Isolation, characterization and biologica! evaiuation of bioactive compounds from compiex matrices with high nutraceutica! vaiue

2018 - 2019My PhD project based its main objective on the isolation, the characterization and the biological evaluation of bioactive compounds deriving from food matrices of animal and vegetable origin typical of the Mediterranean Area. During the first year, the research activity was focused on the identification of bioaccessible peptides generated as a result of a simulated gastrointestinal digestion process of six commercial buffalo milk dairy products. The scientific approach allowed us to identify 342 peptides deriving from the hydrolysis of caseins and whey protein of buffalo milk. However, as far as we know, only one third of the identified peptides reported a biological activity in the literature, so we conducted a rational biological characterization of the six gastro-enteric digests under study. In this regard, the six digested gastro-enteric buffalo-based milk were tested on intestinal epithelial cells (IEC-6) under induced oxidative stress conditions by using hydrogen peroxide. Among all the matrices investigated, at the maximum concentration tested, the buffalo Ricotta determined an increased reduction in the release of Radical oxygen species (ROS). In order to identify the main peptide molecules responsible for the biological activity, the GI digest of buffalo Ricotta has been simplified using both membrane filters with cut-off lower than 1 kDa and the use of semi-prep RP-HPLC. The obtained fractions were tested in the experimental conditions described above and the most active fractions were subjected to high-performance liquid chromatography experiments coupled with high-resolution mass spectrometry studies, through which it was possible to identify two peptides: BRP1 and BRP2 deriving from hydrolysis of β-lactoglobulin. Both peptides were able to reduce the release of ROS by determining the nuclear translocation of NRF-2 and increasing the expression of cytoprotective enzymes regulated by itself. Based on the results obtained during my second year of the PhD, the research activity was focused on the study of the bioavailability of the BRP1 and BRP2 peptides, using, for this purpose, Caco-2 cell monolayers. The results obtained showed that unlike BRP1, the BRP2 peptide permeated through the cell monolayer mainly using a passive diffusion mechanism. From ex vivo biological assays, the peptide showed an antioxidant and hypotensive activity through the reduction of the Ang II-induced vasoconstriction process, and by both the decrease of NADPH-oxidase activity and with the nuclear translocation of Nrf-2, that on the other hand, increased the expression of cytoprotective enzymes. The research activity of my third year of the PhD was focused on the synthesis and subsequent biological evaluation in ex vivo and in vivo experimental models of the most abundant peptide identified of the gastro-enteric digested buffalo milk ice cream. This matrix from previous ex vivo studies has determined a reduction in ROS release. Thus, in order to identify the main peptide molecules responsible of the biological activity observed, my research activity focused on the most abundant pentapeptide (PG1) identified in GI of buffalo ice cream. In order to ensure a sufficient amount of peptide for further biological tests in ex vivo and in vivo, the PG1 peptide was synthesized by solid phase peptide synthesis experiments in according to the Fmoc strategy. The next phase of the study was centered on the evaluation of the pharmacokinetic properties of the PG1 peptide. In particular, the pentapeptide showed good stability to digestive and microsomal enzymes. The bioavailability of the PG1 peptide was again evaluated using a Caco-2 cell monolayer and the results obtained showed that the peptide permeated the cell monolayer, mainly using a passive diffusion mechanism. We moved to evaluate the local activity of the PG1 peptide, through ex vivo studies conducted on mesenteric arteries, to evaluate a potential reduction in the Ang II-induced vasoconstriction process. The results showed that at increasing doses of Ang II the vasoconstrictor response was inhibited in a dose-dependent manner after pretreatment with the PG1 peptide. The next phase of the study then included an in vivo evaluation for peptide. Specifically, the efficacy of the peptide on wild type mice under conditions of Ang II-induced hypertension was evaluated. The results obtained showed that the administration of the PG1 8mg / Kg peptide for 14 days, via gavage, determined a normalization of blood pressure values. During the second phase of my third year of my PhD project, the research activity focused on the evaluation of the anti-inflammatory and antioxidant properties of Solanum Tuberosum. The bioaccessible peptides generated following the oral ingestion of the plant matrix determined a reduction in the enzymatic activity of iNOS and COX2 in IEC-6 cell lines in inflammatory conditions induced by LPS and INT- ℽ. Finally, the evaluation of the antioxidant activity of dehydrated potato polyphenol extract was carried out using pre-column off-line DPPH.XXXII cicl

The effects of topical application of melatonin on periodontal Disease in diabetic patients

Periodontal disease is a chronic disease, affects the supporting tissues of the tooth. The clinical manifestation varies from gingivitis to periodontitis. Periodontal disease is caused by bacterial infection that release toxins. The imbalance between oxidants and antioxidants determines the progression of periodontal disease. Melatonin (MEL), (N-Acetyl-5-methoxytrittamine) is a hormone in the human body. Its production takes place in various organs including the retina, gastrointestinal tract, bone marrow, leukocytes, lymphocytes, skin, and principally pineal gland. Its main function is the regulation of the circadian and seasonal rhythm, body weight, reproduction, bone metabolism, and tumor growth. An important function of melatonin is the ability to reduce oxidative stress. The aim of this review is to evaluate the necessary effects on melatonin on the progression of periodontal disease and diabetes. The purpose of this review is to answer to the following questions using a PICO method (P: patient problem/population; I: intervention; C: comparison; O: outcome): (1) Can adding melatonin in diabetic and periodontal patients lead to improved periodontal health? (2) Does melatonin also affect the control of blood sugar levels in the diabetic patient? The study was conducted utilizing the main scientific databases (PubMed, MEDLINE, and WEB of SCIENCE). The time window considered for the electronic search was from March 1, 2007, to March 1, 2020. The following inclusion criterion was used: Articles in English, human studies and clinical trials. Two independent people search with the same keywords all article and select the article founding. The risk of bias in this phase is solved by an independent author that conduct the same search. We can hypothesize that melatonin may indirectly help control blood sugar levels. Further studies will be needed to evaluate a direct healing effect of melatonin on diabetes

Cocaine and aortic dissection: the need for collaboration to overcome the underreporting bias

The dissection of the aorta is a serious and potentially fatal consequence of cocaine use. Nonetheless, the underlying mechanisms and characteristics of this phenomenon remain to be deeply studied. The autopsy case of a 46-year-old white male found irresponsive and unconscious in his house and had a history of abusing cocaine is presented. Autopsy findings showed aortic arch and thoracic aorta of regular calibre, with evidence of a mid-adventitial dissecting aneurysm of the ascending intrapericardial portion of the aorta. Forensic toxicological analyses evidenced a positivity to cocaine and its main metabolite, benzoylecgonine, in all fluids (peripheral blood, bile and urine), liver and brain homogenates. Data also evidenced a positivity to alcohol, confirming a past history of intake. The presented case confirms the connection between cocaine addiction and the risk of aortic dissection, emphasizing the need for increased knowledge about the risks connected to this drug. It is emphasized the need of prompt examination and effective treatment of patients exhibiting cardiac symptoms linked to cocaine use, as well as the need of an accurate anamnesis to evidence use/abuse of other cardiotoxic substances

Effect of Toothpaste on the Surface Roughness of the Resin-Contained CAD/CAM Dental Materials: A Systematic Review

Background: The purpose of this review is to describe the possible effect of toothbrushing on surface roughness of resin-contained CAD/CAM materials. Methods: Systematic literature search for articles published in peer-reviewed journals between January 2000 and February 2020 has been conducted, which evaluated the effect of brushing on surface roughness of resin-contained CAD/CAM dental materials. The research was conducted in Scopus, PubMed/Medline, Web of Science, Embase, and Science Direct using a combination of the following MeSH/Emtree terms: “brushing”, “resin-based”, “dental”, “CAD/CAM”, and “surface roughness”. Results: A total of 249 articles were found in the search during initial screening. Fifty-five articles were selected for the full-text evaluation after the steps of reading of abstract/title and remotion of duplicate. Only six articles fulfilled the inclusion criteria. The Cohen’s Kappa agreement test showed an index of 0.91 for full-text. Discussion: Four of five selected articles identified an increase of surface roughness on resin-contained CAD/CAM materials after toothbrushing. Although all the articles examined used different toothpastes with no homogeneous relative dentine abrasivity (RDA) and cycles of brushing, the findings are about the same. The possible reason is attributable to the compositions of the resin-contained CAD/CAM materials. Conclusions: The surface roughness of most resin-contained CAD/CAM materials was affected by artificial toothbrushing. Correct knowledge of the composition of the dental material and toothpastes is fundamental to avoid an increase of surface roughness on prosthetic rehabilitation

Distinct mechanisms mediate X chromosome dosage compensation in Anopheles and Drosophila

Sex chromosomes induce potentially deleterious gene expression imbalances that are frequently corrected by dosage compensation (DC). Three distinct molecular strategies to achieve DC have been previously described in nematodes, fruit flies, and mammals. Is this a consequence of distinct genomes, functional or ecological constraints, or random initial commitment to an evolutionary trajectory? Here, we study DC in the malaria mosquito Anopheles gambiae. The Anopheles and Drosophila X chromosomes evolved independently but share a high degree of homology. We find that Anopheles achieves DC by a mechanism distinct from the Drosophila MSL complex–histone H4 lysine 16 acetylation pathway. CRISPR knockout of Anopheles msl-2 leads to embryonic lethality in both sexes. Transcriptome analyses indicate that this phenotype is not a consequence of defective X chromosome DC. By immunofluorescence and ChIP, H4K16ac does not preferentially enrich on the male X. Instead, the mosquito MSL pathway regulates conserved developmental genes. We conclude that a novel mechanism confers X chromosome up-regulation in Anopheles. Our findings highlight the pluralism of gene-dosage buffering mechanisms even under similar genomic and functional constraints

Pediatric poisoning management: How clinical practice can benefit from forensic approach

Pediatric population represents the most vulnerable and at risk for unintentional poisoning, with children younger than 6 years old accounting for nearly half of poison exposures. Poisoning is a time-dependent emergency. The need to reach a scientific agreement on diagnostic protocol and treatment seems to be crucial to reduce morbidity and mortality. Starting from a buprenorphine pediatric intoxication case, this article highlights the limits and pitfalls of the traditional diagnostic approach. Diagnosis of drug intoxication was achieved after several days when an in-depth diagnostic investigation became necessary and complete forensic toxicological analyses were performed. Results evidenced an alarming lack of an unequivocal diagnostic protocol in case of suspect intoxication in structures not provided with a forensic toxicological service/unit. Collection of biological specimens according to forensic protocols at hospitalization plays a paramount role in the definitive diagnosis of intoxication. A diagnostic algorithm that focuses on medical history and biological specimen collection timing is herein proposed, in order to unify emergency approaches to the suspected poisoned child

Oral Adverse Events Associated with BRAF and MEK Inhibitors in Melanoma Treatment: A Narrative Literature Review

Background: Melanoma cancer represents the most lethal type of skin cancer originating from the malignant transformation of melanocyte cells. Almost 50% of melanomas show the activation of BRAF mutations. The identification and characterization of BRAF mutations led to the development of specific drugs that radically changed the therapeutic approach to melanoma. Methods: We conducted a narrative review of the literature according to a written protocol before conducting the study. This article is based on previously conducted studies. We identified articles by searching electronic databases (Medline, Google Scholar and PubMed). We used a combination of “melanoma”, “Braf-Mek inhibitors”, “ targeted therapy” and “oral side effects”. Results: Eighteen studies were reported in this article showing the relationship between the use of targeted therapy in melanoma cancer and the development of oral side effects, such as mucositis, hyperkeratosis and cellular proliferation. Conclusion: Targeted therapy plays an important role in the treatment of melanoma cancer, showing a notable increase in response rate, prolonged progression-free survival and overall survival in BRAF-mutated melanoma patients. Oral side effects represent a common finding over the course of treatment. However, these adverse effects can be easily managed in a multidisciplinary approach involving collaboration between medical oncologists and dental doctors

Novel Anticancer Drug 5H-pyro[3,2-a] Phenoxazin-5-one (PPH) Regulates lncRNA HOTAIR and HOXC genes in Human MCF-7 Cells

Breast cancer in women is the second most commonly cancer, after skin cancer. The percentage of mortalityrisk for breast cancer is 1 in 37 women (2.7%), which makes breast cancer represent the second cause of cancerdeath in women. Recently, new research based on previously published work in systemic chemotherapy andendocrine therapy field, have improved the incidence rates. The quinonic nucleus is common to many naturaland synthetic products associated with anticancer and antibacterial activities, these compounds are typicallyDNA-intercalating agents. The Class I Homeobox genes (HOX in human and hox in mouse) control embryonicdevelopment and specific determination of positional identity anteroposterior axis of the human body. The HOXgenes, are 39 transcription factors related to morphological, physiological disease. It has been demonstratedthat any deregulation into the network is able to induce neoplastic transformation. Particularly, HOXC locuscollaborating with lncRNA HOTAIR play a key role in breast cancer. In this study, our group evaluated the chemical and metabolic stability of new anticancer molecule 5H-pyro[3,2-a] phenoxazin-5-one (PPH). In a recent paper, we have already demonstrated that a new and potent anticancersynthetic iminoquinone, the 5H-pyrido[3,2-a]phenoxazin-5-one (PPH), is able to inhibit a large number oflymphoblastoid and solid-tumor-derived cells at submicromolar concentrations. Based on our previous research, we decided to analyze the cytotoxic effect and capability of PPH to control thelncRNA HOTAIR and HOXC locus gene expression in human breast cancer cells MCF-7, in order to demonstrateits role like potential new breast cancer antitumor drug

Investigation of the Accuracy of Four Intraoral Scanners in Mandibular Full-Arch Digital Implant Impression: A Comparative In Vitro Study

background: we compare the accuracy of new intraoral scanners (IOSs) in full-arch digital implant impressions. methods: a master model with six scan bodies was milled in poly(methyl methacrylate), measured by using a coordinate measuring machine, and scanned 15 times with four IOSs: primescan, medit i500, Vatech EZ scan, and iTero. the software was developed to identify the position points on each scan body. The 3D position and distance analysis were performed. Results: The average and ± standard deviation of the 3D position analysis was 29 μm ± 6 μm for PrimeScan, 39 μm ± 6 μm for iTero, 48 μm ± 18 μm for mediti500, and 118 μm ± 24 μm for Vatech EZ scan (p < 0.05). conclusions: all IOSs are able to make a digital complete implant impression in vitro according to the average misfit value reported in literature (150 μm); however, the 3D distance analysis showed that only the primescan and iTero presented negligible systematic error sources