Genome-Wide Analysis Reveals Mucociliary Remodeling of the Nasal Airway Epithelium Induced by Urban PM2.5.

Air pollution particulate matter <2.5 µm (PM2.5) exposure is associated with poor respiratory outcomes. Mechanisms underlying PM2.5-induced lung pathobiology are poorly understood but likely involve cellular and molecular changes to the airway epithelium. We extracted and chemically characterized the organic and water-soluble components of air pollution PM2.5 samples, then determined the whole transcriptome response of human nasal mucociliary airway epithelial cultures to a dose series of PM2.5 extracts. We found that PM2.5 organic extract (OE), but not water-soluble extract, elicited a potent, dose-dependent transcriptomic response from the mucociliary epithelium. Exposure to a moderate OE dose modified the expression of 424 genes, including activation of aryl hydrocarbon receptor signaling and an IL-1 inflammatory program. We generated an OE-response gene network defined by eight functional enrichment groups, which exhibited high connectivity through CYP1A1, IL1A, and IL1B. This OE exposure also robustly activated a mucus secretory expression program (>100 genes), which included transcriptional drivers of mucus metaplasia (SPDEF and FOXA3). Exposure to a higher OE dose modified the expression of 1,240 genes and further exacerbated expression responses observed at the moderate dose, including the mucus secretory program. Moreover, the higher OE dose significantly increased the MUC5AC/MUC5B gel-forming mucin expression ratio and strongly downregulated ciliated cell expression programs, including key ciliating cell transcription factors (e.g., FOXJ1 and MCIDAS). Chronic OE stimulation induced mucus metaplasia-like remodeling characterized by increases in MUC5AC+ secretory cells and MUC5AC mucus secretions. This epithelial remodeling may underlie poor respiratory outcomes associated with high PM2.5 exposure.

Assessment of gonadal and thyroid histology in Gulf killifish (Fundulus grandis) from Barataria Bay Louisiana one year after the Deepwater Horizon oil spill.

We examined gonads and thyroid glands of Gulf killifish (Fundulus grandis) 1yr after the Deepwater Horizon oil spill. F. grandis were trapped from two impacted sites in Barataria Bay (Bayou St. Denis, Bay Jimmy) and an un-impacted site in East Texas (Sabine Pass). The greatest number of F. grandis were collected at Sabine Pass. F. grandis collected at Bayou St. Denis were smaller and had smaller Fulton condition factor scores than fish collected at Sabine Pass. Sex ratios were biased roughly 2:1 in favor of females at Sabine Pass and Bayou St. Denis. Gonad-somatic index (GSI) in males from Sabine Pass was double that of fish from Bay Jimmy while germinal epithelium thickness of the testes was 2.7 fold smaller in males from the impacted site. GSI and oocyte diameters in females from Bayou St. Denis were significantly smaller than females from Bay Jimmy or the reference site. There were no differences in thyroid follicle cell height. While total polyaromatic hydrocarbons at the impacted sites were no different from the reference site, the impacted sites did have greater concentrations of benzo[a]pyrene in sediment pore water. The finding of smaller GSI and testicular germinal epithelium in males from an impacted site suggest that exposure to a combination of oil and dispersants may adversely impact testicular function.

Enhanced anticancer effects of Scutellaria barbata D. Don in combination with traditional Chinese medicine components on non-small cell lung cancer cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Experience-based herbal medicine as a complementary to modern western medicine has triggered an array of studies in quest of novel anticancer drugs. Scutellaria barbata D. Don (SB) is commonly used to treat different types of cancers, but its molecular mechanism of action is not clearly understood. In this study, we attempted to elucidate the mode of action of a traditional Chinese medicine prescription with a total of 14 components, named Lian-Jia-San-Jie-Fang (LJSJF, in Chinese), where SB works as the "principle" against non-small cell lung cancer (NSCLC) cells. MATERIALS AND METHODS: Four different NSCLC cell lines (A549, H460, H1650, and H1975) were used. Cytotoxicity, in vitro tumorigenicity, gene expression, and protein expression were analyzed by MTT assay, soft agar assay, real-time PCR, and Western blots, respectively. RESULTS: Among the 14 components in LJSJF, SB was the only one to possess cytotoxic effects at its pharmacologically relevant doses. Additionally, we observed synergistically dose-dependent cytotoxic effects of SB in combination with other LJSJF components. After SB or LJSJF treatment, significant reductions in colony number and/or size were observed in A549 and H460; a notable dose-dependent decrease in EGFR was observed in A549, H460, and H1650; significant downregulation in EGFR and its downstream signaling targets mTOR and p38MAPK were also observed in A549 and H460; and p53 and p21 were significantly increased while survivin, cyclin D1, and MDM2 were significantly decreased in A549. Additionally, p53, p21, and Mettl7b were decreased, but p73 was increased in H460. Neither EGFR nor p53 was changed in H1975. Therefore, SB or LJSJF may induce cytotoxic effects by regulating multiple and/or distinct apoptotic pathways in different NSCLC cells. CONCLUSION: LJSJF exerts more pronounced cytotoxic effects against NSCLC cells than SB does by synergistically regulating the underlining molecular mechanisms including EGFR and/or p53 signaling pathways.

Chlorogenic Acid Ameliorates Experimental Colitis by Promoting Growth of Akkermansia in Mice.

Chlorogenic acid (ChA)-one of the most abundant polyphenol compounds in the human diet-exerts anti-inflammatory activities. The aim of this study was to investigate the effect of ChA on gut microbiota in ulcerative colitis (UC). Colitis was induced by 2.5% dextran sulfate sodium (DSS) in C57BL/6 mice, which were on a control diet or diet with ChA (1 mM). The histopathological changes and inflammation were evaluated. Fecal samples were analyzed by 16S rRNA gene sequencing. ChA attenuated several effects of DSS-induced colitis, including weight loss, increased disease activity index, and improved mucosal damage. Moreover, ChA could significantly suppress the secretion of IFNγ, TNFα, and IL-6 and the colonic infiltration of F4/80⁺ macrophages, CD3⁺ T cells, and CD177⁺ neutrophils via inhibition of the active NF-κB signaling pathway. In addition, ChA decreased the proportion of Firmicutes and Bacteroidetes. ChA also enhanced a reduction in fecal microbiota diversity in DSS treated mice. Interestingly, ChA treatment markedly increased the proportion of the mucin-degrading bacterium Akkermansia in colitis mice. ChA acted as the intestine-modifying gut microbial community structure, resulting in a lower intestinal and systemic inflammation and also improving the course of the DSS-induced colitis, which is associated with a proportional increase in Akkermansia.

Epigallocatechin-3-gallate augments the therapeutic effects of benzo[a]pyrene-mediated lung carcinogenesis.

Our previous study found curcumin and vitamin E to have protective effects against benzo[a]pyrene (BaP) exposure in human normal lung epithelial BEAS-2B cells. The first objective of this study was to determine whether epigallocatechin-3-gallate (EGCG) elicited the same response. Co-treatment with 5 µM BaP and 20 µM EGCG in BEAS-2B promoted a significant reduction in cell viability and greater G2/M cell cycle arrest, induction of ROS, and reductions in BaP-induced CYP1A1/CYP1B1/COMT, EGFR, p-Akt (Ser473), p-p53 (Thr55), and survivin mRNA/protein expression, as well as an increase in p-p53 (Ser15). Based on these findings, the second objective was to extend the investigation by developing a novel BaP-transformed BEAS-2B cell line, BEAS-2BBaP , to examine the effects of EGCG when co-administered with gefitinib, an EGFR tyrosine kinase inhibitor. Cell colony formation assay demonstrated in vitro tumorigenic potential of BEAS-2BBaP , which had an overexpression of EGFR. Viability testing revealed gefitinib co-treatment with EGCG resulted in more cell death compared with gefitinib alone. Co-treated cells had greater reductions in gefitinib-induced CYP1A1/CYB1B1, EGFR, cyclin D1, p-Akt (Ser473), and survivin mRNA/protein expression, as well as an increase in p-p53 (Ser15). Therefore, EGCG was found to promote greater cytotoxicity to BEAS-2B co-treated with BaP and BEAS-2BBaP upon gefitinib co-treatment through regulating metabolism enzymes and signaling pathways involving EGFR and p53. These findings suggest that EGCG did not act as a protective compound in BEAS-2B after acute BaP exposure, but has the potential to be a useful adjuvant chemotherapeutic compound when coupled with gefitinib for chemosensitization. © 2017 BioFactors, 43(4):529-539, 2017.

Therapeutic properties of green tea against environmental insults.

Pesticides, smoke, mycotoxins, polychlorinated biphenyls (PCBs), and arsenic are the most common environmental toxins and toxicants to humans. These toxins and toxicants may impact on human health at the molecular (DNA, RNA, or protein), organelle (mitochondria, lysosome, or membranes), cellular (growth inhibition or cell death), tissue, organ, and systemic levels. Formation of reactive radicals, lipid peroxidation, inflammation, genotoxicity, hepatotoxicity, embryotoxicity, neurological alterations, apoptosis, and carcinogenic events are some of the mechanisms mediating the toxic effects of the environmental toxins and toxicants. Green tea, the nonoxidized and nonfermented form of tea that contains several polyphenols, including green tea catechins, exhibits protective effects against these environmental toxins and toxicants in preclinical studies and to a much-limited extent, in clinical trials. The protective effects are collectively mediated by antioxidant, antiinflammatory, antimutagenic, hepatoprotective and neuroprotective, and anticarcinogenic activities. In addition, green tea modulates signaling pathway including NF-κB and ERK pathways, preserves mitochondrial membrane potential, inhibits caspase-3 activity, down-regulates proapoptotic proteins, and induces the phase II detoxifying pathway. The bioavailability and metabolism of green tea and its protective effects against environmental insults induced by pesticides, smoke, mycotoxins, PCBs, and arsenic are reviewed in this paper. Future studies with emphasis on clinical trials should identify biomarkers of green tea intake, examine the mechanisms of action of green tea polyphenols, and investigate potential interactions of green tea with other toxicant-modulating dietary factors.

BDE47 induces rat CYP3A1 by targeting the transcriptional regulation of miR-23b.

Cytochrome P450 3A (CYP3A) is the most abundant CYP450 enzyme in the liver and is involved in the metabolism of over 50% of xenobiotics. Our previous studies revealed that 2,2',4,4'-tetrabromodiphenyl ether (BDE47) could induce rat CYP3A1 expression, but the molecular basis remains unclear. Using in silico analysis, we identified a potential miR-23b recognition element (MRE23b) in the 3'-UTR region of CYP3A1 mRNA, which was verified by the luciferase assay. The miR-23b mimic and inhibitor significantly down- and up-regulated the expression of CYP3A1, respectively. Additionally, BDE47 significantly down-regulated the expression of miR-23b in rats and in hepatic H4IIE cells. Induction or blockage of CYP3A1 by a miR-23b inhibitor or mimic could correspondingly alter BDE47-induced expression of CYP3A1 and cytotoxicity in H4IIE cells. Furthermore, LV-anti-miR-23b significantly decreased endogenous levels of miR-23b and increased the expression and activity of CYP3A1 in rat liver. LV-anti-miR-23b also significantly increased the hydroxylated metabolites of BDE47 (3-OH-BDE47, 4-OH-BDE42, and 4'-OH-BDE49) in rat serum. In conclusion, we first found that BDE47 induced rat CYP3A1 expression by targeting the transcriptional regulation of miR-23b. This study helps provide a better understanding of CYP3A regulation and offers novel clues for the role of miRNAs in the metabolism and distribution of environmental pollutants.

The classic EDCs, phthalate esters and organochlorines, in relation to abnormal sperm quality: a systematic review with meta-analysis.

The association between endocrine disrupting chemicals (EDCs) and human sperm quality is controversial due to the inconsistent literature findings, therefore, a systematic review with meta-analysis was performed. Through the literature search and selection based on inclusion criteria, a total of 9 studies (7 cross-sectional, 1 case-control, and 1 pilot study) were analyzed for classic EDCs (5 studies for phthalate esters and 4 studies for organochlorines). Funnel plots revealed a symmetrical distribution with no evidence of publication bias (Begg's test: intercept = 0.40; p = 0.692). The summary odds ratios (OR) of human sperm quality associated with the classic EDCs was 1.67 (95% CI: 1.31-2.02). After stratification by specific chemical class, consistent increases in the risk of abnormal sperm quality were found in phthalate ester group (OR = 1.52; 95% CI: 1.09-1.95) and organochlorine group (OR = 1.98; 95% CI: 1.34-2.62). Additionally, identification of official data, and a comprehensive review of the mechanisms were performed, and better elucidated the increased risk of these classic EDCs on abnormal sperm quality. The present systematic review and meta-analysis helps to identify the impact of classic EDCs on human sperm quality. However, it still highlights the need for additional epidemiological studies in a larger variety of geographic locations.

Twist1-mediated 4E-BP1 regulation through mTOR in non-small cell lung cancer.

Twist1 overexpression corresponds with poor survival in non-small cell lung cancer (NSCLC), but the underlining mechanism is not clear. The objective of the present study was to investigate the tumorigenic role of Twist1 and its related molecular mechanisms in NSCLC. Twist1 was overexpressed in 34.7% of NSCLC patients. The survival rate was significantly lower in patients with high Twist1 expression than low expression (P < 0.05). Twist1 expression levels were higher in H1650 cells, but relatively lower in H1975 cells. H1650 with stable Twist1 knockdown, H1650shTw, demonstrated a significantly slower rate of wound closure; however, H1975 with stable Twist1 overexpression, H1975Over, had an increased motility velocity. A significant decrease in colony number and size was observed in H1650shTw, but a significant increase in colony number was found in H1975Over (P < 0.05). Tumor growth significantly decreased in mice implanted with H1650shTw compared to H1650 (P < 0.05). 4E-BP1 and p53 gene expressions were increased, but p-4E-BP1 and p-mTOR protein expressions were decreased in H1650shTw. However, 4E-BP1 gene expression was decreased, while p-4E-BP1 and p-mTOR protein expressions were increased in H1975Over. p-4E-BP1 was overexpressed in 24.0% of NSCLC patients. Survival rate was significantly lower in patients with high p-4E-BP1 expression than low p-4E-BP1 (P < 0.01). A significant correlation was found between Twist1 and p-4E-BP1 (P < 0.01). A total of 13 genes in RT-PCR array showed significant changes in H1650shTw. Altogether, Twist1 is correlated with p-4E-BP1 in predicting the prognostic outcome of NSCLC. Inhibition of Twist1 decreases p-4E-BP1 expression possibly through downregulating p-mTOR and increasing p53 expression in NSCLC.

Epigallocatechin-3-gallate enhances the therapeutic effects of leptomycin B on human lung cancer a549 cells.

Our previous studies have shown Leptomycin B (LMB) is a promising antilung cancer drug. Epigallocatechin-3-gallate (EGCG) has antitumor properties but a debatable clinical application. The objective of this study is to evaluate the combination therapeutic effect of LMB and EGCG and its molecular mechanisms in human lung cancer A549 cells. Increased cytotoxicity was observed in LMB+EGCG-treated cells compared to LMB-treated cells. Elevated ROS was maximized 2 h after treatment, and LMB+EGCG-treated cells had higher ROS levels compared to LMB. N-Acetyl-L-cysteine (NAC) studies confirmed the oxidative role of LMB and/or EGCG treatment. In comparison to the control, CYP3A4, SOD, GPX1, and p21 mRNA expression levels were increased 7.1-, 2.0-, 4.6-, and 13.1-fold in LMB-treated cells, respectively, while survivin was decreased 42.6-fold. Additionally, these increases of CYP3A4, SOD, and GPX1 were significantly reduced, while p21 was significantly increased in LMB+EGCG-treated cells compared to LMB-treated cells. The qRT-PCR results for p21 and survivin were further confirmed by Western blot. Our study first shows that LMB produces ROS and is possibly metabolized by CYP3A4, GPX1, and SOD in A549 cells, and combination treatment of LMB and EGCG augments LMB-induced cytotoxicity through enhanced ROS production and the modulation of drug metabolism and p21/survivin pathways.

Curcumin and vitamin E protect against adverse effects of benzo[a]pyrene in lung epithelial cells.

Benzo[a]pyrene (BaP), a well-known environmental carcinogen, promotes oxidative stress and DNA damage. Curcumin and vitamin E (VE) have potent antioxidative activity that protects cells from oxidative stress and cellular damage. The objectives of the present study were to investigate the adverse effects of BaP on normal human lung epithelial cells (BEAS-2B), the potential protective effects of curcumin and VE against BaP-induced cellular damage, and the molecular mechanisms of action. MTT assay, flow cytometry, fluorescence microplate assay, HPLC, qRT-PCR, and western blot were performed to analyze cytotoxicity, cell cycle, reactive oxygen species (ROS), BaP diol-epoxidation (BPDE)-DNA adducts, gene expression, and protein expression, respectively. Curcumin or VE prevented cells from BaP-induced cell cycle arrest and growth inhibition, significantly suppressed BaP-induced ROS levels, and decreased BPDE-DNA adducts. While CYP1A1 and 1B1 were induced by BaP, these inductions were not significantly reduced by curcumin or VE. Moreover, the level of activated p53 and PARP-1 were significantly induced by BaP, whereas this induction was markedly reduced after curcumin and VE co-treatment. Survivin was significantly down-regulated by BaP, and curcumin significantly restored survivin expression in BaP-exposed cells. The ratio of Bax/Bcl-2 was also significantly increased in cells exposed to BaP and this increase was reversed by VE co-treatment. Taken together, BaP-induced cytotoxicity occurs through DNA damage, cell cycle arrest, ROS production, modulation of metabolizing enzymes, and the expression/activation of p53, PARP-1, survivin, and Bax/Bcl-2. Curcumin and VE could reverse some of these BaP-mediated alterations and therefore be effective natural compounds against the adverse effects of BaP in lung cells.

A low blood volume LC-MS/MS assay for the quantification of fentanyl and its major metabolites norfentanyl and despropionyl fentanyl in children.

Preterm and term neonates often require surgical procedures and analgesia. However, our knowledge about neonatal pharmacokinetics of fentanyl, the most commonly used drug for these procedures, and its metabolites is still incomplete. To facilitate pharmacokinetic studies of fentanyl and its metabolites in neonates and other children, we developed and validated an LC-MS/MS method based on minimally invasive, low blood volume sampling. LC-MS/MS was used for the simultaneous analysis of fentanyl, despropionyl fentanyl (DPF), and norfentanyl from dried blood samples (DBS) collected on filter paper. Positive ions were monitored using multiple reaction monitoring. Since the standard matrix for measuring fentanyl blood concentrations is plasma, the assay was developed and validated in plasma, whole blood, and then DBS. Our method was able to measure clinically relevant levels of fentanyl and its metabolites. In DBS, the lower limits of quantification were 100 pg/mL for fentanyl with a range of reliable response from 0.1 to 100 ng/mL (r(2)>0.99) and 250 pg/mL for both DPF and norfentanyl with a range of reliable response from 0.25 to 100 ng/mL (r(2)>0.99). In plasma and in DBS inter-day accuracy and precisions of fentanyl met predefined acceptance criteria and also indicated comparable assay performance in both matrices.