Search Results (183)

Two-pore-domain K⁺ channels (K2p), known previously as leak channels, are responsible for maintaining the resting membrane potential of cells. Fifteen subtypes are known to exist in humans and eleven are known in Drosophila melanogaster, as well as six subfamilies; however, little is known about the expression of these subtypes in various animal tissues or the impact of altered expression on cellular physiology. The Drosophila melanogaster model allows for selective misexpression of certain neuron subsets, providing insight into individual cell types and the animal’s physiology more generally. Prior research on the overexpression of K2p channels and the resulting behavioral and neuronal effects is limited. This project expanded upon this prior research by using Drosophila motor neurons to examine the effects of K2p overexpression on behavior and physiology. After conducting various assays, it was concluded that K2p overexpression in motor neurons had the most prominent effects on Drosophila functioning, with sensory, cardiac, and chordotonal neurons also generating differences in behavior. Altered expression levels of K2p channels could result in tissue-specific and/or whole-animal dysfunction. Full article

Pteridium aquilinum (bracken fern) poses a global threat to biodiversity and to the health of both animals and humans due to its toxic metabolites and aggressive ecological expansion. In northern Spain, particularly in regions of intensive livestock farming, these risks may be exacerbated, calling for urgent assessment and monitoring strategies. In this study, we implemented a multidisciplinary approach to evaluate the toxicological and ecological relevance of P. aquilinum through four key actions: (a) quantification of pterosins A and B in young fronds (croziers) using ultra-high-performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS); (b) analysis of in vivo genotoxicity of aqueous extracts using Drosophila melanogaster as a model organism; (c) a large-scale survey of local livestock farmers to assess awareness and perceived impact of bracken; and (d) the development and field application of a drone-based mapping tool to assess the spatial distribution of the species at the regional level. Our results confirm the consistent presence of pterosins A and B in croziers, with concentrations ranging from 0.17 to 2.20 mg/g dry weight for PtrB and 13.39 to 257 µg/g for PtrA. Both metabolite concentrations and genotoxicity levels were found to correlate with latitude and, importantly, with each other. All tested samples exhibited genotoxic activity, with notable differences among them. The farmer survey (n = 212) revealed that only 50% of respondents were aware of the toxic risks posed by bracken, indicating a need for targeted outreach. The drone-assisted mapping approach proved to be a promising tool for identifying bracken-dominated areas and provides a scalable foundation for future ecological monitoring and land management strategies. Altogether, our findings emphasize that P. aquilinum is not merely a local concern but a globally relevant toxic species whose monitoring and control demand coordinated scientific and policy-based efforts. Full article

Conditional transgenic systems and multi-copy target transgenes were used to produce transient fluorescent protein expression in adult Drosophila melanogaster, with the goal of developing an in vivo assay of protein turnover. Free-moving flies were assayed at multiple time points using video, and decay in fluorescence was used to calculate protein half-life. Additional experiments involved image capture of anesthetized flies. The half-life of eGFP was increased by the proteasome inhibitor bortezomib, both in vivo and in vitro, indicating proteasomal degradation of eGFP. The accumulation of eGFP in vivo was decreased by the protein synthesis inhibitor cycloheximide, without affecting half-life. The half-lives of several fluorescent proteins were determined, using both tissue-general and tissue-specific expression, in flies of both sexes and varying ages. Typical half-life values varied by fluorescent protein. DsRED showed a greater half-life than eGFP, and little if any degradation was detected for mCherry. Half-life also varied by tissue, with greater eGFP half-life observed in muscle relative to other tissues. Increased half-life with age was detected for DsRED but not for eGFP. Limited effects were observed for sex and female mating status. Taken together, the data indicate the in vivo assays are promising tools for the study of protein degradation regulated by protein sequence, subcellular compartment, tissue and small molecules. Full article

Objectives: Comparative analyses of anesthetic agents on mutants with altered anesthetic sensitivity remain limited in the current literature. This study examines the sensitivity of various Drosophila melanogaster wild-type strains and mutants to the volatile anesthetics chloroform and isoflurane. We utilized recently identified mutants in ion channel-encoding genes and others historically selected for anesthetic resistance, such as AGAR (autosomal general anesthesia resistant) and har (halothane-resistant). Method: Based on the principles of the conventional inebriometer assay used to isolate these mutants, we developed a new, simpler method to measure the anesthetic response in these flies. Results: Interestingly, we discovered that wild-type flies exhibit varying levels of anesthetic resistance. Contrary to previous reports, AGAR and har mutants showed little resistance to anesthesia using our method. Several ion channel mutants displayed increased resistance or sensitivity. Across all strains, isoflurane was more potent than chloroform. To ensure objectivity, all experiments were conducted double-blind. These findings highlight the variability in anesthetic sensitivity among both wild-type and mutant flies and underscore the importance of assay design in assessing resistance. Full article

Bisphenol A (BPA) is one of the most produced compounds worldwide. It acts as an endocrine disruptor and can cause adverse effects in the body, even at low-dose exposures. By interacting with estrogen receptors, it can play an important role in the pathogenesis of several endocrine diseases, such as infertility, hormone-dependent tumours and various metabolic disorders. Exposure in humans, especially early in life, is of particular concern, since it can have a big impact on an individual’s development and growth. The objective of this study was to evaluate, in vivo, the genotoxicity of BPA and its effects on longevity, prolificacy and behaviour in Drosophila melanogaster. To evaluate the biological and behavioural effects, flies were crossed and subjected to different concentrations of BPA (0.5 mM, 1 mM, 2 mM, 5 mM and 10 mM) in Drosophila instant “Carolina” medium hydrated with the BPA solution for 3 days, and then transferred to a non-treated medium, where they continued to lay eggs. This procedure was repeated to obtain the F₂ generation. To evaluate genotoxicity, the somatic mutation and recombination test (SMART) and Comet assays were performed. In these cases, higher concentrations of BPA were chosen (1 mM, 10 mM, 20 mM and 50 mM). The results showed that this compound caused changes in longevity and prolificacy, and that these changes also affected subsequent generations. They also showed that BPA affected Drosophila’s behaviour and social interaction, but at the exposure levels investigated here, it did not cause significant genotoxic effects. Full article

Human angiotensin-I-converting enzyme (ACE) is involved in vasoregulation, inflammation, and neurodegenerative disorders. The enzyme is formed of two domains; the C-domain (cACE) is primarily involved in blood pressure regulation, whereas the N-domain (nACE) is strongly linked to fibrosis; hence, designing domain-specific inhibitors could make a difference between treating one condition without having a negative effect on another. AnCE (a close homologue of ACE) is derived from Drosophila melanogaster and has a high similarity specifically to cACE. Due to high similarity and ease of crystallisation, AnCE has been chosen as a model protein for ACE studies and for the design of ACE inhibitors. In this study, enzyme kinetic assays and X-ray crystallography techniques revealed the significance of using dipeptides as selective inhibitors for AnCE and how this knowledge could be applied to cACE and nACE. All the dipeptides tested in this study were shown to bind AnCE in two distinct locations, i.e., the non-prime and prime subsites. It was found that a hydrophobic residue at the S1 and S1′ subsites, with a tryptophan at the S2 and S2′ subsites, showed highest affinity towards AnCE. It was also observed that a key pocket within the S2′ subsite had a major influence on the binding orientation within the prime subsites and could potentially explain ACE’s dipeptidyl carboxypeptidase activity. Importantly these dipeptides are found in functional foods, making them potentially available from diets. Knowledge of the dipeptide binding presented here could aid in the development of ACE domain-specific inhibitors. Full article

Background/Objectives: Intestinal alkaline phosphatase (IAP) is an enzyme expressed in the intestinal brush border, which may exert anti-inflammatory effects by detoxifying lipopolysaccharides (LPSs), thereby preventing metabolic disorders. Various food components have been reported to influence IAP activity. However, few studies have evaluated the effects of fermented milk on IAP activity. In this study, we aimed to investigate fermented milk with high IAP-activating capacity and investigate its effect. Methods: We screened a skim milk culture (SC), a fermented milk model, using differentiated Caco-2 cells. We investigated the effect of SC on IAP activity and gene expression in the Drosophila midgut. Quantitative PCR and immunoblot assays were conducted to examine gene and protein levels. Results: Among the SC samples from different lactic acid bacteria or bifidobacteria, the SC of Lactobacillus johnsonii SBT0309 (LJ0309 SC) demonstrated a particularly strong capacity to activate IAP in Caco-2 cells, demonstrated by significantly increased IAP gene expression and protein levels in Caco-2 cells. Additionally, LJ0309 SC inhibited increased secretion of IL-8 in LPS-stimulated Caco-2 cells. Finally, in Drosophila melanogaster fed LJ0309 SC, we observed an increase in both IAP activity and gene expression in the midgut. Conclusions: LJ0309 SC increased IAP activity and gene expression in both Caco-2 cells and the Drosophila midgut, and inhibited the inflammatory response in LPS-stimulated Caco-2 cells. Although further in vivo studies are required, LJ0309 SC might help to ameliorate LPS-induced inflammation and disease via IAP activation. Full article

Alpha-amylase is an extracellular enzyme abundantly produced from fungal sources. The catalytic activity of microbial enzymes is higher, more stable, and economical compared to plant and animal enzymes; they can be produced in large quantities in a short time and do not produce unwanted by-products. In this study, the genotoxic effect of different concentrations (25 mg/mL, 50 mg/mL, and 100 mg/mL) of a native fungal thermostable alpha-amylase enzyme, produced from the Aspergillus niger G2-1 isolate with an enzyme activity of 38.6 U/mg, was investigated on the Drosophila melanogaster model organism. The effect of the alpha-amylase enzyme added to the culture medium on the developmental performance of D. melanogaster was assessed through larval toxicity analysis, its effect on DNA damage through the comet assay, and its response to oxidative stress through various biochemical parameters. As a result, it was determined that low-dose alpha-amylase enzyme concentration (25 mg/mL) did not cause intracellular oxidative stress, did not cause genotoxicity, and did not adversely affect growth performance, although feeding with alpha-amylase at 50 mg/mL and 100 mg/mL concentrations caused a significant decrease in the survival rate of D. melanogaster larvae and an increase in DNA damage rate in imagos. However, oxidative stress parameters in adult D. melanogaster did not change after the same alpha-amylase application. Full article

Fructose has been considered as an additive from soda beverages. For the approval of new additives or to extend the usage of an approved one, it is necessary to conduct toxicological studies in order to evaluate the DNA damage induced by these compounds. Our study is based on evaluating the safety and the nutraceutical potential of Fructose (FRU), a soda cola beverage (Pepsi-cola, PEP), and a diet soda cola (Diet Coke, DCC), characterizing the DNA changes induced in the Drosophila melanogaster organism model and in the human leukemia HL-60 cells performing different assays. Our results showed neither the toxicity nor mutagenic activity of FRU, PEP, and DCC in Drosophila melanogaster, while only PEP exhibited protective effects in the antitoxity assay, showing an 80% survival rate in combined treatments. FRU, but not PEP, enhanced lifespan parameters by up to 23 more days at the 5 mg/mL concentration. All three substances exhibited chemopreventive properties in some of the checkpoints carried out related to clastogenicity and methylation patterns in HL-60 cells. In conclusion, the tested compounds were safe at tested concentrations in Drosophila and showed moderate chemopreventive activity. Full article

(1) Background: Advances in food processing practices and health care are some of the most significant advances in modern daily life. The goal of this study is to evaluate the safety of potassium and sodium nitrates and nitrites when they are used as fertilizers in agriculture and food additives, as well as the known conversion of nitrate to nitrite in humans. (2) Methods: Various bioassays were conducted to investigate the effects of nitrates and nitrites in the Drosophila melanogaster genetic tester system. These assays focused on the modulation of degenerative processes at the molecular, cellular, individual, and population levels. Additionally, we assessed the chemopreventive potential and the ability to induce DNA strand breaks in HL-60 tumour cells. (3) Results: All nitrate and nitrite concentrations tested were shown to not be toxic or genotoxic in Drosophila since none of the compounds reached the LD₅₀ and significant genetic mutation. A positive or null protective capacity against a toxic agent was found for nitrates, not for nitrites, showing that sodium nitrite has a synergistic effect when combined with the oxidant toxin hydrogen peroxide; and a nutraceutical potential in the lifespan only for sodium nitrate to improve the quality of life in 5 days at ADI concentration. The in vitro results in human leukemia cells showed a chemopreventive potential only for potassium nitrate and sodium nitrite due to reducing the viability of HL-60 cells growth to 18% and 29%, respectively, compared to the controls at ADI (acceptable daily intake) concentrations. However, neither of these showed DNA damage or methylation modifications. (4) Conclusions: The tested compounds were shown to be safe to use during in vivo and in vitro tests when used at the extrapolated ADI concentrations. Full article

In addition to the immature edible flower heads, the cultivation of globe artichoke (Cynara cardunculus L. var. scolymus (L.) Fiori) generates substantial quantities of by-products, including leaves, stems, and roots, which constitute potential sources of bioactive compounds and prebiotic dietary fiber. Preserving agricultural biodiversity and promoting socioeconomic development are essential for enhancing domestic production and fostering innovation. In the search for new biomolecules with antioxidant properties, this research focused on a globe artichoke landrace at risk of genetic erosion, still cultivated in the northern part of the Lazio region, known as the “Carciofo Ortano”. To investigate the antioxidant properties of various globe artichoke tissues from the “Carciofo Ortano” landrace, methanolic extracts were prepared from the immature main and secondary flower heads, stems, and leaves of representative genotypes of this landrace. Additionally, extracts were obtained from the same tissues of four landraces/clones included in the varietal platform of the PGI “Carciofo Romanesco del Lazio”, which served as reference genotypes: Campagnano, Castellammare, C3, and Grato 1. The antioxidant properties of these extracts were assessed using FRAP, ABTS, DPPH assays, and total phenolic content (TPC). The stem and secondary flower head extracts of two representative “Carciofo Ortano” genotypes and the Grato 1 clone, which have higher phenolic content, demonstrated the highest antioxidant activity. These extracts were therefore studied for their chemical profile using HPLC-DAD and SPME-GC/MS analysis. Additionally, the same extracts were investigated in vitro for their antioxidant capacity in differentiated SH-SY5Y cells, assessing their effects on ROS levels and the restoration of GSH levels. Furthermore, the in vivo beneficial effects of counteracting oxidative stress were evaluated in high sucrose-fed Drosophila melanogaster, as oxidative stress is a typical hallmark of hyperglycemic status. Overall, the results indicated that the edible immature inflorescences of the “Carciofo Ortano” landrace, along with the byproducts of its cultivation, are sources of raw materials containing biomolecules whose properties can be exploited for further applications in the pharmaceutical and medical sectors. Full article

Drosophila suzukii severely damages the production of berry and stone fruits in large parts of the world. Unlike D. melanogaster, which reproduces on overripe and fermenting fruits on the ground, D. suzukii prefers to lay its eggs in ripening fruits still on the plants. Flies locate fruit hosts by their odorant volatiles, which are detected and encoded by a highly specialised olfactory system before being translated into behaviour. The exact information-processing pathway is not yet fully understood, especially the evaluation of odour attractiveness. It is also unclear what differentiates the brains of D. suzukii and D. melanogaster to cause the crucial difference in host selection. We hypothesised that the basis for different behaviours is already formed at the level of the antennal lobe of D. suzukii and D. melanogaster by different neuronal responses to volatiles associated with ripe and fermenting fruit. We thus investigated by 3D in vivo two-photon calcium imaging how both species encoded odours from ripe fruits, leaves, fermented fruits, bacteria, and their mixtures in the antennal lobe. We then assessed their behavioural responses to mixtures of ripe and fermenting odours. The neural responses reflect species-dependent shifts in the odour code. In addition to this, morphological differences were also observed. However, this was not directly reflected in different behavioural responses to the odours tested. Full article

Parasitoid elimination in Drosophila melanogaster involves special hemocytes, called lamellocytes, which encapsulate the eggs or larvae of the parasitoid wasps. The capsules are melanized, and metabolites of the melanization reaction may play a potential role in parasitoid killing. We have observed a variation in the melanization capacity of different, commonly used D. melanogaster strains, such as Canton-S, Oregon-R, and BL5905, BL6326. In this work, we aimed to clarify a possible connection between the effectiveness of capsule melanization and the success of parasitoid elimination following infection with Leptopilina parasitoid wasps. Circulating hemocytes and lamellocyte attachment were visualized by confocal and epifluorescence microscopy using indirect immunofluorescence. Expression profiles of the PPO2 and PPO3 prophenoloxidase genes, which encode key enzymes in the melanization reaction, were detected by qRT-PCR. Parasitization assays were used to analyze fly and wasp eclosion success. Active encapsulation and melanization reactions against Leptopilina boulardi were observed in the BL5905 and the BL6326 strains, though restricted to the dead supernumerary parasitoids, while fly and wasp eclosion rates were essentially the same in the four examined D. melanogaster strains. We conclude that encapsulation and melanization carried out by D. melanogaster following L. boulardi infection have no impact on survival. Full article

G protein-coupled receptors (GPCRs) are essential cell surface proteins involved in transducing extracellular signals into intracellular responses, regulating various physiological processes. This study validated the use of the Tango assay, a sensitive method for detecting GPCR activation, in Drosophila Schneider 2 (S2) cells, focusing on the human Dopamine Receptor D4 (DRD4). Plasmids encoding the LexA-tagged human DRD4 receptor and a luciferase reporter were co-transfected into Drosophila S2 cells and stimulated with dopamine. Receptor activation was measured by quantifying the luciferase activity. The system showed high specificity for dopamine, with no activation in response to octopamine, a non-ligand for DRD4. Furthermore, the system effectively detects activation by a novel compound. These results demonstrate that Drosophila S2 cells, coupled with the Tango assay, provide a viable model for studying human GPCR function and ligand specificity. This system enables the rapid screening of potential GPCR ligands in a cost-effective cellular model. Full article

The controversial results of research on monosodium glutamate demand a new data corpus for the overall safety evaluation. Both animal and cellular model systems have been used to add a multilevel scope on its biological effects. The Drosophila melanogaster animal model has been used to test a wide range of concentrations for safety purposes: toxicity, genotoxicity, longevity and health span. Medium concentrations corresponding to the human acceptable daily intake (ADI) (0.06 mg/mL) were not toxic nor genotoxic for Drosophila and safe for the lifespan parameters. Once safety was determined, the possible nutraceutical effects of monosodium glutamate was monitored in terms of antitoxicity, antigenotoxicity assays and health span. The results for protective activity against hydrogen peroxide were positive in terms of the medium concentration, antitoxic and antigenotoxic in terms of inhibiting the genotoxicity induced by the oxidative toxin up to 43.7% and increasing the health span expectancy by 32% in terms of days. Monosodium glutamate has been demonstrated to be cytotoxic against the model tumour cell line HL-60, not only in a necrotic way but through internucleosomal DNA fragmentation antitumour activity. The significant LINE1 DNA sequence methylation of HL-60 tumour cells induced by monosodium glutamate is a molecular marker for chemoprevention. Conclusions: the slight or non-significant positive nutraceutical and chemo preventive potential showed by monosodium glutamate at its ADI concentration can be considered as a safe dose for a moderate consumption. Full article