determination of chemical reactivities attributed to the induction of oxidative stress by air pollutants. The assays used were dithiothreitol (DTT)- and dihydrobenzoic acid (DHBA)-based procedures for prooxidant content and a glyceraldehyde-3-phosphate dehydrogenase (GAPDH) assay for electrophiles. Prooxidants and electrophiles have been proposed as the reactive chemical species responsible for the induction of oxidative stress by air pollution mixtures. The PM2.5 samples from CM and LB sites showed seasonal differences in reactivities, with higher levels in the winter, whereas the SB sample differences were reversed. The reactivities in the vapor samples were all very similar, except for the summer SB samples, which contained higher levels of both prooxidants and electrophiles. The results suggest that the observed reactivities reflect general geographical differences rather than direct effects of the railyards. Distributional differences in reactivities were also observed, with PM2.5 fractions containing most of the prooxidants (74-81%) and the vapor phase most of the electrophiles (82-96%). The high levels of the vapor-phase electrophiles and their potential for adverse biological effects point out the importance of the vapor phase in assessing the potential health effects of ambient air., Implications: PM2.5 and its corresponding vapor phase, containing semivolatile organics, were collected in three communities in the Los Angeles Basin and examined with toxicologically relevant chemical assays. The PM2.5 phase contained most of the prooxidants and the vapor phase contained most of the electrophiles, whose content was highest in summer samples from a receptor site that reflected greater photochemical processing of the air parcel during its transport. As electrophiles initiate both adverse and adaptive responses to foreign substances by biological systems, their presence in the vapor phase emphasizes the importance of this phase in the overall health effects of ambient air.

Particulate matter (PM) has been the primary focus of studies aiming to understand the relationship between the chemical properties of ambient aerosols and adverse health effects. Size and chemical composition of PM have been linked to their oxidative capacity which has been postulated to promote or exacerbate pulmonary and cardiovascular diseases. But in the last few years, new studies have suggested that volatile and semi-volatile components may also contribute to many adverse health effects. The objectives of this study were: (i) assess for the first time the redox and electrophilic potential of vapor-phase components of ambient aerosols and (ii) evaluate the relative contributions of particle- and vapor-fractions to the hazard of a given aerosol. To achieve these objectives vapor- and particle-phase samples collected in Riverside (CA) were subjected to three chemical assays to determine their redox and electrophilic capacities. The results indicate that redox active components are mainly associated with the particle-phase, while electrophilic compounds are found primarily in the vapor-phase. Vapor-phase organic extracts were also capable of inducing the stress responding protein, heme-oxygenase-1 (HO-1), in RAW264.7 murine macrophages. These results demonstrate the importance of volatile components in the overall oxidative and electrophilic capacity of aerosols, and point out the need for inclusion of vapors in future health and risk assessment studies., ((c) 2010 Elsevier Inc. All rights reserved.)

Background: It has been demonstrated that ambient particulate matter (PM) can act as an adjuvant for allergic sensitization. Redox-active organic chemicals on the particle surface play an important role in PM adverse health effects and may determine the adjuvant effect of different particle types according to their potential to perturb redox equilibrium in the immune system., Objectives: We determined whether the adjuvant effect of ambient fine particles versus ultrafine particles (UFPs) is correlated to their prooxidant potential., Methods: We have established an intranasal sensitization model that uses ambient PM as a potential adjuvant for sensitization to ovalbumin (OVA), which enhances the capacity for secondary OVA challenge to induce allergic airway inflammation., Results: UFPs with a greater polycyclic aromatic hydrocarbon (PAH) content and higher oxidant potential enhanced OVA sensitization more readily than did fine particles. This manifests as enhanced allergic inflammation upon secondary OVA challenge, leading to eosinophilic inflammation and mucoid hyperplasia starting at the nasal turbinates all the way down to the small pulmonary airways. The thiol antioxidant N-acetyl cysteine was able to suppress some of these sensitization events., Conclusions: The adjuvant effects of ambient UFP is determined by their oxidant potential, which likely plays a role in changing the redox equilibrium in the mucosal immune system.

The adverse health effects of air pollutants have been associated with their redox and electrophilic properties. Although the specific chemical species involved in these effects are not known, the characterization of their general physical and chemical properties is important to our understanding of the mechanisms by which they cause health problems. This manuscript describes results of a study examining the partition properties of these activities in aqueous and organic media. The water and dichloromethane (DCM) solubility of redox active and electrophilic constituents of seven diesel exhaust particle (DEP) samples were determined with assays developed earlier in this laboratory. The constituents exhibiting redox activity, which included both metals and nonmetal species, were associated with the particles in the aqueous suspensions. Portions of the redox active compounds were also DCM-soluble. In contrast, the electrophilic constituents included both water-soluble and DCM-soluble species. The role of quinones or quinone-like compounds in redox and electrophilic activities of the DCM-soluble constituents was assessed by reductive acetylation, a procedure that inactivates quinones. The results from this experiment indicated that most of the activities in the organic extract were associated with quinone-like substances. The partition properties of the reactive species are important in exposure assessment since the toxicokinetics of particles and solutes are quite distinct.

Epidemiologic studies have shown associations between ambient particulate matter (PM) and adverse health outcomes including increased mortality, emergency room visits, and time lost from school and work. The mechanisms of PM-related health effects are still incompletely understood, but a hypothesis under investigation is that many of the adverse health effects may derive from oxidative stress, initiated by the formation of reactive oxygen species (ROS) within affected cells. While the adverse effects from PM have historically been associated with the airborne concentration of PM and more recently fine-particle PM, we considered it relevant to develop an assay to quantitatively measure the ability of PM to catalyze ROS generation as the initial step in the induction of oxidative stress. This ability of PM could then be related to different sources, chemical composition, and physical and spatial/temporal characteristics in the ambient environment. The measurement of ROS-forming ability in relation to sources and other factors will have potential relevance to control of redox-active PM. If oxidative stress represents a relevant mechanism of toxicity from PM, the measurement of redox activity represents a first step in the elucidation of the subsequent downstream processes. We have developed an assay for PM redox activity, utilizing the reduction of oxygen by dithiothreitol which serves as an electron source. We have found that PM will catalyze the reduction of oxygen and have examined the distribution and chemical characteristics of the redox activity of PM fractions collected in different sites in the Los Angeles Basin. Samples of concentrated coarse, fine, and ultrafine PM, obtained with aerosol concentrators, were studied with regard to their chemical properties and redox activity. Redox activity was highest in the ultrafine fraction, in agreement with results indicating ultrafines were the most potent toward inducing that heme oxygenase expression and depleting intracellular glutathione, which has relevance to induction of oxidative stress. Comparison of the redox activity with chemical composition showed a reasonable correlation of redox activity with elemental carbon (r(2)=0.79), organic carbon (r(2)=0.53), and with benzo[ghi]perylene (r(2)=0.82), consistent with species typically found in mobile emission sources.

The toxicity of diesel exhaust particles (DEP) can be due to the particle itself, extractable components, or both. Many studies focus on the biological properties of DEP-extractable components although it is possible that chemical properties inherent to the DEP itself can lead to toxicity. Thus, an examination of the chemistry inherent to DEP was carried out. Herein, we report that DEP are capable of catalyzing the consumption of O2 (monitored using a Clarke electrode) by ascorbate and thiols leading to the generation of reactive oxygen species. Consistent with the idea that DEP are capable of catalyzing the generation of reactive oxygen species, they were also found to catalyze DNA strand breakage via an O2- and reductant-dependent process. Significantly, extraction of DEP with either organic solvent (methylene chloride) or acid (aqueous HCl) did little to abrogate this chemistry. Finally, using electron paramagnetic spectrometry (EPR), DEP were found to have paramagnetic properties. The paramagnetic character of DEP may be important to their ability to catalyze the formation of reactive oxygen species and at least partially responsible for their toxicity. These findings indicate that studies that primarily consider or examine particle extracts as the toxic components of DEP may be insufficient in describing the toxicity associated with DEP exposure.

We compared the behavioral and caudate-putamen extracellular dopamine responses following intravenous (3.6 mg/kg) and subcutaneous (8 mg/kg) amphetamine administration using 2-min microdialysate sampling intervals, and doses of the drug selected to achieve comparable maximal brain concentrations. Following intravenous amphetamine, dopamine peaked within the first 2 min, then declined with a first-order decay rate of 0.018+/-0.007 min(-1). Following subcutaneous amphetamine, dopamine achieved maximum concentrations at 9 min and remained near peak levels for about 30 min before declining with a first-order decay rate of 0.019+/-0.008 min(-1). Maximal brain amphetamine levels and peak dopamine concentrations were equivalent following either route of drug administration. In contrast to the short latency to maximal extracellular dopamine, the onset of oral stereotypies was delayed until about 30 min following both routes of drug administration. Furthermore, in contrast to the behavioral response to amphetamine, apomorphine administration resulted in the rapid appearance of oral stereotypies within 5-10 min after drug administration. These results suggest that although caudate-putamen dopamine receptor activation may be a critical factor in the expression of focused oral stereotypies, other effects of amphetamine may interfere with the ability of animals to exhibit these behaviors.

Methamphetamine (MeAmp) abuse has recently experienced a resurgence and approaches to the treatment of its addiction similar to those used with cocaine have been considered. As the treatment regimes are likely to use drugs whose metabolism is related to that of MeAmp, studies were initiated to establish the enzymology of the fate of MeAmp. This report describes investigations of the role of CYP2D6, the human isoform of the enzyme that catalyzes debrisoquine hydroxylation, in the 4-hydroxylation and N-demethylation of MeAmp. The results of studies with human liver microsomes including those from a genetically poor metabolizer with respect to CYP2D6, showing correlation between MeAmp and metoprolol hydroxylation and MDMA demethylenation, were consistent with a major involvement of CYP2D6 in the aromatic 4-hydroxylation of MeAmp. This was confirmed by studies with recombinant CYP2D6 expressed in yeast, which was also shown to effect the N-demethylation of MeAmp. The rate of the 4-hydroxylation reaction was substantially slower than the demethylenation of MDMA. In contrast to MeAmp, MDMA was not N-demethylated by CYP2D6. Since CYP2D6 participates in the major steps of MeAmp metabolism, pharmacokinetic interactions are likely with other drug substrates proposed for the treatment of MeAmp addiction. Furthermore, the genetic polymorphism associated with the enzyme could manifest itself in abnormal responses to MeAmp.

Rat aortic endothelial cells were found to contain both constitutive and lipopolysaccharide (LPS)-inducible arginase activity. Studies were performed to determine whether induction of nitric oxide synthase (NOS) by LPS and cytokines is accompanied by sufficient arginase induction to render arginine concentrations rate limiting for high-output NO production. Unactivated cells contained abundant arginase activity accompanied by continuous urea formation. LPS induced the formation of both inducible NOS (iNOS) and arginase, and this was accompanied by increased production of NO, citrulline, and urea. Immunoprecipitation experiments revealed the constitutive presence of arginase-I in both unactivated and LPS-activated cells and arginase-II induction by LPS. Arginase-I and iNOS were verified by reverse transcriptase-polymerase chain reaction. Induction of large amounts of iNOS by LPS plus several cytokines resulted in large quantities of NO, citrulline, and NG-hydroxy-L-arginine (NOHA), but urea production was markedly diminished. Decreased urea production was attributed to increased formation of NOHA, the precursor to NO and citrulline and a potent inhibitor of arginase-I activity with an inhibitory constant of 10-12 microM. Inhibition of iNOS activity by NG-methyl-L-arginine decreased NO and NOHA production and increased urea production. This study reveals for the first time that substantial arginase activity is present constitutively in rat aortic endothelial cells, a different isoform of arginase is induced by LPS, and intracellular arginase activity can be markedly inhibited during cytokine induction of iNOS because of NOHA formation. The inhibition of arginase activity that occurs by NOHA during marked iNOS induction may be a mechanism to ensure sufficient arginine availability for high-output production of NO.

The high selectivity of amphetamine and its derivatives for CYP2D-mediated oxidations suggested the use of the phenylisopropylamine skeleton as a template for a selective inhibitor of this important enzyme. Accordingly, 4-allyloxymethamphetamine-amine (ALLMA) was synthesized and its ability to selectively inactivate CYP2D was investigated both in in vitro and in vivo experiments. Incubation studies with rat liver microsomes demonstrated that this compound suppressed the CYP2D-mediated methylenedioxymethamphetamine (MDMA) demethylation in time- and dose-dependent manner and that the inhibition required the presence of NADPH. The development of irreversible inhibition was associated with oxidation at position 4 of the aromatic ring, the common site of CYP2D-mediated oxidation of this group of compounds. In in vivo studies doses of ALLMA (1-10 mg/kg) were administered to adult male Sprague-Dawley rats and liver microsomes were obtained 3 hr later. Methamphetamine p-hydroxylation and low Km MDMA demethylation activities, both mediated by CYP2D, were reduced by more than 80% after a dose of 10 mg/kg. Cytochrome P-450 reactions attributed to P-450s other than CYP2D, such as aniline p-hydroxylation, the high Km system of MDMA demethylation and the N-demethylation of methamphetamine, benzphetamine, aminopyrine and erythromycin, all appeared to be minimally affected. The importance of aromatic ring oxidation in the metabolism is such that inhibition of CYP2D would be expected to cause a significant change in the pharmacokinetics of these compounds. The kinetics of MDMA metabolic activity in microsomes from ALLMA-pretreated rats were comparable to those from female Dark-Agouti rats, an animal model for CYP2D1 deficiency.

To establish whether the actions of D-amphetamine (Amp) and D-methamphetamine (MeAmp) on the striatal dopamine system were equipotent, pharmacokinetic profiles of each drug were applied to an analysis of their respective induced dopamine efflux profiles. Amp or MeAmp (1 and 5 mg/kg i.v.) was administered to chloral hydrate-anesthetized rats; plasma and brain kinetics were then assessed from 5 to 60 min. Dose-dependent increases in Amp and MeAmp plasma levels resulted in proportional increases in striatum levels that were equivalent for both drugs; elimination rates also were similar and were characterized by a first-order decay process. After MeAmp administration, low levels of brain MeAmp metabolites were detected throughout the 1-hr time period; relative to MeAmp, Amp and p-hydroxy-MeAmp levels were less than 10 and 1%, respectively. The drug-induced dopamine efflux profiles in the striatum were characterized by microdialysis; Amp and MeAmp (1, 2.5 and 5 mg/kg i.v.) effected equivalent, dose-dependent increases in extracellular dopamine levels. For both drugs at 5- and 10-min postinjection, increases in drug striatum levels preceded increases in dopamine efflux. In contrast, from the time of the peak dopamine responses observed at 10 to 20 min until the end of the study at 90 min, changes in striatal drug levels were correlated with extracellular dopamine levels; this correlation was similar for both drugs. These results indicate that Amp and MeAmp pharmacokinetics and their subsequent dopamine responses in the striatum are equivalent. The pharmacokinetic analysis can be extended to the interpretation of other comparative studies that assess effects of Amp and MeAmp.(ABSTRACT TRUNCATED AT 250 WORDS)

The purpose of this study was to determine whether phencyclidine (PCP) inhibits constitutive hepatic cytochrome P450 (CYP) isozymes when administered to naive adult male Sprague-Dawley rats. Animals were pretreated with PCP (25 mg/kg/day for 2 days), killed 3 and 16 hr after the last dose, and liver microsomes prepared. The washed microsomes were then assayed for benzphetamine, methamphetamine (MA), and methylenedioxymethamphetamine (MDMA) N-demethylation together with MDMA demethylenation and MA 4-hydroxylation activities. MDMA demethylenation (low substrate concentration), MA 4-hydroxylation, and metoprolol alpha-hydroxylation reactions, which are catalyzed by CYP2D isozymes, were reduced > 74% 3 hr after the last PCP dose and were only partially restored 13 hr later. Benzphetamine and (-)-MDMA N-demethylation activities were restored to control values 16 hr after the last dose. These results indicate that PCP suppresses constitutive isozymes, including CYP2C11 and members of the CYP2D subfamily. The suppression of cytochromes P450 activity by PCP in vivo is consistent with its in vitro actions found in this and other studies, and demonstrates that alteration of CYP activity is another pharmacological effect of this compound.