Article

Evaluation of hydroxyapatite nanoparticles - induced in vivo toxicity in Drosophila melanogaster

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Abstract

Hydroxyapatite (HAp), a compound similar in composition to human hard tissues such as bone and teeth, has widespread applications in bulk as well as nanoparticle form in numerous fields due to excellent biocompatibility and stability. The increased use of this nanoparticle thus warrants the study of its toxic potential in in vivo models. In this study, the toxicity induced by HAp NPs administered via the oral route using Drosophila as an in vivo model was investigated. Biochemical assays such as lipid peroxidation and nitro blue tetrazolium assay did not reveal toxicity at lowest (10 μg/ml) as well as highest (1000 μg/ml) doses. The HAp nanoparticle-treated flies and larvae did not exhibit signs of behavioral change as evidenced from the negative geotaxis assay and larvae crawling assay, respectively. Survival assay to assess the chronic toxicity of HAp nanoparticles also did not reveal any signs of toxicity at both lowest and highest doses. This demonstrates that hydroxyapatite nanoparticles do not elicit significant in vivo toxicity. Considering the paucity of studies available to support the non-toxic effect of HAp nanoparticles, further investigations are warranted in Drosophila and other in vivo models. Moreover, such HAp nanoparticles can also serve as reliable carrier for drug delivery system.

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... The results showed that chronic exposure of the nanoparticles can lead to behavioral effects, phenotypic abnormalities in progeny flies, absence of appendages, expression of proteins related to alterations in the morphogenesis of the digestive tract, striated differentiation of muscle cells, chromatin organization regulation, phototransduction, and unwinding of DNA duplex (Anand et al., 2019). Dan et al. (2019) assessed the in vivo toxicity of hydroxyapatite nanoparticles using Drosophila and the result revealed that they are nontoxic even at high concentration and did not affect their climbing and crawling patterns in larvae and adult fruit flies (Dan et al., 2019). All the studies showed that Drosophila can be a potential in vivo model to evaluate the toxicity of nanomedicines for diabetes therapy. ...
... The results showed that chronic exposure of the nanoparticles can lead to behavioral effects, phenotypic abnormalities in progeny flies, absence of appendages, expression of proteins related to alterations in the morphogenesis of the digestive tract, striated differentiation of muscle cells, chromatin organization regulation, phototransduction, and unwinding of DNA duplex (Anand et al., 2019). Dan et al. (2019) assessed the in vivo toxicity of hydroxyapatite nanoparticles using Drosophila and the result revealed that they are nontoxic even at high concentration and did not affect their climbing and crawling patterns in larvae and adult fruit flies (Dan et al., 2019). All the studies showed that Drosophila can be a potential in vivo model to evaluate the toxicity of nanomedicines for diabetes therapy. ...
Chapter
In vivo toxicological analyses are crucial in demonstrating the efficacy of drugs in the treatment of diseases. Animal models are commonly used for in vivo toxicological and efficacy analysis of drugs. Nanoparticles-based drug formulations or nanomedicines are characterized with varying degrees of toxicity to target and/or nontarget cells in animal models. Hence, it is necessary to comprehensively evaluate nanomedicines to assess their toxicology and therapeutic indices in specific animal models as part of the drug discovery pipeline. Various animals have been used as live models for in vivo assessment of nanomedicines. This chapter discusses different types of in vivo animal models that are used to evaluate toxicity and efficacy of nanomedicines in diabetes treatment.
... In Drosophila melanogaster new born flies presented toxicity symptoms such as imperceptible movement and abnormal wing and bristle phenotypes after treatment with zinc oxide. 36,37 Barik & Mirsha 38 studying the effects of silver nanoparticles (AgNPs) in Drosophila concluded that the NPs distresses larva to pupa and pupa to adult transitions. Overlapping of both species developmental stages along with magnetite damage made difficult to recognize if a larvae or a pupa corresponds to C. capitata or to A. fraterculus (Figures 3&4). ...
... 2-The delay in growth and development, high necrosis and mortality of the larvae, along with phenotypic alterations such as abnormalities of wings and bristles, and the alteration and interruption of the life cycle, are in tune with the changes observed in model species such as Drosophila melanogaster, when exposed to nanoparticles. [36][37][38][39][40] ...
... HApNP produces ROS, which was detected by NBT assay [99]. This study was supported by Dan et al. [100], who also finds more amounts of ROS and after HApNP treatment. Besides ROS, more amount of lipid peroxidation was also observed after the oral intake of HApNP treatment [100]. ...
... This study was supported by Dan et al. [100], who also finds more amounts of ROS and after HApNP treatment. Besides ROS, more amount of lipid peroxidation was also observed after the oral intake of HApNP treatment [100]. Oral administration of nCeO 2 to Drosophila increases the production of oxidative stress or ROS within the body [101]. ...
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... In this context, non-mammalian animals emerge as an option, since they present a series of advantages, such as low cost, easy handling, small size and short life span. Among these, Acanthamoeba castellanii, Dictyostellium discoideum, Caenorhabditis elegans, Drosophila melanogaster, Galleria mellonella and Danio rerio have been increasingly employed to study various human diseases, as well as the toxicity and efficiency of new chemical substances [15][16][17][18]. ...
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... The most stable and biocompatible form of calcium phosphate is HAp with the chemical formula of [Ca 10 (PO 4 ) 6 (OH) 2 ] which is the main human bone mineral constituent [15]. It has promising properties for utilization in orthopedic surgeries and dental implants due to its high biocompatibility [16], osteoconductivity [17], bioactivity [18], non-inflammatory, non-immunogenic, and non-toxic nature [19,20]. According to previous research on alkaline phosphatase activity, HAp derived from natural resources could be effectively deployed for cell proliferation and differentiation enhancement [21,22]. ...
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... Furthermore, the toxicity of various nanoparticles was analysed using Drosophila as a model (Pompa et al., 2011;Posgai et al., 2011;Vales et al., 2013). HApNP toxicity was recently investigated in Drosophila using a concentration of 10-1000 mg l −1 (Dan et al., 2019). The metabolism of HAp is conserved for Drosophila and human being (Zhao et al., 2013). ...
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... The glycogen content in flies exposed to different concentrations of nCeO 2 for a period of one month was estimated according to a previously described protocol [49]. Briefly, 5 flies were homogenized in 500 μL of 2% sodium sulfate. ...
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Abstract Drosophila melanogaster has been used as an in vivo model organism for the study of genetics and development since 100 years ago. Recently, the fruit fly Drosophila was also developed as an in vivo model organism for toxicology studies, in particular, the field of nanotoxicity. The incorporation of nanomaterials into consumer and biomedical products is a cause for concern as nanomaterials are often associated with toxicity in many in vitro studies. In vivo animal studies of the toxicity of nanomaterials with rodents and other mammals are, however, limited due to high operational cost and ethical objections. Hence, Drosophila, a genetically tractable organism with distinct developmental stages and short life cycle, serves as an ideal organism to study nanomaterial-mediated toxicity. This review discusses the basic biology of Drosophila, the toxicity of nanomaterials, as well as how the Drosophila model can be used to study the toxicity of various types of nanomaterials.
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Nanomaterials are at the leading edge of the rapidly developing field of nanotechnology. Their unique size-dependent properties make these materials superior and indispensable in many areas of human activity. This brief review tries to summarise the most recent developments in the field of applied nanomaterials, in particular their application in biology and medicine, and discusses their commercialisation prospects.
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The present study refers to the preparation and characterization of porous hydroxyapatite scaffolds to be used as matrices for bone regeneration or as specific release vehicles. Ceramics are widely used for bone tissue engineering purposes and in this study, hydroxyapatite porous scaffolds were produced using the polymer replication method. Polyurethane sponges were used as templates and impregnated with a ceramic slurry at different ratios, and sintered at 1300°C following a specific thermal cycle. The characteristics of the hydroxyapatite porous scaffolds and respective powder used as starting material, were investigated by using scanning electron microscopy, particle size distribution, X-ray diffraction, Fourier transformed infrared spectroscopy and compressive mechanical testing techniques. It was possible to produce highly porous hydroxyapatite scaffolds presenting micro and macropores and pore interconnectivity.
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Although the toxicity/biocompatibility of hydroxyapatite nanoparticles (nano HA), a prospective nano biomaterial is extensively studied, its interaction on biological systems following chronic exposure is less exploited. In the present study, Wistar rats were given various concentrations of nano HA in the diet to determine the chronic toxicity and potential carcinogenicity. Altogether 140 rats were used for the study under various administration dosages along with control. The animals were sacrificed after 12 months of controlled continuous dosing. All in-life parameters, including body weight, food consumption, clinical observations, survival, biochemical and hematology, were unaffected by the chronic exposure of nano HA orally. Similarly, gross and histopathological evaluation was also unchanged following exposure to nano HA. No evidence of nano HA-related lesions or Nano HA-induced neoplasia was suggested in this rodent bioassay study.
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In this study, porous scaffolds were produced by a thermal crosslinking of polycaprolactone diacrylate in the presence of hydroxyapatite (HA) and particulate leaching technique with sodium chloride as the water soluble porogen for bone tissue engineering applications. The prepared scaffolds were characterized using techniques such as Field Emission Scanning Electron Microscopy, Differential Scanning Calorimetry, and Attenuated Total Reflectance-Fourier Transform Infrared Spectroscopy. Moreover, dynamic mechanical properties were investigated using Dynamic Mechanical Thermal Analysis. The obtained scaffolds present a porous structure with interconnected pores and porosity around 73%. It was found that the incorporation of HA particles to polycaprolactone (PCL) matrix resulted in an increased crystallinity. Moreover, both the storage modulus (E’) and glass transition temperature (Tg) increased, while the loss factor (tan δ) decreased due to the hindrance of the HA particles to the mobility of polymer segments. Cytocompatability of the scaffolds was assessed by MTT assay and cell attachment studies. Osteoconductivity of the scaffolds was investigated with cells alkaline phosphatase extraction. The levels of alkaline phosphatase activity were found to be higher for PCL/HA network scaffold than for PCL network scaffold. In addition, cytocompatibility of the PCL/HA network scaffold indicated no toxicity, and cells were attached and spread to the scaffold walls.
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The objective of this study was to improve the therapeutic efficacy of methylprednisolone acetate (MPA) in the treatment of rheumatoid arthritis (RA) by incorporating the drug into the hydroxyapatite (HAp) nanoparticles. The nanoparticles were synthesized using chemical precipitation technique and their size and morphology were evaluated by dynamic light scattering and scanning electron microscopy (SEM). The solid-state behavior of the nanoparticles was also characterized operating X-ray powder diffraction (XRPD), differential scanning calorimetry (DSC) and Fourier-transform infrared spectroscopy (FTIR). The Brunauer–Emmett–Teller and Barrett–Joyner–Halenda N2 adsorption/desorption analyses were also performed to determine the surface area, Vm (the volume of the N2 adsorbed on the one gram of the HAp when the monolayer is complete) and the pore size of the samples. Furthermore, the therapeutic efficacy of the prepared nanoformulation on the adjuvant induced arthritic rats was assessed. HAp mesoporous nanoparticles with a particle size of 70.45 nm, pore size of 2.71 nm and drug loading of 44.53% were obtained. The specific surface area of HAp as well as the Vm values were decreased after the drug loading process. The nanoformulation revealed the slower drug release profile compared to the pure drug. The MTT assay indicated that the MPA-loaded nanoparticles had lower cytotoxic effect on NIH-3 T3 and CAOV-4 cell lines compared to the pure drug. Interestingly, the in vivo study confirmed that the drug-loaded nanoparticles could considerably decrease the paw volume and normalize the haematological abnormalities in the arthritic rats.
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Despite being a relatively new field, nanoscience has been in forefront among many scientific areas. Nanoparticle materials (NM) present interesting physicochemical characteristics not necessarily found in their bulky forms, and alterations in their size or coating markedly modifies their physical, chemical, and biological properties. Due to these novel properties there is a general trend to exploit these NM in several fields of science, particularly in medicine and industry. The increased presence of NM in the environment warrants evaluation of potential harmful effects in order to protect both environment and human exposed populations. Although in vitro approaches are commonly used to determine potential adverse effects of NM, in vivo studies generate data expected to be more relevant for risk assessment. As an in vivo model Drosophila melanogaster was previously found to possess reliable utility in determining the biological effects of NM, and thus its usage increased markedly over the last few years. The aim of this review was to present a comprehensive overview of all apparent studies carried out with NM and Drosophila, to attain a clear and comprehensive picture of the potential risk of NM exposure to health, and demonstrate the advantages of using Drosophila in nanotoxicological investigations.
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In the present study, copper-doped ZnO nanoparticles (doped ZnO NPs Cu) were synthesized, characterized and evaluated for their possible toxic effects in Drosophila melanogaster (Oregon R). X-ray diffraction, Fourier transform infrared spectroscopy, scanning electron microscopy and energy dispersive X-ray spectrometry confirm the formation of doped ZnO NPs Cu. Doped ZnO NPs Cu (3%) were mixed in the diet at final concentrations of 1, 2, 4 and 8 µg/µl. The starved male flies were allowed to feed on it for 4 days. After completion of the desired duration, climbing ability, activity pattern, activity of acetylcholinesterase (AChE), glutathione (GSH), glutathione-S-transferase (GST), lipid peroxidation (LPO), total protein content and caspases were studied. SDS-PAGE was also performed for whole fly homogenate of control as well as treated flies. No loss in the climbing and activity pattern was observed at the selected doses of doped ZnO NPs Cu. No significant change in the levels of AChE, GSH, GST, LPO, caspase 9/3 and total protein content was observed. The brain sections showed no gross changes in the structure and SDS-PAGE patterns also revealed no change in the protein expression. The results suggest that doped ZnO NPs Cu are non-toxic at 1, 2, 4 and 8 µg/µl of concentration in D. melanogaster.
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Photodynamic therapy (PDT) involves the cellular uptake of a photosensitizer (PS) combined with oxygen molecules and light at a specific wavelength to be able to trigger cancer cell death via the apoptosis pathway, which is less harmful and has less inflammatory side effect than necrosis. However, the traditional PDT treatment has two main deficiencies: the dark toxicity of the PS and the poor selectivity of the cellular uptake of PS between the target cells and normal tissues. In this work, methylene blue (MB), a known effective PS, combined with Au nanoparticles (NPs) was prepared using an intermolecular interaction between a polystyrene-alt-maleic acid (PSMA) layer on the Au NPs and MB. The Au@polymer/MB NPs produced a high quantum yield of singlet oxygen molecules, over 50% as much as that of free MB, when they were excited by a dark red light source at 660 nm, but without significant dark toxicity. Furthermore, transferrin (Tf) was conjugated on the Au@polymer/MB NPs via an EDC/NHS reaction to enhance the selectivity to HeLa cells compared to 3T3 fibroblasts. With a hand-held single laser treatment (32 mW/cm) for 4 min, the new Au@polymer/MB-Tf NPs showed a two-fold enhancement of PDT efficiency toward HeLa cells over the use of free MB at 4 times dosage. Cellular staining examinations showed that the HeLa cells reacted with Au@polymer/MB-Tf NPs and the 660 nm-light excitation triggered PDT, which caused the cells to undergo apoptosis (''programmed'' cell death). We propose that applying this therapeutic Au@polymer/MB-Tf nanoagent is facile and safe for delivery and cancer cell targeting to simultaneously minimize side effects and accomplish a significant enhancement of the photodynamic therapeutic efficiency towards next-generation nanomedicine development.
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Nanocrystalline calcium phosphate based bioceramics are the new rage in biomaterials research. Conventionally, calcium phosphates based materials are preferred as bone grafts in hard tissue engineering because of their superior biocompatibility and bioactivity. However, this group of bioceramics exhibits poor mechanical performance, which restricts their uses in load bearing applications. The recent trend in bioceramic research is mainly concentrated on bioactive and bioresorbable ceramics, i.e. hydroxyapatite, bioactive glasses, tricalcium phosphates and biphasic calcium phosphates as they exhibit superior biological properties over other materials. In recent times, the arena of nanotechnology has been extensively studied by various researchers to overcome the existing limitations of calcium phosphates, mainly hydroxyapatite, as well as to fabricate nanostructured scaffolds to mimic structural and dimensional details of natural bone. The bone mineral consists of tiny HAp crystals in the nano-regime. It is found that nanocrystalline HAp powders improve sinterability and densification due to greater surface area, which could improve the fracture toughness and other mechanical properties. Nano-HAp is also expected to have better bioactivity than coarser crystals. Nanocrystalline calcium phosphate has the potential to revolutionize the field of hard tissue engineering from bone repair and augmentation to controlled drug delivery devices. This paper reviews the current state of knowledge and recent developments of various nanocrystalline calcium phosphate based bioceramics from synthesis to characterization.
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Mesenchymal stem cells or multipotent progenitor cells isolated from bone marrow presents close resemblance to the natural in vivo milieu and hence preferred more than the conventional cell culture systems to predict the toxicological behavior of bio-nano interaction. The objective of the present study is to evaluate the molecular toxicity of hydroxyapatite nanoparticles (HANPs) using mouse bone marrow mesenchymal stem cells (BMSCs). In-house synthesized HANPs (50nm) were used to study the cytotoxicity, nano particle uptake, effect on cyto skeletal arrangement, oxidative stress response and apoptotic behavior with the confluent BMSCs as per standard protocols. The results of the MTT assay indicated that HANPs does not induce cytotoxicity up to 800μg/mL. It was also observed that oxidative stress related apoptosis and reactive oxygen species (ROS) production following nanoparticle treatment was similar to that of control (cells without treatment). Hence it can be concluded that the in-house synthesized HANPs are non-toxic/safe at the molecular level suggesting that the HANPs are compatible to BMSCs. Further, the in vitro BMSCs cell culture can be used as a model for evaluating the preliminary toxicity of nanomaterials.
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Methylene blue (MB) has been shown to slow down the progression of the Alzheimer's disease (AD) and other tauopathies; however distribution of MB into the brain is limited due its high hydrophilicity. In this study, we aimed to prepare a novel hydrophobic glutathione coated PLGA nanoparticlesto improve bioavailability of MB in the brain. Glutathione coated poly-(lactide-co-glycolide) (PLGA-b-PEG) nanoparticles (NPs) were prepared and tested in two different cell culture models of AD expressing microtubule associated protein tau (tau). The NPs showed a particle size averaging 136.5±4.4nm, which is suitable for the blood brain barrier (BBB) permeation. The in vitro release profile of the NPs exhibited no initial burst release and showed sustained drug release for up to 144 hours.Interestingly, treatment of newly formulated MB-NPs showed apotent reduction in both endogenous and overexpressed tau protein levels in human neuroblastoma SHSY-5Y cells expressing endogenous tau and transfected HeLa cells over-expressing tau protein, respectively. Furthermore, In vitro BBB TranswellTMstudy showed significantly higher permeation of MB-NP compared to the MB solution through the coculture of rat brain endothelial 4 (RBE4) and C6 astrocytoma cells (p<0.05).The proposed MB loaded nanoparticles could provide a more effective treatment option for AD and many other related disorders.
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In this study, novel polycaprolactone/hydroxyapatite (PCL/HA) scaffolds were prepared to increase mechanical properties and degradation of PCL/HA ones for bone tissue engineering. PCL macromers were synthesized through the reaction of PCL diol (Mn: 530, 1250, and 2000) and PCL triol (Mn: 900) with acryloyl chloride and confirmed using nuclear magnetic resonance spectrometer (NMR) and fourier transform infrared (FTIR). The PCL/HA scaffolds were prepared by cross-linking of PCL macromer in the presence of HA by UV treatment and freeze drying methods. Mechanical property and porosity as well as degradability of the PCL/HA scaffolds were also investigated. PCL/HA scaffolds showed faster degradation and higher compressive modulus than those of PCL itself due to their low crystallinity and modification of terminal groups. The pore morphology and pore sizes of the PCL/HA scaffold were checked by scanning electron microscope (SEM). Cell cytotoxicity and proliferation of MG-63 osteoblast cultured onto the PCL/HA scaffold was assessed by lactate dehydrogenase (LDH) assay and Alamar blue assay, respectively. The novel PCL/HA scaffold appears to be suitable for bone substitutes.
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Natural bone is a kind of nanocomposites composed of orientated hydroxyapatite (HAp) along c-axis and fibrous collagen. Therefore. composites exhibiting composition and structure analogous to those of natural bone have been expected to be useful bone Substitute materials. Organic polymer-HAp composites have been attracted Much attention since they have interesting features such as bone-bonding ability, i.e. bioactivity and flexibility. In the present study, chitosan-HAp nanocomposites were prepared through mechanochemical reaction using conventional ball mill and Subsequent aging. The obtained composites contained carbonate-containing, HAp, and HAp nanocrystals in the composites aged at 25 C for 24 h showed a needle-like Structure. They can form bone-like HAp on their Surfaces after soaking in simulated body fluid (SBF), indicating potential for bioactivity in living body. The prepared chitosan-HAp composites are expected to be one of the useful bone Substitute materials.
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Targeted drug delivery using nanocrystalline materials delivers the drug at the deseased site. This increases the efficacy of the drug in killing the cancer cells. Surface modifications were done to target the drug to a particular receptor on the cell surface. This paper reports synthesis of hydroxyapatite and titanium dioxide nanoparticles and modification of their surface with polyethylene glycol (PEG) followed by folic acid (FA). Paclitaxel, an anticancer drug, is attached to functionalized hydroxyapatite and titanium dioxide nanoparticles. The pure and functionalised nanoparticles are characterised with XRD, TEM and UV spectroscopy. Anticancer analysis was carried out in DEN induced hepatocarcinoma animals. Biochemical, hematological and histopathological analysis show that the surface modified paclitaxel attached nanoparticles have an higher anticancer activity than the pure paclitaxel and surface modified nanoparticles without paclitaxel. This is due to the targeting of the drug to the folate receptor in the cancer cells.
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Acrylamide (ACR) intoxication in its monomeric form leads to neuronal damage in both experimental animals and humans. Oxidative stress is one of the principle mechanisms related to the neurotoxicity of ACR exposure. Hence, the present study aimed to recapitulate the potential of ACR to cause oxidative stress and neurotoxic effects in Drosophila melanogaster. Exposure of adult male flies (Oregon K strain) to ACR (1-10mM, 7 d) in the diet resulted in a concentration and time dependent mortality, while the survivors exhibited significant locomotor deficits. Further, ACR exposure (1-5mM, 3 d) caused robust oxidative stress as evidenced by markedly elevated levels of reactive oxygen species and hypdroperoxides in head/body regions. Enhanced lipid peroxidation, perturbations in the activities of antioxidant enzymes accompanied with depletion of reduced glutathione levels in head region at high concentrations suggested induction of oxidative stress. Further, marked diminution in the activities of complexes I-III, Succinic dehydrogenase, with concomitant reduction in MTT suggested the propensity of ACR to impair mitochondrial function. Furthermore, ACR-induced neurotoxic effects were discernible in terms of diminished ATPase activity, enhanced activity of acetylcholinesterase and dopamine depletion. In a satellite study, employing a co-exposure paradigm, we tested the propensity of spice actives namely eugenol (EU) and isoeugenol (IE) to ameliorate ACR-induced neurotoxicity. EU/IE enriched diet offered marked protection against ACR-induced mortality, locomotor dysfunctions and oxidative stress. Furthermore, the spice actives prevented the depletion of reduced GSH levels, maintained the activity of AChE enzyme and dopamine levels in head region. Collectively, these findings clearly demonstrate that ACR induced neurotoxicity in Drosophila may be mediated through oxidative stress mechanisms and the potential of spice actives to abrogate the condition. These data suggest that Drosophila may serve as a suitable model to understand the possible mechanism/s associated with ACR associated neuropathy.
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A force field for the modeling of hydroxyapatite; Ca10(PO4)6OH2 (HAP) is established based upon transferable potentials. Ca–O, P–O and O–O potentials were transported from those previously published for fluorapatite and based on single crystal experimental data. The interactions of hydroxyl oxygen with calcium and phosphate were re-scaled by fitting to experimental data for CaO and AlPO4, respectively, to account for the reduction in the oxygen charge from −2.0 to −1.426. Force field accuracy is tested by comparing the calculated and experimental values for the cell constant and atom positions in the unit cell. The elastic constants and bulk modulus calculated for HAP are in close agreement with the experimental results. The potentials were also used to calculate the compressibility data of HAP and fluorapatite, and these results also agree with the published experimental data. Using formal charges for metal cations allows modeling the complete solid solution of Cd–Ca hydroxyapatite with a good accuracy.
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This Full Paper investigates the adsorption and desorption of the anticancer drugs cis-diammmedichloroplatmum(II) (CDDP, cisplatin) and the new platinum(II) complex di(ethylenediamineplatinum)medronate (DPM), as well as the clinically relevant bisphosphonate alendronate, towards two biomimetic synthetic HA nanocrystalline materials with either plate-shaped (HAps) or needle-shaped (HAns) morphologies and different chemico-physical properties. The adsorption and desorption kinetics are dependent on the specific properties of the drugs and the morphology of the HA nanoparticles. Adsorption of the platinum complexes occurs with retention of the nitrogen ligands but the chloride ligands of cisplatin are displaced. Despite their opposite charges, the negatively charged alendronate bisphosphonate and the positively charged aquated cisplatin are strongly adsorbed, while the neutral DPM complex shows lower affinity towards the negatively charged apatitic surface. The data suggest that adsorption of the two platinum complexes is driven by electrostatic attractions, while interaction between the alendronate and the HA surface takes place by ligand exchange in which the two phosphonate groups of the drug molecule replace two surface phosphate groups. Significantly, adsorption of positively charged hydrolysis species of cisplatin is more favored on the phosphate-rich HAns surface while adsorption of negatively charged alendronate is more favored on the calcium-rich HAps surface. The latter type of short-range electrostatic interactions also appear to dominate the desorption kinetics; consequently, drug release is greater for neutral DPM than for charged alendronate and aquated cisplatin. Moreover, while the release per unit area of charged species is the same for the two types of HAs, the release of DPM is faster from HAns, which is lower in surface calcium, than for HAps. Overall, this work demonstrates that the properties of HA nanocrystals can be modulated in such a way to produce HA/biomolecule conjugates tailored for specific therapeutic applications.
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In bone disorders infections are common. The concentration of majority of antibiotics is very low in the bone tissue. A high local dose can be obtained from the ciprofloxacin-loaded hydroxyapatite nanoparticles. The present study is aimed at developing the use of hydroxyapatite and zinc-doped hydroxyapatite nanoparticles as a carrier for ciprofloxacin drug delivery system. The ciprofloxacin-loaded hydroxyapatite and zinc-doped hydroxyapatite have a good antibacterial activity against Pseudomonas aeruginosa and Staphylococcus aureus. Hydroxyapatite and zinc-doped hydroxyapatite were prepared and characterized using X-ray diffraction, Transmission electron microscopy and inductively coupled plasma optical emission spectrometry. They were loaded with ciprofloxacin using optimized drug loading parameters. Drug loading, in vitro drug release and antimicrobial activity were analyzed. The influence of zinc on the controlled release of ciprofloxacin was analyzed. The results show that the presence of zinc increases the drug release percentage and that the drug was released in a controlled manner.
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The use of invertebrate model hosts has increased in popularity due to numerous advantages of invertebrates over mammalian models, including ethical, logistical and budgetary features. This review provides an introduction to three model hosts, the nematode Caenorhabditis elegans, the fruit fly Drosophila melanogaster and the larvae of Galleria mellonella, the greater wax moth. It highlights principal experimental advantages of each model, for C. elegans the ability to run high-throughput assays, for D. melanogaster the evolutionarily conserved innate immune response, and for G. mellonella the ability to conduct experiments at 37°C and easily inoculate a precise quantity of pathogen. It additionally discusses recent research that has been conducted with each host to identify pathogen virulence factors, study the immune response, and evaluate potential antimicrobial compounds, focusing principally on fungal pathogens.
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In order to find a new way to slow down the release of drugs and to solve the burst release problem of drugs from traditionally used hydrogel matrices, a series of novel pH-sensitive sodium alginate/hydroxyapatite (SA/HA) nanocomposite beads was prepared by the in situ generation of HA micro-particles in the beads during the sol-gel transition process of SA. The SA/HA nanocomposites were characterized by Fourier transform IR spectroscopy, X-ray fluorescence spectrometry, scanning electron microscopy and field emission SEM in order to reveal their composition and surface morphology as well as the role that the in situ generated HA micro-particles play. The factors influencing the swelling behavior, drug loading and controlled release behavior of the SA/HA nanocomposite beads were also investigated using diclofenac sodium (DS) as the model drug. The HA micro-particles act as inorganic crosslinkers in the nanocomposites, which could contract and restrict the movability of the SA polymer chains, and then change the surface morphology and decrease the swell ratio. Meanwhile, the entrapment efficiency of DS was improved, and the burst release of DS was overcome. The factors (including concentration of Ca(2+), reaction time and temperature) affecting the growth of HA micro-particles have a clear influence on the entrapment efficiency and release rate of DS. In this work, the nanocomposite beads prepared under optimum condition could prolong the release of DS for 8h more compared with the pristine SA hydrogel beads.
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The reaction of lipid peroxides in animal tissues with thiobarbituric acid was dependent on pH of the reaction mixture as was the case for linoleic acid hydroperoxide. The optimum pH was found to be 3.5. Taking this fact into consideration, a standard procedure for the assay of lipid peroxide level in animal tissues by their reaction with thiobarbituric acid was developed as follows. Ten percent ( tissue homogenate was mixed with sodium dodecyl sulfate, acetate buffer (pH 3.5), and aqueous solution of thiobarbituric acid. After heating at 95°C for 60 min, the red pigment produced was extracted with n-butanol-pyridine mixture and estimated by the absorbance at 532nm. As an external standard, tetramethoxy-propane was used, and lipid peroxide level was expressed in terms of nmol malondialdehyde. Using this method, the liped peroxide level in the liver of rats suffering from carbon tetrachloride intoxication was investigated. The results were in good agreement with previously reported data obtained by measuring diene content.
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A protein determination method which involves the binding of Coomassie Brilliant Blue G-250 to protein is described. The binding of the dye to protein causes a shift in the absorption maximum of the dye from 465 to 595 nm, and it is the increase in absorption at 595 nm which is monitored. This assay is very reproducible and rapid with the dye binding process virtually complete in approximately 2 min with good color stability for 1 hr. There is little or no interference from cations such as sodium or potassium nor from carbohydrates such as sucrose. A small amount of color is developed in the presence of strongly alkaline buffering agents, but the assay may be run accurately by the use of proper buffer controls. The only components found to give excessive interfering color in the assay are relatively large amounts of detergents such as sodium dodecyl sulfate, Triton X-100, and commercial glassware detergents. Interference by small amounts of detergent may be eliminated by the use of proper controls.
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Parkinson's disease is a common neurodegenerative syndrome characterized by loss of dopaminergic neurons in the substantia nigra, formation of filamentous intraneuronal inclusions (Lewy bodies) and an extrapyramidal movement disorder. Mutations in the -synuclein gene are linked to familial Parkinson's disease1, 2 and -synuclein accumulates in Lewy bodies and Lewy neurites3, 4, 5. Here we express normal and mutant forms of -synuclein in Drosophila and produce adult-onset loss of dopaminergic neurons, filamentous intraneuronal inclusions containing -synuclein and locomotor dysfunction. Our Drosophila model thus recapitulates the essential features of the human disorder, and makes possible a powerful genetic approach to Parkinson's disease.