scholarly journals Alkyl Galactofuranosides Strongly Interact with Leishmania donovani Membrane and Provide Antileishmanial Activity

2014 ◽  
Vol 58 (4) ◽  
pp. 2156-2166 ◽  
Author(s):  
Muhammad Suleman ◽  
Jean-Pierre Gangneux ◽  
Laurent Legentil ◽  
Sorya Belaz ◽  
Yari Cabezas ◽  
...  
Keyword(s):  
Membrane Fluidity ◽  
Leishmania Donovani ◽  
Electron Paramagnetic Resonance Spectroscopy ◽  
Direct Interaction ◽  
Annexin V ◽  
Antileishmanial Activity ◽  
Resonance Spectroscopy ◽  
Paramagnetic Resonance ◽  
Content Type

ABSTRACTWe investigated thein vitroeffects of four alkyl-galactofuranoside derivatives, i.e., octyl-β-d-galactofuranoside (compound 1), 6-amino-β-d-galactofuranoside (compound 2), 6-N-acetamido-β-d-galactofuranoside (compound 3), and 6-azido-β-d-galactofuranoside (compound 4), onLeishmania donovani. Their mechanism of action was explored using electron paramagnetic resonance spectroscopy (EPR) and nuclear magnetic resonance (NMR), and ultrastructural alterations were analyzed by transmission electron microscopy (TEM). Compound 1 showed the most promising effects by inhibiting promastigote growth at a 50% inhibitory concentration (IC50) of 8.96 ± 2.5 μM. All compounds exhibit low toxicity toward human macrophages. Compound 1 had a higher selectivity index than the molecule used for comparison, i.e., miltefosine (159.7 versus 37.9, respectively). EPR showed that compound 1 significantly reduced membrane fluidity compared to control promastigotes and to compound 3. The furanose ring was shown to support this effect, since the isomer galactopyranose had no effect on parasite membrane fluidity or growth. NMR showed a direct interaction of all compounds (greatest with compound 1, followed by compounds 2, 3, and 4, in descending order) with the promastigote membrane and with octyl-galactopyranose and octanol, providing evidence that then-octyl chain was primarily involved in anchoring with the parasite membrane, followed by the putative crucial role of the furanose ring in the antileishmanial activity. A morphological analysis of compound 1-treated promastigotes by TEM revealed profound alterations in the parasite membrane and organelles, but this was not the case with compound 3. Quantification of annexin V binding by flow cytometry confirmed that compound 1 induced apoptosis in >90% of promastigotes. The effect of compound 1 was also assessed on intramacrophagic amastigotes and showed a reduction in amastigote growth associated with an increase of reactive oxygen species (ROS) production, thus validating its promising effect.

scholarly journals In Vitro Evaluation of Antileishmanial Activity of Computationally Screened Compounds against Ascorbate Peroxidase To Combat Amphotericin B Drug Resistance

2017 ◽  
Vol 61 (7) ◽  
Author(s):  
Rani Mansuri ◽  
Ashish Kumar ◽  
Sindhuprava Rana ◽  
Bhavana Panthi ◽  
M. Yousuf Ansari ◽  
...  
Keyword(s):  
Amphotericin B ◽  
Ascorbate Peroxidase ◽  
Leishmania Donovani ◽  
Harmful Effect ◽  
Annexin V ◽  
Antileishmanial Activity ◽  
New Drugs ◽  
Content Type ◽  
Lipinski’S Rule

ABSTRACT In visceral leishmaniasis (VL), the host macrophages generate oxidative stress to destroy the pathogen, while Leishmania combats the harmful effect of radicals by redox homeostasis through its unique trypanothione cascade. Leishmania donovani ascorbate peroxidase (LdAPx) is a redox enzyme that regulates the trypanothione cascade and detoxifies the effect of H2O2. The absence of an LdAPx homologue in humans makes it an excellent drug target. In this study, the homology model of LdAPx was built, including heme, and diverse compounds were prefiltered (PAINS, ADMET, and Lipinski's rule of five) and thereafter screened against the LdAPx model. Compounds having good affinity in terms of the Glide XP (extra precision) score were clustered to select diverse compounds for experimental validation. A total of 26 cluster representatives were procured and tested on promastigote culture, yielding 12 compounds with good antileishmanial activity. Out of them, six compounds were safer on the BALB/c peritoneal macrophages and were also effective against disease-causing intracellular amastigotes. Three out of six compounds inhibited recombinant LdAPx in a noncompetitive manner and also demonstrated partial reversion of the resistance property in an amphotericin B (AmB)-resistant strain, which may be due to an increased level of reactive oxygen species (ROS) and decrease of glutathione (GSH) content. However, inhibition of LdAPx in resistant parasites enhanced annexin V staining and activation of metacaspase-like protease activity, which may help in DNA fragmentation and apoptosis-like cell death. Thus, the present study will help in the search for specific hits and templates of potential therapeutic interest and therefore may facilitate the development of new drugs for combination therapy against VL.

scholarly journals Nitric oxide interactions with cobalamins: biochemical and functional consequences

Blood ◽  
1996 ◽  
Vol 88 (5) ◽  
pp. 1857-1864 ◽  
Author(s):  
M Brouwer ◽  
W Chamulitrat ◽  
G Ferruzzi ◽  
DL Sauls ◽  
JB Weinberg
Keyword(s):  
Nitric Oxide ◽  
Electron Paramagnetic Resonance Spectroscopy ◽  
Resonance Spectroscopy ◽  
Paramagnetic Resonance ◽  
No Formation ◽  
Iron Nitrosyl ◽  
Wide Range ◽  
Functional Consequences

Abstract Nitric oxide (NO) is a paramagnetic gas that has been implicated in a wide range of biologic functions. The common pathway to evoke the functional response frequently involves the formation of an iron- nitrosyl complex in a target (heme) protein. In this study, we report on the interactions between NO and cobalt-containing vitamin B12 derivatives. Absorption spectroscopy showed that of the four Co(III) derivatives (cyanocobalamin [CN-Cbl], aquocobalamin [H2O-Cbl], adenosylcobalamin [Ado-Cbl], and methylcobalamin [MeCbl]), only the H2O- Cbl combined with NO. In addition, electron paramagnetic resonance spectroscopy of H2O-Cbl preparations showed the presence of a small amount of Cob-(II)alamin that was capable of combining with NO. The Co(III)-NO complex was very stable, but could transfer its NO moiety to hemoglobin (Hb). The transfer was accompanied by a reduction of the Co(III) to Co(II), indicating that NO+ (nitrosonium) was the leaving group. In accordance with this, the NO did not combine with the Hb Fe(II)-heme, but most likely with the Hb cysteine-thiolate. Similarly, the Co(III)-NO complex was capable of transferring its NO to glutathione. Ado-Cbl and Me-Cbl were susceptible to photolysis, but CN- Cbl and H2O-Cbl were not. The homolytic cleavage of the Co(III)-Ado or Co(III)-Me bond resulted in the reduction of the metal. When photolysis was performed in the presence of NO, formation of NO-Co(II) was observed. Co(II)-nitrosyl oxidized slowly to form Co(III)-nitrosyl. The capability of aquocobalamin to combine with NO had functional consequences. We found that nitrosylcobalamin had diminished ability to serve as a cofactor for the enzyme methionine synthase, and that aquocobalamin could quench NO-mediated inhibition of cell proliferation. Our in vitro studies therefore suggest that interactions between NO and cobalamins may have important consequences in vivo.

scholarly journals In Vitro Evaluation of Curcumin- and Quercetin-Loaded Nanoemulsions for Intranasal Administration: Effect of Surface Charge and Viscosity

Pharmaceutics ◽  
2022 ◽  
Vol 14 (1) ◽  
pp. 194
Author(s):  
Gustavo Vaz ◽  
Adryana Clementino ◽  
Evgenia Mitsou ◽  
Elena Ferrari ◽  
Francesca Buttini ◽  
...  
Keyword(s):  
Neurodegenerative Diseases ◽  
Water Solubility ◽  
Electron Paramagnetic Resonance Spectroscopy ◽  
Drug Uptake ◽  
Chemical Degradation ◽  
Nasal Administration ◽  
Resonance Spectroscopy ◽  
Paramagnetic Resonance ◽  
Cytotoxicity Studies

The nose-to-brain delivery of neuroprotective natural compounds is an appealing approach for the treatment of neurodegenerative diseases. Nanoemulsions containing curcumin (CUR) and quercetin (QU) were prepared by high-pressure homogenization and characterized physicochemically and structurally. A negative (CQ_NE−), a positive (CQ_NE+), and a gel (CQ_NEgel) formulation were developed. The mean particle size of the CQ_NE− and CQ_NE+ was below 120 nm, while this increased to 240 nm for the CQ_NEgel. The formulations showed high encapsulation efficiency and protected the CUR/QU from biological/chemical degradation. Electron paramagnetic resonance spectroscopy showed that the CUR/QU were located at the interface of the oil phase in the proximity of the surfactant layer. The cytotoxicity studies showed that the formulations containing CUR/QU protected human nasal cells from the toxicity evidenced for blank NEs. No permeation across an in vitro model nasal epithelium was evidenced for CUR/QU, probably due to their poor water-solubility and instability in physiological buffers. However, the nasal cells’ drug uptake showed that the total amount of CUR/QU in the cells was related to the NE characteristics (CQ_NE− > CQ_NE+ > CQ_NEgel). The method used allowed the obtainment of nanocarriers of an appropriate size for nasal administration. The treatment of the cells showed the protection of cellular viability, holding promise as an anti-inflammatory treatment able to prevent neurodegenerative diseases.

Estradiol replacement reverses ovariectomy-induced muscle contractile and myosin dysfunction in mature female mice

2007 ◽  
Vol 102 (4) ◽  
pp. 1387-1393 ◽  
Author(s):  
Amy L. Moran ◽  
Steven A. Nelson ◽  
Rachel M. Landisch ◽  
Gordon L. Warren ◽  
Dawn A. Lowe
Keyword(s):  
Skeletal Muscle ◽  
Contractile Function ◽  
Electron Paramagnetic Resonance Spectroscopy ◽  
Eccentric Contraction ◽  
Mature Female ◽  
Resonance Spectroscopy ◽  
Paramagnetic Resonance ◽  
Tetanic Force ◽  
Female Mice

Skeletal muscle contractility and myosin function decline following ovariectomy in mature female mice. In the present study we tested the hypothesis that estradiol replacement can reverse those declines. Four-month-old female C57BL/6 mice ( n = 69) were ovariectomized (OVX) or sham operated. Some mice were treated immediately with placebo or 17β-estradiol (OVX + E2) while other mice were treated 30 days postsurgery. Thirty or sixty days postsurgery, soleus muscles were assessed in vitro for contractile function and susceptibility to eccentric contraction-induced injury. Myosin structural dynamics was analyzed in extensor digitorum longus (EDL) muscles by electron paramagnetic resonance spectroscopy. Maximal isometric tetanic force was affected by estradiol status ( P < 0.001) being ∼10% less in soleus muscles from OVX compared with sham-operated mice [168 mN (SD 16.7) vs. 180 mN (SD 14.4)] and was restored in OVX + E2 mice [187 mN (SD 17.6)]. The fraction of strong-binding myosin during contraction was also affected ( P = 0.045) and was ∼15% lower in EDL muscles from OVX compared with OVX + E2 mice [0.263 (SD 0.034) vs. 0.311 (SD 0.022)]. Plasma estradiol levels were correlated with maximal isometric tetanic force ( r = 0.458; P < 0.001) and active stiffness ( r = 0.329; P = 0.044), indicating that circulating estradiol influenced muscle and myosin function. Estradiol was not effective in protecting muscle against an acute eccentric contraction-induced injury ( P ≥ 0.401) but did restore ovariectomy-induced increases in muscle wet mass caused by fluid accumulation. Collectively, estradiol had a beneficial effect on female mouse skeletal muscle.

scholarly journals Besifloxacin liposomes with positively charged additives for an improved topical ocular delivery

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Giselly Almeida dos Santos ◽  
Ricardo Ferreira-Nunes ◽  
Luciana Facco Dalmolin ◽  
Ana Carolina dos Santos Ré ◽  
Jorge Luiz Vieira Anjos ◽  
...  
Keyword(s):  
Drug Delivery ◽  
Defense Mechanisms ◽  
Electron Paramagnetic Resonance Spectroscopy ◽  
Resonance Spectroscopy ◽  
Drug Penetration ◽  
Current Application ◽  
Paramagnetic Resonance ◽  
Electron Paramagnetic ◽  
Positively Charged

Abstract Topical ophthalmic antibiotics show low efficacy due to the well-known physiological defense mechanisms of the eye, which prevents the penetration of exogenous substances. Here, we aimed to incorporate besifloxacin into liposomes containing amines as positively charged additives and to evaluate the influence of this charge on drug delivery in two situations: (i) iontophoretic and (ii) passive treatments. Hypothesis are (i) charge might enhance the electromigration component upon current application improving penetration efficiency for a burst drug delivery, and (ii) positive charge might prolong formulation residence time, hence drug penetration. Liposomes elaborated with phosphatidylcholine (LP PC) or phosphatidylcholine and spermine (LP PC: SPM) were stable under storage at 6 ºC for 30 days, showed mucoadhesive characteristics, and were non-irritant, according to HET-CAM tests. Electron paramagnetic resonance spectroscopy measurements showed that neither the drug nor spermine incorporations produced evident alterations in the fluidity of the liposome's membranes, which retained their structural stability even under iontophoretic conditions. Mean diameter and zeta potential were 177.2 ± 2.7 nm and − 5.7 ± 0.3 mV, respectively, for LP PC; and 175.4 ± 1.9 nm and + 19.5 ± 1.0 mV, respectively, for LP PC:SPM. The minimal inhibitory concentration (MIC) and the minimal bactericide concentration (MBC) of the liposomes for P. aeruginosa showed values lower than the commercial formulation (Besivance). Nevertheless, both formulations presented a similar increase in permeability upon the electric current application. Hence, liposome charge incorporation did not prove to be additionally advantageous for iontophoretic therapy. Passive drug penetration was evaluated through a novel in vitro ocular model that simulates the lacrimal flow and challenges the formulation resistance in the passive delivery situation. As expected, LP PC: SPM showed higher permeation than the control (Besivance). In conclusion, besifloxacin incorporation into positively charged liposomes improved passive topical delivery and can be a good strategy to improve topical ophthalmic treatments.

scholarly journals Tabotamp®, Respectively, Surgicel®, Increases the Cell Death of Neuronal and Glial Cells In Vitro

Materials ◽  
2020 ◽  
Vol 13 (11) ◽  
pp. 2453
Author(s):  
Sandra Leisz ◽  
Marie-Luise Trutschel ◽  
Karsten Mäder ◽  
Christian Scheller ◽  
Christian Strauss ◽  
...  
Keyword(s):  
Fluorescence Microscopy ◽  
Schwann Cells ◽  
Cell Lines ◽  
Ph Value ◽  
Electron Paramagnetic Resonance Spectroscopy ◽  
Regenerated Cellulose ◽  
Resonance Spectroscopy ◽  
Paramagnetic Resonance ◽  
Eloquent Areas

Oxidized regenerated cellulose (ORC) is an approved absorbable hemostat in neurosurgery, and contains 18–21% carboxylic acid groups. This modification leads to a low pH in aqueous solutions. Therefore, the aim of study was to analyze the pH-dependent effects of the ORC Tabotamp® on astrocytes, Schwann cells, and neuronal cells in vitro to investigate whether Tabotamp® is a suitable hemostat in cerebral eloquent areas. The ORC-dependent pH value changes were measured with (i) a pH meter, (ii) electron paramagnetic resonance spectroscopy, using pH-sensitive spin probes, and (iii) with fluorescence microscopy. Cell lines from neurons, astrocytes, and Schwann cells, as well as primary astrocytes were incubated with increasing areas of Tabotamp®. Cytotoxicity was detected using a fluorescence labeled DNA-binding dye. In addition, the wounding extent was analyzed via crystal violet staining of cell layers. The strongest pH reduction (to 2.2) was shown in phosphate buffered saline, whereas culture medium and cerebrospinal fluid demonstrated a higher buffer capacity during Tabotamp® incubation. In addition, we could detect a distance-dependent pH gradient by fluorescence microscopy. Incubation of Tabotamp® on cell monolayers led to detachment of covered cells and showed increased cytotoxicity in all tested cell lines and primary cells depending on the covered area. These in vitro results indicate that Tabotamp® may not be a suitable hemostat in cerebral eloquent areas.

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