Tendons are highly specialized load-bearing tissues with very limited healing capacity. Given their mechanosensitive nature, the combination of tendon mimetic scaffolds with remote mechanical ...actuation could synergistically contribute to the fabrication of improved tissue engineered alternatives for the functional regeneration of tendons. Here, hybrids of cellulose nanocrystals decorated with magnetic nanoparticles were produced to simultaneously reinforce and confer magnetic responsiveness to tendon mimetic hierarchical fibrous scaffolds, resulting in a system that enables remote stimulation of cells
in vitro
and, potentially,
in vivo
after construct transplantation. The biological performance and functionality of these scaffolds were evaluated using human adipose stem cells (hASCs) cultured under or in the absence of magnetic actuation. It was demonstrated that magneto-mechanical stimulation of hASCs promotes higher degrees of cell cytoskeleton anisotropic organization and steers the mechanosensitive YAP/TAZ signaling pathway. As feedback, stimulated cells show increased expression of tendon-related markers, as well as a pro-healing profile in genes related to their inflammatory secretome. Overall, these results support the use of the proposed magnetic responsive fibrous scaffolds as remote biointegrated actuators that can synergistically boost hASC tenogenesis through mechanosensing mechanisms and may modulate their pro-healing paracrine signaling, thus collectively contributing to the improvement of the regenerative potential of engineered tendon grafts.
Topographical cues of magnetically responsive tendon mimetic 3D scaffolds in combination with magneto-mechanical stimulation of human adipose stem cells synergistically boost their tenogenesis.
In the single mitochondrion of protozoan trypanosomatid parasites there are several sites for the generation and elimination of reactive oxygen species (ROS), a class of molecules that exhibit a dual ...role in cells, either as regulatory mediators or as cytotoxic effectors.
Formation of ROS in trypanosomatid mitochondria can be induced by various drug compounds. Importantly, it can also be triggered by specific physiologic stimuli, indicating that this phenomenon may occur in living parasites as well. Elimination of ROS in these organelles is attributed to the activity of two iron-dependent superoxide dismutases (FeSODs) and up to three different peroxidases (a cytochrome c peroxidase and two thiol peroxidases).
Data regarding the formation of ROS in trypanosomatid mitochondria are limited and nonsystematic. Another critical issue refers to the exact contribution of mitochondrial FeSODs and peroxidases for ROS removal, given that their antioxidant activity is not essential when abrogated individually. This suggests some level of functional overlapping or that ROS produced in mitochondria under normal conditions can be removed noncatalytically. Also still unsolved is the mechanism by which mitochondrial thiol peroxidases are regenerated to their reduced (active) form.
The production of intramitochondrial ROS under physiologic conditions and their implication in parasite biology should be further clarified. The relative importance of enzymatic versus nonenzymatic mechanisms for ROS elimination in trypanosomatid mitochondria also requires investigation. Simultaneous depletion of several redundant antioxidant enzymes and determination of noncatalytic antioxidants are possible ways to achieve this.
Significance Peroxiredoxins (Prxs) are highly abundant proteins, which serve two seemingly mutually exclusive roles as peroxidases and molecular chaperones. Little is known about the precise ...mechanism of Prxs’ activation as chaperone and the physiological significance of this second function. Here we demonstrate that in Leishmania infantum , reduced Prx provides a crucial, stress-specific chaperone reservoir, which is activated rapidly upon exposure to unfolding stress conditions. Once activated, Prx protects a wide range of different clients against protein unfolding. Clients are bound in the center of the decameric ring, providing experimental evidence for previous claims that Prxs serve as likely ancestors of chaperonins. Interference with client binding impairs Leishmania infectivity, providing compelling evidence for the in vivo importance of Prx’s chaperone function.
Cytosolic eukaryotic 2-Cys-peroxiredoxins have been widely reported to act as dual-function proteins, either detoxifying reactive oxygen species or acting as chaperones to prevent protein aggregation. Several stimuli, including peroxide-mediated sulfinic acid formation at the active site cysteine, have been proposed to trigger the chaperone activity. However, the mechanism underlying this activation and the extent to which the chaperone function is crucial under physiological conditions in vivo remained unknown. Here we demonstrate that in the vector-borne protozoan parasite Leishmania infantum , mitochondrial peroxiredoxin (Prx) exerts intrinsic ATP-independent chaperone activity, protecting a wide variety of different proteins against heat stress-mediated unfolding in vitro and in vivo. Activation of the chaperone function appears to be induced by temperature-mediated restructuring of the reduced decamers, promoting binding of unfolding client proteins in the center of Prx’s ringlike structure. Client proteins are maintained in a folding-competent conformation until restoration of nonstress conditions, upon which they are released and transferred to ATP-dependent chaperones for refolding. Interference with client binding impairs parasite infectivity, providing compelling evidence for the in vivo importance of Prx’s chaperone function. Our results suggest that reduced Prx provides a mitochondrial chaperone reservoir, which allows L. infantum to deal successfully with protein unfolding conditions during the transition from insect to the mammalian hosts and to generate viable parasites capable of perpetuating infection.
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Amphotericin B (AmB) is a highly hydrophobic drug with significant leishmanicidal activity whose use is limited by its poor water solubility and adverse effects. Polymer-drug conjugates are proposed ...as a delivery system designed to overcome those limitations while improving drug bioavailability, safety, and activity. Here, AmB was covalently linked to periodate-oxidized hyaluronic acid (HA) (oxidation degree of 30.1 ± 5.6%) via a Schiff base (HA-AmB imine). The conjugate presents high water solubility and self-assembles into particles with a mean size of 88.2 ± 17.6 nm, a negative charge (−28.3 ± 0.9 mV), and a drug content of 17.8 ± 1.4%. Spectroscopic studies revealed the presence of AmB in aggregate and super-aggregated forms in the conjugate, which could explain the significant reduction of the in vitro cytotoxicity and hemolytic activity. The formulation showed not only in vitro anti-leishmanial activity against L. infantum-infected macrophages (IC50 = 0.023 μM) but also against an in vivo infected mouse model, promoting a 1.32- and a 4.98-log10 suppression of the L. infantum burden in the spleens and liver, respectively, without toxic effects. In summary, this study describes the safe and effective use of water-soluble HA-AmB imine conjugates for leishmaniasis treatment.
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The contribution of trypanosomatid mitochondrial complex I for energy transduction has long been debated. Herein, we summarize current knowledge on the composition and relevance of this enzyme. ...Bioinformatic and proteomic analyses allowed the identification of many conserved and trypanosomatid-specific subunits of NADH:ubiquinone oxidoreductase, revealing a multifunctional enzyme capable of performing bioenergetic activities and possibly, also of functioning in fatty acid metabolism. A multimeric structure organized in 5 domains of more than 2 MDa is predicted, in contrast to the 1 MDa described for mammalian complex I. The relevance of mitochondrial complex I within the Trypanosomatidae family is quite diverse with its NADH oxidation activity being dispensable for both procyclic and bloodstream
Trypanosoma brucei
, whereas in
Phytomonas serpens
the enzyme is the only respiratory complex able to sustain membrane potential. Aside from complex I, trypanosomatid mitochondria contain a type II NADH dehydrogenase and a NADH-dependent fumarate reductase as alternative electron entry points into the respiratory chain and thus, some trypanosomatids may have bypassed the need for complex I. The involvement of each of these enzymes in the maintenance of the mitochondrial redox balance in trypanosomatids is still an open question and requires further investigation.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OBVAL, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
There is an urgent need for the discovery of new antileishmanial drugs with a new mechanism of action. Type 2 NADH dehydrogenase from
(
NDH2) is an enzyme of the parasite's respiratory system, which ...catalyzes the electron transfer from NADH to ubiquinone without coupled proton pumping. In previous studies of the related NADH: ubiquinone oxidoreductase crystal structure from
, two ubiquinone-binding sites (UQ
and UQ
) were identified and shown to play an important role in the NDH-2-catalyzed oxidoreduction reaction. Based on the available structural data, we developed a three-dimensional structural model of
NDH2 using homology detection methods and performed an in silico virtual screening campaign to search for potential inhibitors targeting the
NDH2 ubiquinone-binding site 1-UQ
. Selected compounds displaying favorable properties in the computational screening experiments were assayed for inhibitory activity in the structurally similar recombinant NDH-2 from
and leishmanicidal activity was determined in the wild-type axenic amastigotes and promastigotes of
. The identified compound, a substituted 6-methoxy-quinalidine, showed promising nanomolar leishmanicidal activity on wild-type axenic promastigotes and amastigotes of
and the potential for further development.
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The objective of this study was to investigate the knowledge, attitudes and behavior regarding antibiotics of the general population.
The study sample consisted of adult subjects who consulted ...general practitioners at health centers in Serbia and agreed to complete the questionnaire. A total of 668 questionnaires were distributed; 500 respondents completed the entire questionnaire (response rate 74.85%).
The average age was 51.65 ± 16.56 years, 60.80% of the respondents were women. The median antibiotic knowledge score was 9. Predictors of adequate antibiotic knowledge were higher education level and a family member whose ocuppation was related to health-care. Overall, 58.4% of respondents believed that antibiotics could be used to treat common cold. Around a half of the respondents (47.2%) self-medicated with antibiotics at least once during their life-time, and around a quarter (24.2%) during the last treatment of infection. Patients with inadequate knowledge had 3 times higher chances of self-medicating with antibiotics compared to those with adequate knowledge. Although 98.20% of respondents claimed that antibiotic treatment should be started after a visit to a doctor and receiving a prescription, only 65.8% obtained antibiotics with prescription from a doctor during the last infection.
Although these results demonstrate that respondents had a relatively adequate level of knowledge regarding antibiotics use, some areas of misconceptions and improper behavior were identified. Therefore, further rationalization should be focused on educational campaigns targeting the behavior of patients with regard to antibiotic use.
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This article provides an overview about the recent advances in the dissection of the peroxide metabolism of Trypanosomatidae. This family of protozoan organisms comprises the medically relevant ...parasites Trypanosoma brucei, Trypanosoma cruzi, and Leishmania spp. Over the past 10 years, three major families of peroxidases have been identified in these organisms: (a) 2-cysteine peroxiredoxins, (b) nonselenium glutathione peroxidases, and (c) ascorbate peroxidases. In trypanosomatids, these enzymes display the unique feature of using reducing equivalents derived from trypanothione, a dithiol found exclusively in these protozoa. The electron transfer between trypanothione and the peroxidases is mediated by a redox shuttle, which can either be tryparedoxin, ascorbate, or even glutathione. The preference for the intermediate molecule differs among each peroxidase and so does the specificity for the peroxide substrate. These observations, added to the fact that these peroxidases are distributed throughout different subcellular compartments, point to the existence of an elaborate peroxide metabolism in trypanosomatids. With the completion of the trypanosomatids genome, other molecules displaying peroxidase activity might be added to this list in the future.
Attitudes towards conventional and complementary medicine among future healthcare professionals can impact their future pharmacotherapy practice. This study aimed to determine the prevalence and ...predisposing factors related to self-medication among medical and pharmacy students.
This cross-sectional questionnaire-based study was performed at the Faculty of Medicine, University of Novi Sad, Serbia, on first- and final-year students of medicine and pharmacy. The multivariate Poisson regression model with robust variance was used to identify the main predictors of self-medication.
The overall self-medication prevalence in the past year was 81.3%. Independent risk factors for self-medication identified in the regression analysis were the final study year, housing condition, i.e., living in a leased apartment or in a student dormitory in comparison to living with parents, and cigarette consumption. The conventional drugs were the most frequently used, mostly for the symptoms of cold and pain. Final-year students had more confidence in conventional medicines than in herbal drugs and were more aware of the risks of their concomitant use.
Self-medication is highly prevalent among students of medical sciences, especially among final-year students. Increased medical knowledge led to the higher awareness of the drug interaction risks.
Abstract
Bacteria are an active and diverse component of pelagic communities. The identification of main factors governing microbial diversity and spatial distribution requires advanced mathematical ...analyses. Here, the bacterial community composition was analysed, along with a depth profile, in the open Adriatic Sea using amplicon sequencing of bacterial 16S rRNA and the Neural gas algorithm. The performed analysis classified the sample into four best matching units representing heterogenic patterns of the bacterial community composition. The observed parameters were more differentiated by depth than by area, with temperature and identified salinity as important environmental variables. The highest diversity was observed at the deep chlorophyll maximum, while bacterial abundance and production peaked in the upper layers. The most of the identified genera belonged to Proteobacteria, with uncultured AEGEAN-169 and SAR116 lineages being dominant Alphaproteobacteria, and OM60 (NOR5) and SAR86 being dominant Gammaproteobacteria. Marine
Synechococcus
and
Cyanobium
-related species were predominant in the shallow layer, while
Prochlorococcus
MIT 9313 formed a higher portion below 50 m depth. Bacteroidota were represented mostly by uncultured lineages (NS4, NS5 and NS9 marine lineages). In contrast, Actinobacteriota were dominated by a candidatus genus
Ca.
Actinomarina. A large contribution of Nitrospinae was evident at the deepest investigated layer. Our results document that neural network analysis of environmental data may provide a novel insight into factors affecting picoplankton in the open sea environment.
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