BACKGROUND: Fluoroquinolones (FQs) are widely used in livestock and poultry industry because of their satisfactory effects in preventing and treating bacterial infection. However, due to irrational use and poor biodegradability, FQs can easily remain in food animals and further enter the human body through the food chain. Therefore, accurate and sensitive detection of FQs residues in animal-origin food is significant. The traditional methods commonly used for FQs detection have some limitations. Ratiometric fluorescence detection technology has the advantages of fast, sensitive, self-correcting, and easy visualization. However, the reports on the use of ratiometric fluorescence probes for FQs detection are limited.
RESULTS: In this work, a novel probe was proposed for ratiometric fluorescent analysis of FQs. In this probe, the fluorescence of dithioerythritol stabilized copper nanoclusters (DTE-Cu NCs) was significantly enhanced due to the Tb3+ triggered aggregation-induced emission effect. FQs bound Tb3+ in Tb3+/DTE-Cu NCs through carboxyl and carbonyl groups, so that Tb3+ was effectively sensitized to emit green fluorescence. However, the red fluorescence of DTE-Cu NCs was not interfered. The fluorescence of the probe transformed from red to green with the increase of FQs concentration. Using norfloxacin (NOR), difloxacin (DIF), and enrofloxacin (ENR) as FQs simulants, this probe showed a sensitive linear response ranged from 0.025 to 22.5 μM, with the limits of detection of 9.6 nM, 9.3 nM, and 7.7 nM. The application potential for FQs detection was verified via a standard addition assay of egg samples with the recovery rate of 90.4 %-114.7 %.
CONCLUSIONS: The fluorescence probe based on Tb3+/DTE-Cu NCs is expected to realize the ratiometric fluorescence sensitive detection of FQs. The establishment of this simple, effective, and rapid detection platform opens up a new way for the detection of FQs residues in animal-origin foods, and also provides a new idea for the design of rapid detection platforms for other hazard factors.

The dithiol reagents phenylarsine oxide (PAO) and dibromobimane (DBrB) have opposite effects on the F1FO-ATPase activity. PAO 20% increases ATP hydrolysis at 50 μM when the enzyme activity is activated by the natural cofactor Mg2+ and at 150 μM when it is activated by Ca2+. The PAO-driven F1FO-ATPase activation is reverted to the basal activity by 50 μM dithiothreitol (DTE). Conversely, 300 μM DBrB decreases the F1FO-ATPase activity by 25% when activated by Mg2+ and by 50% when activated by Ca2+. In both cases, the F1FO-ATPase inhibition by DBrB is insensitive to DTE. The mitochondrial permeability transition pore (mPTP) formation, related to the Ca2+-dependent F1FO-ATPase activity, is stimulated by PAO and desensitized by DBrB. Since PAO and DBrB apparently form adducts with different cysteine couples, the results highlight the crucial role of cross-linking of vicinal dithiols on the F1FO-ATPase, with (ir)reversible redox states, in the mPTP modulation.

HBL-100 breast epithelial cells were cultured with a blocker (N-ethylmaleimide) and protector (1,4-dithioerythritol) of SH groups. The study assessed changes in redox potential of glutathione and thioredoxin systems, intensity of oxidative modification of proteins, ROS production, and cell proliferation. The roles of thioredoxin system and protein oxidative modification in HBL-100 cell proliferation under redox status modulation were established. The role of carbonylated thioredoxin in arrest of the cell cycle in S-phase was demonstrated, which could be used for targeted therapy of the diseases accompanied by oxidative stress and disturbed redox status.

A colorimetric assay is described for highly selective and sensitive determination of Hg(II) ions by using gold nanoparticles (AuNPs) functionalized with dithioerythritol (DETL). This method relies on the unique optical properties of DETL-functionalized AuNPs as well as the thiophilicity of both AuNPs and Hg(II). In the presence of DETL, the AuNPs aggregate due to ligand exchange between thiol groups of DETL and the citrate ions on the surface of AuNPs. This induces a color change from red to blue. On addition of Hg(II), the thiol groups preferably interact with Hg(II) rather than with AuNPs. Thus, the DETL is released from the surface of the AuNPs and binds to Hg(II). This triggers the redispersion of the AuNPs. The ratio of absorbances at 650 and 525 nm drops linearly in two Hg(II) concentration ranges (viz. from 0.1 to 0.5 μM, and from 0.5 to 5 μM). The ions Cu(II), Pb(II), and Cd(II) do not interfere even in the absence of masking agents. The detection limit is as low as 24 nM. Graphical abstract A highly selective colorimetric method based on gold nanoparticles via double ligand exchange reaction is described for determination of Hg2+. This assay can selective detect Hg2+ with no response to major interfering metal ions such as Cu2+, Pb2+, and Cd2+ in the absence of masking agents compared with previous works.

The aim of this study was to establish the role of redox modification of proteins and redox status in the realization of apoptosis of MCF-7 breast adenocarcinoma cells du-ing cultivation with the SH-group blocker N-ethylmaleimide (NEM) and the SH-group protector 1,4-dithioerythritol (DTE). The activation of apoptosis in MCF-7 breast adenocarcinoma cells was shown to be due to the irreversible modification of redox sensitive protein molecules. The presence of DTE in the culture medium of cancer.cells caused reversible glutathionylation of protein molecules and did not change the: number of apoptotic MCF-7 cells.

Translocation of phosphatidylserine from the inner leaflet to the outer leaflet of the endothelial membrane via phospholipid scramblase-1 (PLSCR1) is an apoptotic signal responsible for the loss of endothelial barrier integrity after ischemia-reperfusion injury (IRI). We hypothesized that inhibiting phosphatidylserine expression on endothelial cells would attenuate IRI induced increases in hydraulic permeability (Lp).
Mesenteric Lp was measured in rat post-capillary mesenteric venules subjected to IRI via superior mesenteric artery (SMA) occlusion (45 minutes) and release (300 minutes) in conjunction with several inhibitors of phosphatidylserine exposure as follows: (1) inhibition of PLSCR1 translocation (dithioerythritol, n = 3), (2) inhibition of PLSCR1 membrane trafficking (2-bromopalmitate [2-BP], n = 3), and (3) inhibition of ion exchange necessary for PLSCR1 function (4,4\'-Diisothiocyano-2,2\'-stilbenedisulfonic acid [DIDS], n = 3). Under the same IRI conditions, rats were also administered targeted inhibitors of phosphatidylserine exposure including knockdown of PLSCR1 (n = 3) using RNA interference (RNAi), and as a potential therapeutic tool Diannexin, a selective phosphatidylserine blocker (n = 3).
During IRI net Lp increased by 80% (p < 0.01). Net reductions of Lp were accomplished by 2-BP (46% reduction, p = 0.005), combined DET + 2-BP + DIDS (32% reduction, p = 0.04), RNAi (55% reduction, p = 0.002), Diannexin administered pre-SMA artery occlusion (73% reduction, p = 0.001), and post-SMA occlusion (70% reduction, p = 0.002).
Phosphatidylserine exposure is a key event in the pathogenesis of microvascular dysfunction during IRI. Clinically, inhibition of phosphatidylserine exposure is a promising strategy that may 1 day be used to mitigate the effects of IRI.

S-nitrosylation of the mitochondrial carnitine/acylcarnitine transporter (CACT) has been investigated on the native and the recombinant proteins reconstituted in proteoliposomes, and on intact mitochondria. The widely-used NO-releasing compound, GSNO, strongly inhibited the antiport measured in proteoliposomes reconstituted with the native CACT from rat liver mitochondria or the recombinant rat CACT over-expressed in E. coli. Inhibition was reversed by the reducing agent dithioerythritol, indicating a reaction mechanism based on nitrosylation of Cys residues of the CACT. The half inhibition constant (IC50) was very similar for the native and recombinant proteins, i.e., 74 and 71μM, respectively. The inhibition resulted to be competitive with respect the substrate, carnitine. NO competed also with NEM, correlating well with previous data showing interference of NEM with the substrate transport path. Using a site-directed mutagenesis approach on Cys residues of the recombinant CACT, the target of NO was identified. C136 plays a major role in the reaction mechanism. The occurrence of S-nitrosylation was demonstrated in intact mitochondria after treatment with GSNO, immunoprecipitation and immunostaining of CACT with a specific anti NO-Cys antibody. In parallel samples, transport activity of CACT measured in intact mitochondria, was strongly inhibited after GSNO treatment. The possible physiological and pathological implications of the post-translational modification of CACT are discussed.

Activation of free radical oxidation in different cell types, including breast epithelial cells, may result in damage to macromolecules, in particular, proteins taking part in regulation of cell proliferation and apoptosis. The glutathione, glutaredoxin and thioredoxin systems play an essential role in maintaining intracellular redox homeostasis. Due to this fact, modulation of cellular redox status under the effect of an SH group inhibitor and an SH group protector may be used as a model for studying the role of redox proteins and glutathione in regulating cell proliferation in different pathological processes. In this study we have evaluated the state of the thioredoxin, glutaredoxin and glutathione systems as well as their role in regulating proliferation of HBL-100 breast epithelial cells under redox status modulation with N-ethylmaleimide (NEM) and 1,4-dithioerythriol (DTE). Modulating the redox status of breast epithelial cells under the effect of NEM and DTE influences the functional activity of glutathione-dependent enzymes, glutaredoxin, thioredoxin, and thioredoxin reductase through changes in the GSH and GSSG concentrations. In HBL-100 cells under redox-status modulation, we have found an increase in the number of cells in the S-phase of the cell cycle and a decrease in the number of cells in the G0/G1 and G2/М phases, as opposed to the values in the intact culture. The proposed model of proliferative activity of cells under redox status modulation may be used for development of new therapeutic approaches for treatment of diseases accompanied by oxidative stress generation.
Aktivatsiia svobodnoradikal\'nogo okisleniia v razlichnykh tipakh kletok, v tom chisle kletkakh épiteliia molochnoĭ zhelezy, sposobna privodit\' k povrezhdeniiu makromolekul, v chastnosti belkov, uchastvuiushchikh v reguliatsii proliferatsii i programmirovannoĭ kletochnoĭ gibeli. V podderzhanii vnutrikletochnogo redoks-gomeostaza vazhnuiu rol\' igraiut sistemy glutationa, tioredoksina i glutaredoksina. V sviazi s étim moduliatsiia redoks-statusa kletok pri deĭstvii blokatora i protektora SH-grupp belkov mozhet byt\' ispol\'zovana v kachestve modeli dlia izucheniia roli redoks-belkov i glutationa v reguliatsii proliferatsii kletok pri razvitii razlichnykh patologicheskikh protsessov. V dannoĭ rabote issledovali sostoianie sistem tioredoksina, glutaredoksina, glutationa i ikh rol\' v reguliatsii proliferatsii kletok épiteliia molochnoĭ zhelezy linii HBL-100 pri moduliatsii redoks-statusa s pomoshch\'iu N-étilmaleimida i 1,4-ditioéritritola. Moduliatsiia redoks-statusa kletok épiteliia molochnoĭ zhelezy pri deĭstvii blokatora (N-étilmaleimida) i protektora (1,4-ditioéritritola) tiolovykh grupp belkov i peptidov, sposobstvuet izmeneniiu funktsional\'noĭ aktivnosti glutationzavisimykh fermentov, glutaredoksina, tioredoksina i tioredoksinreduktazy posredstvom izmeneniia kontsentratsii GSH i GSSG. V kletkakh linii HBL-100 pri moduliatsii redoks-statusa obnaruzheno uvelichenie kolichestva kletok v S faze kletochnogo tsikla i snizhenie – v G0/G1 i G2/M fazakh po sravneniiu so znacheniiami pokazateleĭ v intaktnoĭ kul\'ture. Predlozhennyĭ sposob otsenki proliferativnoĭ aktivnosti kletok pri moduliatsii ikh redoks-sostoianiia mozhet byt\' ispol\'zovan pri razrabotke novykh terapevticheskikh podkhodov dlia lecheniia zabolevaniĭ, soprovozhdaiushchikhsia razvitiem okislitel\'nogo stressa.

In this study we explore the preparation of core-crosslinked micelles of linear-dendritic methoxy-poly(ethylene glycol) (MPEG)-co-poly(ester-sulfide) (PES) polymers to improve the stability of such polymeric micelle systems against premature disintegration and drug release. A series of MPEG-PES copolymers were synthesised via stepwise reactions of acetylation and thiol-ene photoreaction. Surface tension measurement showed that the copolymers with ethenyl surface groups could self-associate in dilute aqueous solutions to form micelles. Crosslinking within the micelle cores in the presence of dithioerythritol (DTT) linker was initiated under UV radiation. The formation of core-crosslinked micelles was confirmed by HPLC in combination with charged aerosol detection (CAD). The copolymers were found to readily hydrolyse under acidic conditions due to the ester-containing dendrons. Drug solubilisation capacities of the micellar solutions were determined using griseofulvin as a poorly water-soluble model drug. The solubility of griseofulvin showed a 10-fold enhancement in 1% w/v micelle solution and increased with the concentration of the copolymers. Drug release studies indicated that a more sustained release of griseofulvin was achieved for the core-crosslinked micelles compared to the non-crosslinked micelles, attributable to greater stability of the crosslinked core structure. The findings of this study present a new pathway towards developing biodegradable polymeric nanocarriers.

In spite of the known widespread toxicity of mercury, its impact on mitochondrial bioenergetics is a still poorly explored topic. Even if many studies have dealt with mercury poisoning of mitochondrial respiration, as far as we are aware Hg2+ effects on individual complexes are not so clear. In the present study changes in swine heart mitochondrial respiration and F1FO-ATPase (F-ATPase) activity promoted by micromolar Hg2+ concentrations were investigated. Hg2+ was found to inhibit the respiration of NADH-energized mitochondria, whereas it was ineffective when the substrate was succinate. Interestingly, the same micromolar Hg2+ doses which inhibited the NADH-O2 activity stimulated the F-ATPase, most likely by interacting with adjacent thiol residues. Accordingly, Hg2+ dose-dependently decreased protein thiols and all the elicited effects on mitochondrial complexes were reversed by the thiol reducing agent DTE. These findings clearly indicate that Hg2+ interacts with Cys residues of these complexes and differently modulate their functionality by modifying the redox state of thiol groups. The results, which cast light on some implications of metal-thiol interactions up to now not fully explored, may contribute to clarify the molecular mechanisms of mercury toxicity to mitochondria.