coli showed that an ompC knockout mutant had increased levels of OmpA [40], however, changes in permeability were not evaluated. Furthermore, this has not been evaluated in a S. Typhimurium or E. coli ∆ompW strain. Figure 2 Bacterial concentration of S . Typhimurium 14028s and Δ ompW exposed to H 2 O 2 or NaOCl. Cultures of 14028s and ΔompW were grown to OD ~ 0.4 and treated with H2O2 4 mM or NaOCl 5 CX-6258 mw mM in LB medium. Time of treatment is indicated. Bacterial concentrations were determined by plating. The values are the concentrations of surviving

SYN-117 mw bacteria after exposure to H2O2 or NaOCl. Experiments were performed in triplicate. Error bars indicate SD. Our data supports the proposed model where OmpW allows the influx of small polar molecules, like H2O2 and HOCl. The crystal structure of OmpW from E. coli mTOR activity revealed that the cross-section of the barrel has approximate dimensions of 17 × 12 Å along the length of the barrel and although the interior of the channel has a hydrophobic character, the observed single channel activities shows that polar molecules traverse the barrel [17]. Taken together, these

results provide biochemical and genetic evidence indicating that both toxic compounds are channeled through OmpW. From our knowledge, this is the first direct evidence of HOCl diffusion through porins. Furthermore, preliminary analyses indicate that H2O2 and HOCl channeling is common for S. Typhimurium OmpD, OmpC and OmpF porins (unpublished data). Hydrogen peroxide and hypochlorous acid exposure results in ompW negative regulation Since the OmpW porin channels H2O2 and HOCl through the OM and exposure to these molecules is detrimental to bacteria, we hypothesized that ompW should be negatively regulated when S. Typhimurium is exposed to H2O2 and HOCl. To study ADP ribosylation factor this effect, wild type S. Typhimurium cells were grown to mid-log

phase, exposed to H2O2 or HOCl and ompW mRNA levels were measured by qRT-PCR. As seen in Figure 3, exposure to H2O2 and HOCl resulted in lower levels of ompW transcripts (0.27 ± 0.04 and 0.156 ± 0.079, respectively) relative to control untreated cells. In agreement with our results of ompW negative regulation, similar results were observed by Wang et al. (2010) who showed that S. Enteritidis and Typhimurium cells exposed to HOCl results in modulation of ompD, ompC, ompF (negatively) and ompA (positively) expression. Furthermore, Calderón et al. (2011) demonstrated that the S. Typhimurium ompD gene is negatively regulated in response to H2O2. Therefore, our and all the published data suggest that in the presence of OCl- or H2O2 there might be a general lowering in the concentration of porins in the outer membrane, in order to diminish the permeability. To assess the specificity of our assay, we evaluated ompD, ompC and arcB transcript levels as positive (ompD and ompC) and negative controls (arcB).

However, we did find that high force production

improved with betaine supplementation which reflects some similarity to the study by Hoffman and see more coworkers. While the muscle groups in the two studies were apparently different in their mediating mechanisms, both studies provide evidence for the potential positive influence of B supplementation for strength, power and local muscular endurance in the context of demanding strength/power exercise protocols. In the present study, the larger lower-body muscle group data was more varied within the subject sample and significant differences were less obvious, although patterns of B mediated increases may be suggested. AZD1480 For example, isometric squat

force was enhanced by B supplementation. The REC protocol utilized maximal vertical jumps prior to the squat exercises which might have impaired the neuromuscular performance of high power production as recently noted by Drinkwater et al. [14], indicating that order of exercises is an important element in Akt inhibitor training program design. In this case, the betaine supplement was likely not able to offset the neural effect and partially explains the lack of improved power production in the squat. However, force production may have been facilitated via a post activation potentiation effect of some type [15]. While speculative, the upper body musculature was not inhibited by such an inhibitory neuromuscular influence of high velocity power movements as was the lower body in this exercise testing sequence. Thus, it Suplatast tosilate appears that the mediating mechanisms of betaine supplementation may be more operational in the absence of high frequency neural fatigue. From the non-significant differences in body fluid related variables between the B and P trials, due to the experimental controls for hydration employed in this study, it seems that betaine’s established role as an osmoprotectant

[2, 7, 8] was not a likely candidate for any ergogenicity. This does not, however, minimize the potential role of betaine given the intensity of the REC, as organic osmolytes have been shown to accumulate in cells under varying stressful conditions to help maintain biochemical function [16–18]. Additionally, plasma glucose and lactate results in this study indicate that betaine was either 1) not acting through glucose or lactate processing, or 2) the pre-existing differences among subjects masked any betaine effects on these dependent variables. The use of the very demanding REC might have overwhelmed the ability of betaine to offer any measureable differences, which in the case of the enhanced performances would most likely be related to phosphagen metabolism.

22-μm filter, and stored at −20°C until use. Bacterial strain and growth conditions P. gingivalis strain W83 (kindly supplied by Dr. Koji Nakayama, Nagasaki University Graduate School of Biomedical Sciences) was cultured at 37°C anaerobically (85% N2, 10% H2, and 5% CO2) in

half-strength brain heart AZD1390 concentration infusion www.selleckchem.com/products/blz945.html (BHI) broth (Becton Dickinson, Sparks, MD) supplemented with 0.5% yeast extract (Difco Laboratories, Detroit, MI), 5 μg/ml of hemin (Sigma), and 1 μg/ml of vitamin K1 (Sigma). RNA isolation and cDNA synthesis Use of high concentrations of antibacterial agents for extended periods of time changes the expression of a large set of genes and the effect may be secondary to the action of the drug [46]. Meanwhile, at sub-lethal concentrations, bacteria may sense antibiotics as extracellular chemicals to trigger different cellular responses such as an altered antibiotic resistance/tolerance profile [47]. Hence, we performed the full-genome gene expression microarrays of P. gingivalis W83 exposed to polyP75 at a concentration of 0.03%, which was previously determined to be MIC against the bacterium [16], for a short period of time. P. gingivalis culture grown to early exponential phase (OD600 = 0.3) was divided in half. One aliquot was left untreated, while the other one was treated with 0.03% polyP75. After incubation of both the bacterial cultures for 2 h under anaerobic

conditions, the bacterial cells were harvested, and total RNA was extracted from the cells using Trizol Reagent (Invitrogen, Carlsbad, CA). RNA quality was monitored by Agilent 2100 Bioanalyzer (Agilent Technologies, Selleck PARP inhibitor Santa Clara, CA), and RNA quantity was measured by spectrophotometer.

All the samples used in this study exhibited A260/A280 ratio of at least 1.8. cDNA was synthesized with 20 μg of total RNA using SuperScript® II Reverse Transcriptase (Invitrogen). Microarray analysis Two individual Cy3-labeled cDNA samples were hybridized into DNA microarrays (Nimblegen Systems, Inc., Madison, WI) containing the whole genome of 1,909 genes of aminophylline P. gingivalis W83 for 16 h at 42°C. Five replicates of the genome were included per chip. An average of 19 different 60-mer probes which had at least three mismatches compared to other 60-mers represented each gene in the genome. A quality control check (hybridization) was performed for each array, which contained on-chip control oligonucleotides. Data were extracted from the scanned images using an Axon GenePix 4000B microarray scanner and NimbleScan Version 2.3. Quantile normalization was performed across replicate arrays, and RMA (Robust Multichip Average) analysis was performed to generate gene expression values. Genes evidencing statistically significant changes in expression (>1.5-fold difference) were identified via t-tests (P < 0.05). Assessment of array data quality To confirm the microarray results using qRT-PCR, 10 genes were selected, and specific primers for the selected genes (Table 6) were designed using Primer3 (http://​fokker.

b GDC 0449 Number of pSfr64a ORFs in each category, with highest similarity to ORFs from pRet42d. c Number of pSfr64a ORFs in each category, with highest similarity to ORFs from pRet42a. d Number of pSfr64a ORFs in each category, with highest similarity VX-689 chemical structure to ORFs from the chromosome of NGR234. Among the ORFs shared between pSfr64a and pRet42a, the self-transmissible plasmid of CFN42, most are related to conjugative transfer (20 ORFs), only two were ascribed to macromolecular metabolism. Interestingly, both are related to DNA metabolism, one was classified as a putative nuclease, and the other as a probable DNA methylase. In Figure 3, it can be appreciated

that the genomic region shared between pRet42a and pSfr64a is markedly colinear. Colinearity is disrupted by the absence of an homolog to the regulatory gene cinR of pRet42a, and the presence of pSfr64a ORFs 147 and 148, which encode hypothetical proteins. The correspondence between pSfr64a and pRetCFN42 ORFs C59 wnt mouse is presented in Additional File 1. Figure 2 shows that the segment of pSfr64a shared with pRet42a has a high GC content, compared to the rest of the plasmid. This feature is also present in the similar pRet42a sequence. Figure 3 Colinearity between pSfr64a and other replicons. Dot matrix view of BLASTN comparisons of pSfr64a vs pRet42a, pRet42d and the chromosome

of NGR234. The ORFs similar to the pSym of CFN42 (pRet42d) include the repABC genes (Figure 2, Table 3). This is congruent with our finding that pSfr64a and pRet42d are incompatible (data not shown). The pSfr64a-pRet42d-shared ORFs are mainly involved in small Casein kinase 1 molecule metabolism (26 ORFs), and carbohydrate transport (13 ORFs). It is noteworthy that, in spite of the fact that pRet42d carries genes engaged in symbiotic functions, none

of these are present in pSfr64a. Within the region similar to pRet42d (ORFs 46 to 110), the colinearity is restricted to small segments, some of them in inverse orientation. (Figure 3, Additional file 1). The repABC genes (pSfr64a ORFs 164 to 166) were adjacent to the transfer region, separated from the other pRet42d genes. It has been amply documented that plasmid pRet42d is subject to frequent genomic rearrangements, due to the presence of reiterations and a high density of insertion sequences [16–20]. R. etli ORFs encoding transposon-related proteins located near to the sites where colinearity is disrupted are indicated in Figure 2 (purple arrows) and Additional File 1. For example, pSfr64a ORFs 122 to 146 are colinear with pRet42a ORFs 139 to 162. The adjacent ORF on pRet42a (ORF 138) encodes a transposon-related protein. It is possible that these sequences are related to the generation of rearrangements, causing the interruptions in colinearity. ORFs 114, 115, 116, 117, 118 and 121 show homology to ORFs encoded in another Rhizobium etli strain; IE4771 [21].

(C) upper panel depicts detection of gp340 in parotid saliva alone and after incubation with five different L. gasseri isolates and the L. gasseri type strain; (D) upper panel depicts detection of gp340 and lower panel detection of MUC7 in submandibular/sublingual saliva alone and after incubation with five different L. gasseri isolates and the type strain. Numbers below lanes in panels C and D refer to the following contents: (1) Saliva alone (+ve control), (2) Saliva after L. gasseri CCUG31451T incubation, (3) Saliva after L. gasseri isolate A241 incubation, (4) Saliva after L. gasseri

isolate A274 incubation, (5) Saliva after L. gasseri isolate B1 incubation, (6) Saliva after L. gasseri isolate NVP-HSP990 ic50 B16 incubation, (7) Saliva after L. gasseri isolate L10 incubation. MUC7 (mw ≈150 kDa) was detected using Western blot analysis with mAb LUM7-2 antibodies in submandibular saliva (Figure 4, lower panels A and B, lane 6, lower panel D lane 1) but not in parotid saliva (data not shown). MUC7 levels were reduced in submandibular saliva after incubation with L. gasseri (Figure 4, AZD9291 solubility dmso lower

panel A, lane 7) and S. NCT-501 in vitro mutans (Figure 4, lower panels B, lane 7). MUC7 was detected bound to L. gasseri (Figure 4, lower panel A, lane 8) and S. mutans (Figure 4, lower panel B, lane 8) after incubation with submandibular saliva. SDS treatment

released the MUC7 bound to L. gasseri (Figure 4, lower panel A, lane 9) and to S. mutans (Figure 4, lower panels B, lane 9). Similar results were observed for MUC7 binding to six additional isolates of L. gasseri (Figure 4D, lower panel). L. gasseri binds to human epithelial cells Adherence of FITC-tagged L. gasseri strains was detected by fluorescence microscopy as illustrated for strain A274 (Figure 5). All L gasseri strains were observed only adjacent to epithelial cells. Figure 5 Adhesion Clomifene of L. gasseri to human epithelial cells. Field of view containing differentiated human gingival epithelial cells (HGEP.05) and fluorescently stained L. gasseri A274 (in green). Bacteria were detected only in association with gingival epithelial cells. Images were captured using a Zeiss imager Z1 upright microscope. Bars in panels equal 20 μm. Discussion In this study lactobacilli were detected more frequently in breastfed than formula-fed 4 month-old infants in saliva and mucosal swab samples as we previously observed in a different population of infants [13]. L. gasseri was the dominant Lactobacillus species detected, which was identified from 16S RNA gene sequences of isolates. Probiotic potential of L. gasseri was found to include growth inhibition of F. nucleatum, A. naeslundii, A. oris, S. sobrinus and C.

To further verify the microarray data, we have used qRT-PCR to test expression of 17 genes with decreased expression

in one or both mutants (putative sporulation-induced genes). This overall expression pattern was confirmed for several genes, with eleven out of the 17 tested genes showing a significantly lower expression in the whiA mutant compared to the wildtype at at least one of the two sporulation time points 36 h and 48 h (Additional file 2: Figure S2). Thus, a large fraction of this group are developmentally regulated genes correctly identified by the array analysis. Further investigations of several of these genes are described in the Belinostat clinical trial following sections. For the genes that appeared overexpressed in the whiH mutant, i.e. that were putative candidates for being repressed by WhiH, six genes were tested by qRT-PCR. Five Epigenetics Compound Library appeared to be false positives and only one had its microarray expression profile confirmed by qRT-PCR experiments (Additional file 2: Figure S3). This is the Selleck Poziotinib previously described gene eshB (SCO5249) encoding a putative cyclic nucleotide-binding protein [29]. The qRT-PCR indicated higher eshB expression during development of the whiH mutant compared to the parent

strain. In an S1 nuclease protection assay (Additional file 2: Figure S4), the eshB promoter was found to be similarly up-regulated during development in both the parent and the whiH mutant, and the level of transcript was only 1.4-fold higher in the mutant at the 36 h time point and not different from wildtype at 48 h (after normalisation to the hrdB promoter as internal control). Also the eshB paralogoue eshA (SCO7699) [29] was significantly up-regulated L-NAME HCl in the whiH mutant according to the arrays (Additional file 2: Figure S3), but S1 nuclease protection assays showed that eshA is strongly up-regulated during developmental in both strains, with only subtle difference in mRNA level between the whiH mutant and the wild-type (Additional file 2: Figure S4). Overall, our analyses did not reveal any clear candidates for repression by the WhiH transcription factor. Analysis of expression

and mutant phenotypes of new sporulation genes We have specifically investigated seven potential sporulation loci emerging from the microarray analysis (Figure  4). Expression of these loci has been monitored using qRT-PCR (Figure  5), S1 nuclease mapping (Figure  6), and promoter fusions to a reporter gene encoding the fluorescent protein mCherry (Figure  7 and Table  1). For the latter experiments, we constructed a new vector, pKF210, used this to construct “promoter probe” fusions, and introduced them into Streptomyces strains (described in Materials and Methods). Furthermore, deletion mutants have been constructed for these seven loci and examined to detect phenotypes associated with sporulation and maturation of spores.

At lower pressures, a greater abundance of smaller nanoparticles has been observed in the sub-monolayer analysis in Figure 1, again increasing the absorption at higher energies; this is likely due to a decrease in redeposition of ablated material, and therefore, more nanoparticles are deposited on the substrate in lower pressures over time. Figure 4 Relationship between absorption coefficient and laser fluence and background gas pressure used during deposition. Decreasing trend in absorption coefficient with respect to increasing gas pressure and decreasing laser fluence. Conclusion To conclude, femtosecond pulsed laser deposition

has been used to fabricate solid state nanoparticulate silicon thin films on a fused silica substrate. Fabrication parameters have been studied in order to form high-quality thin films with a continuous film profile and a smooth surface, FG-4592 order ideal for optical and optoelectronic applications. The inclusion of hydrogen in a background gas of argon and the heating of the substrate during deposition have both been shown to dramatically improve the as-deposited film quality. To further this work, it would be appropriate to carry out a quantitative assessment of how properties such as the emission characteristics from a doped lanthanide or the electrical conductivity would vary depending

on the fabrication processes described above. Vorinostat in vitro The conclusions drawn here are also not limited to the fabrication of silicon thin films but can be utilised for better refining the deposition process of different materials. Acknowledgements The authors would like to thank the EPSRC for funding on this research as well as Adam S. HTS assay Qaisar Janus kinase (JAK) for the assistance with Latex. References 1. Kim MK, Takao T, Oki Y, Maeda M: Thin-layer ablation of metals and silicon by femtosecond laser pulses for application to surface analysis. Japanese J Appl Phys 2000,39(11):6277–6280. [http://​jjap.​ipap.​jp/​link?​JJAP/​39/​6277/​]CrossRef 2. Perrière J, Boulmer-Leborgne C, Benzerga R, Tricot S: Nanoparticle formation by femtosecond laser ablation. J Phys D Appl

Phys 2007,40(22):7069–7076. [http://​stacks.​iop.​org/​0022-3727/​40/​i=​22/​a=​031?​key=​crossref.​3dbee54d3aabd962​39b697e75c5e1261​]CrossRef 3. Linde D, Sokolowski-Tinten K, Von Der: Laser-solid interaction in the femtosecond time regime. Appl Surf Sci 1997, 1–10. [http://​linkinghub.​elsevier.​com/​retrieve/​pii/​S016943329600611​3] 4. Cavalleri A, Sokolowski-Tinten K, Bialkowski J, Schreiner M, von der Linde D: Femtosecond melting and ablation of semiconductors studied with time of flight mass spectroscopy. J Appl Phys 1999,85(6):3301. [http://​link.​aip.​org/​link/​JAPIAU/​v85/​i6/​p3301/​s1&​Agg=​doi]CrossRef 5. Sundaram SK, Mazur E: Inducing and probing non-thermal transitions in semiconductors using femtosecond laser pulses. Nat Mater 2002,1(4):217–224.

During the rotational GLAD process, the lateral component of deposition flux with respect to the surface Poziotinib purchase normal of the substrate contributes to the formation of columnar structures due to the shadowing effect, while the rotation of the substrate eliminates the preferred orientation growth, thus controls the shape of the structures. In the past few decades, there is considerable effort of both experimental investigation and atomistic simulations taken to investigate the fundamental mechanisms of the rotational GLAD [7–11]. Since nucleated islands acting as shadowing centers are essentially required for the formation of columnar structures in the initial period of the rotational

GLAD, recently placing nano-sized templates on the bare substrate is proposed to replace the nucleated AZD3965 islands, in such a way both deposition period and deposition

flux can be reduced significantly. Most importantly, by designing the geometry and the alignment of the templates, ordered arrays of columnar structures with pre-designed https://www.selleckchem.com/products/ulixertinib-bvd-523-vrt752271.html shapes can be fabricated under the intensified shadowing effect [12, 13]. Although the template-assisted rotational GLAD has been demonstrated to be one promising nanostructuring technique for the fabrication of 1D nanostructures, our fundamental understanding of the deposition process, particularly the deposition-induced deformation of the templates, is still limited: will the templates deform during the deposition? If yes, what are the underlying

deformation mechanisms of the templates? And how does the deformation behavior of the templates influence the geometry of the fabricated columnar structures? In this letter, we address the above questions by performing three-dimensional molecular dynamics (MD) simulations of the template-assisted Phosphoprotein phosphatase rotational GLAD of 1D Al columnar structures on Cu substrate. Our simulations demonstrate that the presence of templates significantly intensifies the shadowing effect to form 1D columnar structures when deposition flux is small, as compared to the template-free rotational GLAD. Furthermore, the morphology of the fabricated columnar structures by the template-assisted rotational GLAD strongly depends on the deformation behaviors of the templates. Methods Figure 1a illustrates the MD model of the template-assisted rotational GLAD utilized in the present work. The Cu substrate has a dimension of 11.6, 11.6, and 0.7 nm in X, Y, and Z directions, respectively. Periodic boundary condition (PBC) is imposed in the transverse X and Y directions of the substrate to simulate an infinitely wide thin film. There are nine equally spaced Cu templates of square cylinder placed on the substrate. The lattice constant a for Cu is 0.3615 nm. The width d for each template is 6a, and the distance s between each template is 10a. To investigate the influence of the template height h on the deposition process, two height values of 8a and 14a are considered.

Our analysis indicates that the risk for cardiac events is increased in patients

with these contraindications. Indeed, in the case–control analysis of hospitalisation with MI, 12 % of the cases and 4 % of the controls had had a history of previous hospitalisation with MI before index date. Similar elevated risks were found for history of ischaemic heart disease (71 % in cases versus 24 % in controls), peripheral BIBW2992 artery disease (18 % in cases versus 7 % in controls), and cerebrovascular disease (23 % in cases versus 15 % in controls). In line with this, exclusion of patients with the contraindications from the pooled analyses of the randomised-controlled trials with strontium ranelate completely mitigated the risk for MI (data on file). The new contraindications for strontium ranelate are therefore expected to reduce any potential cardiovascular

risk associated with use of this treatment. Conclusion The results of this nested case–control study in the CPRD indicate no evidence for a higher risk of MI or cardiovascular death associated with the use of strontium ranelate in women treated for osteoporosis compared with non-use of this agent in routine medical practice in the UK. Acknowledgments The interpretation and conclusions contained in this report are those of the authors alone. This BMS202 study was funded by Servier. Study data were obtained from the CPRD under license from the UK MHRA to the Acceptability Data and Pharmacoepidemiology Department of Servier. The authors would like to thank Karine Marinier and Nicolas Deltour (Servier) for help with study design and conduct and statistical analysis. KF is an NIHR Senior Investigator supported by the NIHR Cardiovascular Biomedical Research Unit at the Royal Brompton Hospital. Conflicts of interest All authors have disclosed receiving fees, honoraria, and research grants from Servier.

References 1. Meunier PJ, Roux C, Seeman E et al (2004) The effects Resminostat of strontium ranelate on the risk of vertebral fracture in women with postmenopausal osteoporosis. N Engl J Med 350:459–468PubMedCrossRef 2. Reginster J-Y, Felsenberg D, Boonen S et al (2008) Effects of long-term strontium ranelate https://www.selleckchem.com/products/bay-11-7082-bay-11-7821.html treatment on the risk of nonvertebral and vertebral fractures in postmenopausal osteoporosis: results of a five-year, randomized, placebo-controlled trial. Arthritis Rheum 58:1687–1695PubMedCrossRef 3. Kaufman JM, Audran M, Bianchi G et al (2013) Efficacy and safety of strontium ranelate in the treatment of osteoporosis in men. J Clin Endocrinol Metab 98:592–601PubMedCrossRef 4. Reginster JY, Badurski J, Bellamy N et al (2013) Efficacy and safety of strontium ranelate in the treatment of knee osteoarthritis: results of a double-blind, randomised placebo-controlled trial.

The HMCs were plated onto gelatine-coated glass coverslips in 24-well plates (105 cells/well) and infected at a 10:1 parasite:host cell ratio after 24 h. Afterwards LY294002 concentration the cultures were washed, and the NQs (0.5

to 20 μM) were added. At specified intervals, the cultures were fixed in Bouin’s solution, stained with Giemsa and counted to assess the following parameters: percentage of cells infected, number of parasites/infected cell and the endocytic index (EI), which refers to the number of parasites/100 cells [52]. The IC50 values for the different days of treatment, corresponding to the concentration that led to 50% inhibition of each parameter, were calculated. To determine the possible toxic selleck products effects of the compounds on the host cells, uninfected macrophages and HMCs were incubated at 37°C with the NQs. After 2 days, the viability of the cells was measured using the MTT colorimetric assay [53]. The absorbance was measured at 490 nm with a spectrophotometer (VERSAmax Tunable, Molecular Devices, USA), allowing for the determination of an LC50 value, which is the concentration that reduces cellular viability by 50%. Transmission and scanning electron microscopy analysis Epimastigotes (5×106 cells/mL)

were treated for 24 h with the selected NQs at their respective IC50/24 h values in LIT medium at 28°C. Afterward, they were fixed with 2.5% glutaraldehyde in 0.1 M Na-cacodylate buffer (pH 7.2) for 40 min at 25°C and post-fixed with 1% OsO4, 0.8% potassium ferricyanide and 2.5 mM CaCl2 in the same buffer for 20 min at 25°C. The cells were dehydrated in an ascending acetone series and embedded in PolyBed 812 resin. Ultrathin sections were stained with uranyl acetate and lead citrate and examined in a Jeol JEM1011

transmission electron AZD1152 in vivo microscope (Tokyo, Japan). Alternatively, dehydrated samples were dried by the critical point method with CO2, mounted on aluminum stubs, coated with a 20 nm thick gold layer and examined on a Jeol JSM6390LV scanning electron microscope (Tokyo, Japan). Both electron microscopes Chorioepithelioma are located in Plataforma de Microscopia Eletrônica at Instituto Oswaldo Cruz (Fiocruz). Flow cytometry analysis Epimastigotes were treated for 24 h with the NQs at concentrations up to their IC50 values. We then determined the mitochondrial membrane potential (ΔΨm) and reactive oxygen species (ROS) production. For ΔΨm analysis, the parasites were incubated with 50 nM tetramethylrhodamine (TMRE) (Molecular Probes, Carlsbad, USA) for 15 min at 28°C, using 10 μM carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone (FCCP) (Sigma-Aldrich Chemical Co.) as a control for ΔΨm dissipation. Alterations in TMRE fluorescence were quantified using an index of variation (IV), which was calculated using the equation (MT – MC)/MC, where MT is the median of fluorescence for treated parasites and MC is the median of fluorescence of the control parasites.