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Recent evidence indicates that the gut microbiota (GM) has a significant impact on the inflammatory bowel disease (IBD) progression. Our aim was to investigate the GM profiles, the Microbial Dysbiosis Index (MDI) and the intestinal microbiota-associated markers in relation to IBD clinical characteristics and disease state. We performed 16S rRNA metataxonomy on both stools and ileal biopsies, metabolic dysbiosis tests on urine and intestinal permeability and mucosal immunity activation tests on the stools of 35 IBD paediatric patients. On the GM profile, we assigned the MDI to each patient. In the statistical analyses, the MDI was correlated with clinical parameters and intestinal microbial-associated markers. In IBD patients with high MDI, Gemellaceae and Enterobacteriaceae were increased in stools, and Fusobacterium, Haemophilus and Veillonella were increased in ileal biopsies. Ruminococcaceae and WAL_1855D were enriched in active disease condition; the last one was also positively correlated to MDI. Furthermore, the MDI results correlated with PUCAI and Matts scores in ulcerative colitis patients (UC). Finally, in our patients, we detected metabolic dysbiosis, intestinal permeability and mucosal immunity activation. In conclusion, the MDI showed a strong association with both severity and activity of IBD and a positive correlation with clinical scores, especially in UC. Thus, this evidence could be a useful tool for the diagnosis and prognosis of IBD.

Gut microbiota (GM) modulation can be investigated as possible solution to enhance recovery after COVID-19. An open-label, single-center, single-arm, pilot, interventional study was performed by enrolling twenty patients recently recovered from COVID-19 to investigate the role of a mixed probiotic, containing Lactobacilli, Bifidobacteria and Streptococcus thermophilus, on gastrointestinal symptoms, local and systemic inflammation, intestinal barrier integrity and GM profile. Gastrointestinal Symptom Rating Scale, cytokines, inflammatory, gut permeability, and integrity markers were evaluated before (T0) and after 8 weeks (T1) of probiotic supplementation. GM profiling was based on 16S-rRNA targeted-metagenomics and QIIME 2.0, LEfSe and PICRUSt computational algorithms. Multiple machine learning (ML) models were trained to classify GM at T0 and T1. A statistically significant reduction of IL-6 (p < 0.001), TNF-α (p < 0.001) and IL-12RA (p < 0.02), citrulline (p value < 0.001) was reported at T1. GM global distribution and microbial biomarkers strictly reflected probiotic composition, with a general increase in Bifidobacteria at T1. Twelve unique KEGG orthologs were associated only to T0, including tetracycline resistance cassettes. ML classified the GM at T1 with 100% score at phylum level. Bifidobacteriaceae and Bifidobacterium spp. inversely correlated to reduction of citrulline and inflammatory cytokines. Probiotic supplementation during post-COVID-19 may trigger anti-inflammatory effects though Bifidobacteria and related-metabolism enhancement.

Although several studies have explored the molecular landscape of metastatic melanoma, the genetic determinants of therapy resistance are still largely unknown. Here, we aimed to determine the contribution of whole-exome sequencing and circulating free DNA (cfDNA) analysis in predicting response to therapy in a consecutive real-world cohort of 36 patients, undergoing fresh tissue biopsy and followed during treatment. Although the underpowered sample size limited statistical analysis, samples from non-responders had higher copy number variations and mutations in melanoma driver genes compared to responders in the BRAF V600+ subset. In the BRAF V600− subset, Tumor Mutational Burden (TMB) was twice that in responders vs. non-responders. Genomic layout revealed commonly known and novel potential intrinsic/acquired resistance driver gene variants. Among these, RAC1, FBXW7, GNAQ mutations, and BRAF/PTEN amplification/deletion were present in 42% and 67% of patients, respectively. Both Loss of Heterozygosity (LOH) load and tumor ploidy were inversely associated with TMB. In immunotherapy-treated patients, samples from responders showed higher TMB and lower LOH and were more frequently diploid compared to non-responders. Secondary germline testing and cfDNA analysis proved their efficacy in finding germline predisposing variants carriers (8.3%) and following dynamic changes during treatment as a surrogate of tissue biopsy, respectively.

Type 1 diabetes (T1D) is a chronic autoimmune metabolic disorder with onset in pediatric/adolescent age, characterized by insufficient insulin production, due to a progressive destruction of pancreatic β-cells. Evidence on the correlation between the human gut microbiota (GM) composition and T1D insurgence has been recently reported. In particular, 16S rRNA-based metagenomics has been intensively employed in the last decade in a number of investigations focused on GM representation in relation to a pre-disease state or to a response to clinical treatments. On the other hand, few works have been published using alternative functional omics, which is more suitable to provide a different interpretation of such a relationship. In this work, we pursued a comprehensive metaproteomic investigation on T1D children compared with a group of siblings (SIBL) and a reference control group (CTRL) composed of aged matched healthy subjects, with the aim of finding features in the T1D patients’ GM to be related with the onset of the disease. Modulated metaproteins were found either by comparing T1D with CTRL and SIBL or by stratifying T1D by insulin need (IN), as a proxy of β-cells damage, showing some functional and taxonomic traits of the GM, possibly related to the disease onset at different stages of severity.

ATM germline pathogenic variants were recently found enriched in high-risk melanoma patients. However, ATM loss of heterozygosity (LOH) has never been investigated in melanoma and, therefore, a causal association with melanoma development has not been established yet. The purpose of this study was to functionally characterize 13 germline ATM variants found in high-risk melanoma patients—and classified by in silico tools as pathogenic, uncertain significance, or benign—using multiple assays evaluating ATM/pATM expression and/or LOH in melanoma tissues and cell lines. We assessed ATM status by Immunohistochemistry (IHC), Western Blot, Whole-Exome Sequencing/Copy Number Variation analysis, and RNA sequencing, supported by Sanger sequencing and microsatellite analyses. For most variants, IHC results matched those obtained with in silico classification and LOH analysis. Two pathogenic variants (p.Ser1135_Lys1192del and p.Ser1993ArgfsTer23) showed LOH and complete loss of ATM activation in melanoma. Two variants of unknown significance (p.Asn358Ile and p.Asn796His) showed reduced expression and LOH, suggestive of a deleterious effect. This study, showing a classic two-hit scenario in a well-known tumor suppressor gene, supports the inclusion of melanoma in the ATM-related cancer spectrum.

Type 1 diabetes (T1D) is a multifactorial autoimmune disease driven by T-cells against the insulin-producing islet β-cells, resulting in a marked loss of β-cell mass and function. Although a genetic predisposal increases susceptibility, the role of epigenetic and environmental factors seems to be much more significant. A dysbiotic gut microbial profile has been associated with T1D patients. Moreover, new evidence propose that perturbation in gut microbiota may influence the T1D onset and progression. One of the prominent features in clinically silent phase before the onset of T1D is the presence of a microbiota characterized by low numbers of commensals butyrate producers, thus negatively influencing the gut permeability. The loss of gut permeability leads to the translocation of microbes and microbial metabolites and could lead to the activation of immune cells. Moreover, microbiota-based therapies to slow down disease progression or reverse T1D have shown promising results. Starting from this evidence, the correction of dysbiosis in early life of genetically susceptible individuals could help in promoting immune tolerance and thus in reducing the autoantibodies production. This review summarizes the associations between gut microbiota and T1D for future therapeutic perspectives and other exciting areas of research.

Goat cheese is an important element of the Mediterranean diet, appreciated for its health-promoting features and unique taste. A pivotal role in the development of these characteristics is attributed to the microbiota and its continuous remodeling over space and time. Nevertheless, no thorough study of the cheese-associated microbiota using two metaomics approaches has previously been conducted. Here, we employed 16S rRNA gene sequencing and metaproteomics to explore the microbiota of a typical raw goat milk cheese at various ripening timepoints and depths of the cheese wheel. The 16S rRNA gene-sequencing and metaproteomics results described a stable microbiota ecology across the selected ripening timepoints, providing evidence for the microbiologically driven fermentation of goat milk products. The important features of the microbiota harbored on the surface and in the core of the cheese mass were highlighted in both compositional and functional terms. We observed the rind microbiota struggling to maintain the biosafety of the cheese through competition mechanisms and/or by preventing the colonization of the cheese by pathobionts of animal or environmental origin. The core microbiota was focused on other biochemical processes, supporting its role in the development of both the health benefits and the pleasant gustatory nuances of goat cheese.

Alterations of gut microbiota have been identified before clinical manifestation of type 1 diabetes (T1D). To identify the associations amongst gut microbiome profile, metabolism and disease markers, the 16S rRNA-based microbiota profiling and 1H-NMR metabolomic analysis were performed on stool samples of 52 T1D patients at onset, 17 T1D siblings and 57 healthy subjects (CTRL). Univariate, multivariate analyses and classification models were applied to clinical and -omic integrated datasets. In T1D patients and their siblings, Clostridiales and Dorea were increased and Dialister and Akkermansia were decreased compared to CTRL, while in T1D, Lachnospiraceae were higher and Collinsella was lower, compared to siblings and CTRL. Higher levels of isobutyrate, malonate, Clostridium, Enterobacteriaceae, Clostridiales, Bacteroidales, were associated to T1D compared to CTRL. Patients with higher anti-GAD levels showed low abundances of Roseburia, Faecalibacterium and Alistipes and those with normal blood pH and low serum HbA1c levels showed high levels of purine and pyrimidine intermediates. We detected specific gut microbiota profiles linked to both T1D at the onset and to diabetes familiarity. The presence of specific microbial and metabolic profiles in gut linked to anti-GAD levels and to blood acidosis can be considered as predictive biomarker associated progression and severity of T1D.

It remains controversial whether exposure to environmental radiofrequency signals (RF) impacts cell status or response to cellular stress such as apoptosis or autophagy. We used two label-free techniques, cellular impedancemetry and Digital Holographic Microscopy (DHM), to assess the overall cellular response during RF exposure alone, or during co-exposure to RF and chemical treatments known to induce either apoptosis or autophagy. Two human cell lines (SH-SY5Y and HCT116) and two cultures of primary rat cortex cells (astrocytes and co-culture of neurons and glial cells) were exposed to RF using an 1800 MHz carrier wave modulated with various environmental signals (GSM: Global System for Mobile Communications, 2G signal), UMTS (Universal Mobile Telecommunications System, 3G signal), LTE (Long-Term Evolution, 4G signal, and Wi-Fi) or unmodulated RF (continuous wave, CW). The specific absorption rates (S.A.R.) used were 1.5 and 6 W/kg during DHM experiments and ranged from 5 to 24 W/kg during the recording of cellular impedance. Cells were continuously exposed for three to five consecutive days while the temporal phenotypic signature of cells behavior was recorded at constant temperature. Statistical analysis of the results does not indicate that RF-EMF exposure impacted the global behavior of healthy, apoptotic, or autophagic cells, even at S.A.R. levels higher than the guidelines, provided that the temperature was kept constant.

Genomic studies have identified some of the most relevant genetic players in Neuroendocrine Neoplasm (NEN) tumorigenesis. However, we are still far from being able to draw a model that encompasses their heterogeneity, elucidates the different biological effects consequent to the identified molecular events, or incorporates extensive knowledge of molecular biomarkers and therapeutic targets. Here, we reviewed recent insights in NEN tumorigenesis from selected basic research studies on animal models, highlighting novel players in the intergenic cooperation and peculiar mechanisms including splicing dysregulation, chromatin stability, or cell dedifferentiation. Furthermore, models of tumorigenesis based on composite interactions other than a linear progression of events are proposed, exemplified by the involvement in NEN tumorigenesis of genes regulating complex functions, such as MEN1 or DAXX. Although limited by interspecies differences, animal models have proved helpful for the more in-depth study of every facet of tumorigenesis, showing that the identification of driver mutations is only one of the many necessary steps and that other mechanisms are worth investigating.

Background: Strongyloidiasis is a neglected tropical disease caused by the intestinal nematode Strongyloides stercoralis and characterized by gastrointestinal and pulmonary involvement. We report a pediatric case of strongyloidiasis to underline the response of the host microbiota to the perturbation induced by the nematode. Methods: We performed a 16S rRNA-metagenomic analysis of the gut microbiota of a 7-year-old female during and after S. stercolaris infection, investigating three time-point of stool samples’ ecology: T0- during parasite infection, T1- a month after parasite infection, and T2- two months after parasite infection. Targeted-metagenomics were used to investigate ecology and to predict the functional pathways of the gut microbiota. Results: an increase in the alpha-diversity indices in T0-T1 samples was observed compared to T2 and healthy controls (CTRLs). Beta-diversity analysis showed a shift in the relative abundance of specific gut bacterial species from T0 to T2 samples. Moreover, the functional prediction of the targeted-metagenomics profiles suggested an enrichment of microbial glycan and carbohydrate metabolisms in the T0 sample compared with CTRLs. Conclusions: The herein report reinforces the literature suggestion of a putative direct or immune-mediated ability of S. stercolaris to promote the increase in bacterial diversity.

Food allergy (FA) and, in particular, IgE-mediated cow’s milk allergy is associated with compositional and functional changes of gut microbiota. In this study, we compared the gut microbiota of cow’s milk allergic (CMA) infants with that of cow’s milk sensitized (CMS) infants and Healthy controls. The effect of the intake of a mixture of Bifidobacterium longum subsp. longum BB536, Bifidobacterium breve M-16V and Bifidobacterium longum subsp. infantis M-63 on gut microbiota modulation of CMA infants and probiotic persistence was also investigated. Gut microbiota of CMA infants resulted to be characterized by a dysbiotic status with a prevalence of some bacteria as Haemophilus, Klebsiella, Prevotella, Actinobacillus and Streptococcus. Among the three strains administered, B.longum subsp. infantis colonized the gastrointestinal tract and persisted in the gut microbiota of infants with CMA for 60 days. This colonization was associated with perturbations of the gut microbiota, specifically with the increase of Akkermansia and Ruminococcus. Multi-strain probiotic formulations can be studied for their persistence in the intestine by monitoring specific bacterial probes persistence and exploiting microbiota profiling modulation before the evaluation of their therapeutic effects.

Several studies in recent times have linked gut microbiome (GM) diversity to the pathogenesis of cancer and its role in disease progression through immune response, inflammation and metabolism modulation. This study focused on the use of network analysis and weighted gene co-expression network analysis (WGCNA) to identify the biological interaction between the gut ecosystem and its metabolites that could impact the immunotherapy response in non-small cell lung cancer (NSCLC) patients undergoing second-line treatment with anti-PD1. Metabolomic data were merged with operational taxonomic units (OTUs) from 16S RNA-targeted metagenomics and classified by chemometric models. The traits considered for the analyses were: (i) condition: disease or control (CTRLs), and (ii) treatment: responder (R) or non-responder (NR). Network analysis indicated that indole and its derivatives, aldehydes and alcohols could play a signaling role in GM functionality. WGCNA generated, instead, strong correlations between short-chain fatty acids (SCFAs) and a healthy GM. Furthermore, commensal bacteria such as Akkermansia muciniphila, Rikenellaceae, Bacteroides, Peptostreptococcaceae, Mogibacteriaceae and Clostridiaceae were found to be more abundant in CTRLs than in NSCLC patients. Our preliminary study demonstrates that the discovery of microbiota-linked biomarkers could provide an indication on the road towards personalized management of NSCLC patients.

Autism spectrum disorders (ASDs) are neurodevelopmental disorders characterized by behavioral alterations and currently affect about 1% of children. Significant genetic factors and mechanisms underline the causation of ASD. Indeed, many affected individuals are diagnosed with chromosomal abnormalities, submicroscopic deletions or duplications, single-gene disorders or variants. However, a range of metabolic abnormalities has been highlighted in many patients, by identifying biofluid metabolome and proteome profiles potentially usable as ASD biomarkers. Indeed, next-generation sequencing and other omics platforms, including proteomics and metabolomics, have uncovered early age disease biomarkers which may lead to novel diagnostic tools and treatment targets that may vary from patient to patient depending on the specific genomic and other omics findings. The progressive identification of new proteins and metabolites acting as biomarker candidates, combined with patient genetic and clinical data and environmental factors, including microbiota, would bring us towards advanced clinical decision support systems (CDSSs) assisted by machine learning models for advanced ASD-personalized medicine. Herein, we will discuss novel computational solutions to evaluate new proteome and metabolome ASD biomarker candidates, in terms of their recurrence in the reviewed literature and laboratory medicine feasibility. Moreover, the way to exploit CDSS, performed by artificial intelligence, is presented as an effective tool to integrate omics data to electronic health/medical records (EHR/EMR), hopefully acting as added value in the near future for the clinical management of ASD.

The human gut contains trillions of microbes that play a central role in host biology, including the provision of key nutrients from the diet. Food is a major source of precursors for metabolite production; in fact, diet modulates the gut microbiota (GM) as the nutrients, derived from dietary intake, reach the GM, affecting both the ecosystem and microbial metabolic profile. GM metabolic ability has an impact on human nutritional status from childhood. However, there is a wide variability of dietary patterns that exist among individuals. The study of interactions with the host via GM metabolic pathways is an interesting field of research in medicine, as microbiota members produce myriads of molecules with many bioactive properties. Indeed, much evidence has demonstrated the importance of metabolites produced by the bacterial metabolism from foods at the gut level that dynamically participate in various biochemical mechanisms of a cell as a reaction to environmental stimuli. Hence, the GM modulate homeostasis at the gut level, and the alteration in their composition can concur in disease onset or progression, including immunological, inflammatory, and metabolic disorders, as well as cancer. Understanding the gut microbe–nutrient interactions will increase our knowledge of how diet affects host health and disease, thus enabling personalized therapeutics and nutrition.

Extra virgin olive oil (EVOO) with its nutraceutical characteristics substantially contributes as a major nutrient to the health benefit of the Mediterranean diet. Unfortunately, the adulteration of EVOO with less expensive oils (e.g., peanut and corn oils), has become one of the biggest source of agricultural fraud in the European Union, with important health implications for consumers, mainly due to the introduction of seed oil-derived allergens causing, especially in children, severe food allergy phenomena. In this regard, revealing adulterations of EVOO is of fundamental importance for health care and prevention reasons, especially in children. To this aim, effective analytical methods to assess EVOO purity are necessary. Here, we propose a simple, rapid, robust and very sensitive method for non-specialized mass spectrometric laboratory, based on the matrix-assisted laser desorption/ionization mass spectrometry (MALDI-TOF MS) coupled to unsupervised hierarchical clustering (UHC), principal component (PCA) and Pearson’s correlation analyses, to reveal corn oil (CO) adulterations in EVOO at very low levels (down to 0.5%).

Due to the incompleteness of animal genome sequencing, the analysis and characterization of serum proteomes of most farm animals are still in their infancy, compared to the already well-documented human serum proteome. This review focuses on the implications of the farm animal serum proteomics in order to identify novel biomarkers for animal welfare, early diagnosis, prognosis and monitoring of infectious disease treatment, and develop new vaccines, aiming at determining the reciprocal benefits for humans and animals.

She-donkey’s milk (DM) and goat’s milk (GM) are commonly used in newborn and infant feeding because they are less allergenic than other milk types. It is, therefore, mandatory to avoid adulteration and contamination by other milk allergens, developing fast and efficient analytical methods to assess the authenticity of these precious nutrients. In this experimental work, a sensitive and robust matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) profiling was designed to assess the genuineness of DM and GM milks. This workflow allows the identification of DM and GM adulteration at levels of 0.5%, thus, representing a sensitive tool for milk adulteration analysis, if compared with other laborious and time-consuming analytical procedures.

The Mediterranean diet (MD) is considered one of the healthiest dietary models. Many of the characteristic components of the MD have functional features with positive effects on health and wellness. The MD adherence, calculated through various computational scores, can lead to a reduction of the incidence of major diseases (e.g., cancers, metabolic and cardiovascular syndromes, neurodegenerative diseases, type 2 diabetes and allergy). Furthermore, eating habits are the main significant determinants of the microbial multiplicity of the gut, and dietary components influence both microbial populations and their metabolic activities from the early stages of life. For this purpose, we present a study proposal relying on the generation of individual gut microbiota maps from MD-aware children/adolescents. The maps, based on meta-omics approaches, may be considered as new tools, acting as a systems biology-based proof of evidence to evaluate MD effects on gut microbiota homeostasis. Data integration of food metabotypes and gut microbiota “enterotypes” may allow one to interpret MD adherence and its effects on health in a new way, employable for the design of targeted diets and nutraceutical interventions in childcare and clinical management of food-related diseases, whose onset has been significantly shifted early in life.

Polyunsaturated fatty acids (PUFAs) are required to maintain the fluidity, permeability and integrity of cell membranes. Maternal dietary supplementation with ω-3 PUFAs during pregnancy has beneficial effects, including increased gestational length and reduced risk of pregnancy complications. Significant amounts of ω-3 docosahexaenoic acid (DHA) are transferred from maternal to fetal blood, hence ensuring high levels of DHA in the placenta and fetal bloodstream and tissues. Fetal DHA demand increases exponentially with gestational age, especially in the third trimester, due to fetal development. According to the World Health Organization (WHO) and the Food and Agriculture Organization of the United Nations (FAO), a daily intake of DHA is recommended during pregnancy. Omega-3 PUFAs are involved in several anti-inflammatory, pro-resolving and anti-oxidative pathways. Several placental disorders, such as intrauterine growth restriction, premature rupture of membranes (PROM) and preterm-PROM (pPROM), are associated with placental inflammation and oxidative stress. This pilot study reports on a preliminary evaluation of the significance of the daily DHA administration on PROM and pPROM events in healthy pregnant women. Further extensive clinical trials will be necessary to fully elucidate the correlation between DHA administration during pregnancy and PROM/pPROM occurrence, which is related in turn to gestational duration and overall fetal health.

Carbapenem-resistant Acinetobacter baumannii strains cause life-threatening infections due to the lack of therapeutic options. Although the main mechanisms underlying antibiotic-resistance have been extensively studied, the general response to maintain bacterial viability under antibiotic exposure deserves to be fully investigated. Since the periplasmic space contains several proteins with crucial cellular functions, besides carbapenemases, we decided to study the periplasmic proteome of the multidrug-resistant (MDR) A. baumannii AB5075 strain, grown in the absence and presence of imipenem (IMP). Through the proteomic approach, 65 unique periplasmic proteins common in both growth conditions were identified: eight proteins involved in protein fate, response to oxidative stress, energy metabolism, antibiotic-resistance, were differentially expressed. Among them, ABUW_1746 and ABUW_2363 gene products presented the tetratricopeptide repeat motif, mediating protein-protein interactions. The expression switch of these proteins might determine specific protein interactions to better adapt to changing environmental conditions. ABUW_2868, encoding a heat shock protein likely involved in protection against oxidative stress, was upregulated in IMP-exposed bacteria. Accordingly, the addition of periplasmic proteins from A. baumannii cultured with IMP increased bacterial viability in an antioxidant activity assay. Overall, this study provides the first insights about the composition of the periplasmic proteins of a MDR A. baumannii strain, its biological response to IMP and suggests possible new targets to develop alternative antibiotic drugs.

The increase of microorganisms multi-drug resistant (MDR) to antibiotics (ATBs) is becoming a global emergency, especially in frail subjects. In chronic liver disease (LD) with indications for liver transplantation (LT), MDR colonization can significantly affect the LT outcome. However, no clear guidelines for microbial management are available. A novel approach toward MDR-colonized patients undergoing LT was developed at our Center refraining from ATBs use during the transplant waiting list, and use of an intensive perioperative prophylaxis cycle. This study aimed to couple clinical evaluation with monitoring of gut microbiota in a pediatric LD patient colonized with MDR Klebsiella pneumoniae (KP) who underwent LT. No peri-transplant complications were reported, and a decontamination from the MDR bacteria occurred during follow-up. Significant changes in gut microbiota, especially during ATB treatment, were reported by microbiota profiling. Patterns of Klebsiella predominance and microbiota diversity revealed opposite temporal trends, with Klebsiella ecological microbiota niches linked to ATB-driven selection. Our infection control program appeared to control complications following LT in an MDR-KP-colonized patient. The perioperative ATB regimen, acting as LT prophylaxis, triggered MDR-KP overgrowth and gut dysbiosis, but buffered infectious processes. Mechanisms modulating the gut ecosystem should be taken into account in MDR colonization clinical management.

Septic shock, occurring in about 1% of neonates hospitalized in neonatal intensive care unit (NICU), is a major cause of death in the neonatal period. In the 1980s and 90s, exchange transfusion (ET) was reported by some authors to be effective in the treatment of neonatal sepsis and septic shock. The main aim of this retrospective study was to compare the mortality rate of neonates with septic shock treated only with standard care therapy (ScT group) with the mortality rate of those treated with ScT and ET (ET group). All neonates with septic shock admitted to our NICU from 2005 to 2015 were included in the study. Overall, 101/9030 (1.1%) neonates had septic shock. Fifty neonates out of 101 (49.5%) received one or more ETs. The mortality rate was 36% in the ET group and 51% in the ScT group (p = 0.16). At multivariate logistic regression analysis, controlling for potentially confounding factors significantly associated with death (gestational age, serum lactate, inotropic drugs, oligoanuria), ET showed a marked protective effect (Odds Ratio 0.21, 95% Confidence Interval: 0.06–0.71; p = 0.01). The lack of observed adverse events should encourage the use of this procedure in the treatment of neonates with septic shock.

Acute myeloid leukemia (AML) is an aggressive hematologic neoplasia with a complex polyclonal architecture. Among driver lesions, those involving the FLT3 gene represent the most frequent mutations identified at diagnosis. The development of tyrosine kinase inhibitors (TKIs) has improved the clinical outcomes of FLT3-mutated patients (Pt). However, overcoming resistance to these drugs remains a challenge. To unravel the molecular mechanisms underlying therapy resistance and clonal selection, we conducted a longitudinal analysis using a single-cell DNA sequencing approach (MissionBioTapestri® platform, San Francisco, CA, USA) in two patients with FLT3-mutated AML. To this end, samples were collected at the time of diagnosis, during TKI therapy, and at relapse or complete remission. For Pt #1, disease resistance was associated with clonal expansion of minor clones, and 2nd line TKI therapy with gilteritinib provided a proliferative advantage to the clones carrying NRAS and KIT mutations, thereby responsible for relapse. In Pt #2, clonal architecture was less complex, and 1st line TKI therapy with midostaurin was able to eradicate the leukemic clones. Our results corroborate previous findings about clonal selection driven by TKIs, highlighting the importance of a deeper characterization of individual clonal architectures for choosing the best treatment plan for personalized approaches aimed at optimizing outcomes.

To analyze in vivo scleral changes induced by MicroPulse transscleral laser therapy (MP-TLT) in refractory glaucoma using anterior segment–optical coherence tomography (AS-OCT). Forty-two candidate patients for MP-TLT were consecutively enrolled and underwent AS-OCT at baseline and after six months. MP-TLT success was defined as an intraocular pressure (IOP) reduction by one-third. The main outcome measures were the mean superior (S-), inferior (I-), and total (T-) intra-scleral hypo-reflective space area (MISHA: mm2) and scleral reflectivity (S-SR, I-SR, T-SR; arbitrary scale) as in vivo biomarkers of uveoscleral aqueous humor (AH) outflow. The IOP was the secondary outcome. The relations between the baseline-to-six months differences (D) of DS-MISHA, DI-MISHA, and DT-MISHA and DS-SR, DI-SR, DT-SR, and DIOP, were investigated. At 6 months, the median IOP reduction was 21% in the failures and 38% in the successes. The baseline S-MISHA, I-MISHA, and T-MISHA did not differ between the groups, while S-SR and T-SR were higher in the successes (p < 0.05). At six months, successful and failed MP-TLTs showed a 50% increase in S-MISHA (p < 0.001; p = 0.037), whereas I-SR and T-SR reduced only in the successes (p = 0.002; p = 0.001). When comparing DS-MISHA, DI-MISHA, and DT-MISHA and DS-SR, DI-SR, and DT-SR, there were no significant differences between the groups. In the successful procedures, DIOP was positively correlated with DT-MISHA and DI-MISHA (ρ = 0.438 and ρ = 0.490; p < 0.05). MP-TLT produced potentially advantageous modifications of the sclera in refractory glaucoma. Given the partial correlation between these modifications and post-treatment IOP reduction, our study confirmed that the activation of the uveoscleral AH outflow route could significantly contribute to the IOP lowering after MP-TLT.

HER2-positive breast cancer is associated with aggressive behavior and reduced survival rates. Calcitriol restores the antiproliferative activity of antiestrogens in estrogen receptor (ER)-negative breast cancer cells by re-expressing ERα. Furthermore, calcitriol and its analog, EB1089, enhance responses to standard anti-cancer drugs. Therefore, we aimed to investigate EB1089 effects when added to the combined treatment of lapatinib and antiestrogens on the proliferation of HER2-positive breast cancer cells. BT-474 (ER-positive/HER2-positive) and SK-BR-3 (ER-negative/HER2-positive) cells were pre-treated with EB1089 to modulate ER expression. Then, cells were treated with EB1089 in the presence of lapatinib with or without the antiestrogens, and proliferation, phosphorylation array assays, and Western blot analysis were performed. The results showed that EB1089 restored the antiproliferative response to antiestrogens in SK-BR-3 cells and improved the inhibitory effects of the combination of lapatinib with antiestrogens in the two cell lines. Moreover, EB1089, alone or combined, modulated ERα protein expression and reduced Akt phosphorylation in HER2-positive cells. EB1089 significantly enhanced the cell growth inhibitory effect of lapatinib combined with antiestrogens in HER2-positive breast cancer cells by modulating ERα expression and Akt phosphorylation suppression. These results highlight the potential of this therapeutic approach as a promising strategy for managing HER2-positive breast cancer.

This work analyzes the role of the tight junction (TJ) protein ZO-2 on mechanosensation. We found that the lack of ZO-2 reduced apical membrane rigidity measured with atomic force microscopy, inhibited the association of γ-actin and JAM-A to the cell border, and instead facilitated p114RhoGEF and afadin accumulation at the junction, leading to an enhanced mechanical tension at the TJ measured by FRET, with a ZO-1 tension probe, and increased tricellular TJ tension. Simultaneously, adherens junction tension measured with an E-cadherin probe was unaltered. The stability of JAM-A and ZO-2 binding was assessed by a collaborative in silico study. The absence of ZO-2 also impacted the cell response to the substrate, as monolayers plated in 20 kPa hydrogels developed holes not seen in parental cultures and displayed a retarded elongation and formation of cell aggregates. The absence of ZO-2 was sufficient to induce YAP and Snail nuclear accumulation in cells cultured over glass, but when ZO-2 KD cells were plated in nanostructured ridge arrays, they displayed an increased abundance of nuclear Snail and conspicuous internalization of claudin-4. These results indicate that the absence of ZO-2 also impairs the response of cells to substrate stiffness and exacerbates transformation triggered by substrate topography.

Candida spp. periprosthetic joint infections are rare but difficult-to-treat events, with a slow onset, unspecific symptoms or signs, and a significant relapse risk. Treatment with antifungals meets with little success, whereas prosthesis removal improves the outcome. In fact, Candida spp. adhere to orthopedic devices and grow forming biofilms that contribute to the persistence of this infection and relapse, and there is insufficient evidence that the use of antifungals has additional benefits for anti-biofilm activity. To date, studies on the direct antifungal activity of silver against Candida spp. are still scanty. Additionally, polycaprolactone (PCL), either pure or blended with calcium phosphate, could be a good candidate for the design of 3D scaffolds as engineered bone graft substitutes. Thus, the present research aimed to assess the antifungal and anti-biofilm activity of PCL-based constructs by the addition of antimicrobials, for instance, silver, against C. albicans and C. auris. The appearance of an inhibition halo around silver-functionalized PCL scaffolds for both C. albicans and C. auris was revealed, and a significant decrease in both adherent and planktonic yeasts further demonstrated the release of Ag+ from the 3D constructs. Due to the combined antifungal, osteoproliferative, and biodegradable properties, PCL-based 3D scaffolds enriched with silver showed good potential for bone tissue engineering and offer a promising strategy as an ideal anti-adhesive and anti-biofilm tool for the reduction in prosthetic joints of infections caused by Candida spp. by using antimicrobial molecule-targeted delivery.

ETV6::ABL1 rearranged neoplasms are rare hematological diseases. To date, about 80 cases have been reported, including myeloid and lymphoid leukemias. The ETV6 gene codes for an ETS family transcription factor and several fusion partners have been described. When translocated, ETV6 causes the constitutive activation of the partner genes. Here, we report the case of a 54-year-old woman with a cryptic insertion of the 3′ region of ABL1 in the ETV6 gene. The patient was first diagnosed with idiopathic hypereosinophilic syndrome, according to the clinical history, conventional cytogenetics, standard molecular analyses and pathologist description. Next generation sequencing of diagnosis samples unexpectedly detected both ETV6::ABL1 type A and B fusion transcripts, which were then confirmed by FISH. The diagnosis was Myeloid/Lymphoid neoplasm with ETV6::ABL1 fusion, and the patient received imatinib mesylate treatment. In a follow-up after more than one year, the patient still maintained the molecular and complete hematological responses. This case highlights the importance of timely and proper diagnostics and prompt tyrosine kinase inhibitor treatment.

In both healthcare and agriculture, antibiotic resistance is an alarming issue. Biocompatible and biodegradable ingredients (e.g., curcumin) are given priority in “green” criteria supported by the Next Generation EU platform. The solubility and stability of curcumin would be significantly improved if it were enclosed in nanobubbles (NB), and photoactivation with the correct wavelength of light can increase its antibacterial efficacy. A continuous release of curcumin over a prolonged period was provided by using innovative chitosan-shelled carriers, i.e., curcumin-containing nanobubbles (Curc-CS-NBs) and oxygen-loaded curcumin-containing nanobubbles (Curc-Oxy-CS-NBs). The results demonstrated that after photoactivation, both types of NBs exhibited increased effectiveness. For Staphylococcus aureus, the minimum inhibitory concentration (MIC) for Curc-CS-NBs remained at 46 µg/mL following photodynamic activation, whereas it drastically dropped to 12 µg/mL for Curc-Oxy-CS-NBs. Enterococcus faecalis shows a decreased MIC for Curc-CS-NB and Curc-Oxy-CS-NB (23 and 46 µg/mL, respectively). All bacterial strains were more effectively killed by NBs that had both oxygen and LED irradiation. A combination of Curc-Oxy-CS-NB and photodynamic stimulation led to a killing of microorganisms due to ROS-induced bacterial membrane leakage. This approach was particularly effective against Escherichia coli. In conclusion, this work shows that Curc-CS-NBs and Curc-Oxy-CS-exhibit extremely powerful antibacterial properties and represent a potential strategy to prevent antibiotic resistance and encourage the use of eco-friendly substitutes in agriculture and healthcare.

Spinal muscular atrophy (SMA) is a genetic disorder characterized by the loss of spinal motor neurons leading to muscle weakness and respiratory failure. Mitochondrial dysfunctions are found in the skeletal muscle of patients with SMA. For obvious ethical reasons, the diaphragm muscle is poorly studied, notwithstanding the very important role that respiratory involvement plays in SMA mortality. The main goal of this study was to investigate diaphragm functionality and the underlying molecular adaptations in SMNΔ7 mice, a mouse model that exhibits symptoms similar to that of patients with intermediate type II SMA. Functional, biochemical, and molecular analyses on isolated diaphragm were performed. The obtained results suggest the presence of an intrinsic energetic imbalance associated with mitochondrial dysfunction and a significant accumulation of reactive oxygen species (ROS). In turn, ROS accumulation can affect muscle fatigue, cause diaphragm wasting, and, in the long run, respiratory failure in SMNΔ7 mice. Exposure to the antioxidant molecule ergothioneine leads to the functional recovery of the diaphragm, confirming the presence of mitochondrial impairment and redox imbalance. These findings suggest the possibility of carrying out a dietary supplementation in SMNΔ7 mice to preserve their diaphragm function and increase their lifespan.

Trisomy X is the most frequent sex chromosome anomaly in women, but it is often underdiagnosed postnatally because most patients do not show any clinical manifestation. It is estimated that only 10% of patients with trisomy X are diagnosed by clinical findings. Thus, it has been proposed that the clinical spectrum is not yet fully delimited, and additional uncommon or atypical clinical manifestations could be related to this entity. The present report describes a female carrying trisomy X but presenting atypical manifestations, including severe intellectual disability, short stature, thymus hypoplasia, and congenital hypothyroidism (CH). These clinical findings were initially attributed to trisomy X. However, chromosome microarray analysis (CMA) subsequently revealed that the patient also bears a heterozygous 304-kb deletion at 16p11.2. This pathogenic copy-number variant (CNV) encompasses 13 genes, including TUFM. Some authors recommend that when a phenotype differs from that described for an identified microdeletion, the presence of pathogenic variants in the non-deleted allele should be considered to assess for an autosomal recessive disorder; thus, we used a panel of 697 genes to rule out a pathogenic variant in the non-deleted TUFM allele. We discuss the possible phenotypic modifications that might be related to an additional CNV in individuals with sex chromosome aneuploidy (SCA), as seen in our patient. The presence of karyotype-demonstrated trisomy X and CMA-identified 16p11.2 deletion highlights the importance of always correlating a patient’s clinical phenotype with the results of genetic studies. When the phenotype includes unusual manifestations and/or exhibits discrepancies with that described in the literature, as exemplified by our patient, a more extensive analysis should be undertaken to enable a correct diagnosis that will support proper management, genetic counseling, and medical follow-up.

Vasculogenic mimicry (VM), a process in which aggressive cancer cells form tube-like structures, plays a crucial role in providing nutrients and escape routes. Highly plastic tumor cells, such as those with the triple-negative breast cancer (TNBC) phenotype, can develop VM. However, little is known about the interplay between the cellular components of the tumor microenvironment and TNBC cells’ VM capacity. In this study, we analyzed the ability of endothelial and stromal cells to induce VM when interacting with TNBC cells and analyzed the involvement of the FGFR/PI3K/Akt pathway in this process. VM was corroborated using fluorescently labeled TNBC cells. Only endothelial cells triggered VM formation, suggesting a predominant role of paracrine/juxtacrine factors from an endothelial origin in VM development. Via immunocytochemistry, qPCR, and secretome analyses, we determined an increased expression of proangiogenic factors as well as stemness markers in VM-forming cancer cells. Similarly, endothelial cells primed by TNBC cells showed an upregulation of proangiogenic molecules, including FGF, VEGFA, and several inflammatory cytokines. Endothelium-dependent TNBC-VM formation was prevented by AZD4547 or LY294002, strongly suggesting the involvement of the FGFR/PI3K/Akt axis in this process. Given that VM is associated with poor clinical prognosis, targeting FGFR/PI3K/Akt pharmacologically may hold promise for treating and preventing VM in TNBC tumors.

Alzheimer’s disease (AD) and epilepsy are common neurological disorders in the elderly. A bi-directional link between these neurological diseases has been reported, with patients with either condition carrying almost a two-fold risk of contracting the other compared to healthy subjects. AD/epilepsy adversely affects patients’ quality of life and represents a severe public health problem. Thus, identifying the relationship between epilepsy and AD represents an ongoing challenge and continuing need. Seizures in AD patients are often unrecognized because they are often nonconvulsive and sometimes mimic some behavioral symptoms of AD. Regarding this, it has been hypothesized that epileptogenesis and neurodegeneration share common underlying mechanisms. Targeted treatment to decrease epileptiform activity could represent a valuable strategy for delaying the neurodegenerative process and related cognitive impairment. Several preclinical studies have shown that some antiseizure medications (ASMs) targeting abnormal network hyperexcitability may change the natural progression of AD. However, to date, no guidelines are available for managing seizures in AD patients because of the paucity of randomized clinical trials sufficient for answering the correlated questions. Future AD clinical studies are mandatory to update clinicians about the symptomatic treatment of seizures in AD patients and recognize whether ASM therapy could change the natural progression of the disease, thereby rescuing cognitive performance.

The genome sequencing of the tardigrade Ramazzottius varieornatus revealed a unique nucleosome-binding protein named damage suppressor (Dsup), which was discovered to be crucial for the extraordinary abilities of tardigrades in surviving extreme stresses, such as UV. Evidence in Dsup-transfected human cells suggests that Dsup mediates an overall response in DNA damage signaling, DNA repair, and cell cycle regulation, resulting in an acquired resistance to stress. Given these promising outcomes, our study attempts to provide a wider comprehension of the molecular mechanisms modulated by Dsup in human cells and to explore the Dsup-activated molecular pathways under stress. We performed a differential proteomic analysis of Dsup-transfected and control human cells under basal conditions and at 24 h recovery after exposure to UV-C. We demonstrate via enrichment and network analyses, for the first time, that even in the absence of external stimuli, and more significantly, after stress, Dsup activates mechanisms involved with the unfolded protein response, the mRNA processing and stability, cytoplasmic stress granules, the DNA damage response, and the telomere maintenance. In conclusion, our results shed new light on Dsup-mediated protective mechanisms and increases our knowledge of the molecular machineries of extraordinary protection against UV-C stress.

Despite the recent breakthroughs in targeted and immunotherapy for melanoma, the overall survival rate remains low. In recent years, considerable attention has been paid to the gut microbiota and other modifiable patient factors (e.g., diet and body composition), though their role in influencing therapeutic responses has yet to be defined. Here, we characterized a cohort of 31 patients with unresectable IIIC-IV-stage cutaneous melanoma prior to initiation of targeted or first-line immunotherapy via the following methods: (i) fecal microbiome and metabolome via 16S rRNA amplicon sequencing and gas chromatography/mass spectrometry, respectively, and (ii) anthropometry, body composition, nutritional status, physical activity, biochemical parameters, and immunoprofiling. According to our data, patients subsequently classified as responders were obese (i.e., with high body mass index and high levels of total, visceral, subcutaneous, and intramuscular adipose tissue), non-sarcopenic, and enriched in certain fecal taxa (e.g., Phascolarctobacterium) and metabolites (e.g., anethole), which were potentially endowed with immunostimulatory and oncoprotective activities. On the other hand, non-response was associated with increased proportions of Streptococcus, Actinomyces, Veillonella, Dorea, Fusobacterium, higher neutrophil levels (and a higher neutrophil-to-lymphocyte ratio), and higher fecal levels of butyric acid and its esters, which also correlated with decreased survival. This exploratory study provides an integrated list of potential early prognostic biomarkers that could improve the clinical management of patients with advanced melanoma, in particular by guiding the design of adjuvant therapeutic strategies to improve treatment response and support long-term health improvement.

Understanding the complexities of the human brain and its associated disorders poses a significant challenge in neuroscience. Traditional research methods have limitations in replicating its intricacies, necessitating the development of in vitro models that can simulate its structure and function. Three-dimensional in vitro models, including organoids, cerebral organoids, bioprinted brain models, and functionalized brain organoids, offer promising platforms for studying human brain development, physiology, and disease. These models accurately replicate key aspects of human brain anatomy, gene expression, and cellular behavior, enabling drug discovery and toxicology studies while providing insights into human-specific phenomena not easily studied in animal models. The use of human-induced pluripotent stem cells has revolutionized the generation of 3D brain structures, with various techniques developed to generate specific brain regions. These advancements facilitate the study of brain structure development and function, overcoming previous limitations due to the scarcity of human brain samples. This technical review provides an overview of current 3D in vitro models of the human cortex, their development, characterization, and limitations, and explores the state of the art and future directions in the field, with a specific focus on their applications in studying neurodevelopmental and neurodegenerative disorders.

Vitamin D along with its active metabolite calcitriol and its metabolic and signaling system, known as the vitamin D endocrine system, have been widely recognized as a pivotal regulator of calcium homeostasis in addition to non-calcemic antitumoral effects in a variety of human cancers, including cervical cancer. Several studies have found an inverse relationship between the incidence of cervical neoplasia and vitamin D levels. This narrative review updates the current evidence supporting the notion that the vitamin D endocrine system has a preventive role on cervical cancer, mainly in the early phases of the disease, acting at the level of suppressing cell proliferation, promoting apoptosis, modulating inflammatory responses, and probably favoring the clearance of human papillomavirus-dependent cervical lesions. Although an optimal vitamin D status helps in the prevention and regression of low-grade squamous intraepithelial lesions of the cervix, it appears that vitamin D alone or combined with chemotherapeutic agents has little effectivity once advanced cervical cancer is established. These observations suggest that an optimal vitamin D status might exert beneficial actions in the early phases of cervical cancer by preventing its onset and progression.

Placentas from gestational diabetes mellitus (GDM) patients undergo significant metabolic and immunologic adaptations due to hyperglycemia, which results in an exacerbated synthesis of proinflammatory cytokines and an increased risk for infections. Insulin or metformin are clinically indicated for the treatment of GDM; however, there is limited information about the immunomodulatory activity of these drugs in the human placenta, especially in the context of maternal infections. Our objective was to study the role of insulin and metformin in the placental inflammatory response and innate defense against common etiopathological agents of pregnancy bacterial infections, such as E. coli and S. agalactiae, in a hyperglycemic environment. Term placental explants were cultivated with glucose (10 and 50 mM), insulin (50–500 nM) or metformin (125–500 µM) for 48 h, and then they were challenged with live bacteria (1 × 105 CFU/mL). We evaluated the inflammatory cytokine secretion, beta defensins production, bacterial count and bacterial tissue invasiveness after 4–8 h of infection. Our results showed that a GDM-associated hyperglycemic environment induced an inflammatory response and a decreased beta defensins synthesis unable to restrain bacterial infection. Notably, both insulin and metformin exerted anti-inflammatory effects under hyperglycemic infectious and non-infectious scenarios. Moreover, both drugs fortified placental barrier defenses, resulting in reduced E. coli counts, as well as decreased S. agalactiae and E. coli invasiveness of placental villous trees. Remarkably, the double challenge of high glucose and infection provoked a pathogen-specific attenuated placental inflammatory response in the hyperglycemic condition, mainly denoted by reduced TNF-α and IL-6 secretion after S. agalactiae infection and by IL-1β after E. coli infection. Altogether, these results suggest that metabolically uncontrolled GDM mothers develop diverse immune placental alterations, which may help to explain their increased vulnerability to bacterial pathogens.

Magnetic resonance imaging (MRI) is a non-invasive powerful modern clinical technique that is extensively used for the high-resolution imaging of soft tissues. To obtain high-definition pictures of tissues or of the whole organism this technique is enhanced by the use of contrast agents. Gadolinium-based contrast agents have an excellent safety profile. However, over the last two decades, some specific concerns have surfaced. Mn(II) has different favorable physicochemical characteristics and a good toxicity profile, which makes it a good alternative to the Gd(III)-based MRI contrast agents currently used in clinics. Mn(II)-disubstituted symmetrical complexes containing dithiocarbamates ligands were prepared under a nitrogen atmosphere. The magnetic measurements on Mn complexes were carried out with MRI phantom measurements at 1.5 T with a clinical magnetic resonance. Relaxivity values, contrast, and stability were evaluated by appropriate sequences. Studies conducted to evaluate the properties of paramagnetic imaging in water using a clinical magnetic resonance showed that the contrast, produced by the complex [Mn(II)(L’)2] × 2H2O (L’ = 1.4-dioxa-8-azaspiro[4.5]decane-8-carbodithioate), is comparable to that produced by gadolinium complexes currently used in medicine as a paramagnetic contrast agent.

Skeletal muscle atrophy is a condition characterized by a loss of muscle mass and muscle strength caused by an imbalance between protein synthesis and protein degradation. Muscle atrophy is often associated with a loss of bone mass manifesting as osteoporosis. The aim of this study was to evaluate if chronic constriction injury (CCI) of the sciatic nerve in rats can be a valid model to study muscle atrophy and consequent osteoporosis. Body weight and body composition were assessed weekly. Magnetic resonance imaging (MRI) was performed on day zero before ligation and day 28 before sacrifice. Catabolic markers were assessed via Western blot and Quantitative Real-time PCR. After the sacrifice, a morphological analysis of the gastrocnemius muscle and Micro-Computed Tomography (Micro-CT) on the tibia bone were performed. Rats that underwent CCI had a lower body weight increase on day 28 compared to the naive group of rats (p < 0.001). Increases in lean body mass and fat mass were also significantly lower in the CCI group (p < 0.001). The weight of skeletal muscles was found to be significantly lower in the ipsilateral hindlimb compared to that of contralateral muscles; furthermore, the cross-sectional area of muscle fibers decreased significantly in the ipsilateral gastrocnemius. The CCI of the sciatic nerve induced a statistically significant increase in autophagic and UPS (Ubiquitin Proteasome System) markers and a statistically significant increase in Pax-7 (Paired Box-7) expression. Micro-CT showed a statistically significant decrease in the bone parameters of the ipsilateral tibial bone. Chronic nerve constriction appeared to be a valid model for inducing the condition of muscle atrophy, also causing changes in bone microstructure and leading to osteoporosis. Therefore, sciatic nerve constriction could be a valid approach to study muscle–bone crosstalk and to identify new strategies to prevent osteosarcopenia.

Cervical cancer is the fourth most common cancer among women worldwide. The main factor associated with the onset and progression of this neoplasia is the human papillomavirus (HPV) infection. The HPV-oncogenes E6 and E7 are critical drivers of cellular transformation, promoting the expression of oncogenes such as KCNH1. The phytochemical α-mangostin (AM) is a potent antineoplastic and antiviral compound. However, its effects on HPV oncogenes and KCNH1 gene expression remain unknown. This study evaluated the effects of AM on cell proliferation, cell cycle distribution and gene expression, including its effects on tumor growth in xenografted mice. AM inhibited cell proliferation in a concentration-dependent manner, being the most sensitive cell lines those with the highest number of HPV16 copies. In addition, AM promoted G1-cell cycle arrest in CaSki cells, while led to cell death in SiHa and HeLa cells. Of interest was the finding of an AM-dependent decreased gene expression of E6, E7 and KCNH1 both in vitro and in vivo, as well as the modulation of cytokine expression, Ki-67, and tumor growth inhibition. On these bases, we suggest that AM represents a good option as an adjuvant for the treatment and prevention of cervical cancer.

Streptococcus pyogenes causes a wide spectrum of diseases varying from mild to life threatening, despite antibiotic treatment. Nanoparticle application could facilitate the foreign pathogen fight by increasing the antimicrobial effectiveness and reducing their adverse effects. Here, we designed and produced erythromycin-loaded chitosan nanodroplets (Ery-NDs), both oxygen-free and oxygen-loaded. All ND formulations were characterized for physico-chemical parameters, drug release kinetics, and tested for biocompatibility with human keratinocytes and for their antibacterial properties or interactions with S. pyogenes. All tested NDs possessed spherical shape, small average diameter, and positive Z potential. A prolonged Ery release kinetic from Ery-NDs was demonstrated, as well as a favorable biocompatibility on human keratinocytes. Confocal microscopy images showed ND uptake and internalization by S. pyogenes starting from 3 h of incubation up to 24 h. According to cell counts, NDs displayed long-term antimicrobial efficacy against streptococci significantly counteracting their proliferation up to 24 h, thanks to the known chitosan antimicrobial properties. Intriguingly, Ery-NDs were generally more effective (104–103 log10 CFU/mL), than free-erythromycin (105 log10 CFU/mL), in the direct killing of streptococci, probably due to Ery-NDs adsorption by bacteria and prolonged release kinetics of erythromycin inside S. pyogenes cells. Based on these findings, NDs and proper Ery-NDs appear to be the most promising and skin-friendly approaches for the topical treatment of streptococcal skin infections allowing wound healing during hypoxia.

Obesity is an epidemic condition linked to cardiovascular disease severity and mortality. Fat localization and type represent cardiovascular risk estimators. Importantly, visceral fat secretes adipokines known to promote low-grade inflammation that, in turn, modulate its secretome and cardiac metabolism. In this regard, IL-33 regulates the functions of various immune cells through ST2 binding and—following its role as an immune sensor to infection and stress—is involved in the pro-fibrotic remodeling of the myocardium. Here we further investigated the IL-33/ST2 effects on cardiac remodeling in obesity, focusing on molecular pathways linking adipose-derived IL-33 to the development of fibrosis or hypertrophy. We analyzed the Zucker Fatty rat model, and we developed in vitro models to mimic the adipose and myocardial relationship. We demonstrated a dysregulation of IL-33/ST2 signaling in both adipose and cardiac tissue, where they affected Epac proteins and myocardial gene expression, linked to pro-fibrotic signatures. In Zucker rats, pro-fibrotic effects were counteracted by ghrelin-induced IL-33 secretion, whose release influenced transcription factor expression and ST2 isoforms balance regulation. Finally, the effect of IL-33 signaling is dependent on several factors, such as cell types’ origin and the balancing of ST2 isoforms. Noteworthy, it is reasonable to state that considering IL-33 to have a unique protective role should be considered over-simplistic.

Cholesterol accumulation in macrophages leads to the formation of foam cells and increases the risk of developing atherosclerosis. We have verified whether hydroxytyrosol (HT), a phenolic compound with anti-inflammatory and antioxidant properties, can reduce the cholesterol build up in THP-1 macrophage-derived foam cells. We have also investigated the potential mechanisms. Oil Red O staining and high-performance liquid chromatography (HPLC) assays were utilized to detect cellular lipid accumulation and cholesterol content, respectively, in THP-1 macrophages foam cells treated with HT. The impact of HT on cholesterol metabolism-related molecules (SR-A1, CD36, LOX-1, ABCA1, ABCG1, PPARγ and LRX-α) in foam cells was assessed using real-time PCR (RT-qPCR) and Western blot analyses. Finally, the effect of HT on the adhesion of THP-1 monocytes to human vascular endothelial cells (HUVEC) was analyzed to study endothelial activation. We found that HT activates the PPARγ/LXRα pathway to upregulate ABCA1 expression, reducing cholesterol accumulation in foam cells. Moreover, HT significantly inhibited monocyte adhesion and reduced the levels of adhesion factors (ICAM-1 and VCAM-1) and pro-inflammatory factors (IL-6 and TNF-α) in LPS-induced endothelial cells. Taken together, our findings suggest that HT, with its ability to interfere with the import and export of cholesterol, could represent a new therapeutic strategy for the treatment of atherosclerotic disease.

Endothelial dysfunction (ED) causes worse prognoses in heart failure (HF) patients treated with cardiac resynchronization therapy (CRTd). ED triggers the downregulation of microRNA-130 (miR-130a-5p), which targets endothelin-1 (ET-1). Thus, we evaluated ED and the response to CRTd by assessing miR-130a-5p and ET-1 serum levels. We designed a prospective multi-center study with a 1-year follow-up to evaluate ED, ET-1, and miR-130a-5p in CRTd patients with ED (ED-CRTd) vs. patients without ED (NED-CRTd). Clinical outcomes were CRTd response, HF hospitalization, cardiac death, and all-cause death. At 1-year follow-up, NED-CRTd (n = 541) vs. ED-CRTd (n = 326) patients showed better clinical statuses, lower serum values of B type natriuretic peptide (BNP: 266.25 ± 10.8 vs. 297.43 ± 16.22 pg/mL; p < 0.05) and ET-1 (4.57 ± 0.17 vs. 5.41 ± 0.24 pmol/L; p < 0.05), and higher values of miR-130a-5p (0.51 ± 0.029 vs. 0.41 ± 0.034 A.U; p < 0.05). Compared with NED-CRTd patients, ED-CRTd patients were less likely to be CRTd responders (189 (58%) vs. 380 (70.2%); p < 0.05) and had higher rates of HF hospitalization (115 (35.3%) vs. 154 (28.5%); p < 0.05) and cardiac deaths (30 (9.2%) vs. 21 (3.9%); p < 0.05). Higher miR-130a-5p levels (HR 1.490, CI 95% [1.014–2.188]) significantly predicted CRTd response; the presence of hypertension (HR 0.818, CI 95% [0.669–0.999]), and displaying higher levels of ET-1 (HR 0.859, CI 98% [0.839–0.979]), lymphocytes (HR 0.820, CI 95% [0.758–0.987]), LVEF (HR 0.876, CI 95% [0.760–0.992]), and ED (HR 0.751, CI 95% [0.624–0.905]) predicted CRTd non-response. Higher serum miR-130a-5p levels (HR 0.332, CI 95% [0.347–0.804]) and use of ARNI (HR 0.319, CI 95% [0.310–0.572]) predicted lower risk of HF hospitalization, whereas hypertension (HR 1.818, CI 95% [1.720–2.907]), higher BNP levels (HR 1.210, CI 95% [1.000–1.401]), and presence of ED (HR 1.905, CI 95% [1.238–2.241]) predicted a higher risk of HF hospitalization. Hence, serum miR-130a-5p could identify different stages of ED and independently predict CRTd response, therefore representing a novel prognostic HF biomarker.

Idiopathic pulmonary fibrosis (IPF) is a form of chronic and irreversible fibrosing interstitial pneumonia of unknown etiology. Although antifibrotic treatments have shown a reduction of lung function decline and a slow disease progression, IPF is characterize by a very high mortality. Emerging evidence suggests that IPF increases the risk of lung carcinogenesis. Both diseases show similarities in terms of risk factors, such as history of smoking, concomitant emphysema, and viral infections, besides sharing similar pathogenic pathways. Lung cancer (LC) diagnosis is often difficult in IPF patients because of the diffuse lung injuries and abnormalities due to the underlying fibrosis. This is reflected in the lack of optimal therapeutic strategies for patients with both diseases. For this purpose, we performed a proteomic study on bronchoalveolar lavage fluid (BALF) samples from IPF, LC associated with IPF (LC-IPF) patients, and healthy controls (CTRL). Molecular pathways involved in inflammation, immune response, lipid metabolism, and cell adhesion were found for the dysregulated proteins in LC-IPF, such as TTHY, APOA1, S10A9, RET4, GDIR1, and PROF1. The correlation test revealed a relationship between inflammation- and lipid metabolism-related proteins. PROF1 and S10A9, related to inflammation, were up-regulated in LC-IPF BAL and serum, while APOA1 and APOE linked to lipid metabolism, were highly abundant in IPF BAL and low abundant in IPF serum. Given the properties of cytokine/adipokine of the nicotinamide phosphoribosyltransferase, we also evaluated its serum abundance, highlighting its down-regulation in LC-IPF. Our retrospective analyses of BAL samples extrapolated some potential biomarkers of LC-IPF useful to improve the management of these contemporary pathologies. Their differential abundance in serum samples permits the measurement of these potential biomarkers with a less invasive procedure.

Climate change is deeply impacting the food chain production, lowering quality and yield. In this context, the international scientific community has dedicated many efforts to enhancing resilience and sustainability in agriculture. Italy is among the main European producers of several fruit trees; therefore, national research centers and universities undertook several initiatives to maintain the specificity of the ‘Made in Italy’ label. Despite their importance, fruit crops are suffering from difficulties associated with the conventional breeding approaches, especially in terms of financial commitment, land resources availability, and long generation times. The ‘new genomic techniques’ (NGTs), renamed in Italy as ‘technologies for assisted evolution’ (TEAs), reduce the time required to obtain genetically improved cultivars while precisely targeting specific DNA sequences. This review aims to illustrate the role of the Italian scientific community in the use of NGTs, with a specific focus on Citrus, grapevine, apple, pear, chestnut, strawberry, peach, and kiwifruit. For each crop, the key genes and traits on which the scientific community is working, as well as the technological improvements and advancements on the regeneration of local varieties, are presented. Lastly, a focus is placed on the legal aspects in the European and in Italian contexts.

Living organisms do not disregard the laws of thermodynamics and must therefore consume energy for their survival. In this way, cellular energy exchanges, which aim above all at the production of ATP, a fundamental molecule used by the cell for its metabolisms, favor the formation of waste products that, if not properly disposed of, can contribute to cellular aging and damage. Numerous genes have been linked to aging, with some favoring it (gerontogenes) and others blocking it (longevity pathways). Animal model studies have shown that calorie restriction (CR) may promote longevity pathways, but given the difficult application of CR in humans, research is investigating the use of CR-mimetic substances capable of producing the same effect. These include some phytonutrients such as oleuropein, hydroxytyrosol, epigallo-catechin-gallate, fisetin, quercetin, and curcumin and minerals such as magnesium and selenium. Some of them also have senolytic effects, which promote the apoptosis of defective cells that accumulate over the years (senescent cells) and disrupt normal metabolism. In this article, we review the properties of these natural elements that can promote a longer and healthier life.

Plant biomass is the most abundant renewable resource in nature. In a circular economy perspective, the implementation of its bioconversion into fermentable sugars is of great relevance. Lytic Polysaccharide MonoOxygenases (LPMOs) are accessory enzymes able to break recalcitrant polysaccharides, boosting biomass conversion and subsequently reducing costs. Among them, auxiliary activity of family 9 (AA9) acts on cellulose in synergism with traditional cellulolytic enzymes. Here, we report for the first time, the production of the AA9 LPMOs from the mesophilic Trichoderma reesei (TrAA9B) and the thermophilic Thermoascus aurantiacus (TaAA9B) microorganisms in tobacco by plastid transformation with the aim to test this technology as cheap and sustainable manufacture platform. In order to optimize recombinant protein accumulation, two different N-terminal regulatory sequences were used: 5′ untranslated region (5′-UTR) from T7g10 gene (DC41 and DC51 plants), and 5′ translation control region (5′-TCR), containing the 5′-UTR and the first 14 amino acids (Downstream Box, DB) of the plastid atpB gene (DC40 and DC50 plants). Protein yields ranged between 0.5 and 5% of total soluble proteins (TSP). The phenotype was unaltered in all transplastomic plants, except for the DC50 line accumulating AA9 LPMO at the highest level, that showed retarded growth and a mild pale green phenotype. Oxidase activity was spectrophotometrically assayed and resulted higher for the recombinant proteins without the N-terminal fusion (DC41 and DC51), with a 3.9- and 3.4-fold increase compared to the fused proteins.