Novel PLGA-based nanoformulation decreases doxorubicin-induced cardiotoxicity.

Nanotechnology has the potential to provide formulations of antitumor agents with increased selectivity towards cancer tissue thereby decreasing systemic toxicity. This in vivo study evaluated the potential of novel nanoformulation based on poly(lactic-co-glycolic acid) (PLGA) to reduce the cardiotoxic potential of doxorubicin (DOX). In vivo toxicity of PLGADOX was compared with clinically approved non-PEGylated, liposomal nanoformulation of DOX (LipoDOX) and conventional DOX form (ConvDOX). The study was performed using Wistar Han rats of both sexes that were treated intravenously for 28 days with 5 doses of tested substances at intervals of 5 days. Histopathological analyses of heart tissues showed the presence of myofiber necrosis, degeneration processes, myocytolysis, and hemorrhage after treatment with ConvDOX, whereas only myofiber degeneration and hemorrhage were present after the treatment with nanoformulations. All DOX formulations caused an increase in the troponin T with the greatest increase caused by convDOX. qPCR analyses revealed an increase in the expression of inflammatory markers IL-6 and IL-8 after ConvDOX and an increase in IL-8 expression after lipoDOX treatments. The mass spectra imaging (MSI) of heart tissue indicates numerous metabolic and lipidomic changes caused by ConvDOX, while less severe cardiac damages were found after treatment with nanoformulations. In the case of LipoDOX, autophagy and apoptosis were still detectable, whereas PLGADOX induced only detectable mitochondrial toxicity. Cardiotoxic effects were frequently sex-related with the greater risk of cardiotoxicity observed mostly in male rats.

Spectroscopic study of L-DOPA and dopamine binding on novel gold nanoparticles towards more efficient drug-delivery system for Parkinson's disease.

Nano-drug delivery systems may potentially overcome current challenges in the treatment of Parkinson's disease (PD) by enabling targeted delivery and more efficient blood-brain penetration ability. This study investigates novel gold nanoparticles (AuNPs) to be used as delivery systems for L-DOPA and dopamine by considering their binding capabilities in the presence and absence of a model protein, bovine serum albumin (BSA). Four different AuNPs were prepared by surface functionalization with polyethylene glycol (PEG), 1-adamantylamine (Ad), 1-adamantylglycine (AdGly), and peptidoglycan monomer (PGM). Fluorescence and UV-Vis measurements demonstrated the strongest binding affinity and L-DOPA/dopamine loading efficiency for PGM-functionalized AuNPs with negligible impact of the serum protein presence. Thermodynamic analysis revealed a spontaneous binding process between L-DOPA or dopamine and AuNPs that predominantly occurred through van der Waals interactions/hydrogen bonds or electrostatic interactions. These results represent PGM-functionalized AuNPs as the most efficient at L-DOPA and dopamine binding with a potential to become a drug-delivery system for neurodegenerative diseases. Detailed investigation of L-DOPA/dopamine interactions with different AuNPs was described here for the first time. Moreover, this study highlights a cost- and time-effective methodology for evaluating drug binding to nanomaterials.

Sex affects the response of Wistar rats to polyvinyl pyrrolidone (PVP)-coated silver nanoparticles in an oral 28 days repeated dose toxicity study.

BACKGROUND: Silver nanoparticles (AgNPs) are widely used in biomedicine due to their strong antimicrobial, antifungal, and antiviral activities. Concerns about their possible negative impacts on human and environmental health directed many researchers towards the assessment of the safety and toxicity of AgNPs in both in vitro and in vivo settings. A growing body of scientific information confirms that the biodistribution of AgNPs and their toxic effects vary depending on the particle size, coating, and dose as well as on the route of administration and duration of exposure. This study aimed to clarify the sex-related differences in the outcomes of oral 28 days repeated dose exposure to AgNPs. METHODS: Wistar rats of both sexes were gavaged daily using low doses (0.1 and 1 mg Ag/kg b.w.) of polyvinylpyrrolidone (PVP)-coated small-sized (10 nm) AgNPs. After exposure, blood and organs of all rats were analysed through biodistribution and accumulation of Ag, whereas the state of the liver and kidneys was evaluated by the levels of reactive oxygen species (ROS) and glutathione (GSH), catalase (CAT) activity, superoxide dismutase (SOD) and glutathione peroxidase (GPx), expression of metallothionein (Mt) genes and levels of Mt proteins. RESULTS: In all animals, changes in oxidative stress markers and blood parameters were observed indicating the toxicity of AgNPs applied orally even at low doses. Sex-related differences were noticed in all assessed parameters. While female rats eliminated AgNPs from the liver and kidneys more efficiently than males when treated with low doses, the opposite was observed for animals treated with higher doses of AgNPs. Female Wistar rats exposed to 1 mg PVP-coated AgNPs/kg b.w. accumulated two to three times more silver in the blood, liver, kidney and hearth than males, while the accumulation in most organs of digestive tract was more than ten times higher compared to males. Oxidative stress responses in the organs of males, except the liver of males treated with high doses, were less intense than in the organs of females. However, both Mt genes and Mt protein expression were significantly reduced after treatment in the liver and kidneys of males, while they remained unchanged in females. CONCLUSIONS: Observed toxicity effects of AgNPs in Wistar rats revealed sex-related differences in response to an oral 28 days repeated exposure.

In vitro study on the immunomodulatory effects of differently functionalized silver nanoparticles on human peripheral blood mononuclear cells.

The interaction of silver nanoparticles (AgNPs) with the immune system has not yet been sufficiently elucidated even though they belong to the most investigated and exploited group of nanomaterials. This study aimed to evaluate immunomodulatory effect of four different AgNPs on human peripheral blood mononuclear cells (hPBMCs). Fresh hPBMCs were exposed to the small sized (~ 10 nm) AgNPs immediately after isolation from the whole blood of healthy volunteers. The study considered coating-, time- and dose-dependent response of hPBMSc and stimulation of both early and intermediate activation of lymphocytes and monocytes using flow cytometry. The AgNPs differed in surface charge and were stabilised with polyvinyl pyrrolidone (PVP), poly-L-lysine (PLL), bis(2-ethylhexyl) sulfosuccinate sodium (AOT) or blood serum albumin (BSA). Response of hPBMCs to coating agents and ionic Ag form was evaluated to distinguish their effect from the AgNPs action as they may be released from the nanosurface. There was no significant effect of any tested AgNPs on relative count of hPBMCs subpopulations. The T-cells and monocytes were not activated after treatment with AgNPs, but the highest concentration of PLL- and BSA-AgNPs decreased density of CD4 and CD8 markers on T-helper and T-cytotoxic cells, respectively. The same AgNPs activated B- and NK-cells. Ionic Ag activated T-, B- and NK-cells, but at very higher concentration, whereas only PLL exhibited immunomodulatory activity. This study evidenced immunomodulatory activity of AgNPs that may be fine-tuned by the design of their surface functionalization.

Precipitation at Room Temperature as a Fast and Versatile Method for Calcium Phosphate/TiO2 Nanocomposites Synthesis.

The constantly growing need for advanced bone regeneration materials has motivated the development of calcium phosphates (CaPs) composites with a different metal or metal-oxide nanomaterials and their economical and environmentally friendly production. Here, two procedures for the synthesis of CaPs composites with TiO2 nanoplates (TiNPl) and nanowires (TiNWs) were tested, with the immersion of TiO2 nanomaterials (TiNMs) in corrected simulated body fluid (c-SBF) and precipitation of CaP in the presence of TiNMs. The materials obtained were analyzed by powder X-ray diffraction, spectroscopic and microscopic techniques, Brunauer-Emmett-Teller surface area analysis, thermogravimetric analysis, dynamic and electrophoretic light scattering, and their hemocompatibility and ability to induce reactive oxygen species were evaluated. After 28 days of immersion in c-SBF, no significant CaP coating was formed on TiNMs. However, the composites with calcium-deficient apatite (CaDHA) were obtained after one hour in the spontaneous precipitation system. In the absence of TiNMs, CaDHA was also formed, indicating that control of the CaP phase formed can be accomplished by fine-tuning conditions in the precipitation system. Although the morphology and size of crystalline domains of CaDHA obtained on the different nanomaterials differed, no significant difference was detected in their local structure. Composites showed low reactive oxygen species (ROS) production and did not induce hemolysis. The results obtained indicate that precipitation is a suitable and fast method for the preparation of CaPs/TiNMs nanocomposites which shows great potential for biomedical applications.

Fate and transformation of silver nanoparticles in different biological conditions.

The exploitation of silver nanoparticles (AgNPs) in biomedicine represents more than one third of their overall application. Despite their wide use and significant amount of scientific data on their effects on biological systems, detailed insight into their in vivo fate is still lacking. This study aimed to elucidate the biotransformation patterns of AgNPs following oral administration. Colloidal stability, biochemical transformation, dissolution, and degradation behaviour of different types of AgNPs were evaluated in systems modelled to represent biological environments relevant for oral administration, as well as in cell culture media and tissue compartments obtained from animal models. A multimethod approach was employed by implementing light scattering (dynamic and electrophoretic) techniques, spectroscopy (UV-vis, atomic absorption, nuclear magnetic resonance) and transmission electron microscopy. The obtained results demonstrated that AgNPs may transform very quickly during their journey through different biological conditions. They are able to degrade to an ionic form and again reconstruct to a nanoparticulate form, depending on the biological environment determined by specific body compartments. As suggested for other inorganic nanoparticles by other research groups, AgNPs fail to preserve their specific integrity in in vivo settings.

Interaction of Differently Sized, Shaped, and Functionalized Silver and Gold Nanoparticles with Glycosylated versus Nonglycosylated Transferrin.

Exposure of nanomaterials (NMs) to biological medium results in their direct interaction with biomolecules and the formation of a dynamic biomolecular layer known as the biomolecular corona. Despite numerous published data on nano-biointeractions, the role of protein glycosylation in the formation, characteristics, and fate of such nano-biocomplexes has been almost completely neglected, although most serum proteins are glycosylated. This study aimed to systematically investigate the differences in interaction of metallic NPs with glycosylated vs nonglycosylated transferrin. To reach this aim, we compared interaction mechanisms between differently sized, shaped, and surface-functionalized silver NMs and gold NMs to commercially available human transferrin (TRF), a glycosylated protein, and to its nonglycosylated recombinant form (ngTRF). Bovine serum albumin (BSA) was also included in the study for comparative purposes. Characterization of NMs was performed using transmission electron microscopy and dynamic and electrophoretic light scattering techniques. Fluorescence quenching and circular dichroism methods were used to evaluate protein binding constants on the nanosurface and conformational changes after the protein-NM interactions, respectively. Competitive binding of TRF, ngTRF, and BSA to the surface of different NMs was evaluated by separating them after extraction from protein corona by gel electrophoresis following quantification with a commercial protein assay. The results showed that the binding strength between NMs and transferrin and the changes in the secondary protein structure largely depend not only on NM physicochemical properties but also on the protein glycosylation status. Data gained by this study highlight the relevance of protein glycosylation for all future design, development, and efficacy and safety assessment of NMs.

Interaction of Differently Coated Silver Nanoparticles With Skin and Oral Mucosal Cells.

Silver nanoparticles (AgNP) can be found in different consumer products and various medical devices due to their excellent biocidal properties. Despite extensive scientific literature reporting biological effects of AgNP, there is still a lack of scientific evidence on how different surface functionalization affects AgNP interaction with the human skin and the oral epithelium. This study aimed to investigate biological consequences following the treatment of HaCaT and TR146 cells with AgNP stabilized with negatively charged sodium bis(2-ethylhexyl)-sulfosuccinate (AOT), neutral polyvinylpyrrolidone (PVP), and positively charged poly-l-lysine (PLL). All AgNP were characterized by means of size, shape and surface charge. Interactions with biological barriers were investigated in vitro by determining cell viability, particle uptake, oxidative stress response and DNA damages following AgNP treatment. Results showed a significant difference in cytotoxicity depending on the surface coating used for AgNP stabilization. All three types of AgNP induced apoptosis, oxidative stress response and DNA damages in cells, but AOT- and PVP-coated AgNP exhibited lower toxicity than positively charged PLL-AgNP. Considering the number of data gaps related to the safe use of nanomaterials in biomedicine, this study highlights the importance of particle surface functionalization that should be considered during design and development of future AgNP-based medical products.

Application of Localized Surface Plasmon Resonance Spectroscopy to Investigate a Nano-Bio Interface.

The accurate determination of events at the interface between a biological system and nanomaterials is necessary for efficacy and safety evaluation of novel nano-enabled medical products. Investigating the interaction of proteins with nanoparticles (NPs) and the formation of protein corona on nanosurfaces is particularly challenging from the methodological point of view due to the multiparametric complexity of such interactions. This study demonstrated the application of localized surface plasmon resonance (LSPR) spectroscopy as a low-cost and rapid biosensing technique that can be used in parallel with other sophisticated methods to monitor nano-bio interplay. Interaction of citrate-coated gold NPs (AuNPs) with human plasma proteins was selected as a case study to evaluate the applicability and value of scientific data acquired by LSPR as compared to fluorescence spectroscopy, which is one of the most used techniques to study NP interaction with biomolecules. LSPR results obtained for interaction of AuNPs with bovine serum albumin, glycosylated human transferrin, and non-glycosylated recombinant human transferrin correlated nicely with the adsorption constants obtained by fluorescence spectroscopy. This ability, complemented by its fast operation and reliability, makes the LSPR methodology an attractive option for the investigation of a nano-bio interface.

Sex-related response in mice after sub-acute intraperitoneal exposure to silver nanoparticles.

Silver nanoparticles (AgNPs) are among the most commercialized nanomaterials in biomedicine due to their antimicrobial and anti-inflammatory properties. Nevertheless, possible health hazards of exposure to AgNPs are yet to be understood and therefore raise public concern in regards of their safety. In this study, sex-related differences, role of steroidal hormones and influence of two different surface stabilizing agents (polymer vs. protein) on distribution and adverse effects of AgNPs were investigated in vivo. Intact and gonadectomised male and female mice were treated with seven AgNPs doses administered intraperitoneally during 21 days. After treatment, steroid hormone levels in serum, accumulation of Ag levels and oxidative stress biomarkers in liver, kidneys, brain and lungs were determined. Sex-related differences were observed in almost all tissues. Concentration of Ag was significantly higher in the liver of females compared to males. No significant difference was found for AgNP accumulation in lungs between females and males, while the lungs of intact males showed significantly higher Ag accumulation compared to gonadectomised group. Effect of surface coating was also observed, as Ag accumulation was significantly higher in kidneys and liver of intact females, as well as in kidneys and brain of intact males treated with protein-coated AgNPs compared to polymeric AgNPs. Oxidative stress response to AgNPs was the most pronounced in kidneys where protein-coated AgNPs induced stronger effects compared to polymeric AgNPs. Interestingly, protein-coated AgNPs reduced generation of reactive oxygen species in brains of females and gonadectomised males. Although there were no significant differences in levels of hormones in the AgNP-exposed animals compared to controls, sex-related differences in oxidative stress parameters were observed in all organs. Results of this study highlight the importance of including the sex-related differences and effects of protein corona in biosafety evaluation of AgNPs exposure.

Effect of differently coated silver nanoparticles on hemostasis.

With the emergence of nano-enabled medical devices (MDs) for the use in human medicine, ensuring their safety becomes of crucial importance. Hemocompatibility is one of the major criteria for approval of all MDs in contact with blood (e.g. vascular grafts, stents, or valves). Silver nanoparticles (AgNPs) are among the most used nanomaterials for MDs due to their biocidal activity; however, detailed knowledge on their hemostatic effects is still lacking.This study aimed to evaluate comprehensively AgNPs effects on hemostasis in human blood by exploiting combination of affordable and clinically relevant techniques.Differently stabilized AgNPs were prepared using sodium bis(2-ethylhexyl)sulphosuccinate (AOT), polyvinylpyrrolidone (PVP), poly-L-lysine (PLL), and bovine serum albumin (BSA) as coating agents. They were tested for hemolytic activity, induction of platelet aggregation, plasmatic coagulation, thrombin generation, and hemostasis in whole blood.All AgNPs were found to cause dose-dependent hemolysis. The BSA-, AOT-, and PVP-coated AgNPs delayed plasmatic coagulation, while only PLL-AgNPs inhibited plasmatic coagulation, induced platelet activation, and interfered with hemostasis by delaying clotting time and decreasing clot firmness in whole blood.Obtained results demonstrate that a combination of different techniques should be used for reliable assessment of AgNPs hemostatic effects highlighting the need for a standardized approach in sampling and experimental protocols.

Interaction of silver nanoparticles with plasma transport proteins: A systematic study on impacts of particle size, shape and surface functionalization.

Metallic nanoparticles are an important and widely used materials in development of nano-enabled medicine. For that reason, their interaction with biological molecules has to be systematically examined, as use of nanoparticles can lead to altered biological functions. In this study, we evaluated the interaction between silver nanoparticles (AgNPs) and two important plasma transport proteins - albumin and α-1-acid glycoprotein. To investigate comprehensively how different physico-chemical properties impact interaction of proteins with nanosurface, AgNPs of different size, shape and surface coating was prepared. The study was conducted using UV-Vis absorption, fluorescence, inductively coupled plasma mass spectrometry, circular dichroism spectroscopy, transmission electron microscopy, dynamic and electrophoretic light scattering techniques. The results showed significant complexities of the nano-bio interface and binding affinities of proteins onto surface of different AgNPs, which were affected by both AgNPs and protein properties. The most significant role on AgNPs-protein interaction had the coating agents used for AgNPs surface stabilization. Our findings should improve safe-by-design approach to development of the metallic nanomaterials for medical use.

Impact of surface functionalization on the toxicity and antimicrobial effects of selenium nanoparticles considering different routes of entry.

Selenium nanoparticles (SeNPs) were first designed as nutritional supplements, but they are attractive also for use in diagnostic and therapeutic systems owing to their biocompatibility and protective effects. This study aimed to examine if different SeNPs stabilization strategies affect their (i) antimicrobial activity against bacteria Escherichia coli and Staphylococcus aureus and yeast Saccharomyces cerevisiae and (ii) toxicity to human cells of different biological barriers i.e., skin, oral and intestinal mucosa. For surface stabilization, polyvinylpyrrolidone (PVP), poly-L-lysine (PLL) and polyacrylic acid (PAA) were used rendering neutral, positively and negatively charged SeNPs, respectively. The SeNPs (primary size ~80 nm) showed toxic effects in human cells in vitro and in bacteria S. aureus, but not in E. coli and yeast S. cerevisiae. Toxicity of SeNPs (24 h IC50) ranged from 1.4 to >100 mg Se/L, depending on surface functionalization (PLL > PAA > PVP) and was not caused by ionic Se. At subtoxic concentrations, all SeNPs were taken up by all human cell types, induced oxidative stress response and demonstrated genotoxicity. As the safety profile of SeNPs was dependent not only on target cells (mammalian cells, bacteria, yeast), but also on surface functionalization, these aspects should be considered during development of novel SeNPs-based biomedical products.

Association between arsenic exposure and biomarkers of type 2 diabetes mellitus in a Croatian population: A comparative observational pilot study.

Chronic exposure to high inorganic As levels in drinking water has been related to many diseases, including type 2 diabetes mellitus (T2D). The association with low and moderate As levels, however, remains controversial and has yet not been studied in European populations. This study aimed to investigate possible association between As exposure and biomarkers of T2D in Croatian population. Observation recruited 86 adults from Eastern Croatia, where groundwater is contaminated with inorganic As, and 116 adults from Western Croatia, where As levels in drinking water are low. Both populations were divided in patient groups (T2D or prediabetes) and healthy controls. Exposure was assessed by determining total As in blood and urine and As metabolites in urine. Eastern Croatian population had a significantly higher content of As in urine than Western, whereas the opposite was true for arsenobetain. Total As and As metabolites in urine positively correlated with hemoglobin A1c (HbA1c) and negatively with albuminuria. This study provides important preliminary data on the levels of As in urine and blood and their association with biomarkers of T2D in Croatian population exposed to low or moderate levels of As through drinking water as a solid basis for further research of the pathophysiological effects of such As exposure on the status and complications of diabetes.

Stability and toxicity of differently coated selenium nanoparticles under model environmental exposure settings.

This study, motivated to fill the knowledge gap on environmental safety of selenium nanoparticles (SeNPs), provides information on the stability and environmental safety of four differently coated SeNPs rendering both positive and negative surface charges. The stability and dissolution behaviour of SeNPs were determined in an aquatic model media of different ionic strength to provide information regarding the environmental fate of SeNPs in different environmental conditions. The environmental safety of SeNPs was evaluated by acute regulatory toxicity tests using Daphina magna and Vibrio fischeri as model organisms. Agglomeration was observed for all studied SeNPs in test media with higher ionic strength caused by the disruption of surface charge leading to electrostatic instability. Toxicity of SeNPs on both aquatic species was dose-dependent and increased with exposure time. The obtained data indicated that all of the tested SeNPs could be classified as harmful to the natural bacteria V. fischeri and harmful to toxic to crustaceans D. magna, but dependent on the coating agent used for SeNPs stabilization. Although SeNPs have attracted great interest for use in biomedicine, this study demonstrated that their ecotoxicological effects should be considered during the design of new of SeNPs-based products.

Antifungal activities of silver and selenium nanoparticles stabilized with different surface coating agents.

BACKGROUND: Extensive and growing use of different chemical pesticides that affect both the environment and human health raises a need for new and more suitable methods to deal with plant pathogens. Nanotechnology has enabled the use of materials at the nanoscale with exceptional functionality in different economic domains including agricultural production. This study aimed to evaluate antifungal potential of selenium nanoparticles (SeNPs) and silver nanoparticles (AgNPs) stabilized with different surface coatings and characterized by different surface charge on plant pathogenic fungi Macrophomina phaseolina, Sclerotinia sclerotiorum and Diaporthe longicolla. RESULTS: AgNPs were coated with three different stabilizing agents: mono citrate (MC-AgNPs), cetyltrimethyl ammonium bromide (CTAB-AgNPs) and polyvinylpyrrolidon (PVP-AgNPs). SeNPs were coated with poly-l-lysine (PLL-SeNPs), polyacrylic acid (PAA-SeNPs), and polyvinylpyrrolidon (PVP-SeNPs). Seven different concentrations (0.1, 0.5, 1, 5, 10, 50 and 100 mg L-1 ) of nanoparticles were applied. All AgNPs and SeNPs significantly inhibited the growth of the tested fungi. Among the tested NPs, PVP-AgNPs showed the best inhibitory effect on the tested plant pathogenic fungi, especially against S. sclerotiorum. The similar inhibition of the sclerotia formation was observed for S. sclerotiorum treated with PLL-SeNPs. CONCLUSION: Obtained results provides new insights on fungicide effect of AgNPs and SeNPs stabilized with different coating agents on different plant pathogens. Further work should focus on detailed risk/benefit ratio assessment of using SeNPs or AgNPs in agriculture taking into account whole agroecosystem. © 2020 Society of Chemical Industry.