Search

Nitro-containing compounds such as nitrofuran and nitroimidazole are drugs used for the treatment of infectious diseases. However, many of these nitro-containing drugs are positive in the bacterial reverse mutation assay (Ames test). The recently approved anti-multidrug-resistant tuberculosis (MDR-TB) drug, delamanid (Deltyba™; OPC-67683), a derivative of 4-nitroimidazole, was negative for mutagenicity in the Ames assay. In Salmonella typhimurium , mutagenicity of nitro compounds has been closely associated with the ability of nitroreductase to metabolize (degradation)these compounds. To explore the lack of mutagenicity for delamanid, we examined the initial metabolic rate and mutagenic-specific activity of a series of nitro compounds in S. typhimurium TA100. The order of maximum mutagenic-activity was nitrofuran > 2-nitroimidazole > 5-nitroimidazole ≥ 4-nitroimidazole, which is very similar to the order of initial metabolic rate, i.e., the Pearson's correlation coefficient (r = 0.85) showed a correlation between metabolic rate and mutagenic-activity. No metabolism of delamanid was detected even after 60 h of treatment. In addition, delamanid was not reduced by two human nitroreductases. These facts may explain the absence of genotoxicity of delamanid in both in vitro and in vivo tests. [ABSTRACT FROM AUTHOR]

Abstract: RELAXIN (RLN) is a polypeptide hormone of the insulin-like hormone family; it facilitates birth by softening and widening the pubic symphysis and cervix in many mammals, including humans. The role of RLN in bone metabolism was recently suggested by its ability to induce osteoclastogenesis and activate osteoclast function. RLN binds to RELAXIN/INSULIN-LIKE FAMILY PEPTIDE 1 (RXFP1) and 2 (RXFP2), with varying species-specific affinities. Young men with mutated RXFP2 are at high risk for osteoporosis, as RXFP2 influences osteoblast metabolism by binding to INSULIN-LIKE PEPTIDE 3 (INSL3). However, there have been no reports on RLN function in osteoblast differentiation and mineralization or on the functionally dominant receptors for RLN in osteoblasts. We previously described Rxfp1 and 2 expression patterns in developing mouse oral components, including the maxillary and mandibular bones, Meckel's cartilage, tongue, and tooth primordia. We hypothesized that Rln/Rxfp signaling is a key mediator of skeletal development and metabolism. Here, we present the gene expression patterns of Rxfp1 and 2 in developing mouse calvarial frontal bones as determined by in situ hybridization. In addition, RLN enhanced osteoblastic differentiation and caused abnormal mineralization and extracellular matrix metabolism through Rxfp2, which was predominant over Rxfp1 in MC3T3-E1 mouse calvarial osteoblasts. Our data suggest a novel role for Rln in craniofacial skeletal development and metabolism through Rxfp2. [Copyright &y& Elsevier]

Abstract: Prostate cancer, a leading cause of cancer-related deaths in males, is well recognized as having late disease on-set (mostly at age 60–70) and showing slow/latent disease development, and strategies to prevent cancer formation in late manhood may have significant health impacts. Cucurbitacin B (CuB) is a naturally occurring compound that is found abundantly in cucumbers and other vegetables, and it is known to exert anti-cancer activities (primarily via apoptosis-induction) in several human cancers. However, its chemopreventive potential for prostate cancer has not yet been investigated. Here, we reported that CuB significantly and specifically inhibited prostate cancer cell growth with low IC50 (∼0.3μM; PC-3 and LNCaP), accompanied by marked apoptosis (Caspase 3/7 activation, PARP cleavage, increase of Annexin V-Alexa Fluor 488 (Alexa488)+ cells and accumulation of Sub-G0/G1 population), whereas normal human prostate epithelial cells (PrEC) were CuB-insensitive. Using a chemopreventive model, pre-treatment of mice with CuB (2weeks before PC-3 prostate cancer cell implantation) significantly reduced the rate of in vivo tumor-formation. A 79% reduction in tumor size (accompanied by marked in situ apoptosis) was observed in the CuB-treated group (with no noticeable toxicity) vs. controls at day 31. Strikingly, mechanistic investigations demonstrated that CuB drove dose-dependent inhibition of ATP citrate lyase phosphorylation (ACLY; an important enzyme for cancer metabolism) both in vitro and in the CuB-chemopreventive mouse model. Importantly, ACLY over-expression abrogated CuB’s apoptotic effects in prostate cancer cells, confirming ACLY as a direct target of CuB. Thus, CuB harbors potent chemopreventive activity for prostate cancer, and we revealed a novel anti-tumor mechanism of CuB via inhibition of ACYL signaling in human cancer. [Copyright &y& Elsevier]

Abstract: Rheumatoid arthritis (RA) patients usually exhibit immune complex (IC) deposition and increased neutrophil activation in the joint. In this study, we assessed how four flavonols (galangin, kaempferol, quercetin, and myricetin) modulate the effector functions of healthy individuals' and active RA patients' IC-stimulated neutrophils. We measured superoxide anion and total reactive oxygen species production using lucigenin (CL-luc)- and luminol (CL-lum)-enhanced chemiluminescence assays, respectively. Galangin, kaempferol, and quercetin inhibited CL-lum to the same degree (mean IC50 =2.5μM). At 2.5μM, quercetin and galangin suppressed nearly 65% CL-lum of active RA patients' neutrophils. Quercetin inhibited CL-luc the most effectively (IC50 =1.71±0.36μM). The four flavonols diminished myeloperoxidase activity, but they did not decrease NADPH oxidase activity, phagocytosis, microbial killing, or cell viability of neutrophils. The ability of the flavonols to scavenge hypochlorous acid and chloramines, but not H2O2, depended on the hydroxylation degree of the flavonol B-ring. Therefore, at physiologically relevant concentrations, the flavonols partially inhibited the oxidative metabolism of IC-stimulated neutrophils without affecting the other investigated effector functions. Using these compounds to modulate IC-mediated neutrophil activation is a promising safe therapeutic strategy to control inflammation in active RA patients. [Copyright &y& Elsevier]

Highlights: [•] Milk was investigated for estrogenic endocrine disruptor activity. [•] A novel estrogenic reporter gene assay was developed for quantifying total estrogenic load in milk. [•] The validated method is suitable for screening a range of milk samples, except soy. [•] Testing real samples revealed various levels of estrogenic activity. [Copyright &y& Elsevier]

Abstract: Glucocorticosteroids (GCs) are widely used to treat different kinds of chronic inflammatory and immune diseases through transcriptional regulation of inflammatory genes. Modulation of gene expression by GCs is known to occur through diverse mechanisms of varying relevance to specific classes of genes. Epigenetic modifications are indeed a pivotal regulatory feature of glucocorticoid receptor and other transcription factors. In this study, histone post-translational modifications were investigated for their involvement in the regulation of selected pro-inflammatory genes – expressed in human monocyte-derived macrophages – in response to treatment with synthetic GC dexamethasone (DEX). We show that histone tail acetylation status is modified following DEX administration, through distinct and alternative mechanisms at the promoters of interleukin-8 and interleukin-23. In addition to histone H3 acetylation, our results demonstrate that H3 lysine 4 trimethylation is affected following drug treatment. [Copyright &y& Elsevier]

Highlights: [•] Toxic effects of the combination of carbon nanotubes and nonylphenol are examined. [•] No generalized evidences of synergistic toxicity were reported for most conditions. [•] More stable dispersions of nanoparticles were achieved by adding serum to the medium. [•] The serum increased the tolerance of 3T3-L1 cells to all treatments tested. [•] Intracellular distribution of nanoparticles was analyzed by Raman spectroscopy. [Copyright &y& Elsevier]

Highlights: [•] Effects of protein oxidation by haze PM2.5 were investigated. [•] Oxidative stress was increased by the haze PM2.5. [•] Haze episodes to albumin resulted in oxidation of methionine moieties. [•] Oxidation of methionine associated with oxidative stress and PAHs in the PM2.5. [ABSTRACT FROM AUTHOR]

Abstract: ABC transporters including MRP2, MDR1 and BCRP play a major role in tissue defense. Epidemiological and experimental studies suggest a cytoprotective role of estrogens in intestine, though the mechanism remains poorly understood. We evaluated whether pharmacologic concentrations of ethynylestradiol (EE, 0.05pM to 5nM), or concentrations of genistein (GNT) associated with soy ingestion (0.1–10μM), affect the expression and activity of multidrug resistance proteins MRP2, MDR1 and BCRP using Caco-2 cells, an in vitro model of intestinal epithelium. We found that incubation with 5pM EE and 1μM GNT for 48h increased expression and activity of both MRP2 and MDR1. Estrogens did not affect expression of BCRP protein at any concentration studied. Irrespective of the estrogen tested, up-regulation of MDR1 and MRP2 protein was accompanied by increased levels of MDR1 mRNA, whereas MRP2 mRNA remained unchanged. Cytotoxicity assays demonstrated association of MRP2 and MDR1 up-regulation with increased resistance to cell death induced by 1-chloro-2,4-dinitrobenzene, an MRP2 substrate precursor, and by paraquat, an MDR1 substrate. Experiments using an estrogen receptor (ER) antagonist implicate ER participation in MRP2 and MDR1 regulation. GNT but not EE increased the expression of ERβ, the most abundant form in human intestine and in Caco-2 cells, which could lead in turn to increased sensitivity to estrogens. We conclude that specific concentrations of estrogens can confer resistance against cytotoxicity in Caco-2 cells, due in part to positive modulation of ABC transporters involved in extrusion of their toxic substrates. Although extrapolation of these results to the in vivo situation must be cautiously done, the data could explain tentatively the cytoprotective role of estrogens against chemical injury in intestine. [Copyright &y& Elsevier]

Abstract: The synthesis of new substituted E-3-(3-indolylmethylene)1,3-dihydroindol-2-ones is reported. The antiproliferative activity was evaluated according to protocols available at the National Cancer Institute (NCI), Bethesda, MD. The action of the most active compound 10 was further investigated in HL-60 leukemia cells. Results obtained show that it causes a block in cell cycle progression, with cell arrest in the G2/M phase, associated with activation of apoptosis accompanied with increased oxidative stress and deregulation of the homeostasis of divalent cations, with significant increase in the cellular concentrations of free Ca2+ and Mg2+. [Copyright &y& Elsevier]

Highlights: [•] NM exposure caused growth arrest and increase in DNA double strand breaks in JB6 cells. [•] NM exposure caused the activation of both DNA DSB repair pathways NHEJ and HRR. [•] Unlike NHEJ, inactivation of HRR sensitized the cells towards NM-induced cytotoxicity. [ABSTRACT FROM AUTHOR]

Abstract: The epidermis of skin is the first line of defense against the environment. A three dimensional model of human skin was used to investigate tissue-specific phenotypes induced by the environmental contaminant, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Continuous treatment of organotypic cultures of human keratinocytes with TCDD resulted in intracellular spaces between keratinocytes of the basal and immediately suprabasal layers as well as thinning of the basement membrane, in addition to the previously reported hyperkeratinization. These tissue remodeling events were preceded temporally by changes in expression of the extracellular matrix degrading enzyme, matrix metalloproteinase-10 (MMP-10). In organotypic cultures MMP-10 mRNA and protein were highly induced following TCDD treatment. Q-PCR and immunoblot results from TCDD-treated monolayer cultures, as well as indirect immunofluorescence and immunoblot analysis of TCDD-treated organotypic cultures, showed that MMP-10 was specifically contributed by the epidermal keratinocytes but not the dermal fibroblasts. Keratinocyte-derived MMP-10 protein accumulated over time in the dermal compartment of organotypic cultures. TCDD-induced epidermal phenotypes in organotypic cultures were attenuated by the keratinocyte-specific expression of tissue inhibitor of metalloproteinase-1, a known inhibitor of MMP-10. These studies suggest that MMP-10 and possibly other MMP-10-activated MMPs are responsible for the phenotypes exhibited in the basement membrane, the basal keratinocyte layer, and the cornified layer of TCDD-treated organotypic cultures. Our studies reveal a novel mechanism by which the epithelial–stromal microenvironment is altered in a tissue-specific manner thereby inducing structural and functional pathology in the interfollicular epidermis of human skin. [Copyright &y& Elsevier]

Highlights: [•] Reverse-phase evaporation method was chosen as the optimal method for preparation of RGPL. [•] Based on single factor investigation, RSM was employed to determine the optimum preparation conditions of RGPL. [•] RGPL significantly stimulated proliferation of murine splenic lymphocyte in vitro. [Copyright &y& Elsevier]

Abstract: In this study, we tested the feasibility of magnetic liposomes as a carrier for pulmonary preferential accumulation of fasudil, an investigational drug for the treatment of pulmonary arterial hypertension (PAH). To develop an optimal inhalable formulation, various magnetic liposomes were prepared and characterized for physicochemical properties, storage stability and in vitro release profiles. Select formulations were evaluated for uptake by pulmonary arterial smooth muscle cells (PASMCs) – target cells – using fluorescence microscopy and HPLC. The efficacy of the magnetic liposomes in reducing hyperplasia was tested in 5-HT-induced proliferated PASMCs. The drug absorption profiles upon intratracheal administration were monitored in healthy rats. Optimized spherical liposomes – with mean size of 170nm, zeta potential of −35mV and entrapment efficiency of 85% – exhibited an 80% cumulative drug release over 120h. Fluorescence microscopic study revealed an enhanced uptake of liposomes by PASMCs under an applied magnetic field: the uptake was 3-fold greater compared with that observed in the absence of magnetic field. PASMC proliferation was reduced by 40% under the influence of the magnetic field. Optimized liposomes appeared to be safe when incubated with PASMCs and bronchial epithelial cells. Compared with plain fasudil, intratracheal magnetic liposomes containing fasudil extended the half-life and area under the curve by 27- and 14-fold, respectively. Magnetic-liposomes could be a viable delivery system for site-specific treatment of PAH. [Copyright &y& Elsevier]

Highlights: [•] PCB quinone is genotoxic positive to HepG2 cells. [•] PCB quinone exposure causes increasing of DNA breaks and micronuclei. [•] PCB quinone induced 8-OHdG formation. [•] PCB quinone induced γ-H2AX expression. [•] ROS scavengers inhibited the genotoxicity of PCB quinone. [ABSTRACT FROM AUTHOR]

Highlights: [•] Impulse noise leads to OHC pathology within 24h after exposure. [•] In the normal cochlea phospho-p53 (Ser 15) was lightly expressed in the cochlea. [•] P53 is overexpressed in both OHC and Hensen cells in the early stage of damage. [•] OHC damage could be reduced by PFT, a p53 inhibitor; or by KX1-004. [•] A greater protection by KX1-004 suggests a role of Src-dependent pathway in noise. [Copyright &y& Elsevier]

Abstract: Calcium/calmodulin-stimulated protein kinase II (CaMKII) is a multi-functional serine/threonine protein kinase that controls a range of cellular functions, including proliferation. The biological properties of CaMKII are regulated by multi-site phosphorylation and targeting via interactions with specific proteins. To investigate the role specific CaMKII phosphorylation sites play in controlling cell proliferation and cell cycle progression, we examined phosphorylation of CaMKII at two sites (T253 and T286) at various stages of the cell cycle, and also examined the effects of overexpression of wild-type (WT), T286D phosphomimic, T253D phosphomimic and T253V phosphonull forms of CaMKIIα in MDA-MB-231 breast cancer and SHSY5Y neuroblastoma cells on cellular proliferation and cell cycle progression. We demonstrate herein that whilst there is no change in total CaMKII expression or T286 phosphorylation throughout the cell cycle, a marked dephosphorylation of CaMKII at T253 occurs during the G2 and/or M phases. Additionally, we show by molecular inhibition, as well as pharmacological activation, that protein phosphatase 2A (PP2A) is the phosphatase responsible for this dephosphorylation. Furthermore, we show that inducible overexpression of WT, T286D and T253V forms of CaMKIIα in MDA-MB-231 and SHSY5Y cells increases cellular proliferation, with no alteration in cell cycle profiles. By contrast, overexpression of a T253D phosphomimic form of CaMKIIα significantly decreases proliferation, and cells accumulate in mitosis, specifically in metaphase. Taken together, these results strongly suggest that the dephosphorylation of CaMKII at T253 is involved in controlling the cell cycle, specifically the metaphase–anaphase transition. [Copyright &y& Elsevier]

The aim of this study was to prepare and characterize cholesterol-poly(ethylene) glycol (chol-PEG) nanocarriers of two different molecular weights (1 and 5 kDa) and to determine their effect on the transscleral retention and permeation of a lipophilic multi-therapeutic agent, sirolimus (rapamycin), with potential application in angiogenic and immunogenic ocular diseases. Sirolimus-containing nanocarriers were prepared using the thin-film hydration method and characterized for their physicochemical properties including size, drug entrapment (EE) and loading (DL) efficiencies, stability, surface charge, morphology, critical micelle concentration (CMC) and thermal properties. Ussing chambers were used to determine the retention and permeability of sirolimus-containing nanocarriers in porcine sclera followed by ultrastructural tissue examination. Sirolimus-containing nanocarriers had an average size of 11.7 nm (chol-PEG 1 kDa) and 13.8 nm (chol-PEG 5 kDa) and zeta potentials of 0.41 and −1.05, respectively. Both nanocarriers had similar transscleral permeabilities (chol-PEG 1 kDa 6.44 × 10−7 and 5 kDa 6.16 × 10−7 cm2 s−1), and very high scleral retention compared with a free solution of sirolimus (chol-PEG 1 kDa 16.9 μg/g; chol-PEG 5 kDa 7.48 μg/g; free sirolimus 0.57 μg/g). The DL (EE) for chol-PEG 1 and 5 kDa were 2.93% (77.4%) and 3.10% (81.6%), respectively. The CMC values for the nanocarriers were similar to those previously reported in literature (3.85 × 10−7 M for chol-PEG 1 kDa; 4.26 × 10−7 M for chol-PEG 5 kDa). In conclusion, chol-PEG nanocarriers successfully loaded sirolimus and resulted in scleral permeation and high retention, which shows potential utility for the topical delivery of lipophilic ocular drugs. [ABSTRACT FROM AUTHOR]

Highlights: [•] Plasma treated titanium is better to cell viability than untreated titanium. [•] Cellular antioxidant response was more efficient on plasma treated titanium. [•] Plasma treatment could improve the titanium biocompatibility. [ABSTRACT FROM AUTHOR]

Abstract: Neuropathological evidence indicates that dopaminergic cell death in Parkinson׳s disease (PD) involves impairment of mitochondrial complex I, oxidative stress, microglial activation, and the formation of Lewy bodies. Epidemiological findings suggest that the consumption of berries rich in anthocyanins and proanthocyanidins may reduce PD risk. In this study, we investigated whether extracts rich in anthocyanins, proanthocyanidins, or other polyphenols suppress the neurotoxic effects of rotenone in a primary cell culture model of PD. Dopaminergic cell death elicited by rotenone was suppressed by extracts prepared from blueberries, grape seed, hibiscus, blackcurrant, and Chinese mulberry. Extracts rich in anthocyanins and proanthocyanidins exhibited greater neuroprotective activity than extracts rich in other polyphenols, and a number of individual anthocyanins interfered with rotenone neurotoxicity. The blueberry and grape seed extracts rescued rotenone-induced defects in mitochondrial respiration in a dopaminergic cell line, and a purple basal extract attenuated nitrite release from microglial cells stimulated by lipopolysaccharide. These findings suggest that anthocyanin- and proanthocyanidin-rich botanical extracts may alleviate neurodegeneration in PD via enhancement of mitochondrial function. [Copyright &y& Elsevier]

Abstract: Cigarette smoking is a key factor for the development and progression of different cancers including mammary tumor in women. Resveratrol (Res) is a promising natural chemotherapeutic agent that regulates many cellular targets including p21, a cip/kip family of cyclin kinase inhibitors involved in DNA damage-induced cell cycle arrest and blocking of DNA replication and repair. We have recently shown that cigarette smoke condensate (CSC) prepared from commercially available Indian cigarette can cause neoplastic transformation of normal breast epithelial MCF-10A cell. Here we studied the mechanism of Res mediated apoptosis in CSC transformed (MCF-10A-Tr) cells in vitro and in vivo. Res mediated apoptosis in MCF-10A-Tr cells was a p21 dependent event. It increased the p21 protein expression in MCF-10A-Tr cells and MCF-10A-Tr cells-mediated tumors in xenograft mice. Res treatment reduced the tumor size(s) and expression of anti-apoptotic proteins (e.g. PI3K, AKT, NFκB) in solid tumor. The expressions of cell cycle regulatory (Cyclins, CDC-2, CDC-6, etc.), BER associated (Pol-β, Pol-δ, Pol-ε, Pol-η, RPA, Fen-1, DNA-Ligase-I, etc.) proteins and LP-BER activity decreased in MCF-10A-Tr cells but remain significantly unaltered in isogenic p21 null MCF-10A-Tr cells after Res treatment. Interestingly, no significant changes were noted in SP-BER activity in both the cell lines after Res exposure. Finally, it was observed that increased p21 blocks the LP-BER in MCF-10A-Tr cells by increasing its interaction with PCNA via competing with Fen-1 after Res treatment. Thus, Res caused apoptosis in CSC-induced cancer cells by reduction of LP-BER activity and this phenomenon largely depends on p21. [Copyright &y& Elsevier]

Highlights: [•] Bile acid toxicity: glycine conjugated>unconjugated>taurine conjugated. [•] (Cheno)deoxycholate more toxic in day-1 SCRH than in day-3 SCRH. [•] Lower glycine conjugate levels in day-3 SCRH may explain this toxicity difference. [•] Decreased conjugates formation in day-3 SCRH drives lower glycine conjugate levels. [•] Canalicular/sinusoidal efflux contributes to decreased intracellular exposure. [Copyright &y& Elsevier]

Abstract: The main purpose of this research was to prepare and to characterize ferulic acid-loaded nanostructured lipid carrier (FA-NLC) to evaluate the cytotoxic effect on human glioblastoma cancer U87MG cells. First of all, the influence of different materials on mean size and homogeneity of NLC prepared by a low energy organic solvent-free method was investigated. Technological characterization (encapsulation efficiency, mean particle size, homogeneity and in vitro release profile) was performed on the selected NLC in comparison to others lipid carriers, nanoemulsion and SLN. Furthermore, the thermal behavior of NLC and SLN was investigated using Differential Scanning Calorimetry (DSC) in order to evaluate their structure. Biological studies (MTT bioassay and caspase-3 cleavage) on the selected NLC showed no cytotoxic effects of the unloaded tested NLC. Besides, the effectiveness of FA-loaded NLC was higher compared to the free drug. Cells treated with FA or FA-loaded NLC showed a greater effect compared to idebenone (IDE) or IDE-loaded NLC, respectively. These results strongly support that FA-loaded NLC could be potentially used for the treatment of glioblastoma. [Copyright &y& Elsevier]

Highlights: [•] BPA for 24h promoted dendritic development of hippocampal neurons. [•] BPA for 24h increased the level of F-actin in dendrites of hippocampal neurons. [•] BPA promoted the expression of Rac1/Cdc42 but inhibited that of RhoA. [•] These BPA-induced effects were blocked by ERs antagonist and were suppressed by MEK1/2 inhibitor. [Copyright &y& Elsevier]

Abstract: The Cancer/Testis Antigen (CTA), Prostate-associated Gene 4 (PAGE4), is a stress-response protein that is upregulated in prostate cancer (PCa) especially in precursor lesions that result from inflammatory stress. In cells under stress, translocation of PAGE4 to mitochondria increases while production of reactive oxygen species decreases. Furthermore, PAGE4 is also upregulated in human fetal prostate, underscoring its potential role in development. However, the proteins that interact with PAGE4 and the mechanisms underlying its pleiotropic functions in prostatic development and disease remain unknown. Here, we identified c-Jun as a PAGE4 interacting partner. We show that both PAGE4 and c-Jun are overexpressed in the human fetal prostate; and in cell-based assays, PAGE4 robustly potentiates c-Jun transactivation. Single-molecule Förster resonance energy transfer experiments indicate that upon binding to c-Jun, PAGE4 undergoes conformational changes. However, no interaction is observed in presence of BSA or unilamellar vesicles containing the mitochondrial inner membrane diphosphatidylglycerol lipid marker cardiolipin. Together, our data indicate that PAGE4 specifically interacts with c-Jun and that, conformational dynamics may account for its observed pleiotropic functions. To our knowledge, this is the first report demonstrating crosstalk between a CTA and a proto-oncogene. Disrupting PAGE4/c-Jun interactions using small molecules may represent a novel therapeutic strategy for PCa. [Copyright &y& Elsevier]

Purpose: c-MYC is a promising target for cancer therapy but its use is restricted by unwanted, devastating side effects. We explored whether intravesical instillation of the c-MYC inhibitor KSI-3716 could suppress tumor growth in murine orthotopic bladder xenografts. Materials and Methods: The small molecule KSI-3716, which blocks c-MYC/MAX binding to target gene promoters, was used as an intravesical chemotherapy agent. KSI-3716 action was assessed by electrophoretic mobility shift assay, chromatin immunoprecipitation, transcription reporter assay and quantitative reverse transcriptase-polymerase chain reaction. Inhibition of cell proliferation and its mechanism was monitored by cell cytotoxicity assay, EdU incorporation assay and flow cytometry. The in vivo efficacy of KSI-3716 was examined by noninvasive luminescence imaging and histological analysis after intravesical instillation of KSI-3716 in murine orthotopic bladder xenografts. Results: KSI-3716 blocked c-MYC/MAX from forming a complex with target gene promoters. c-MYC mediated transcriptional activity was inhibited by KSI-3716 at concentrations as low as 1 μM. The expression of c-MYC target genes, such as cyclin D2, CDK4 and hTERT, was markedly decreased. KSI-3716 exerted cytotoxic effects on bladder cancer cells by inducing cell cycle arrest and apoptosis. Intravesical instillation of KSI-3716 at a dose of 5 mg/kg significantly suppressed tumor growth with minimal systemic toxicity. Conclusions: The c-MYC inhibitor KSI-3716 could be developed as an effective intravesical chemotherapy agent for bladder cancer. [ABSTRACT FROM AUTHOR]

Abstract: Current antibiotics for treating Clostridium difficile infections (CDI), that is, metronidazole, vancomycin and more recently fidaxomicin, are mostly effective but treatment failure and disease relapse remain as significant clinical problems. The shortcomings of these agents are attributed to their low selectivity for C. difficile over normal gut microflora and their ineffectiveness against C. difficile spores. This Letter reports that certain diarylacylhydrazones identified during a high-throughput screening/counter-screening campaign show selective activity against two Clostridium species (C. difficile and Clostridium perfringens) over common gut commensals. Representative examples are shown to possess activity similar to vancomycin against clinical C. difficile strains and to kill stationary-phase C. difficile cells, which are responsible for spore production. Structure–activity relationships with additional synthesised analogues suggested a protonophoric mechanism may play a role in the observed activity/selectivity and this was supported by the well-known protonophore carbonyl cyanide m-chlorophenyl hydrazone (CCCP) showing selective anti-Clostridium effects and activity similar to diarylacylhydrazones against stationary-phase C. difficile cells. Two diarylacylhydrazones were shown to be non-toxic towards human FaDu and Hep G2 cells indicating that further studies with the class are warranted towards new drugs for CDI. [Copyright &y& Elsevier]

Highlights: [•] We show anisomycin (ANS) induces cell death in drug-resistant breast cancer cells. [•] We identify death-inducing ANS analogues. [•] Our data do not fit the current model for ANS-induced cell death. [•] We propose that a ribosome-ANS complex is a death signal or ANS has an unknown target. [Copyright &y& Elsevier]

Highlights: [•] S. cervi is widely used as a model to screen antifilarials. [•] Polyphenol rich EtOH extract of A. indica leaves were optimized by RSM. [•] EEA was tested against mf and adults of S. cervi cultured in vitro. [•] Morphological, molecular and biochemical alterations were examined. [•] Apoptosis and ROS up regulation are the probable cause of effectiveness. [ABSTRACT FROM AUTHOR]

In order to evaluate the effects of meiotic stage on survival of vitrified goat oocytes, the Cryoloop method was used to cryopreserve goat immature germinal vesicle (GV) and meiosis II (MII) oocytes following in vitro maturation (IVM). The oocytes collected from local slaughter house were divided into two parts. In the first part, the immature cumulus oocyte complexes (COCs) were further divided into three groups: (1) untreated (control), (2) exposed to the vitrification and dilution solutions but without being plunged into liquid nitrogen (toxicity), or (3) vitrified by the Cryoloop method (vitrification). In the second part, the MII oocytes produced by IVM of immature COCs were also divided into three groups as the above immature COCs. Oocytes survival was assessed by morphological appearance, nuclear maturation, and developmental capability following parthenogenetic activation (PA). The experiment was repeated three times. Our data indicated the rates of oocytes with normal appearance and developmental capability in the toxicity or vitrification group were decreased as compared to the control oocytes. In the toxicity group, the rate of GV oocytes with normal morphology was 76.25%±3.37% and significantly less than that of MII oocytes (92.18%±1.94%, P <0.05). However, the cleavage rate of MII oocytes was not significantly different from that of GV oocytes in the toxicity group (68.23%±1.71% vs 67.59%±3.51%, P >0.05). Additionally, the percentage of GV oocytes developing to blastocyst was significantly less than that of MII oocytes in the toxicity group (13.84%±2.81% vs 29.78%±4.17%, P <0.05). The rate of vitrified/thawed GV oocytes with normal morphology was 60.37%±2.91% and significantly less than that of MII oocytes (82.91%±3.01%, P <0.05). Additionally, the cleavage rate of GV oocytes was also significantly less than that of MII oocytes in the vitrification group (42.81%±2.94% vs 57.91±1.06%, P <0.05). However, the blastocyst rate of vitrified/thawed GV oocytes was not significantly different from that of MII oocytes (8.51%±1.46% vs 12.41%±3.74%, P >0.05). In conclusion, although vitrification can greatly damage the structure and development capability of goat oocytes, the Cryoloop method can result in acceptable levels of survival and development of goat oocytes. Additionally, compared to immature GV goat oocytes, mature MII oocytes may be more tolerant to the vitrification process. [ABSTRACT FROM AUTHOR]

Abstract: Early detection of drug-induced cholestasis remains a challenge during drug development. We have developed and validated a biorelevant sandwich-cultured hepatocytes- (SCH) based model that can identify compounds causing cholestasis by altering bile acid disposition. Human and rat SCH were exposed (24–48h) to known cholestatic and/or hepatotoxic compounds, in the presence or in the absence of a concentrated mixture of bile acids (BAs). Urea assay was used to assess (compromised) hepatocyte functionality at the end of the incubations. The cholestatic potential of the compounds was expressed by calculating a drug-induced cholestasis index (DICI), reflecting the relative residual urea formation by hepatocytes co-incubated with BAs and test compound as compared to hepatocytes treated with test compound alone. Compounds with clinical reports of cholestasis, including cyclosporin A, troglitazone, chlorpromazine, bosentan, ticlopidine, ritonavir, and midecamycin showed enhanced toxicity in the presence of BAs (DICI≤0.8) for at least one of the tested concentrations. In contrast, the in vitro toxicity of compounds causing hepatotoxicity by other mechanisms (including diclofenac, valproic acid, amiodarone and acetaminophen), remained unchanged in the presence of BAs. A safety margin (SM) for drug-induced cholestasis was calculated as the ratio of lowest in vitro concentration for which was DICI≤0.8, to the reported mean peak therapeutic plasma concentration. SM values obtained in human SCH correlated well with reported % incidence of clinical drug-induced cholestasis, while no correlation was observed in rat SCH. This in vitro model enables early identification of drug candidates causing cholestasis by disturbed BA handling. [Copyright &y& Elsevier]

Abstract: Ureaplasma spp. are members of the family Mycoplasmataceae and have been considered to be associated with chorioamnionitis and preterm delivery. However, it is unclear whether Ureaplasma spp. have virulence factors related to these manifestations. The purpose of the present study was to determine whether the immunogenic protein multiple-banded antigen (MBA) from Ureaplasma parvum is a virulence factor for preterm delivery. We partially purified MBA from a type strain and clinical isolates of U. parvum, and also synthesized a diacylated lipopeptide derived from U. parvum, UPM-1. Using luciferase assays, both MBA-rich fraction MRF and UPM-1 activated the NF-κB pathway via TLR2. UPM-1 upregulated IL-1β, IL-6, IL-12p35, TNF-α, MIP2, LIX, and iNOS in mouse peritoneal macrophage. MRF or UPM-1 was injected into uteri on day 15 of gestation on pregnant C3H/HeN mice. The intrauterine MRF injection group had a significantly higher incidence of intrauterine fetal death (IUFD; 38.5%) than the control group (14.0%). Interestingly, intrauterine injection of UPM-1 caused preterm deliveries at high concentration (80.0%). In contrast, a low concentration of UPM-1 induced a significantly higher rate of fetal deaths (55.2%) than the control group (14.0%). The placentas of the UPM-1 injection group showed neutrophil infiltration and increased iNOS protein expression. Our data indicate that MBA from the clinical isolate of U. parvum is a potential virulence factor for IUFD and preterm delivery in mice and that the N-terminal diacylated lipopeptide is essential for the initiation of inflammation. [Copyright &y& Elsevier]

Abstract: Introduction : Adverse drug reactions are a major cause for failures of drug development programs, drug withdrawals and use restrictions. Early hazard identification and diligent risk avoidance strategies are therefore essential. For drug-induced liver injury (DILI), this is difficult using conventional safety testing. To reduce the risk for DILI, drug candidates with a high risk need to be identified and deselected. And, to produce drug candidates without that risk associated, risk factors need to be assessed early during drug discovery, such that lead series can be optimized on safety parameters. This requires methods that allow for medium-to-high throughput compound profiling and that generate quantitative results suitable to establish structure–activity-relationships during lead optimization programs. Methods : We present the validation of such a method, a novel high content screening assay based on six parameters (nuclei counts, nuclear area, plasma membrane integrity, lysosomal activity, mitochondrial membrane potential (MMP), and mitochondrial area) using ~100 drugs of which the clinical hepatotoxicity profile is known. Results discussion : We find that a 100-fold TI between the lowest toxic concentration and the therapeutic Cmax is optimal to classify compounds as hepatotoxic or non-hepatotoxic, based on the individual parameters. Most parameters have ~50% sensitivity and ~90% specificity. Drugs hitting ≥2 parameters at a concentration below 100-fold their Cmax are typically hepatotoxic, whereas non-hepatotoxic drugs typically hit <2 parameters within that 100-fold TI. In a zone classification model, based on nuclei count, MMP and human Cmax, we identified an area without a single false positive, while maintaining 45% sensitivity. Hierarchical clustering using the multi-parametric dataset roughly separates toxic from non-toxic compounds. We employ the assay in discovery projects to prioritize novel compound series during hit-to-lead, to steer away from a DILI risk during lead optimization, for risk assessment towards candidate selection and to provide guidance of safe human exposure levels. [Copyright &y& Elsevier]

Highlights: [•] We synthesized carboxylated and PEGylated bifunctionalized InP/ZnS QDs (QInP). [•] Cell-penetrating peptides (CPPs) could deliver QInP into human A549 cells. [•] Five hundred nM of CPP/QInP and QInP at or below 1μM did not reduce cell viability. [•] QInP are biocompatible nanoparticles for use in biomedical applications. [ABSTRACT FROM AUTHOR]

Abstract: Changes in mitochondrial ATP synthesis can affect the function of tumor cells due to the dependence of the first step of glycolysis on mitochondrial ATP. The oxidative phosphorylation (OXPHOS) system is responsible for the synthesis of approximately 90% of the ATP in normal cells and up to 50% in most glycolytic cancers; therefore, inhibition of the electron transport chain (ETC) emerges as an attractive therapeutic target. We studied the effect of a lipophilic isoprenylated catechol, 3-hydroxybakuchiol (3-OHbk), a putative ETC inhibitor isolated from Psoralea glandulosa. 3-OHbk exerted cytotoxic and anti-proliferative effects on the TA3/Ha mouse mammary adenocarcinoma cell line and induced a decrease in the mitochondrial transmembrane potential, the activation of caspase-3, the opening of the mitochondrial permeability transport pore (MPTP) and nuclear DNA fragmentation. Additionally, 3-OHbk inhibited oxygen consumption, an effect that was completely reversed by succinate (an electron donor for Complex II) and duroquinol (electron donor for Complex III), suggesting that 3-OHbk disrupted the electron flow at the level of Complex I. The inhibition of OXPHOS did not increase the level of reactive oxygen species (ROS) but caused a large decrease in the intracellular ATP level. ETC inhibitors have been shown to induce cell death through necrosis and apoptosis by increasing ROS generation. Nevertheless, we demonstrated that 3-OHbk inhibited the ETC and induced apoptosis through an interaction with Complex I. By delivering electrons directly to Complex III with duroquinol, cell death was almost completely abrogated. These results suggest that 3-OHbk has antitumor activity resulting from interactions with the ETC, a system that is already deficient in cancer cells. [Copyright &y& Elsevier]

Highlights: [•] Our aim was to evaluate the effects of avermectin on pigeon neurons in vitro. [•] Morphologic, biochemical and molecular hallmarks of apoptosis was observed. [•] This is first report from apoptosis of avermectin on bird neurons in vitro. [Copyright &y& Elsevier]

Abstract: Since the inflammatory response and oxidative stress are involved in the stroke cascade, we evaluated here the effects of Phycocyanobilin (PCB, the C-Phycocyanin linked tetrapyrrole) on PC12 cell survival, the gene expression and the oxidative status of hypoperfused rat brain. After the permanent bilateral common carotid arteries occlusion (BCCAo), the animals were treated with saline or PCB, taking samples 24h post-surgery. Global gene expression was analyzed with GeneChip Rat Gene ST 1.1 from Affymetrix; the expression of particular genes was assessed by the Fast SYBR Green RT-PCR Master Mix and Bioplex methods; and redox markers (MDA, PP, CAT, SOD) were evaluated spectrophotometrically. The PCB treatment prevented the H2O2 and glutamate induced PC12 cell injury assessed by the MTT assay, and modulated 190 genes (93 up- and 97 down-regulated) associated to several immunological and inflammatory processes in BCCAo rats. Furthermore, PCB positively modulated 19 genes mostly related to a detrimental pro-inflammatory environment and counteracted the oxidative imbalance in the treated BCCAo animals. Our results support the view of an effective influence of PCB on major inflammatory mediators in acute cerebral hypoperfusion. These results suggest that PCB has a potential to be a treatment for ischemic stroke for which further studies are needed. [Copyright &y& Elsevier]

Abstract: Background: Peroxisome proliferator-activated receptor gamma (PPARγ) agonists are clinically used to counteract hyperglycemia. However, so far experienced unwanted side effects, such as weight gain, promote the search for new PPARγ activators. Methods: We used a combination of in silico, in vitro, cell-based and in vivo models to identify and validate natural products as promising leads for partial novel PPARγ agonists. Results: The natural product honokiol from the traditional Chinese herbal drug Magnolia bark was in silico predicted to bind into the PPARγ ligand binding pocket as dimer. Honokiol indeed directly bound to purified PPARγ ligand-binding domain (LBD) and acted as partial agonist in a PPARγ-mediated luciferase reporter assay. Honokiol was then directly compared to the clinically used full agonist pioglitazone with regard to stimulation of glucose uptake in adipocytes as well as adipogenic differentiation in 3T3-L1 pre-adipocytes and mouse embryonic fibroblasts. While honokiol stimulated basal glucose uptake to a similar extent as pioglitazone, it did not induce adipogenesis in contrast to pioglitazone. In diabetic KKAy mice oral application of honokiol prevented hyperglycemia and suppressed weight gain. Conclusion: We identified honokiol as a partial non-adipogenic PPARγ agonist in vitro which prevented hyperglycemia and weight gain in vivo. General significance: This observed activity profile suggests honokiol as promising new pharmaceutical lead or dietary supplement to combat metabolic disease, and provides a molecular explanation for the use of Magnolia in traditional medicine. [Copyright &y& Elsevier]

Highlights: [•] Aβ fibrillar seed can interact with monomer Aβ at the stoichiometry of 1:2. [•] Aβ fibrillar seed can accelerate fibrils formation and reduce higher-molecular-weight oligomer level and its lifespan. [•] Aβ fibrillar seeds are found to be able to rescue neuronal cells from Aβ cytotoxicity. [ABSTRACT FROM AUTHOR]

Abstract: It has been reported that non-subtype-selective GABAA receptor antagonists injected into the nucleus pontis oralis (PnO) of rats induced long-lasting increases in REM sleep. Characteristics of these REM sleep increases were identical to those resulting from injection of muscarinic cholinergic agonists. Both actions were blocked by the muscarinic antagonist, atropine. Microdialysis of GABAA receptor antagonists into the PnO resulted in increased acetylcholine levels. These findings were consistent with GABAA receptor antagonists disinhibiting acetylcholine release in the PnO to result in an acetylcholine-mediated REM sleep induction. Direct evidence has been lacking for localization in the PnO of the specific GABAA receptor-subtypes mediating the REM sleep effects. Here, we demonstrated a dose-related, long-lasting increase in REM sleep following injection (60nl) in the PnO of the inverse benzodiazepine agonist, methyl-6,7-dimethoxy-4-ethyl-β-carboline (DMCM, 10−2 M). REM sleep increases were greater and more consistently produced than with the non-selective antagonist gabazine, and both were blocked by atropine. Fluorescence immunohistochemistry and laser scanning confocal microscopy, colocalized in PnO vesicular acetylcholine transporter, a presynaptic marker of cholinergic boutons, with the γ2 subunit of the GABAA receptor. These data provide support for the direct action of GABA on mechanisms of acetylcholine release in the PnO. The presence of the γ2 subunit at this locus and the REM sleep induction by DMCM are consistent with binding of benzodiazepines by a GABAA receptor-subtype in control of REM sleep. [Copyright &y& Elsevier]

Abstract: Here, we have discovered CXI-benzo-84 as a potential anticancer agent from a library of benzimidazole derivatives using cell based screening strategy. CXI-benzo-84 inhibited cell cycle progression in metaphase stage of mitosis and accumulated spindle assembly checkpoint proteins Mad2 and BubR1 on kinetochores, which subsequently activated apoptotic cell death in cancer cells. CXI-benzo-84 depolymerized both interphase and mitotic microtubules, perturbed EB1 binding to microtubules and inhibited the assembly and GTPase activity of tubulin in vitro. CXI-benzo-84 bound to tubulin at a single binding site with a dissociation constant of 1.2±0.2μM. Competition experiments and molecular docking suggested that CXI-benzo-84 binds to tubulin at the colchicine-site. Further, computational analysis provided a significant insight on the binding site of CXI-benzo-84 on tubulin. In addition to its potential use in cancer chemotherapy, CXI-benzo-84 may also be useful to screen colchicine-site agents and to understand the colchicine binding site on tubulin. [Copyright &y& Elsevier]

Highlights: [•] Piperlongumine (PL) selectively kills GBM cells but not normal astrocytes. [•] PL kills GBM cells via ROS accumulation. [•] PL activates JNK and p38 differentially in GBM cells via ROS accumulation. [•] JNK and p38 activation contributes to PL’s cytotoxicity in GBM cells. [Copyright &y& Elsevier]

Abstract: Cancer, like other diseases accompanied by metabolic changes, shows characteristic DNA/RNA modifications and activities of modifying enzymes, resulting in fluctuations in nucleoside levels. In this study, we undertook targeted metabolomic analyses of nucleotides in different cancer cell culture models using a sensitive and reproducible ion-pair HPLC method. The experimental data were analyzed by principal component analysis (PCA) to identify potential biomarkers in cancer cells, and statistical significance was determined by one-way analysis of variance. As a result, a clear differentiation of normal and tumor cells into two clusters was shown, indicating abnormal metabolism of nucleotides in tumor cells. Six variables (AMP, UDP, CTP levels with a significance of P<0.05; ATP, UTP and GMP levels with a significance of P<0.01) were considered as potential biomarkers; the content of AMP, UTP, GMP and ATP was significantly higher in cancer cells. The receiver operating characteristic (ROC) curve analysis allowed us to discriminate normal cells from tumor cells based on area under the curve (AUC). The sequence of their AUC values were: ATP (0.979)>UTP (0.938)>CTP=GMP (0.896)>AMP (0.812)>UDP (0.792), so we conclude that ATP and UTP are the best potential biomarkers in tumor cells. This study may provide a valuable tool for studying minute alterations of intracellular nucleotide pools induced by anticancer/antiviral drugs, diseases or environmental factors. [Copyright &y& Elsevier]

Highlights: [•] p,p′-Methoxyl-diphenyl diselenide [(MeOPhSe)2] protected against amyloid-β induced cytotoxicity. [•] (MeOPhSe)2 prevented Aβ-induced cell death by inhibiting apoptosis and downregulating JNK phosphorylation. [•] (MeOPhSe)2 treatment attenuated Aβ-induced memory deficits in mice. [ABSTRACT FROM AUTHOR]

Peptide-based hydrogel nanoparticles represent a promising alternative to current drug delivery approaches. We have previously demonstrated that the Fmoc-FF aromatic dipeptide building block can self-assemble in aqueous solutions to form nano-scaled ordered hydrogels of remarkable mechanical rigidity. Here, we present a scalable process for the assembly of this peptide into hydrogel nanoparticles (HNPs) aimed to be utilized as potential drug delivery carriers. Fmoc-FF based HNPs were formulated via modified inverse-emulsion method using vitamin E-TPGS as an emulsion stabilizer and high speed homogenization. The formed HNPs exhibited two distinguishable populations with an average size of 21.5±1.3 and 225.9±0.8nm. Gold nanoparticles were encapsulated within the hydrogel nanoparticles as contrast agents to monitor the formation of the assemblies and their ultrastructural properties. Next, we demonstrated a robust experimental procedure developed and optimized for the formulation, purification, storage and handling procedures of HNPs. Encapsulation of doxorubicin (Dox) and 5-flourouracil (5-Fu) within the HNPs matrix showed release kinetics of the drugs depending on their chemical structure, molecular weight and hydrophobicity. The results clearly indicate that Fmoc-FF based hydrogel nanoparticles have the potential to be used as encapsulation and delivery system of various drugs and bioactive molecules. [ABSTRACT FROM AUTHOR]

Highlights: [•] Inhibiting HIV-1 integrase (IN) DNA binding is a way to develop new anti-HIV drugs. [•] The first high-throughput assay is developed to screen IN DNA binding inhibitors. [•] The economical assay shows good agreements with results obtained by other assays. [•] A new IN binding inhibitor nigranoic acid was identified using this assay. [•] Provide a proof-of-concept of using the strategy to develop new drugs against HIV. [Copyright &y& Elsevier]

Abstract: Deoxynivalenol (DON) is a mycotoxin produced by Fusarium sp., and is known to elicit pro-inflammatory responses in the cell. The cells exposed to DON undergo apoptosis as a mechanism to shut down the inflammation. In this study, we tested the cytoprotective effect of the green tea polyphenol epigallocatechin 3-gallate (EGCG) on DON-induced toxicity in HT-29 cells. EGCG prevented DON-induced cytotoxicity to HT-29 cells in a dose–response manner. Even the lowest concentration (5μM) of EGCG showed protection against the highest concentration of DON tried (3.38μM=1000ng/ml). Our study also demonstrates that IC20 value of DON in HT 29 cells were 250ng/ml and pre-treatment with 20μM EGCG yielded 99% cell viability. EGCG also protected against oxidative stress, up regulation of nuclear factor-kB (NF-κB), cyclooxygenase-2 (COX-2) and caspase-3 activated apoptosis. These results suggest that EGCG acts as cytoprotective agent against DON-induced toxicity. [Copyright &y& Elsevier]

Abstract: Reconstructed human epidermis (RHE) is used in non-animal testing for hazard analysis and reconstructed human skin (RHS) gains growing interest in preclinical drug development. RHE and RHS have been characterised regarding their barrier function, but knowledge about biotransformation capacity in these constructs and in human skin remains rather poor. However, metabolising enzymes can be highly relevant for the efficacy of topical dermatics as well as genotoxicity and sensitisation. We have compared the esteratic cleavage of the prednisolone diester prednicarbate and the enzyme kinetic parameters (V max and S 0.5) of the model substrate fluorescein diacetate (FDA) in commercially available RHS and RHE with excised human skin and monolayer cultures of normal and immortalised human keratinocytes and of fibroblasts. Formation of the main metabolite prednisolone and of fluorescein ranked as: RHS∼RHE>excised human skin and keratinocytes>fibroblasts, respectively. Because of the aromatic probe, however, V max of FDA cleavage did not show a linear relationship with prednicarbate metabolism. In conclusion, RHE and RHS may be useful to quantitatively address esterase activity of human skin in drug development and hazard analysis, although an increased activity compared to native human skin has to be taken into account. [Copyright &y& Elsevier]

Highlights: [•] Glycogen synthase kinase 3 (GSK-3) is an important target for the treatment of Alzheimer’s disease. [•] Reducing tau phosphorylation through GSK-3 inhibition has emerged as a target for drug development of AD. [•] We have discovered novel GSK-3 inhibitors (C-7a, -7b) and demonstrated pharmacological validation. [•] New GSK-3 inhibitors (C-7a, -7b) were interfered with Aβ oligomers-induced neuronal cell death and tau phosphorylation in vitro. [•] C-7a has efficacy in reducing the level of tau phosphorylation and improving short-term memory in 3xTg-AD mice. [ABSTRACT FROM AUTHOR]

Abstract: Perfluorinated compounds are ubiquitous chemicals of major concern for their potential adverse effects on the human population. We have used primary rat embryonic neural stem cells (NSCs) to study the effects of perfluorooctane sulfonate (PFOS) on the process of NSC spontaneous differentiation. Upon removal of basic fibroblast growth factor, NSCs were exposed to nanomolar concentrations of PFOS for 48h, and then allowed to differentiate for additional 5days. Exposure to 25 or 50nM concentration resulted in a lower number of proliferating cells and a higher number of neurite-bearing TuJ1-positive cells, indicating an increase in neuronal differentiation. Exposure to 50nM also significantly increased the number of CNPase-positive cells, pointing to facilitation of oligodendrocytic differentiation. PPAR genes have been shown to be involved in PFOS toxicity. By q-PCR we detected an upregulation of PPARγ with no changes in PPARα or PPARδ genes. One of the downstream targets of PPARs, the mitochondrial uncoupling protein 2 (UCP2) was also upregulated. The number of TuJ1- and CNPase-positive cells increased after exposure to PPARγ agonist rosiglitazone (RGZ, 3μM) and decreased after pre-incubation with the PPARγ antagonist GW9662 (5μM). RGZ also upregulated the expression of PPARγ and UCP2 genes. Meanwhile GW9662 abolished the UCP2 upregulation and decreased Ca2+ activity induced by PFOS. Interestingly, a significantly higher expression of PPARγ and UCP3 genes was also detected in mouse neonatal brain after prenatal exposure to PFOS. These data suggest that PPARγ plays a role in the alteration of spontaneous differentiation of NSCs induced by nanomolar concentrations of PFOS. [Copyright &y& Elsevier]