Exploring the role of microRNAs as diagnostic and prognostic biomarkers in canine mammary tumors.

Canine mammary tumors (CMTs) represent a significant health concern in dogs, with a high incidence among intact female dogs. CMTs are a promising comparative model for human breast cancer, due to sharing several pathophysiological features. Additionally, CMTs have a strong genetic correlation with their human counterpart, including the expression of microRNAs (miRNAs). MiRNAs are a class of non-coding RNAs that play important roles in post-translational regulation of gene expression, being implicated in carcinogenesis, tumor progression, and metastasis. Moreover, miRNAs hold promise as diagnostic, prognostic, and metastatic biomarkers. Understanding the molecular mechanisms underlying CMTs is crucial for improving diagnosis, prognosis, and monitoring of treatments. Herein, we provide a comprehensive overview of the current knowledge on miRNAs in CMTs, highlighting their roles in carcinogenesis and their potential as biomarkers. Additionally, we highlight the current limitations and critically discuss the overarching challenges in this field, emphasizing the need for future research to translate miRNA findings into veterinary clinical practice.

NK1 receptor mediates cerebral cellular and extracellular morphological changes during the LPS-induced febrile response.

Fever elicited by bacterial lypopolyssacharide (LPS) is mediated by pro-inflammatory cytokines, which activate central mediators and regulate the hypothalamic temperature setpoint. This response is often accompanied by morphological changes involving the extracellular matrix, neurons and glial cells, with significant health impacts. The NK1 receptor is involved in the febrile response induced by LPS but its effects over the extracellular matrix in the context of neuroinflammation remain unknown. The present work aims to clarify the extracellular changes associated with NK1 signaling in LPS-induced fever. Male Wistar rats were exposed to LPS intraperitoneally. Experimental groups were pre-treated intracerebroventricularly with the NK1 selective inhibitor SR140333B or saline. Histological changes involving the brain extracellular matrix were evaluated using hematoxylin and eosin, Mason's trichrome, picrosirius, alcian blue, periodic acid Schiff's stains. The expression of matrix metalloproteinase 9 (MMP9) was studied using confocal microscopy. Fever was accompanied by edema, perivascular lymphoplamacytic and neutrophylic infiltration, spongiosis and MMP9 overexpression. SR140333B significantly reduced LPS-induced fever (p < 0.0001), MMP9 overexpression (p < 0.01) and associated histological changes. These results contribute to characterize cerebral extracellular matrix changes associated with LPS-induced fever. Overall, the present work supports a role for NK1 receptor in these neuroinflammatory changes, involving MMP9 overexpression, edema and leukocytic infiltration.

Unveiling the RKIP and EGFR Inverse Relationship in Solid Tumors: A Case Study in Cervical Cancer.

Raf Kinase Inhibitor Protein (RKIP) is recognized as a bona fide tumor suppressor gene, and its diminished expression or loss is associated with the progression and poor prognosis of various solid tumors. It exerts multifaceted roles in carcinogenesis by modulating diverse intracellular signaling pathways, including those governed by HER receptors such as MAPK. Given the significance of HER receptor overexpression in numerous tumor types, we investigated the potential oncogenic relationship between RKIP and HER receptors in solid tumors. Through a comprehensive in silico analysis of 30 TCGA PanCancer Atlas studies encompassing solid tumors (10,719 samples), we uncovered compelling evidence of an inverse correlation between RKIP and EGFR expression in solid tumors observed in 25 out of 30 studies. Conversely, a predominantly positive association was noted for the other HER receptors (ERBB2, ERBB3, and ERBB4). In particular, cervical cancer (CC) emerged as a tumor type exhibiting a robust inverse association between RKIP and EGFR expression, a finding that was further validated in a cohort of 202 patient samples. Subsequent in vitro experiments involving pharmacological and genetic modulation of EGFR and RKIP showed that RKIP depletion led to significant upregulation of EGFR mRNA levels and induction of EGFR phosphorylation. Conversely, EGFR overactivation decreased RKIP expression in CC cell lines. Additionally, we identified a common molecular signature among patients depicting low RKIP and high EGFR expression and demonstrated the prognostic value of this inverse correlation in CC patients. In conclusion, our findings reveal an inverse association between RKIP and EGFR expression across various solid tumors, shedding new light on the underlying molecular mechanisms contributing to the aggressive phenotype associated with RKIP and EGFR in cervical cancer.

Transthyretin mutagenesis: impact on amyloidogenesis and disease.

Transthyretin (TTR), a homotetrameric protein found in plasma, cerebrospinal fluid, and the eye, plays a pivotal role in the onset of several amyloid diseases with high morbidity and mortality. Protein aggregation and fibril formation by wild-type TTR and its natural more amyloidogenic variants are hallmarks of ATTRwt and ATTRv amyloidosis, respectively. The formation of soluble amyloid aggregates and the accumulation of insoluble amyloid fibrils and deposits in multiple tissues can lead to organ dysfunction and cell death. The most frequent manifestations of ATTR are polyneuropathies and cardiomyopathies. However, clinical manifestations such as carpal tunnel syndrome, leptomeningeal, and ocular amyloidosis, among several others may also occur. This review provides an up-to-date listing of all single amino-acid mutations in TTR known to date. Of approximately 220 single-point mutations, 93% are considered pathogenic. Aspartic acid is the residue mutated with the highest frequency, whereas tryptophan is highly conserved. "Hot spot" mutation regions are mainly assigned to ß-strands B, C, and D. This manuscript also reviews the protein aggregation models that have been proposed for TTR amyloid fibril formation and the transient conformational states that convert native TTR into aggregation-prone molecular species. Finally, it compiles the various in vitro TTR aggregation protocols currently in use for research and drug development purposes. In short, this article reviews and discusses TTR mutagenesis and amyloidogenesis, and their implications in disease onset.

Effective acid mine drainage remediation in fixed bed column using porous red mud/fly ash-containing geopolymer spheres.

Acid mine drainage (AMD) poses a significant threat to water quality worldwide, being amongst the most problematic environmental concerns of the millennium. This work reports for the first time the remediation of real AMD, from a Portuguese abandoned mine, in fixed bed column using porous red mud/fly ash-based geopolymeric spheres. Porous waste-based spheres (2.6 ± 0.2 mm) were obtained by a suspension-solidification method through the addition of optimum foaming agent dosage. The sorbent capacity in removing cations from AMD was evaluated by targeting selected hazardous elements: Zn, Cu, Co, Pb and Ni, based on their occurrence in the effluent and potential hazard. The spheres exhibited a dual mechanism of action, simultaneously neutralizing the acidic sample while removing cations through adsorption achieving removal efficiencies between 51 % and 80 %. Other elements present in high levels, such as iron were efficiently removed (>96 %). The role of precipitation, due to the pH neutralization, and adsorption was determined. The sorbent regeneration and reusability were evaluated for up to five cycles. Moreover, the effectiveness of waste-based geopolymers treating distinct AMD waters due to seasonal variations was also evaluated, further demonstrating the effectiveness of the proposed strategy to address environmental concerns stemming from mining activities.

Effect of ohmic heating on the structure and properties of flexible multilayer packaging.

Food-packaging-processing interactions define packaging materials' performance properties and product quality. This study evaluated the effect of ohmic heating (OH) processing and different food simulants on the properties of four multilayer flexible packaging materials (PETmet/PE, PETmet/PP, PET/Al/PE, and PET/Al/PA/PP). OH treatment was applied to the sealed packages containing the food simulants using a voltage gradient of 3.7 V/cm at a frequency of 20 kHz, resulting in a thermal process of at 80 °C for 1 min. The structure and performance of the different packages were then evaluated. The materials did not show changes in chemical groups nor thermal properties. However, the simulant-packaging-processing interaction resulted in changes in crystallinity, morphology, mechanical and barrier properties (water and oxygen), especially for metallized films in contact with acidic food simulants. The results indicate that although OH resulted in changes in packaging materials, these materials can be used under the conditions applied in this study.

Açaí (Euterpe oleracea Mart.) Seed Oil and Its Nanoemulsion: Chemical Characterisation, Toxicity Evaluation, Antioxidant and Anticancer Activities.

This study explores a nanoemulsion formulated with açaí seed oil, known for its rich fatty acid composition and diverse biological activities. This study aimed to characterise a nanoemulsion formulated with açaí seed oil and explore its cytotoxic effects on HeLa and SiHa cervical cancer cell lines, alongside assessing its antioxidant and toxicity properties both in vitro and in vivo. Extracted from fruits sourced in Brazil, the oil underwent thorough chemical characterization using gas chromatography-mass spectrometry. The resulting nanoemulsion was prepared and evaluated for stability, particle size, and antioxidant properties. The nanoemulsion exhibited translucency, fluidity, and stability post centrifugation and temperature tests, with a droplet size of 238.37, PDI -9.59, pH 7, and turbidity 0.267. In vitro assessments on cervical cancer cell lines revealed antitumour effects, including inhibition of cell proliferation, migration, and colony formation. Toxicity tests conducted in cell cultures and female Swiss mice demonstrated no adverse effects of both açaí seed oil and nanoemulsion. Overall, açaí seed oil, particularly when formulated into a nanoemulsion, presents potential for cancer treatment due to its bioactive properties and safety profile.

The Effect of Lemon Juice (Citrus limon L.) Treated with Melatonin on the Health Status and Treatment of K14HPV16 Mice.

Lemon is a fruit rich in antioxidant properties and has several health benefits, namely the reduction of skin edema and anticarcinogenic properties, which are due to its high content of bioactive compounds. Melatonin can improve and preserve the properties of lemon for longer and also has health benefits. The aim of this study was to evaluate the effects of oral administration of lemon juice after melatonin treatment on murinometric parameters of wild-type (WT) mice and transgenic mice carrying human papillomavirus (HPV). Two trials were performed for oral administration of the lemon extract compound: in drinking water and in diet. First of all, lemons were treated by immersion with melatonin at 10 mM. Then, lemons were squeezed, and the juice obtained was freeze-dried and stored to be subsequently added to drinking water or diet, according to the assay. Thus, mice were divided into eight groups in the drink assay (each with n = 5): group 1 (G1, WT, control), group 2 (G2, WT, 1 mL lemon), group 3 (G3, WT, 1.5 mL lemon), group 4 (G4, WT, 2 mL lemon), group 5 (G5, HPV16, control), group 6 (G6, HPV16, 1 mL lemon) group 7 (G6, HPV16, 1.5 mL lemon) and group 8 (G6, HPV16, 2 mL lemon). The diet assay was divided into four groups: group 1 (G1, WT, control), group 2 (G2, WT, 4 mL lemon), group 3 (G3, HPV16, control) and group 4 (G4, HPV16, 4 mL lemon). In the drink assay, the highest concentration of melatonin (308 ng/100 mL) was for groups 4 and 8, while in the food assay, there was only one concentration of melatonin (9.96 ng/g) for groups 2 and 4. Both trials lasted 30 days. During this time, body weight, food and water were recorded. Afterward, they were sacrificed, and samples were collected for different analyses. At the concentrations used, the lemon juice with melatonin had no adverse effects on the animals' health and showed a positive outcome in modifying weight gain and enhancing antioxidant activity in mice. Moreover, a reduction in the incidence of histological lesions was observed in treated animals. Further research is needed to better understand the effects of lemon extract on health and treatment outcomes in this animal model.

Comparison among Different Green Extraction Methods of Polyphenolic Compounds from Exhausted Olive Oil Pomace and the Bioactivity of the Extracts.

The use of by-products as a source of bioactive compounds with economic added value is one of the objectives of a circular economy. The olive oil industry is a source of olive pomace as a by-product. The olive pomace used in the present study was the exhausted olive pomace, which is the by-product generated from the air drying and subsequent hexane extraction of residual oil from the olive pomace. The objective was to extract bioactive compounds remaining in this by-product. Various types of green extraction were used in the present study: solvent extraction (water and hydroalcoholic); ultrasound-assisted extraction; Ultra-Turrax-assisted extraction; and enzyme-assisted extraction (cellulase; viscoenzyme). The phenolic profile of each extract was determined using HPLC-DAD and the total phenolic content (TPC) and antioxidant activity (ABTS, DPPH, and ORAC) were determined as well. The results showed significant differences in the yield of extraction among the different methods used, with the enzyme-assisted, with or without ultrasound, extraction presenting the highest values. The ultrasound-assisted hydroethanolic extraction (USAHE) was the method that resulted in the highest content of the identified phenolic compounds: 2.021 ± 0.29 mg hydroxytyrosol/100 mg extract, 0.987 ± 0.09 mg tyrosol/100 mg extract, and 0.121 ± 0.005 mg catechol/100 mg extract. The conventional extraction with water at 50 °C produced the best results for TPC and antioxidant activity of the extracts. The extracts from the USAHE were able to inhibit Gram-positive bacteria, especially Bacillus cereus, showing 67.2% inhibition at 3% extract concentration.

Folding Kinetics and Volume Variation of the ß-Hairpin Peptide Chignolin upon Ultrafast pH-Jumps.

In-depth characterization of fundamental folding steps of small model peptides is crucial for a better understanding of the folding mechanisms of more complex biomacromolecules. We have previously reported on the folding/unfolding kinetics of a model α-helix. Here, we study folding transitions in chignolin (GYDPETGTWG), a short ß-hairpin peptide previously used as a model to study conformational changes in ß-sheet proteins. Although previously suggested, until now, the role of the Tyr2-Trp9 interaction in the folding mechanism of chignolin was not clear. In the present work, pH-dependent conformational changes of chignolin were characterized by circular dichroism (CD), nuclear magnetic resonance (NMR), ultrafast pH-jump coupled with time-resolved photoacoustic calorimetry (TR-PAC), and molecular dynamics (MD) simulations. Taken together, our results present a comprehensive view of chignolin's folding kinetics upon local pH changes and the role of the Tyr2-Trp9 interaction in the folding process. CD data show that chignolin's ß-hairpin formation displays a pH-dependent skew bell-shaped curve, with a maximum close to pH 6, and a large decrease in ß-sheet content at alkaline pH. The ß-hairpin structure is mainly stabilized by aromatic interactions between Tyr2 and Trp9 and CH-π interactions between Tyr2 and Pro4. Unfolding of chignolin at high pH demonstrates that protonation of Tyr2 is essential for the stability of the ß-hairpin. Refolding studies were triggered by laser-induced pH-jumps and detected by TR-PAC. The refolding of chignolin from high pH, mainly due to the protonation of Tyr2, is characterized by a volume expansion (10.4 mL mol-1), independent of peptide concentration, in the microsecond time range (lifetime of 1.15 µs). At high pH, the presence of the deprotonated hydroxyl (tyrosinate) hinders the formation of the aromatic interaction between Tyr2 and Trp9 resulting in a more disorganized and dynamic tridimensional structure of the peptide. This was also confirmed by comparing MD simulations of chignolin under conditions mimicking neutral and high pH.

Sixty years of research on bracken fern (Pteridium spp.) toxins: Environmental exposure, health risks and recommendations for bracken fern control.

Bracken fern (Pteridium spp.) is a highly problematic plant worldwide due to its toxicity in combination with invasive properties on former farmland, in deforested areas and on disturbed natural habitats. The carcinogenic potential of bracken ferns has caused scientific and public concern for six decades. Its genotoxic effects are linked to illudane-type glycosides (ITGs), their aglycons and derivatives. Ptaquiloside is considered the dominating ITG, but with significant contributions from other ITGs. The present review aims to compile evidence regarding environmental pollution by bracken fern ITGs, in the context of their human and animal health implications. The ITG content in bracken fern exhibits substantial spatial, temporal, and chemotaxonomic variation. Consumption of bracken fern as food is linked to human gastric cancer but also causes urinary bladder cancers in bovines browsing on bracken. Genotoxic metabolites are found in milk and meat from bracken fed animals. ITG exposure may also take place via contaminated water with recent data pointing to concentrations at microgram/L-level following rain events. Airborne ITG-exposure from spores and dust has also been documented. ITGs may synergize with major biological and environmental carcinogens like papillomaviruses and Helicobacter pylori to induce cancer, revealing novel instances of chemical and biological co-carcinogenesis. Thus, the emerging landscape from six decades of bracken research points towards a global environmental problem with increasingly complex health implications.

A review of machine learning methods for cancer characterization from microbiome data.

Recent studies have shown that the microbiome can impact cancer development, progression, and response to therapies suggesting microbiome-based approaches for cancer characterization. As cancer-related signatures are complex and implicate many taxa, their discovery often requires Machine Learning approaches. This review discusses Machine Learning methods for cancer characterization from microbiome data. It focuses on the implications of choices undertaken during sample collection, feature selection and pre-processing. It also discusses ML model selection, guiding how to choose an ML model, and model validation. Finally, it enumerates current limitations and how these may be surpassed. Proposed methods, often based on Random Forests, show promising results, however insufficient for widespread clinical usage. Studies often report conflicting results mainly due to ML models with poor generalizability. We expect that evaluating models with expanded, hold-out datasets, removing technical artifacts, exploring representations of the microbiome other than taxonomical profiles, leveraging advances in deep learning, and developing ML models better adapted to the characteristics of microbiome data will improve the performance and generalizability of models and enable their usage in the clinic.

New Perspectives in the Association between Anthropometry and Mortality: The Role of Calf Circumference.

AIMS: Considering the impact of sarcopenia on mortality, and the difficulty to assessment of body composition, the hypothesis of the study is that calf circumference (CC) is closely related to mortality in older patients. The aim of the study was to analyze the potential role of CC to predict mortality in old individuals at 3, 6 and 12 months after discharge from hospital. METHODS: Patients aged >65 years were recruited for this retrospective study from September 2021 to March 2022. Their physical and body composition characteristics (including Body Mass Index-BMI and Mini Nutritional Assessment-MNA) were measured; data on mortality at 3 (T3), 6 (T6) and 12 (T12) months after discharge were recorded. Sarcopenia was diagnosed according to the 2019 European Consensus criteria. RESULTS: Participants were 192 older adults (92 women), with a mean age of 82.8±7.0 years. Sarcopenic people were 41. The mortality rate was higher in sarcopenic people only at T3 and T6. CC had comparable validity in predicting mortality to that of MNA and ASMMI (Appendicular Skeletal Muscle Mass), and was better than BMI and serum albumin at each time point. Youden's index showed that the best cut-off for CC for predicting mortality was 30.6 cm both at T3 (sensitivity: 74%; specificity: 75%) and T6 (sensitivity: 75%; specificity: 67%). At the Cox regression model for mortality, high values of CC (HR 0.73, CI95% 0.60-0.89/p<0.001) and ADL scores (HR 0.72, CI95% 0.54-0.96/p=0.04) were protective factors at T6 and T12 respectively; at T12 high comorbidity rate was a risk factor (HR 1.28, IC95% 1.02-1.62/p=0.04). CONCLUSIONS: CC has a validity comparable to MNA and ASMMI in predicting mortality at 3, 6 and 12 months after hospital discharge. Moreover, it can be considered an independent predictor of medium-term mortality in the hospitalized older population. CC can be an effective method for the prognostic stratification of these patients, due to its simplicity and immediacy.

Impact of high Spirulina diet, extruded or supplemented with enzymes, on blood cells, systemic metabolites, and hepatic lipid and mineral profiles of broiler chickens.

The impact of 15% dietary inclusion of Spirulina (Arthrospira platensis) in broiler chickens was explored, focusing on blood cellular components, systemic metabolites and hepatic lipid and mineral composition. From days 14 to 35 of age, 120 broiler chickens were divided and allocated into four dietary treatments: a standard corn and soybean meal-based diet (control), a 15% Spirulina diet, a 15% extruded Spirulina diet, and a 15% Spirulina diet super-dosed with an enzyme blend (0.20% porcine pancreatin plus 0.01% lysozyme). The haematological analysis revealed no significant deviations (p > 0.05) in blood cell counts across treatments, suggesting that high Spirulina inclusion maintains haematological balance. The systemic metabolic assessment indicated an enhanced antioxidant capacity in birds on Spirulina diets (p < 0.001), pointing toward a potential reduction in oxidative stress. However, the study noted a detrimental impact on growth performance metrics, such as final body weight and feed conversion ratio (both p < 0.001), in the Spirulina-fed treatments, with the super-dosed enzyme blend supplementation failing to alleviate these effects but with extrusion mitigating them. Regarding hepatic composition, birds on extruded Spirulina and enzyme-supplemented diets showed a notable increase in n-3 fatty acids (EPA, DPA, DHA) (p < 0.001), leading to an improved n-6/n-3 PUFA ratio (p < 0.001). Despite this positive shift, a reduction in total hepatic lipids (p = 0.003) was observed without a significant change in cholesterol levels. Our findings underscore the need for further exploration into the optimal inclusion levels, processing methods and potential enzymatic enhancements of Spirulina in broiler diets. Ultimately, this research aims to strike a balance between promoting health benefits and maintaining optimal growth performance in poultry nutrition.

Valorization of Cork Stoppers, Coffee-Grounds and Walnut Shells in the Development and Characterization of Pectin-Based Composite Films: Physical, Barrier, Antioxidant, Genotoxic, and Biodegradation Properties.

The development of sustainable materials from the valorization of waste is a good alternative to reducing the negative environmental impact of plastic packaging. The objectives of this study were to develop and characterize pectin-based composite films incorporated with cork or cork with either coffee grounds or walnut shells, as well as to test the films' genotoxicity, antioxidant properties, and biodegradation capacity in soil and seawater. The addition of cork, coffee grounds, or walnut shells modified the films' characteristics. The results showed that those films were thicker (0.487 ± 0.014 mm to 0.572 ± 0.014 mm), more opaque (around 100%), darker (L* = 25.30 ± 0.78 to 33.93 ± 0.84), and had a higher total phenolic content (3.17 ± 0.01 mg GA/g to 4.24 ± 0.02 mg GA/g). On the other hand, the films incorporated only with cork showed higher values of elongation at break (32.24 ± 1.88% to 36.30 ± 3.25%) but lower tensile strength (0.91 ± 0.19 MPa to 1.09 ± 0.08 MPa). All the films presented more heterogeneous and rougher microstructures than the pectin film. This study also revealed that the developed films do not contain DNA-reactive substances and that they are biodegradable in soil and seawater. These positive properties could subsequently make the developed films an interesting eco-friendly food packaging solution that contributes to the valorization of organic waste and by-products, thus promoting the circular economy and reducing the environmental impact of plastic materials.

Long-Term Follow-up of Patients With Autonomous Thyroid Nodules Treated With Radioiodine.

PURPOSE: This article aims to describe the presentation of Plummer disease and its evolution after radioiodine treatment and determine factors that may influence treatment efficacy. PATIENTS AND METHODS: The sample included retrospective medical records of 165 adult patients with toxic nodular goiter treated with radioiodine between 1997 and 2017, followed up at a single thyroid center. RESULTS: The efficacy of treatment with a single dose of radioiodine was higher than 90%. The mean radioiodine activity was 28.9 ± 3.4 mCi. The mean time between radioiodine performance and hyperthyroidism resolution was 3.6 ± 3.0 months, ranging from 1-12 months. After the first year, 33.9% of the patients were under hypothyroidism, 59.4% under euthyroidism, and 6.7% under hyperthyroidism. Among the nonresponders, the variables that showed statistical difference were the presence of multinodular goiter and the radioiodine activity (mean, 25.5 ± 6.5 mCi; median, 30 [15-30 mCi]). The cumulative rate of hypothyroidism was 48.9% over 20 years of follow-up. CONCLUSIONS: Radioiodine therapy is an effective and safe treatment. In Plummer disease, high rates of euthyroidism are expected after the radioiodine treatment. Therapeutic failure was observed mainly in patients with larger multinodular goiters treated with lower doses of radioiodine. The evolution to hypothyroidism was mostly observed in younger patients with larger and uninodular goiters.

Development of a dry powder insufflation device with application in in vitro cell-based assays in the context of respiratory delivery.

Research on pharmaceutical dry powders has been increasing worldwide, along with increased therapeutic strategies for an application through the pulmonary or the nasal routes. In vitro methodologies and tests that mimic the respiratory environment and the process of inhalation itself are, thus, essential. The literature frequently reports cell-based in vitro assays that involve testing the dry powders in suspension. This experimental setting is not adequate, as both the lung and the nasal cavity are devoid of abundant liquid. However, devices that permit powder insufflation over cells in culture are either scarce or technically complex and expensive, which is not feasible in early stages of research. In this context, this work proposes the development of a device that allows the delivery of dry powders onto cell surfaces, thus simulating inhalation more appropriately. Subsequently, a quartz crystal microbalance (QCM) was used to establish a technique enabling the determination of dry powder deposition profiles. Additionally, the determination of the viability of respiratory cells (A549) after the insufflation of a dry powder using the developed device was performed. In all, a prototype for dry powder insufflation was designed and developed, using 3D printing methods for its production. It allowed the homogenous dispersion of the insufflated powders over a petri dish and a QCM crystal, and a more detailed study on how dry powders disperse over the supports. The device, already protected by a patent, still requires further improvement, especially regarding the method for powder weighing and the efficiency of the insufflation process, which is being addressed. The impact of insufflation of air and of locust bean gum (LBG)-based microparticles revealed absence of cytotoxic effect, as cell viability roughly above 70 % was always determined.

Innovations in Food Packaging for a Sustainable and Circular Economy.

Packaging is fundamental to maintaining the quality of food, but its contribution with a negative footprint to the environment must be completely changed worldwide to reduce pollution and climate change. Innovative and sustainable packaging and new strategies of reutilization are necessary to reduce plastic waste accumulation, maintain food quality and safety, and reduce food losses and waste. The purpose of this chapter is to present innovations in food packaging for a sustainable and circular economy. First, to present the eco-design packaging approach as well as new strategies for recycled or recyclable materials in food packaging. Second, to show current trends in new packaging materials developed from the use of agro-industrial wastes as well as new methods of production, including 3D/4D printing, electrostatic spinning, and the use of nanomaterials.

A metatranscriptomics strategy for efficient characterization of the microbiome in human tissues with low microbial biomass.

The high background of host RNA poses a major challenge to metatranscriptome analysis of human samples. Hence, metatranscriptomics has been mainly applied to microbe-rich samples, while its application in human tissues with low ratio of microbial to host cells has yet to be explored. Since there is no computational workflow specifically designed for the taxonomic and functional analysis of this type of samples, we propose an effective metatranscriptomics strategy to accurately characterize the microbiome in human tissues with a low ratio of microbial to host content. We experimentally generated synthetic samples with well-characterized bacterial and host cell compositions, and mimicking human samples with high and low microbial loads. These synthetic samples were used for optimizing and establishing the workflow in a controlled setting. Our results show that the integration of the taxonomic analysis of optimized Kraken 2/Bracken with the functional analysis of HUMAnN 3 in samples with low microbial content, enables the accurate identification of a large number of microbial species with a low false-positive rate, while improving the detection of microbial functions. The effectiveness of our metatranscriptomics workflow was demonstrated in synthetic samples, simulated datasets, and most importantly, human gastric tissue specimens, thus providing a proof of concept for its applicability on mucosal tissues of the gastrointestinal tract. The use of an accurate and reliable metatranscriptomics approach for human tissues with low microbial content will expand our understanding of the functional activity of the mucosal microbiome, uncovering critical interactions between the microbiome and the host in health and disease.