Search

Bisphenol A (BPA) is one of the world's most widely used plasticizer, and its hazardous impacts have been well studied. However, few studies focused on the effects of parental long-term BPA exposure on the bone development of offspring. In the present study, the bone development of offspring was studied following long-term exposure of parental zebrafish to environmentally relevant 15 and 225 µg/L BPA. The results showed that BPA increased the mortality and deformity rate of offspring and caused craniofacial deformities characterized by changes in various cartilage angles and lengths. The alizarin red and calcein staining showed that BPA could delay bone mineralization and reduce bone mass accumulation. The results of acridine orange staining indicated that BPA induced apoptosis of the skull. The degree of harm of BPA presented a dose-dependent pattern. The results of the comparative transcriptome showed that there were 380 different expression genes (DEGs) in the 15 µg/L BPA group, and 645 DEGs in the 225 µg/L BPA group. MAPK/Wnt/FoxO signaling pathway-related genes were significantly down-regulated in the BPA-exposed groups. The present study demonstrates that long-term parental BPA exposure would severely affect cartilage development and bone mineralization of fish offspring, and MAPK/Wnt/FoxO signaling pathways may be involved in this process., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022. Published by Elsevier Inc.)

Organotin compounds, being biologically active, affect a variety of cellular functions due to their ability to accumulate in and penetrate biological membranes. These compounds influence the distribution of electrostatic charges, alter organization, disrupt molecular dynamics and change mechanical properties of biological membranes. It was found that the membrane/water partition coefficient equals 4, a value significantly higher than octanol/water partition coefficient. In addition, the effect of di- and tri-phenyltin chlorides on the mechanics of model lipid membranes was measured for the first time. It has been determined that phenyltins affect the global model lipid bilayer properties by reducing the membrane expansion modulus, when measured using micromanipulation technique, and elevating the bending rigidity coefficient of the lipid bilayer, as determined with the flickering noise spectroscopy. In addition, the elevated water permeability shows that phenyltins also cause the local defects formation in the lipid bilayer, i.e. lipid pores. These data shows that phenyltins may interfere indirectly with variety cellular processes by altering non-specifically the entire cellular membrane system. Accordingly, when phenyltins are added to macrophages in culture, they inflict massive alterations of cell morphology and interfere with membrane-associated processes, as visualized using fluorescence labelling of selected subcellular compartments., (Copyright © 2017 Elsevier B.V. All rights reserved.)

The aim of this study was to evaluate the anticancer activity of bioinspired silver nanoparticles (AgNPs) and gold nanoparticles (AuNPs) against mouse myoblast cancer cells (C 2 C 12 ). Both AgNPs and AuNPs were biologically synthesized using Spinacia oleracea Linn., aqueous leaves extract. UV-Vis. spectrophotometer, high resolution-transmission electron microscopy (HR-TEM), field emission-scanning electron microscopy (FE-SEM) and X-ray diffraction (XRD) studies supported the successful synthesis of AgNPs and AuNPs. Both these NPs have shown cytotoxicity against C 2 C 12 cells even at very low concentration (5μg/mL). Acridine orange/Ethidium bromide (AO/EB) dual staining confirmed the apoptotic morphological features. The levels of caspase enzymes (caspase-3 and caspase-7) were significantly up-regulated in NPs treated myoblast cells than the plant extract. Furthermore, in zebrafish embryo toxicity study, AgNPs showed 100% mortality at 3μg/mL concentration while AuNPs exhibited the same at much higher concentration (300mg/mL). Taken together, these results provide a preliminary guidance for the development of biomaterials based drugs to fight against the fatal diseases for example cancer., (Copyright © 2016 Elsevier B.V. All rights reserved.)

The surface properties of biomaterials, such as wettability, polar group distribution, and topography, play important roles in the behavior of cell adhesion and proliferation. Gaseous plasma discharges are among the most common means to modify the surface of a polymer without affecting its properties. Herein, we describe the surface modification of poly(styrene) (PS) and poly(methyl methacrylate) (PMMA) films using atmospheric pressure plasma processing through exposure to a dielectric barrier discharge (DBD). After treatment the film surface showed significant changes from hydrophobic to hydrophilic as the water contact angle decreasing from 95° to 37°. All plasma-treated films developed more hydrophilic surfaces compared to untreated films, although the reasons for the change in the surface properties of PS and PMMA differed, that is, the PS showed chemical changes and in the case of PMMA they were topographical. Excellent adhesion and cell proliferation were observed in all films. In vitro studies employing flow cytometry showed that the proliferation of L929 cells was higher in the film formed by a 1:1 mixture of PS/PMMA, which is consistent with the results of a previous study. These findings suggest better adhesion of L929 onto the 1:1 PS/PMMA modified film, indicating that this system is a new candidate biomaterial for tissue engineering., (Copyright © 2012 Elsevier B.V. All rights reserved.)

The interaction of luteolin with calf thymus deoxyribonucleic acid (ctDNA) under physiological conditions (Tris-HCl buffer solutions, pH 7.4) was studied by UV-Vis spectroscopy, fluorescence spectroscopy and viscosity measurement method, respectively. The results indicated that a complex of luteolin with ctDNA can be formed. Spectroscopic techniques together with viscosity determination provided evidences of intercalation mode of binding for the interaction between luteolin and ctDNA. The binding constant of luteolin to DNA calculated based on UV-Vis spectroscopy data was found to be 4.52×10(4)L mol(-1) at 310 K. The thermodynamic parameters of the complex were calculated by a double reciprocal method: Δ(r)H(m)(s)=-8.9×10(3)J mol(-1),Δ(r)S(m)(s)=60.5 JK(-1)mol(-1) and Δ(r)G(m)(s)=-2.76×10(4)J mol(-1) (310 K). The interacting forces between luteolin and DNA mainly included hydrophobic interactions and hydrogen bonds. The acridine orange displacement studies revealed that luteolin had significant effect for acridine orange bounded on DNA, which was indicative of intercalation binding., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Climate change, world population growth, and poverty have led to an increase in the incidence of leptospirosis. Leptospirosis is caused by pathogenic spirochaete bacteria that belong to the genus Leptospira. The bacteria are maintained in the renal tubules of the reservoir hosts (typically a rodent), then shed into the environment via the urine. Water is key for environmental survival and transmission, as leptospires can survive for several weeks in a moist environment. Therefore, environmental epidemiological studies are needed to study the contamination of environmental water sources. However, few such studies have been performed using cultivation of the isolates and PCR assays. But, leptospira cultivation can be easily contaminated by other organisms and takes usually several weeks. Moreover, PCR is a complex and costly analysis for the underdeveloped countries that have the highest incidence of leptospirosis. In this study, we describe two modifications of a fluorescence microscopy assay based on immuno-magnetic separation (IMS) to detect leptospires in environmental water samples that mainly differ in fluorescent dye staining. The first type uses acridine orange fluorescent dye staining combined with multiplexed IMS for sample screening. The detection limit ranged from 10(2) to 10(3) organisms per mL and largely depended on the capture efficiency (CE) of the immuno-magnetic particles. The second type uses serogroup-specific immuno-particles and direct fluorescence antibody staining (DFA) to detect leptospires; the detection limit of this second assay was approximately 10(1) cells per mL. Both assay types were applied to natural and experimentally infected water samples, which were also analysed with DFM and real-time PCR. Our data show that the fluorescent microscopy immunoassay successfully identified experimental leptospire contamination and was as sensitive as PCR. This modified immune-fluorescence assay may therefore enable epidemiological studies of leptospirosis., (Copyright © 2012 Elsevier B.V. All rights reserved.)

By means of UV and fluorescence spectra, the binding ratios between Er(III)-Trp and DNA in physiological pH environment (pH 7.40) were determined as n(Trp):n(Er(III))=3:1 and (n)ER(III)(Trp)(3):(n)(DNA) = 2:1, and the apparent molar absorptivity of epsilon(Er(III)-Trp-DNA) is 4.33 x 10(5) L mol(-1)cm(-1) which was confirmed by molar ratio method. The binding constants at different temperatures K(B25 degrees C)(theta)=1.93 x 10(4)L mol(-1) and K(B37 degrees C)(theta)=5.28 x 10(3)L mol(-1) were obtained by double reciprocal method. Thermodynamic function computation demonstrates that Delta(r)H(m)(theta) is the primary driving power of the interaction between Er(III)(Trp)(3) and DNA. By combination analysis of the Scatchard method and CD spectrometry, we suggested that the interaction mode between Er(III)(Trp)(3) complex and herring sperm DNA is groove and intercalation bindings., (Copyright 2010 Elsevier B.V. All rights reserved.)

Methods to determine viability of taeniid oncospheres following treatments with potential lethality have practical application in efforts to control transmission. Here we investigated several methods, in lieu of infectivity studies, to assess oncosphere viability and determine lethal temperature treatment regimens. In the first experiment, a standard treatment to exshell oncospheres with 0.5% hypochlorite was assessed for influence on oncosphere recovery of Taenia taeniaeformis eggs. Recovery of eggs and exshelled oncospheres decreased with increasing time in hypochlorite, which indicated that hypochlorite can damage eggs and oncospheres, translating into potential overestimation of lethality of experimental treatments. Losses in hypochlorite were accentuated when eggs were pretreated at 75 degrees C, but not lower temperatures, including 65 degrees C, indicating a sharp threshhold between 65 degrees C and 75 degrees C where eggs and oncospheres became hypersensitive to subsequent hypochlorite treatment. To further investigate this change in relation to temperature, non-vital (acridine orange, AO) and vital (propidium iodide, PI; trypan blue, TB) dyes were used to assess staining of oncospheres (exshelled or not) under conditions ranging from room temperature up to 95 degrees C. The behaviors of dyes as related to internal staining of oncospheres were described using non-linear regression and a sigmoid four-parametric model to determine the inflection point (T50). Each of the dyes differed significantly in T50 estimates, e.g. AO (69.22+/-0.53), PI (73.89+/-0.52) and TB (79.43+/-0.45). For these dyes, the T50 increased in relation to the increasing molecular weight of the dyes. Collectively, the results suggested that barriers to chemical permeability exist in eggs that breakdown incrementally with increasing temperatures above 65 degrees C. This staining behavior and the likelihood that the temperatures involved are above a lethal threshhold clarify a basic limitation in the use of vital dyes to assess oncosphere viability. The results may be relevant to other Taenia spp.

L-glutamine (Gln) withdrawal rapidly triggers apoptosis in the murine hybridoma cell line Sp2/0-Ag14 (Sp2/0). In this report, we examined the possibility that Gln deprivation of Sp2/0 cells triggers an oxidative stress which would contribute to the activation of apoptotic pathways. Gln withdrawal triggered an oxidative stress in Sp2/0 cells, as indicated by an increased accumulation of reactive oxygen species (ROS) and an increase in the intracellular content in protein carbonyl groups. Gln starvation also caused a decrease in the intracellular levels of glutathione (GSH). However, a decrease in GSH was not sufficient to induce Sp2/0 cell death since reducing GSH levels with DL-buthionine-[S,R]-sulfoximine did not affect cell viability. The antioxidant N-acetyl-L-cysteine (NAC), while effective in inhibiting ROS accumulation and oxidative stress, did not prevent the loss in cell viability or the processing and activation of caspase-3 triggered by Gln starvation. On the other hand, NAC did reduce the formation of apoptotic bodies in dying cells. Altogether these results indicate that in Sp2/0 cells, Gln deprivation leads to the induction of an oxidative stress which, while involved in the formation of apoptotic bodies, is not essential to the activation of the cell death program.

Non-small cell lung cancer (NSCLC) includes a group of tumors that respond poorly to drugs. cis-Diamminedichloroplatinum(II) (cis-DDP) toxicity still remains problematic, and not completely solved by the improvement of supportive care. Therefore, the cis-Pt(II) complex of 3-aminoflavone was selected from cis-DDP analogues with a more favourable toxic profile towards normal cells and at least similar or better anti-tumor activity in comparison with cis-DDP. The aim of this research is to compare the ability of the cis-Pt(II) complex of 3-aminoflavone and cis-DDP to induce apoptosis and necrosis in the human non-small cancer cell line A549. Trypan blue dye exclusion, fluorochrome staining (acridine orange/ethidium bromide double staining), MTT and TUNEL (TdT-mediated dUTP Nick-End Labeling) assays were used. The results obtained show that the cis-Pt(II) complex of 3-aminoflavone is more active in inducing apoptosis and necrosis and in decreasing viability in A549 cells than cis-DDP, which suggests that it could be a potential chemotherapeutic drug.

We describe an approach to creating a new class of luminophores which display both long wavelength emissions exceeding 600 nm and long lifetimes. These luminophores are based on resonance energy transfer (RET) from a long lifetime donor to a short lifetime but long wavelength acceptor. We demonstrated the possibility of obtaining these desirable spectral properties using donors and acceptors noncovalently bound to DNA. The donor was a ruthenium (Ru) metal-ligand complex in which one of the diimine ligands intercalated into double-helix DNA. The acceptors were either nile blue, TOTO-3, or TO-PRO-3. Upon binding of the acceptor to donor-labeled DNA, we found that the acceptor quantum yield was remarkably enhanced so that the wavelength-integrated intensities of the donor and acceptor bound to DNA were many-fold greater than the intensity of the donor and acceptor alone when separately bound to DNA. The origin of this effect is efficient energy transfer from the donor. Under these conditions the effective overall quantum yield approaches that of the acceptor. Importantly, the increased quantum yield can be obtained while maintaining usefully long apparent acceptor lifetimes of 30 to 80 ns. The effect of an increased quantum yield from a low quantum yield donor may find use in assays to detect macromolecular binding interactions. These results suggest the synthesis of covalently linked donor-acceptor pairs with the desirable spectral properties of long wavelength emission, high quantum yield, and moderately long lifetimes for gated detection., (Copyright 2001 Academic Press.)

It has been known for many years that trypanosomatids require exogenous essential growth factors in order to divide. Two surface domains are involved in starting nutrient endocytosis: the flagellar pocket and the cytostome. Although the flagellar pocket plays a fundamental role in the endocytic process occurring in several trypanosomatids, we have shown the cytostome as the main structure involved in this process in epimastigote forms of T. cruzi. After one minute of endocytosis, cargo is still found at the cytostome entry as well as along the cytopharynx. After two, five and fifteen minutes of endocytosis, cargo was seen inside vesicles and tubules, prior to fusing with reservosomes. Three-dimensional reconstruction of these tubules and vesicles showed they are interconnected, forming an intricate and branched network, distributed from the perinuclear region to the posterior end of the cell. Whole unfixed parasites that had taken up gold-protein conjugates for fifteen minutes were washed and dried on electron microscope grids. Observation with an energy-filtering transmission electron microscope revealed long gold-filled tubules at the posterior end of the cell. Parasites treated with ammonium chloride had their intracellular traffic slowed down, which allowed us to observe many events of vesicle fusion. The acidic nature of this network was evidenced using acridine orange. Based on pH and protein uptake kinetics we propose that the vesicular-tubular network is the early endosome of Trypanosoma cruzi epimastigotes.

We developed an automated image analysis system to obtain objective data for the rodent peripheral blood micronucleus assay with acridine orange (AO) supravital staining. The system was able to identify micronucleated reticulocytes (MNRETs) and to evaluate inhibition of bone marrow cell proliferation by measuring the reticular area of reticulocytes (RETs). We also developed automated equipment to produce homogeneous acridine orange-coated glass slides. This study was designed to compare automated scoring with manual scoring using 4 model clastogens and 2 mouse strains. The MNRET incidence induced by each clastogen was similar for automated and manual scoring, and there was good correlation (r=0.92) between the methods. In addition, an index of bone marrow toxicity based on the reticular area of RETs was compared to the conventional index (% of polychromatic erythrocytes (PCE) to total erythrocytes; PCE ratio) and was similar. The results indicated that our technique for computer-assisted image analysis for the micronucleus assay with AO supravitally stained peripheral blood RETs was comparable to conventional microscopic scoring, and it was superior in objectivity and statistical power., (Copyright 1998 Elsevier Science B.V.)

In several models of lymphocyte apoptosis, two alterations of mitochondrial function precede advanced DNA fragmentation: (1) a reduction of mitochondrial transmembrane potential (delta psi m) and (2) an increase in mitochondrial generation of superoxide anion. Here we show that two fluorochromes allow for the identification of analogous mitochondrial perturbations in circulating T lymphocytes from human immunodeficiency virus (HIV)-1+ donors. The first among these fluorochromes, the cationic lipophilic dye DiOC6(3), measures delta psi m; the second marker, hydroethidine (HE), is nonfluorescent, unless it is oxidized by superoxide anions to the product ethidium (Eth). CD4+ or CD8+ cells from clinically asymptomatic HIV-1 carriers contain a significantly elevated percentage of cells endowed with enhanced HE --> Eth conversion and/or reduced DiOC6(3) uptake as compared with normal controls. Phenotypic characterization of (HE --> Eth)high cells from HIV+ donors shows that these cells possess a low delta psi m, thus demonstrating a functional alteration of mitochondria. In addition, (HE --> Eth)high cells display a reduced incorporation of the cardiolipin-specific dye nonyl-acridine orange (NAO), showing a structural defect of the cardiolipin-containing inner mitochondrial membrane. Control experiments involving rotenone, an inhibitor of the respiratory chain complex I, indicate that the reactive oxygen species responsible for HE --> Eth conversion is generated during mitochondrial electron transport. In synthesis, it appears that mitochondrial alterations occur in a significant percentage of circulating T lymphocytes from HIV-1 carriers. The extent of delta psi m reduction, as determined ex vivo, correlates with the frequency of cells undergoing DNA fragmentation after overnight in vitro culture. These observations may be important for the understanding and for the direct ex vivo quantitation of HIV-triggered lymphocyte destruction.

The interactions of monocytes and endothelial cells were examined employing acridine orange (AO) binding to chromatin as an electron microscopic probe for studying changes in nuclear activity. The number of AO labeled nuclei were determined in adherent and sub-endothelial monocytes, and endothelial cells either associated with or devoid of monocytes in mesenteric arteries of renal hypertensive and normotensive rats. No AO-positive cell nuceli were found in the few adherent monocytes in the normotensive rats while in hypertensive rats 63% of the adherent monocytes and 86% of the subendothelial monocytes were labeled by AO. The number of AO-positive cell nuclei increased in the endothelium from 2% in normotensive rats to 17% in hypertensive animals. Fifty-seven percent of these endothelial cells were associated with monocytes adhering to their surfaces. Thirty-nine (52%) of AO-positive and 24 (52%) of AO-negative cell pairs represented 84% of cell pairs with identical nuclear activity. These findings indicate a sequential interaction of monocytes and adjacent endothelial cells recognized at the nuclear level. According to other experimental work the binding capacity of chromatin for AO increases as cell nuclei are reactivated by stimulation of cell proliferation. In addition, AO labeling of chromatin visualized electron microscopically is confined to three of the four stages of the cell cycle. Therefore AO labeling in the present experiments might indicate the reentrance of monocytes and endothelial cells into the cell cycle.

Two hypotheses have recently been proposed for the thiocyanate inhibition of gastric acid secretion--a protonophore mechanism and an uncoupling model. The mechanistic aspects for the latter scheme have been examined on the following basis: capability of generating verifiable predictions, supporting evidence that is unambiguous, and compatibility with experimental realities. Gastric microsomes bind 5 nmol of SCN-/mg, and a "pure" and highly active fraction of H+,K+-ATPase prepared from gastric microsomes binds about 15 nmol of SCN-/mg. The affinity of SCN- binding to gastric microsomes changes from 10 to 25 mM in the presence of 20 mM K+ suggesting competition between K+ and SCN-. Potassium also displaces the bound SCN- from "pure" H+,K+-ATPase with a Ki of about 25 mM. Of the cations tested--Tl+, K+, Rb+, Cs+, NH4+, Na+, and Li+--Tl+ was the most effective in displacing bound SCN- while Na+ and Li+ were without effect. The effects of anions such as Cl-, NO3-, and gluconate were found to be nonspecific and absolutely dependent on K+ as cocation. Sulfate and OCN-, on the other hand, showed an ability to displace SCN- as both K+ and Na+ salts. For SO4(-2) the K+ form was much more effective than the Na+ salt. Besides these antagonistic effects of K+ and congeners with the H+,K+-ATPase-bound SCN-, a competition between K+ and SCN- was also observed at the level of gastric K+-stimulated pNPPase reaction. The effects of SCN- and two other unrelated anions, F- and NO2-, on artificial delta pH across the microsomal vesicles exhibited a lack of appreciable change up to 5 mM and a small (about 13%) reduction between 10 and 20 mM. However, a combination of CCCP and nigericin or valinomycin completely abolished the delta pH under identical conditions. The present data in conjunction with other reports suggest that the proton impediment model best explains the gastric antisecretory effects of SCN-.