The porcine corpus luteum as a model for studying the effects of nanoplastics.

Nanoplastics (NPs) affect fertility. We evaluated the effects of NPs treatment on luteal and endothelial cells. We examined crucial markers of growth and redox status. NPs treatment did not induce changes in ATP levels in luteal cells, while it increased (p< 0.05) their proliferation. In endothelial cells, no change in proliferation was detected, while an increase (p<0.05) in ATP levels was observed. The increase of reactive oxygen species, superoxide anion (p<0.05) and nitric oxide (p<0.001) was detected in both cell types, which also showed changes in superoxide dismutase enzyme activity as well as an increase of non-enzymatic antioxidant power (p<0.05). A decrease (p<0.05) in progesterone production as well as an increase of vascular endothelial growth factor A levels were detected (p<0.05). In addition, a dose-dependent accumulation of NPs in endothelial cells was shown, that likely occurred through adhesion and internalization. Results underline potential risk of NPs for corpus luteum functionality.

1-Acetoxypinoresinol, a Lignan from Olives: Insight into Its Characterization, Identification, and Nutraceutical Properties.

Extra virgin olive oil (EVOO) is a symbol of the Mediterranean diet, constituting its primary source of fat. The beneficial effect of EVOO is strictly related to the presence of fatty acids and polyphenols, bioactive compounds endowed with nutraceutical properties. Among EVOO polyphenols, lignans possess a steroid-like chemical structure and are part of the phytoestrogen family, which is renowned for its health properties. The natural lignans (+)-pinoresinol and 1-acetoxypinoresinol (1-AP) are commonly present in olives and in EVOO. Although (+)-pinoresinol is found in different edible plants, such as flaxseed, beans, whole-grain cereals, sesame seeds, and certain vegetables and fruit, 1-AP was exclusively identified in olives in 2000. So far, the scientific literature has extensively covered different aspects of (+)-pinoresinol, including its isolation and nutraceutical properties. In contrast, less is known about the olive lignan 1-AP. Therefore, this review aimed to comprehensively evaluate the more important aspects of 1-AP, collecting all the literature from 2016 to the present, exploring its distribution in different cultivars, analytical isolation and purification, and nutraceutical properties.

Quantitative determination of tetracyclines in medicated feed for food-producing animals by HPLC-DAD.

Tetracyclines are a group of antibiotic substances largely administered through medicated feed to control diseases in food-producing animals. Fine dosing of antibiotics contained in medicated feed is crucial for the success of the treatment as well as minimising potential threats such as the spread of antimicrobial resistance and the transfer of antibiotic residues in food. A rapid analytical method based on HPLC with diode array detection (HPLC-DAD) was developed to quantify oxytetracycline, chlortetracycline and doxycycline in medicated feed. The reported method underwent in-house validation and was found to be suitable for the quantification of three target tetracyclines within the concentration range of 40-1000 mg kg-1 in official routine analysis. The method was applied to 103 official samples in the framework of the Italian National Plan on animal feed during the years 2021-2023 and nine non-compliant concentrations were identified in swine and fish feed samples.

Fecal microbiota characterization of an Italian local horse breed.

The Bardigiano horse is a traditional native Italian breed with a rich history and peculiar characteristics. Local breeds are proven to have unique genetic traits developed over generations to adapt to defined geographical regions and/or conditions. The specific microbial communities that coexist within these animals are unraveled by studying their microbiota, which permits a further step in the characterization of local heritage. This work aimed to characterize Bardigiano horse fecal microbiota composition. The data obtained were then compared with published data of a mix of athlete breeds to evaluate potential differences among local and specialized breeds. The study involved 11 Bardigiano mares between 3 and 4 years of age, from which stool was sampled for the study. Samples were processed for 16S rRNA sequencing. Data obtained were analyzed and plotted using R, RStudio, and FastTree software. The samples analyzed were similar to what literature has reported on horses of other breeds and attitudes at higher taxonomic levels (from phylum to genera). While at lower taxonomic levels, the difference was more marked highlighting specific families found in the Bardigiano breed only. Weight, province of origin, and breeding sites significantly affected microbiota composition (p-value ≤0.02, p-value ≤0.04, and p-value ≤0.05, respectively). The comparison with athlete breed showed a significant difference confirming that animal and environmental factors are crucial in determining fecal microbiota composition (p-value <0.001). Understanding the microbiota composition in local breeds like the Bardigiano horse is crucial for preserving biodiversity, managing animal health, and promoting sustainable farming practices.

Identification of New GSK3ß Inhibitors through a Consensus Machine Learning-Based Virtual Screening.

Glycogen synthase kinase-3 beta (GSK3ß) is a serine/threonine kinase that plays key roles in glycogen metabolism, Wnt/ß-catenin signaling cascade, synaptic modulation, and multiple autophagy-related signaling pathways. GSK3ß is an attractive target for drug discovery since its aberrant activity is involved in the development of neurodegenerative diseases such as Alzheimer's and Parkinson's disease. In the present study, multiple machine learning models aimed at identifying novel GSK3ß inhibitors were developed and evaluated for their predictive reliability. The most powerful models were combined in a consensus approach, which was used to screen about 2 million commercial compounds. Our consensus machine learning-based virtual screening led to the identification of compounds G1 and G4, which showed inhibitory activity against GSK3ß in the low-micromolar and sub-micromolar range, respectively. These results demonstrated the reliability of our virtual screening approach. Moreover, docking and molecular dynamics simulation studies were employed for predicting reliable binding modes for G1 and G4, which represent two valuable starting points for future hit-to-lead and lead optimization studies.

Nanoplastics induced oxidative stress and VEGF production in aortic endothelial cells.

Plastic is an important environmental issue and a more critical aspect concerns plastic fragments, mainly in term of nanoplastics (NPs). We demonstrated that NPs interfere with reproductive and adipose stromal cells. Since several research underlined an increased cardiovascular risk due to NPs, present study was undertaken to investigate their effect on aortic endothelial cells (AOC). We explored the specificity of their interaction with endothelial cells, quantifying their load in treated cells. Then, NPs effect was assessed on cell growth, generation of free radicals and antioxidant defence. Our data demonstrate that NPs colocalize with AOC. We found a significant (p < 0.01) increase both in metabolic activity and Vascular Endothelial Growth Factor (VEGF) production (p < 0.01). Redox status appeared to be disrupted (p < 0.05) by NPs. Taken together, the normal function of cultured AOC appeared negatively affected by AOC. Since NPs have been detected in blood, our present data appear of particular interest.

Olive Mill Wastewater as Source of Polyphenols with Nutraceutical Properties.

BACKGROUND: Agrifood waste products are often considered rich sources of bioactive compounds that can be conveniently recovered. Due to these peculiar characteristics, the study of these waste products is attracting great interest in nutraceutical research. Olive mill wastewaters (OMWWs) are generated by extra virgin olive oil (EVOO) production, and they pose environmental challenges due to their disposal. This study aimed to characterize the polyphenolic profile and to evaluate the nutraceutical properties of OMWW extracts from two Tuscan olive cultivars, Leccino (CL) and Frantoio (CF), collected during different time points in EVOO production. METHOD: After a liquid-liquid extraction, the HPLC and LC-MS/MS analysis of OMWW extracts confirmed the presence of 18 polyphenolic compounds. RESULTS: The polyphenol composition varied between the cultivars and during maturation stages. Notably, oleacein was detected at remarkably high levels in CL1 and CF1 extracts (314.628 ± 19.535 and 227.273 ± 3.974 µg/mg, respectively). All samples demonstrated scavenging effects on free radicals (DPPH and ABTS assays) and an anti-inflammatory potential by inhibiting cyclooxygenase (COX) enzymes. CONCLUSIONS: This study highlights the nutraceutical potential of OMWW extracts, emphasizing their antioxidant, antiradical, and anti-inflammatory activities. The results demonstrate the influence of olive cultivar, maturation stage, and extraction process on the polyphenolic composition and the bioactivity of OMWW extracts. These findings support a more profitable reuse of OMWW as an innovative, renewable, and low-cost source of dietary polyphenols with potential applications as functional ingredients in the development of dietary supplements, as well as in the pharmaceutical and cosmetics industries.

Insights into the Antioxidant/Antiradical Effects and In Vitro Intestinal Permeation of Oleocanthal and Its Metabolites Tyrosol and Oleocanthalic Acid.

(1) Background: In recent years, numerous studies have highlighted the beneficial effects of extra virgin olive oil (EVOO) as an active ingredient against chronic diseases. The properties of EVOO are due to its peculiar composition, mainly to its rich content of polyphenols. In fact, polyphenols may contribute to counteract oxidative stress, which often accompanies chronic diseases. In this work, the antioxidant effects of high-value polyphenol oleocanthal (OC) and its main metabolites, tyrosol (Tyr) and oleocanthalic acid (OA), respectively, have been investigated along with their impact on cell viability. (2) Methods: OC, Tyr, and OA have been evaluated regarding antiradical properties in term of scavenging capacity towards biologically relevant reactive species, including O2●-, HOCl, and ROO●, as well as their antioxidant/antiradical capacity (FRAP, DPPH●, ABTS●+). Moreover, the ability to permeate the intestinal membrane was assessed by an intestinal co-culture model composed by Caco-2 and HT29-MTX cell lines. (3) Results: The capacity of OC and Tyr as radical oxygen species (ROS) scavengers, particularly regarding HOCl and O2●-, was clearly demonstrated. Furthermore, the ability to permeate the intestinal co-culture model was plainly proved by the good permeations (>50%) achieved by all compounds. (4) Conclusions: OC, OA, and Tyr revealed promising properties against oxidative diseases.

A New Method for Ivermectin Detection and Quantification through HPLC in Organic Matter (Feed, Soil, and Water).

Ivermectin is a macrocyclic lactone widely used in veterinary medicine for its broad-spectrum antiparasitic properties. It has been proven to be effective and safe. The purpose of this study was to develop a high-performance liquid chromatography method with a diode array detector for ivermectin screening in feed and water for animal consumption. Furthermore, the objective was to quantify ivermectin levels that were higher than 0.5 mg/kg in solid matrixes and 0.1 mg/kg in water. Doramectin was used as process standard. Samples were extracted using solid phase extraction with silica and C-18 columns. The method involved the use of high-performance liquid chromatography (HPLC) with a diode array detector (DAD). The results were interpreted using a calibration curve built with ivermectin standards at multiple concentrations (0.5, 1, 2, 5, and 12.5 mg/kg). Statistical evaluation of data was done using ANOVA. The data analysis showed that the linear regression was highly significant (P < 0.001), the intercept values were not significantly different from zero, and the correlation coefficient values (>0.999) indicated excellent linearity. Further tests demonstrated that this method is also useful when studying soil matrixes. The soil was dried and analyzed in the same way as feed; the same recoveries were realized on the spiked samples. The method is easy, inexpensive, precise, and repeatable; it requires very small amounts of sample.

Enhanced Nutraceutical Properties of Extra Virgin Olive Oil Extract by Olive Leaf Enrichment.

(1) Background: Nowadays, the health-promoting properties of extra virgin olive oil (EVOO), including the antioxidant and anti-inflammatory actions, are well recognized and mainly attributed to the different polyphenols, such as oleocanthal and oleacein. In EVOO production, olive leaves represent a high value by-product, showing a wide spectrum of beneficial effects due to the presence of polyphenols, especially oleuropein. Here we report the study of olive leaf extract (OLE)-enriched EVOO extracts, obtained by adding different percentages of OLE to EVOO in order to ameliorate their nutraceutical activities. (2) Methods: The polyphenolic content of the EVOO/OLE extracts was analyzed by HPLC and the Folin-Ciocalteau assay. For further biological testing, an 8% OLE-enriched EVOO extract was chosen. Therefore, antioxidant effects were evaluated by three different methods (DPPH, ABTS, and FRAP), and the anti-inflammatory properties were assessed in terms of cyclooxygenase activity inhibition. (3) Results: The antioxidant and anti-inflammatory profiles of the new EVOO/OLE extract are significantly improved compared to those of EVOO extract; (4) Conclusions: The combination of OLE and EVOO extract can lead to an extract enriched in terms of bioactive polyphenols and endowed with better biological properties than the singular EVOO extract. Therefore, it may represent a new complement in the nutraceutical field.

Redox Status, Estrogen and Progesterone Production by Swine Granulosa Cells Are Impaired by Triclosan.

Triclosan is a chlorinated biphenolic with a broad spectrum of antiseptic activities used in cosmetics and hygiene products. Continuous exposure can lead to absorption and bioaccumulation of this substance with harmful health effects. In fact, previous studies have shown that Triclosan acts as an endocrine-disrupting chemical on reproductive organs, with consequent negative effects on reproductive physiology. Therefore, to assess potential adverse impacts on fertility, we tested Triclosan on swine granulosa cells, a model of endocrine reproductive cells. We examined its effects on the main features of granulosa cell functions such as cell growth (BrdU incorporation and ATP production) and steroidogenesis (17-ß estradiol and progesterone secretion). Moreover, since oxidant−antioxidant balance plays a pivotal role in follicular function, redox status markers (superoxide, hydrogen peroxide and nitric oxide production, enzymatic and non-enzymatic scavenging activity) were studied. Our results show that Triclosan significantly inhibits cell growth (p < 0.001), steroidogenesis (p < 0.001), superoxide and nitric oxide production (p < 0.001), while it increases (p < 0.05) enzymatic defense systems. Collectively, these data suggest a disruption of the main granulosa cell functions, i.e., proliferation and hormone production, as well as an imbalance in redox status. On these bases, we can speculate that Triclosan would impair granulosa cell functions, thus exerting negative effects on reproductive function. Further studies are needed to explore lower Triclosan concentrations and to unravel its mechanisms of action at gene level.

Machine Learning-Based Virtual Screening for the Identification of Cdk5 Inhibitors.

Cyclin-dependent kinase 5 (Cdk5) is an atypical proline-directed serine/threonine protein kinase well-characterized for its role in the central nervous system rather than in the cell cycle. Indeed, its dysregulation has been strongly implicated in the progression of synaptic dysfunction and neurodegenerative diseases, such as Alzheimer's disease (AD) and Parkinson's disease (PD), and also in the development and progression of a variety of cancers. For this reason, Cdk5 is considered as a promising target for drug design, and the discovery of novel small-molecule Cdk5 inhibitors is of great interest in the medicinal chemistry field. In this context, we employed a machine learning-based virtual screening protocol with subsequent molecular docking, molecular dynamics simulations and binding free energy evaluations. Our virtual screening studies resulted in the identification of two novel Cdk5 inhibitors, highlighting an experimental hit rate of 50% and thus validating the reliability of the in silico workflow. Both identified ligands, compounds CPD1 and CPD4, showed a promising enzyme inhibitory activity and CPD1 also demonstrated a remarkable antiproliferative activity in ovarian and colon cancer cells. These ligands represent a valuable starting point for structure-based hit-optimization studies aimed at identifying new potent Cdk5 inhibitors.

Novel Potent and Selective Agonists of the GPR55 Receptor Based on the 3-Benzylquinolin-2(1H)-One Scaffold.

A growing body of evidence underlines the crucial role of GPR55 in physiological and pathological conditions. In fact, GPR55 has recently emerged as a therapeutic target for several diseases, including cancer and neurodegenerative and metabolic disorders. Several lines of evidence highlight GPR55's involvement in the regulation of microglia-mediated neuroinflammation, although the exact molecular mechanism has not been yet elucidated. Nevertheless, there are only a limited number of selective GPR55 ligands reported in the literature. In this work, we designed and synthesized a series of novel GPR55 ligands based on the 3-benzylquinolin-2(1H)-one scaffold, some of which showed excellent binding properties (with Ki values in the low nanomolar range) and almost complete selectivity over cannabinoid receptors. The full agonist profile of all the new derivatives was assessed using the p-ERK activation assay and a computational study was conducted to predict the key interactions with the binding site of the receptor. Our data outline a preliminary structure-activity relationship (SAR) for this class of molecules at GPR55. Some of our compounds are among the most potent GPR55 agonists developed to date and could be useful as tools to validate this receptor as a therapeutic target.

Content Variations in Oleocanthalic Acid and Other Phenolic Compounds in Extra-Virgin Olive Oil during Storage.

The health benefits of extra-virgin olive oil (EVOO) are strictly linked to the presence of phenolic compounds, which exhibit numerous nutraceutical properties. In EVOO, the most important class of phenolic compounds is represented by secoiridoids (oleacein and oleocanthal). EVOO is constantly subjected to degradation processes, including hydrolytic and oxidative reactions that influence its phenolic composition. In particular, the hydrolytic reactions determine the transformation of oleocanthal and oleacein into the corresponding phenyl-alcohols, tyrosol, and hydroxytyrosol. Furthermore, oleocanthal by oxidation processes can be converted to oleocanthalic acid. In this study, we evaluated the phenolic composition of three EVOO samples kept at different storage conditions for 15 months, focusing on the variation of oleocanthalic acid content. Specifically, the samples were stored at 4 °C in darkness and at 25 °C with light exposure. The results of our analyses highlighted that in EVOOs exposed to light and maintained at 25 °C, the degradation was more marked than in EVOO stored in dark and at 4 °C, due to the greater influence of external factors on storage conditions. Although chemical-physical characteristics of EVOOs are slightly different depending on provenience and treatment time, the results of this study reveal that storage conditions are fundamental to controlling phenol concentration.

Pyridylidene Amide Ru Complex for Selective Oxidation in Organic Synthesis.

The ruthenium(II) bis(PYA) complex 1 (PYA = p-pyridylidene amide) is a powerful catalyst for the oxidation of sulfides to sulfones, of alkenes to carbonyl compounds, and of terminal alkynes to carboxylic acids by using NaIO4 as the terminal oxidant. The catalytic system shows a broad functional group tolerance and rate differences between alkyne and sulfide oxidation that are sufficiently large to effectively achieve selective sulfide oxidation with exquisite selectivity.

CB1 receptor binding sites for NAM and PAM: A first approach for studying, new n­butyl­diphenylcarboxamides as allosteric modulators.

The development of cannabinoid receptor type-1 (CB1R) modulators has been implicated in multiple pathophysiological events ranging from memory deficits to neurodegenerative disorders among others, even if their central psychiatric side effects such as depression, anxiety, and suicidal tendencies, have limited their clinical use. Thus, the identification of ligands which selectively act on peripheral CB1Rs, is becoming more interesting. A recent study reported a class of peripheral CB1R selective antagonists, characterized by a 5-aryl substituted nicotinamide core. These derivatives have structural similarities with the biphenyl compounds, endowed with CB2R antagonist activity, previously synthesized by our research group. In this work we combined the pharmacophoric portion of both classes, in order to obtain novel CBR antagonists. Among the synthesized compounds rather unexpectedly two compounds of this series, C7 and C10, did not show the radioligand ([3H]CP55940) displacement on CB1R but increased binding (∼ 150%), suggesting a possible allosteric behavior. Computational studies were performed to investigate the role of these compounds in CB1R modulation. The analysis of their binding poses in two different binding cavities of the CB1R surface, revealed a preferred interaction with the experimental binding site for negative allosteric modulators.

Heavy Metals in the Muscle and Hepatopancreas of Red Swamp Crayfish (Procambarus clarkii) in Campania (Italy).

The aim of this study was to carry out a quali-quantitative analysis of the presence of non-essential and essential trace elements in freshwater crayfish (Procambarus clarkii) edible tissues to establish the healthiness of this product and to evaluate the pollution status of the sampling sites included in the present study. P. clarkii is one of the most common species of freshwater crustaceans in Italy, regularly consumed by local people. Moreover, the crayfish, due to its trophic position and diet, can be considered as an excellent bioindicator of the health status of the ecosystem. We collected sixty crayfish samples from two different sites in Campania (Italy): Villa Literno and Sessa Aurunca. Concentrations of trace elements were determined by Inductively Coupled Plasma Optical Emission Spectroscopy (ICP-OES). Our data showed low concentrations of Cd, Hg and Pb, with values below the European Commission MRL (Commission Regulation (EC) 1881/2006). We suggest that data obtained from this study showed that crayfish collected from Villa Literno and Sessa Aurunca were safe for human consumption. Furthermore, the results of this research indicated mild contamination of heavy metals of the sampling sites, indicating a good health status of the area's aquatic ecosystem.

Short-Term Effects of COVID-19 Lockdown in Italian Children and Adolescents with Type 1 Diabetes Mellitus: The Role of Separation Anxiety.

In March 2020, the Italian Government imposed mandatory home confinement to limit the spread of COVID-19. Few studies assessed the psychophysical impact of COVID-19 on chronically ill children. This study examined these effects on children with Type 1 Diabetes Mellitus (T1D) and their caregivers. Seventy-one patients (7-13 years) with T1D and their caregivers were administered a survey created ad hoc and some standardized questionnaires, assessing psychological well-being and anxiety. Medical data (physical and biochemical characteristics) were recorded before (T0, January-February) and after (T1, May-June) the lockdown. Paired Student t-test, Spearman two-tailed correlations, and a linear regression model were used for statistical analysis. Children at T1 showed higher BMI (body mass index), daily total and basal insulin dose, and time spent in therapeutic range, and they showed lower HbA1c (glycated hemoglobin), time spent above the therapeutic range, and standard deviations of the mean glucose values than at T0. A total of 32.9% scored in the clinical range for separation anxiety. The increase in separation anxiety was predicted by younger age, female gender, more recent T1D diagnosis, less time spent in therapeutic range at T1, and higher perceived fear of COVID-19 infection. In a pandemic context, separation anxiety may be stronger in younger females, with more recent T1D diagnosis and poor metabolic control, thus affecting the parent's ability to manage diabetes and to support children's autonomy.

Medicinal Chemistry approach, pharmacology and neuroprotective benefits of CB2R modulators in neurodegenerative diseases.

In the last decades, cannabinoid receptor 2 (CB2R) has continued to receive attention as a key therapeutic target in neuroprotection. Indeed, several findings highlight the neuroprotective effects of CB2R through suppression of both neuronal excitability and reactive microglia. Additionally, CB2R seems to be a more promising target than cannabinoid receptor 1 (CB1R) thanks to the lack of central side effects, its lower expression levels in the central nervous system (CNS), and its inducibility, since its expression enhances quickly in the brain following pathological conditions. This review aims to provide a thorough overview of the main natural and synthetic selective CB2R modulators, their chemical classification and their potential therapeutic usefulness in neuroprotection, a crucial aspect for the treatment of neurodegenerative diseases.

Oxo-functionalised mesoionic NHC nickel complexes for selective electrocatalytic reduction of CO2 to formate.

Strategies for the conversion of CO2 to valuable products are paramount for reducing the environmental risks associated with high levels of this greenhouse gas and offer unique opportunities for transforming waste into useful products. While catalysts based on nickel as an Earth-abundant metal for the sustainable reduction of CO2 are known, the vast majority produce predominantly CO as a product. Here, efficient and selective CO2 reduction to formate as a synthetically valuable product has been accomplished with novel nickel complexes containing a tailored C,O-bidentate chelating mesoionic carbene ligand. These nickel(ii) complexes are easily accessible and show excellent catalytic activity for electrochemical H+ reduction to H2 (from HOAc in MeCN), and CO2 reduction (from CO2-saturated MeOH/MeCN solution) with high faradaic efficiency to yield formate exclusively as an industrially and synthetically valuable product from CO2. The most active catalyst precursor features the 4,6-di-tert-butyl substituted phenolate triazolylidene ligand, tolerates different proton donors including water, and reaches an unprecedented faradaic efficiency of 83% for formate production, constituting the most active and selective Ni-based system known to date for converting CO2 into formate as an important commodity chemical.