Eosinophilic pulmonary disease with ulcerative colitis: A case report.

RATIONALE: Eosinophilic pulmonary disease (EPD) is a general term for a large group of diseases with complex etiology. Ulcerative colitis is an inflammatory bowel disease (IBD). Patients with IBD may have pulmonary involvement. We herein present a case of ulcerative colitis complicated with EPD. PATIENT CONCERNS: A 34-year-old woman with ulcerative colitis presented with dry cough. She had peripheral eosinophilia and apical ground glass opacities on CT (computed tomography) of her chest. Antibiotic treatment was ineffective. DIAGNOSES: Lung biopsy revealed eosinophil infiltration in the alveolar space and interstitial space, so EPD was considered. INTERVENTIONS: After oral administration of prednisone, the lung shadow on CT disappeared when the cough symptoms resolved. However, the symptoms recurred after drug withdrawal, and the lung shadow reappeared on imaging. The cough symptoms and lung shadow disappeared after oral prednisone was given again. Prednisone was slowly discontinued after 6 months of treatment. OUTCOMES: The patient stopped prednisone for half a year. No recurrence or abnormal CT findings were detected during the half-year follow-up. LESSONS: The clinical manifestations of EPD are atypical, laboratory and imaging findings are not specific, and it is difficult to make a definite diagnosis before lung biopsy. The diagnosis depends on pathological examination. Glucocorticoid treatment is effective, but some patients may relapse after drug withdrawal. Active follow-up after glucocorticoid treatment is very important for identifying disease recurrence. Patients with IBD are relatively prone to developing EPD. The etiology of EPD is complex. In clinical practice, we need to make a diagnosis and differential diagnosis to clarify its etiology.

Kidney organoid models reveal cilium-autophagy metabolic axis as a therapeutic target for PKD both in vitro and in vivo.

Human pluripotent stem cell-derived kidney organoids offer unprecedented opportunities for studying polycystic kidney disease (PKD), which still has no effective cure. Here, we developed both in vitro and in vivo organoid models of PKD that manifested tubular injury and aberrant upregulation of renin-angiotensin aldosterone system. Single-cell analysis revealed that a myriad of metabolic changes occurred during cystogenesis, including defective autophagy. Experimental activation of autophagy via ATG5 overexpression or primary cilia ablation significantly inhibited cystogenesis in PKD kidney organoids. Employing the organoid xenograft model of PKD, which spontaneously developed tubular cysts, we demonstrate that minoxidil, a potent autophagy activator and an FDA-approved drug, effectively attenuated cyst formation in vivo. This in vivo organoid model of PKD will enhance our capability to discover novel disease mechanisms and validate candidate drugs for clinical translation.

A missing jigsaw within the hygiene hypothesis: Low-dose bisphenol A exposure attenuates lipopolysaccharide-induced asthma protection.

The rising occurrence of allergic asthma in early life across industrialized countries suggests that environmental factors play a crucial role in determining asthma susceptibility and severity. While prior exposure to microbial lipopolysaccharides (LPSs) has been found to offer protection against allergic asthma, infants residing in urban environments are increasingly exposed to environmental pollutants. Utilizing limulus lysate test screens and virtual screening models, we identified pollutants that can modulate LPS bioactivity. This investigation revealed that bisphenol A (BPA), a chemical commonly used in numerous household items and previously implicated in obesity and cancer, effectively neutralizes LPS. In-depth mechanistic analyses showed that BPA specifically binds to the lipid A component of LPS, leading to inactivation. This interaction eliminates the immunostimulatory activity of LPS, making mice more susceptible to house dust mite (HDM)-induced allergic asthma. BPA reactivates lung epithelial cells, consequently amplifying type 2 responses to HDMs in dendritic cells. This chemical interplay provides new insights into the pathophysiology of asthma in relation to human exposure. Understanding the intricate relationships between environmental chemicals and microbial antigens, as well as their impacts on innate immunity, is critical for the development of intervention strategies to address immune disorders resulting from urbanization.

Analogy or fallacy, unsafe chemical alternatives: Mechanistic insights into energy metabolism dysfunction induced by Bisphenol analogs in HepG2 cells.

Bisphenol analogs (BPs) are widely used as industrial alternatives for Bisphenol A (BPA). Their toxicity assessment in humans has mainly focused on estrogenic activity, while other toxicity effects and mechanisms resulting from BPs exposure remain unclear. In this study, we investigated the effects of three BPs (Bisphenol AF (BPAF), Bisphenol G (BPG) and Bisphenol PH (BPPH)) on metabolic pathways of HepG2 cells. Results from comprehensive cellular bioenergetics analysis and nontarget metabolomics indicated that the most important process affected by BPs exposure was energy metabolism, as evidenced by reduced mitochondrial function and enhanced glycolysis. Compared to the control group, BPG and BPPH exhibited a consistent pattern of metabolic dysregulation, while BPAF differed from both, such as an increased ATP: ADP ratio (1.29-fold, p < 0.05) observed in BPAF and significantly decreased ATP: ADP ratio for BPG (0.28-fold, p < 0.001) and BPPH (0.45-fold, p < 0.001). Bioassay endpoint analysis revealed BPG/BPPH induced alterations in mitochondrial membrane potential and overproductions of reactive oxygen species. Taken together these data suggested that BPG/BPPH induced oxidative stress and mitochondrial damage in cells results in energy metabolism dysregulation. By contrast, BPAF had no effect on mitochondrial health, but induced a proliferation promoting effect on cells, which might contribute to the energy metabolism dysfunction. Interestingly, BPPH induced the greatest mitochondrial damage among the three BPs but did not exhibit Estrogen receptor alpha (ERα) activating effects. This study characterized the distinct metabolic mechanisms underlying energy metabolism dysregulation induced by different BPs in target human cells, providing new insight into the evaluation of the emerging BPA substitutes.

Environmental endocrine disruptor Bisphenol A induces metabolic derailment and obesity via upregulating IL-17A in adipocytes.

BACKGROUND: Bisphenol A (BPA), a ubiquitous environmental endocrine disruptor, has been extensively demonstrated to be associated with metabolic disorders, including obesity and type 2 diabetes mellitus. However, the underlying mechanism underpinning the environmental etiology of chronic metabolic disorders has not been sufficiently elucidated. OBJECTIVES: This study is designed to explore the toxicological pathogenesis of chronic inflammation in BPA exposure during obesity. METHODS: We investigated the role of IL-17A in the association of BPA exposure and obesity from human cross-sectional study to animal models, including genetically modified IL-17A-/- mice. RESULTS: Here, our work started from case-control observation that BPA exposure was significantly associated with risk of obesity (odds ratio = 4.72, 95%CI: 3.18 - 11.18, P < 0.01), metabolic disorder and levels of interleukin-17A (IL-17A) in human adipose (estimated changes ß = 0.46, 95%CI: 0.15 - 1.01, P < 0.01) with bariatric surgery. Animal model fed with high-fat diet (HFD) confirmed that BPA exposure aggravated body weight gain and insulin resistance, concurrent with much heightened inflammatory responses in the adipose tissue including increase in IL-17A and macrophage polarization towards M1 stage. Genetically modified IL-17A ablated mice (IL-17A-/-) showed reversed adipose tissue inflammation response, improved macrophage polarization homeostasis, along with insulin sensitivity in both HFD group alone or much more significantly the HFD + BPA group. Moreover, mediation analysis in human epidemiological investigation demonstrated that plasma IL-17A attributed up to 30.01% mediating role in the associations between BPA exposure and obesity risk. DISCUSSION: This research paradigm from human to animal provides strong evidence for the elucidation of IL-17A moderating inflammation and insulin resistance in obesity. Such findings reiterate the obesogenic role of environmental endocrine disruptor BPA in metabolic disorders and unveils the potential toxicological mechanisms underpinning such effect.

Neisseria species as pathobionts in bronchiectasis.

Neisseria species are frequently identified in the bronchiectasis microbiome, but they are regarded as respiratory commensals. Using a combination of human cohorts, next-generation sequencing, systems biology, and animal models, we show that bronchiectasis bacteriomes defined by the presence of Neisseria spp. associate with poor clinical outcomes, including exacerbations. Neisseria subflava cultivated from bronchiectasis patients promotes the loss of epithelial integrity and inflammation in primary epithelial cells. In vivo animal models of Neisseria subflava infection and metabolipidome analysis highlight immunoinflammatory functional gene clusters and provide evidence for pulmonary inflammation. The murine metabolipidomic data were validated with human Neisseria-dominant bronchiectasis samples and compared with disease in which Pseudomonas-, an established bronchiectasis pathogen, is dominant. Metagenomic surveillance of Neisseria across various respiratory disorders reveals broader importance, and the assessment of the home environment in bronchiectasis implies potential environmental sources of exposure. Thus, we identify Neisseria species as pathobionts in bronchiectasis, allowing for improved risk stratification in this high-risk group.

Preclinical investigation of artesunate as a therapeutic agent for hepatocellular carcinoma via impairment of glucosylceramidase-mediated autophagic degradation.

Artesunate (ART) has been indicated as a candidate drug for hepatocellular carcinoma (HCC). Glucosylceramidase (GBA) is required for autophagic degradation. Whether ART regulates autophagic flux by targeting GBA in HCC remains to be defined. Herein, our data demonstrated that the dramatic overexpression of GBA was significantly associated with aggressive progression and short overall survival times in HCC. Subsequent experiments revealed an association between autophagic activity and GBA expression in clinical HCC samples, tumor tissues from a rat model of inflammation-induced HCC and an orthotopic mouse model, and human HCC cell lines. Interestingly, probe labeling identified GBA as an ART target, which was further verified by both a glutathione-S-transferase pulldown assay and surface plasmon resonance analysis. The elevated protein expression of LC3B, the increased numbers of GFP-LC3B puncta and double-membrane vacuoles, and the enhanced expression of SQSTM1/p62 indicated that the degradation of autophagosomes in HCC cells was inhibited by ART treatment. Both the in vitro and in vivo data revealed that autophagosome accumulation through targeting of GBA was responsible for the anti-HCC effects of ART. In summary, this preclinical study identified GBA as one of the direct targets of ART, which may have promising potential to inhibit lysosomal autophagy for HCC therapy.

Non-targeted metabolomics revealing the effects of bisphenol analogues on human liver cancer cells.

Bisphenol analogues (BPs) are widely used in plastics, food packaging and other commercial products as non safer alternative of BPA. As emerging environmental contaminants, BPs have received considerable attention for their adverse effects on human health. However, their effects on liver metabolisms are only marginally understood. In this study, high-resolution mass spectrometry-based global metabolomics and extracellular flux (XF) analysis were applied to characterize the cellular metabolome alterations and reveal the possible mechanisms of the metabolic disorders associated with BPs-induced toxicity in HepG2 cells. BPE, BPB and BPAP with similar chemical structures were selected to compare their interference with different metabolic pathways. A total of 61 key metabolite profiles were significantly altered after exposure to the three BPs. Overall, BPs altered metabolites which are associated with energy metabolism, oxidative stress, cell proliferation and nucleotides synthesis. The primary dysregulated pathways included energy and nucleotides synthesis related Purine and Glycolysis/Gluconeogenesis metabolism. In addition, attenuated mitochondrial function and enhanced glycolysis were found under BPB and BPAP treatment. While attenuated glycolysis was observed under BPE treatment. These findings may provide potential biomarkers indicating the cytotoxicity of BPs and prompt a deeper understanding of the intramolecular metabolic processes induced by BPs exposure.

Dose-response metabolomics and pathway sensitivity to map molecular cartography of bisphenol A exposure.

In the toxicological regime, the toxicological endpoint and its dose-response relationship are two of the most prominent characters in conducting a risk assessment for chemical exposure. Systems biological methods have been used to comprehensively characterize the impact of toxicants on the biochemical pathways. However, the majority of the current studies are only based on single-dose, and limited information can be extrapolated to other doses from these experiments, regardless of the sensitivity of each endpoint. This study aims to understand the dose-response metabolite dysregulation pattern and metabolite sensitivity at the system-biological level. Here, we applied bisphenol A (BPA), an endocrine-disrupting chemical (EDC), as the model chemical. We first employed the global metabolomics method to characterize the metabolome of breast cancer cells (MCF-7) upon exposure to different doses (0, 20, 50, and 100 µM) of BPA. The dysregulated features with a clear dose-response relationship were also effectively picked up with an R-package named TOXcms. Overall, most metabolites were dysregulated by showing a significant dose-dependent behaviour. The results suggested that BPA exposure greatly perturbed purine metabolism and pyrimidine metabolism. Interestingly, most metabolites within the purine metabolism were described as a biphasic dose-response relationship. With the established dose-response relationship, we were able to fully map the metabolite cartography of BPA exposure within a wide range of concentrations and observe some unique patterns. Furthermore, an effective concentration of certain fold changes (e.g., EC+10 means the dose at which metabolite is 10% upregulated) and metabolite sensitivity were defined and introduced to this dose-response omics information. The result showed that the purine metabolism pathway is the most venerable target of BPA, which can be a potential endogenous biomarker for its exposure. Overall, this study applied the dose-response metabolomics method to fully understand the biochemical pathway disruption of BPA treatment at different doses. Both dose-response omics strategy and metabolite sensitivity analysis can be further considered and emphasized in future chemical risk assessments.

Pathway Retrofitting Yields Insights into the Biosynthesis of Anthraquinone-Fused Enediynes.

Anthraquinone-fused enediynes (AQEs) are renowned for their distinctive molecular architecture, reactive enediyne warhead, and potent anticancer activity. Although the first members of AQEs, i.e., dynemicins, were discovered three decades ago, how their nitrogen-containing carbon skeleton is synthesized by microbial producers remains largely a mystery. In this study, we showed that the recently discovered sungeidine pathway is a "degenerative" AQE pathway that contains upstream enzymes for AQE biosynthesis. Retrofitting the sungeidine pathway with genes from the dynemicin pathway not only restored the biosynthesis of the AQE skeleton but also produced a series of novel compounds likely as the cycloaromatized derivatives of chemically unstable biosynthetic intermediates. The results suggest a cascade of highly surprising biosynthetic steps leading to the formation of the anthraquinone moiety, the hallmark C8-C9 linkage via alkyl-aryl cross-coupling, and the characteristic epoxide functionality. The findings provide unprecedented insights into the biosynthesis of AQEs and pave the way for examining these intriguing biosynthetic enzymes.

Polycyclic aromatic hydrocarbons (PAHs) in indoor environments are still imposing carcinogenic risk.

Polycyclic aromatic hydrocarbons (PAHs) are among the most health-relevant air pollutants. Herein, we conducted meta-analysis and experimental validation to evaluate PAHs in our surroundings and carcinogenic risks. We summarized the occurrence of PAHs in outdoors and indoors from 131 studies with 6,766 samples collected in different countries in 1989-2019. The global weighted-median concentration in outdoor air, indoor air and dust of ΣPAHs were 142 ng/m3, 369 ng/m3 and 10,201 ng/g; respectively. ΣPAHs have decreased in indoor air but remained steady in outdoor air and indoor dust. More carcinogenic PAHs in indoor/outdoor air was observed in Asia, while in dust was North America. Monte-Carlo simulation further showed indoor sources for children's exposure from dust and air can exceed outdoor. To further validate the health effect of PAHs from indoors, 15 more recent indoor dust samples were collected to examine their mutagenicity. The results showed that ΣPAHs were found to be significantly correlated with mutagenicity potency in the dust sample metabolically activated with liver S9 subcellular fraction and likely accounted for 0.42-0.50 of the mutagenic activity. Our findings indicated that PAHs are still likely to have carcinogenic activity in indoor environments and exposure risk of children to indoor dust should be emphasized.

Metabolomic and Transcriptomic Analysis of MCF-7 Cells Exposed to 23 Chemicals at Human-Relevant Levels: Estimation of Individual Chemical Contribution to Effects.

BACKGROUND: Humans are constantly being exposed to various xenobiotics at relatively low concentrations. To date, limited evidence is available to ascertain whether a complex xenobiotic mixture at human-relevant levels causes any health effect. Moreover, there is no effective method to pinpoint the contribution of each chemical toward such an effect. OBJECTIVES: This study aims to understand the responses of cells to a mixture containing 23 xenobiotics at human-relevant levels and develop a feasible method to decipher the chemical(s) that contribute significantly to the observed effect. METHODS: We characterized the metabolome and transcriptome of breast cancer cells (MCF-7) before and after exposure to the mixture at human-relevant levels; preexposure levels were derived from existing large-scale biomonitoring data. A high-throughput metabolomics-based "leave-one-out" method was proposed to understand the relative contribution of each component by comparing the metabolome with and without the particular chemical in the mixture. RESULTS: The metabolomic analysis suggested that the mixture altered metabolites associated with cell proliferation and oxidative stress. For the transcriptomes, gene ontology terms and pathways including "cell cycle," "cell proliferation," and "cell division" were significantly altered after mixture exposure. The mixture altered genes associated with pathways such as "genotoxicity" and "nuclear factor erythroid 2-related factor 2 (Nrf2)." Through joint pathways analysis, metabolites and genes were observed to be well-aligned in pyrimidine and purine metabolisms. The leave-one-out results showed that many chemicals made their contributions to specific metabolic pathways. The overall metabolome pattern of the absence of 2,4-dihyroxybenzophenone (DHB) or bisphenol A (BPA) showed great resemblance to controls, suggesting their higher relative contribution to the observed effect. DISCUSSION: The omics results showed that exposure to the mixture at human-relevant levels can induce significant in vitro cellular changes. Also, the leave one out method offers an effective approach for deconvoluting the effects of the mixture. https://doi.org/10.1289/EHP6641.

A Generic Liquid Chromatography-Tandem Mass Spectrometry Exposome Method for the Determination of Xenoestrogens in Biological Matrices.

We are constantly exposed to a variety of environmental contaminants and hormones, including those mimicking endogenous estrogens. These highly heterogeneous molecules are collectively referred to as xenoestrogens and hold the potential to affect and alter the delicate hormonal balance of the human body. To monitor exposure and investigate potential health implications, comprehensive analytical methods covering all major xenoestrogen classes are needed but not available to date. Herein, we describe a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the simultaneous determination of multiple classes of endogenous as well as exogenous estrogens in human urine, serum, and breast milk to enable proper exposure and risk assessment. In total, 75 analytes were included, whereof a majority was successfully in-house validated in the three matrices. Extraction recoveries of validated analytes ranged from 71% to 110% and limits of quantification from 0.015 to 5 µg/L, 0.03 to 14 µg/L, and 0.03 to 4.6 µg/L in urine, serum, and breast milk, respectively. The applicability of the novel method was demonstrated in proof-of-principle experiments by analyzing urine from Austrian individuals and breast milk from Austrian and Nigerian individuals. Thereby, we proved the methods' feasibility to identify and quantify different classes of xenoestrogens simultaneously. The results illustrate the general importance of multiclass exposure assessment in the context of the exposome paradigm. Specifically, they highlight the need for estimating total estrogenic burden rather than single analyte or chemical class measurements and its potential impact in endocrine disruption and hormone related diseases including cancers.

Generation of Human PSC-Derived Kidney Organoids with Patterned Nephron Segments and a De Novo Vascular Network.

Human pluripotent stem cell-derived kidney organoids recapitulate developmental processes and tissue architecture, but intrinsic limitations, such as lack of vasculature and functionality, have greatly hampered their application. Here we establish a versatile protocol for generating vascularized three-dimensional (3D) kidney organoids. We employ dynamic modulation of WNT signaling to control the relative proportion of proximal versus distal nephron segments, producing a correlative level of vascular endothelial growth factor A (VEGFA) to define a resident vascular network. Single-cell RNA sequencing identifies a subset of nephron progenitor cells as a potential source of renal vasculature. These kidney organoids undergo further structural and functional maturation upon implantation. Using this kidney organoid platform, we establish an in vitro model of autosomal recessive polycystic kidney disease (ARPKD), the cystic phenotype of which can be effectively prevented by gene correction or drug treatment. Our studies provide new avenues for studying human kidney development, modeling disease pathogenesis, and performing patient-specific drug validation.

Exposure and risk assessment of volatile organic compounds and airborne phthalates in Singapore's Child Care Centers.

Infants and children under 6 years old spend most of daily time in Child Care Centers (CCCs), especially in the tropical regions like Singapore. Environmental exposure and associated risk during this early critical developmental stage is of great public concern. In this study, seven representative volatile organic compounds (VOCs) and five typical phthalates were analyzed in the indoor and outdoor air samples collected from 32 Singapore CCCs. The median of total VOC and phthalate concentration in indoor air was 19.03 and 5.41 µg m-3; respectively. For both indoors and outdoors environment, benzene, toluene and xylene were the dominant VOC contributors (more than 68%). For indoor air phthalates, di(2-ethylhexyl) phthalate and di-butyl phthalate (DBP) accounts for 60-76%. The level of both VOCs and phthalates in indoor environment was significantly higher than that in outdoor, with an average indoor/outdoor ratio of 1.24 and 1.45; respectively. A strong correlation (r > 0.50, p < 0.05) was observed between indoor and outdoor air compounds. VOC and phthalate levels have no significant difference between CCCs with split-unit and centrally ventilated air conditioners. Monte Carlo simulation was used to estimate exposure uncertainty and variability for the risk assessment. Overall, the concentrations of VOC were below the healthy reference values from either EPA Integrated Risk Information System (IRIS) or Singapore guideline. However, similar to other countries' report, benzene, DBP, ethylbenzene and naphthalene were at levels that could exceed the stringent standards such as Office of Environmental Health Hazard Assessment (OEHHA) cancer and reproductive health-based benchmarks.

Metabolomics Reveals that Dietary Xenoestrogens Alter Cellular Metabolism Induced by Palbociclib/Letrozole Combination Cancer Therapy.

Recently, the palbociclib/letrozole combination therapy was granted accelerated US FDA approval for the treatment of estrogen receptor (ER)-positive breast cancer. Since the underlying metabolic effects of these drugs are yet unknown, we investigated their synergism at the metabolome level in MCF-7 cells. As xenoestrogens interact with the ER, we additionally aimed at deciphering the impact of the phytoestrogen genistein and the estrogenic mycotoxin zearalenone. A global metabolomics approach was applied to unravel metabolite and pathway modifications. The results clearly showed that the combined effects of palbociclib and letrozole on cellular metabolism were far more pronounced than that of each agent alone and potently influenced by xenoestrogens. This behavior was confirmed in proliferation experiments and functional assays. Specifically, amino acids and central carbon metabolites were attenuated, while higher abundances were observed for fatty acids and most nucleic acid-related metabolites. Interestingly, exposure to model xenoestrogens appeared to counteract these effects.

Quantitative metabolomics of photoreceptor degeneration and the effects of stem cell-derived retinal pigment epithelium transplantation.

Photoreceptor degeneration is characteristic of vision-threatening diseases including age-related macular degeneration. Photoreceptors are metabolically demanding cells in the retina, but specific details about their metabolic behaviours are unresolved. The quantitative metabolomics of retinal degeneration could provide valuable insights and inform future therapies. Here, we determined the metabolomic 'fingerprint' of healthy and dystrophic retinas in rat models using optimized metabolite extraction techniques. A number of classes of metabolites were consistently dysregulated during degeneration: vitamin A analogues, fatty acid amides, long-chain polyunsaturated fatty acids, acyl carnitines and several phospholipid species. For the first time, a distinct temporal trend of several important metabolites including DHA (4Z,7Z,10Z,13Z,16Z,19Z-docosahexaenoic acid), all-trans-retinal and its toxic end-product N-retinyl-N-retinylidene-ethanolamine were observed between healthy and dystrophic retinas. In this study, metabolomics was further used to determine the temporal effects of the therapeutic intervention of grafting stem cell-derived retinal pigment epithelium (RPE) in dystrophic retinas, which significantly prevented photoreceptor atrophy in our previous studies. The result revealed that lipid levels such as phosphatidylethanolamine in eyes were restored in those animals receiving the RPE grafts. In conclusion, this study provides insight into the metabolomics of retinal degeneration, and further understanding of the efficacy of RPE transplantation.This article is part of the themed issue 'Quantitative mass spectrometry'.

Thermal Degradation of Small Molecules: A Global Metabolomic Investigation.

Thermal processes are widely used in small molecule chemical analysis and metabolomics for derivatization, vaporization, chromatography, and ionization, especially in gas chromatography mass spectrometry (GC/MS). In this study the effect of heating was examined on a set of 64 small molecule standards and, separately, on human plasma metabolite extracts. The samples, either derivatized or underivatized, were heated at three different temperatures (60, 100, and 250 °C) at different exposure times (30 s, 60 s, and 300 s). All the samples were analyzed by liquid chromatography coupled to electrospray ionization mass spectrometry (LC/MS) and the data processed by XCMS Online ( xcmsonline.scripps.edu ). The results showed that heating at an elevated temperature of 100 °C had an appreciable effect on both the underivatized and derivatized molecules, and heating at 250 °C created substantial changes in the profile. For example, over 40% of the molecular peaks were altered in the plasma metabolite analysis after heating (250 °C, 300s) with a significant formation of degradation and transformation products. The analysis of 64 small molecule standards validated the temperature-induced changes observed on the plasma metabolites, where most of the small molecules degraded at elevated temperatures even after minimal exposure times (30 s). For example, tri- and diorganophosphates (e.g., adenosine triphosphate and adenosine diphosphate) were readily degraded into a mono-organophosphate (e.g., adenosine monophosphate) during heating. Nucleosides and nucleotides (e.g., inosine and inosine monophosphate) were also found to be transformed into purine derivatives (e.g., hypoxanthine). A newly formed transformation product, oleoyl ethyl amide, was identified in both the underivatized and derivatized forms of the plasma extracts and small molecule standard mixture, and was likely generated from oleic acid. Overall these analyses show that small molecules and metabolites undergo significant time-sensitive alterations when exposed to elevated temperatures, especially those conditions that mimic sample preparation and analysis in GC/MS experiments.

Antibody microarray profiling of osteosarcoma cell serum for identifying potential biomarkers.

The aim of the present study was to identify biomarkers in osteosarcoma (OS) cell serum by antibody microarray profiling, which may be used for OS diagnosis and therapy. An antibody microarray was used to detect the expression levels of cytokines in serum samples from 20 patients with OS and 20 healthy individuals. Significantly expressed cytokines in OS serum were selected when P<0.05 and fold change >2. An enzyme-linked immunosorbent assay (ELISA) was used to validate the antibody microarray results. Finally, classification accuracy was calculated by cluster analysis. Twenty one cytokines were significantly upregulated in OS cell serum samples compared with control samples. Expression of interleukin-6, monocyte chemoattractant protein-1, tumor growth factor-ß, growth-related oncogene, hepatocyte growth factor, chemokine ligand 16, Endoglin, matrix metalloproteinase-9 and platelet-derived growth factor-AA was validated by ELISAs. OS serum samples and control samples were distinguished by significantly expressed cytokines with an accuracy of 95%. The results demonstrated that expressed cytokines identified by antibody microarray may be used as biomarkers for OS diagnosis and therapy.

Metabolites of organophosphate flame retardants and 2-ethylhexyl tetrabromobenzoate in urine from paired mothers and toddlers.

As a result of the polybrominated diphenyl ether (PBDE) ban in the mid-2000s, the chemical flame retardant market has moved toward alterative compounds including chlorinated alkyl and nonchlorinated aryl organophosphate flame retardants (OPFRs) as well as aromatic brominated compounds such as Firemaster 550 (FM550). Recent studies have shown that the OPFRs and Firemaster 550 components are frequently detected in polyurethane foams and in indoor dust. Some OPFRs are considered carcinogenic and/or neurodevelopmental toxicants, and children's exposure to these compounds is a concern. OPFRs are readily metabolized and excreted in the urine as their dialkyl and diaryl compounds which function as biomarkers for OPFR exposure. Limited research has shown that adults are broadly exposed to OPFRs, but nothing is known about children's exposure. Similarly, 2-ethylhexyl-2,3,4,5-tetrabromobenzoate (EH-TBB), a FM550 component, is metabolized to tetrabromobenzoic acid (TBBA). The current study measured levels of bis(1,3-dichloro-2-propyl) phosphate (BDCIPP), bis(1-chloro-2-propyl) phosphate (BCIPP), diphenyl phosphate (DPHP), 2 alkylated DPHPs, and TBBA in urine collected in 2013 from 21 US mother-toddler pairs. BDCIPP, DPHP, and ip-DPHP were detected in 100%, 98%, and 96% of all individuals, whereas BCIPP and tert-butyl-DPHP (tb-DPHP) were only detected in 8% and 13%. Further, TBBA was detected in 27% of adults but 70% of children. Overall, children had higher urinary levels of BDCIPP, DPHP, ip-DPHP, and TBBA as compared to their mothers, suggesting higher exposure. For example, on average, BDCIPP levels in children were 4.9 times those of mothers. BDCIPP and DPHP levels in mother's urine were also significantly correlated with levels in children's urine, suggesting similar exposure routes, likely in the home environment. Various potential predictors of OPFR exposure were assessed using a questionnaire. In children some predictors of hand-mouth exposure were associated with elevated BDCIPP and DPHP levels (e.g., less frequent hand washing for BDCIPP). Overall, these trends are consistent with higher flame retardant levels in children as a result of increased hand-mouth behavior and elevated dust exposure.