Pharmacological and Non-Pharmacological Agents versus Bovine Colostrum Supplementation for the Management of Bone Health Using an Osteoporosis-Induced Rat Model

Osteoporosis is defined by loss of bone mass and deteriorated bone microarchitecture. The present study compared the effects of available pharmacological and non-pharmacological agents for osteoporosis [alendronate (ALE) and concomitant supplementation of vitamin D (VD) and calcium (Ca)] with the effects of bovine colostrum (BC) supplementation in ovariectomized (OVX) and orchidectomized (ORX) rats. Seven-month-old rats were randomly allocated to: (1) placebo-control, (2) ALE group (7.5 μg/kg of body weight/day/5 times per week), (3) VD/Ca group (VD: 35 μg/kg of body weight/day/5 times per week; Ca: 13 mg/kg of body weight/day/3 times per week), and (4) BC supplementation (OVX: 1.5 g/day/5 times per week; ORX: 2 g/day/5 times per week). Following four months of supplementation, bone microarchitecture, strength and bone markers were evaluated. ALE group demonstrated significantly higher Ct.OV, Ct.BMC, Tb.Th, Tb.OV and Tb.BMC and significantly lower Ct.Pr, Tb.Pr, Tb.Sp, Ct.BMD and Tb.BMD, compared to placebo (p < 0.05). BC presented significantly higher Ct.Pr, Ct.BMD, Tb.Pr, Tb.Sp, and Tb.BMD and significantly lower Ct.OV, Ct.BMC, Tb.Th, Tb.OV and Tb.BMC compared to ALE in OVX rats (p < 0.05). OVX rats receiving BC experienced a significant increase in serum ALP and OC levels post-supplementation (p < 0.05). BC supplementation may induce positive effects on bone metabolism by stimulating bone formation, but appear not to be as effective as ALE

Ο ρόλος του λιπώδους ιστού στα μεταβολικά νοσήματα και μοριακά μονοπάτια που επηρεάζουν τη λειτουργία του

The aim of this thesis is to expand the current knowledge on the role of adipose tissue (AT) on metabolic disorders and to research the molecular pathways that affect its function. In this light, we performed two narrative reviews. Initially, the crosstalk between AT and the vascular system and how this can affect the onset and progression of metabolic diseases was investigated. In a second narrative review, the angiogenic factors that affect AT and pharmacological approaches that could be used in the treatment of obesity and its accompanying co-morbidities were described. Moreover, current bioengineering approaches that could be employed in the fight against obesity and consequent metabolic diseases are suggested. Hence this information led to testing the properties of a new, natural, Histogel-based bioscaffold to support the proliferation and differentiation of white AT to into brown-like AT, a more metabolically active version of fat. As AT is affected by the cytokines of other tissues, irisin, a myokine, and its role on metabolic disorders was explored. Based on accumulating evidence on the implication of irisin in cancer a systematic review was performed, with the aim to elucidate how irisin is involved in the progression of cancer and if it can constitute a valid biomarker for its diagnosis. Finally, the role of irisin on AT and how it affects its metabolic state was investigated. The results of the current thesis suggest that the communication between vascular system and adipose tissue should be considered, when pharmacological approaches are employed for the treatment of metabolic disorders. Moreover, the use of biological scaffolds is suggested, that could result in increased gain of brown adipose tissue (increased metabolic activity) and be part of the treatment of obesity and accompanying metabolic disorders. The results demonstrate that irisin is involved in cancer by modulating cell proliferation and tumor invasion through several signaling pathways and can also be a good biomarker for the diagnosis of cancer. Finally, the role of irisin in the regulation of lipolysis and thermogenesis in vitro is presented. Taken together, these results indicate that adipose tissue is an excellent medium for pharmacological or biomedical interventions for the treatment of metabolic disorders.Στόχος της παρούσας διατριβής είναι η διεύρυνση της τρέχουσας γνώσης σχετικά με το ρόλο του λιπώδους ιστού στις μεταβολικές διαταραχές και η έρευνα των μοριακών μονοπατιών που επηρεάζουν τη λειτουργία του. Υπό αυτό το πρίσμα, πραγματοποιήθηκαν δύο βιβλιογραφικές μελέτες. Αρχικά, διερευνήθηκε πώς η αλληλεπίδραση μεταξύ του λιπώδους ιστού και του αγγειακού συστήματος μπορεί να επηρεάσει την εμφάνιση και την εξέλιξη των μεταβολικών ασθενειών. Στη δεύτερη ανασκόπησή που πραγματοποιήθηκε, περιγράφονται οι αγγειογόνοι παράγοντες που επηρεάζουν το λιπώδη ιστό και οι φαρμακολογικές προσεγγίσεις που θα μπορούσαν να χρησιμοποιηθούν στη θεραπεία της παχυσαρκίας και των συνοδών νοσηροτήτων της. Επιπλέον, προτείνονται τρέχουσες προσεγγίσεις της εμβιομηχανικής που θα μπορούσαν να χρησιμοποιηθούν για την καταπολέμηση της παχυσαρκίας και των συνακόλουθων μεταβολικών ασθενειών. Αυτές οι πληροφορίες οδήγησαν στη δοκιμή των ιδιοτήτων ενός νέου, φυσικού, βιολογικού ικριώματος με βάση το Histogel που υποστηρίζει τον πολλαπλασιασμό και τη διαφοροποίηση του λευκού λίπους σε μπεζ, μια μεταβολικά πιο ενεργή εκδοχή του λίπους. Καθώς ο λιπώδης ιστός επηρεάζεται από τις κυτοκίνες άλλων ιστών, ερευνήθηκε η ιρισίνη, μια μυοκίνη, και ο ρόλος της στις μεταβολικές διαταραχές. Με βάση τη βιβλιογραφία σχετικά με το ρόλο της ιρισίνης στον καρκίνο, πραγματοποιήθηκε μια συστηματική ανασκόπηση της βιβλιογραφίας, με στόχο να διευκρινιστεί πώς η ιρισίνη εμπλέκεται στην εξέλιξη του καρκίνου και εάν μπορεί να αποτελέσει έγκυρο βιοδείκτη για τη διάγνωσή του. Τέλος, διερευνήθηκε ο ρόλος της ιρισίνης στο λιπώδη ιστό και πώς επηρεάζει τη μεταβολική του κατάσταση. Τα αποτελέσματα της παρούσας διατριβής προτείνουν ότι η επικοινωνία μεταξύ αγγειακού συστήματος και λιπώδους ιστού θα πρέπει να λαμβάνεται υπόψη, όταν χρησιμοποιούνται φαρμακολογικές προσεγγίσεις για τη θεραπεία μεταβολικών διαταραχών. Επιπλέον, προτείνεται η χρήση βιολογικών ικριωμάτων, που θα μπορούσαν να οδηγήσουν σε αύξηση του καφέ λιπώδους ιστού (αυξημένη μεταβολική δραστηριότητα) και να αποτελούν μέρος της θεραπείας της παχυσαρκίας και των συνοδών μεταβολικών διαταραχών. Φαίνεται επίσης ότι η ιρισίνη εμπλέκεται στον καρκίνο, ρυθμίζοντας τον κυτταρικό πολλαπλασιασμό και την μετάσταση των όγκων, μέσω διαφόρων σηματοδοτικών μονοπατιών και μπορεί επίσης να είναι ένας καλός βιοδείκτης για τη διάγνωση του καρκίνου. Τέλος, φαίνεται ότι η ιρισίνη μπορεί να ρυθμίσει τη λιπόλυση και τη θερμογένεση in vitro. Συνοψίζοντας, τα παραπάνω αποτελέσματά δείχνουν ότι ο λιπώδης ιστός είναι ένα εξαιρετικό μέσο για φαρμακολογικές ή βιοϊατρικές παρεμβάσεις σχετικά με τη θεραπεία των μεταβολικών διαταραχών. Lo scopo di questa tesi è di espandere le conoscenze attuali sul ruolo del tessuto adiposo (AT) sui disordini metabolici e di ricercare i pathways molecolari che ne influenzano la funzione. In questo contesto abbiamo eseguito due revisioni narrative. Nella prima review abbiamo indagato su come il crosstalk tra AT e sistema vascolare possa influire sull’esordio e sulla progressione dei disordini metabolici. Nella nostra seconda review, abbiamo descritto i fattori angiogenici che influenzano l’AT e gli approcci farmacologici che potrebbero essere utilizzati nella lotta contro l’obesità e le sue co- morbilità associate. Inoltre, proponiamo degli attuali approcci di bioingegneria che potrebbero essere impiegati nella lotta contro l’obesità e i disordini metabolici associati. Queste informazioni ci hanno portato a testare le proprietà di un nuovo, naturale Histogel-based bioscaffold per supportare la proliferazione e il differenziamento del tessuto adiposo bianco in tessuto adiposo brown-like, un tipo di grasso più metabolicamente attivo. Siccome l’AT è influenzato dalle citochine di altri tessuti, noi abbiamo indagato sull’irisina, una miochina, e sul suo ruolo nei disordini metabolici. Basandoci su evidenze crescenti sul ruolo dell’irisina nel cancro abbiamo eseguito una review sistematica con l’obiettivo di chiarire come l’irisina sia coinvolta nella progressione del cancro e se possa costituire un valido biomarker per la sua diagnosi. Infine, abbiamo indagato sul ruolo dell’irisina nell’AT e su come influisca sul suo stato metabolico. I risultati della presente tesi suggeriscono che la comunicazione tra sistema vascolare e tessuto adiposo dovrebbe essere considerata, quando approcci farmacologici sono impiegati per il trattamento di disordini metabolici. Inoltre, noi proponiamo l’uso di scaffolds biologici nella medicina rigenerativa, che possono risultare nell’incremento di tessuto adiposo bruno (attività metabolica incrementata) ed essere parte del trattamento dell’obesità e dei disordini metabolici associati. Mostriamo inoltre che l’irisina è coinvolta nel cancro modulando la proliferazione cellulare e l’invasività tumorale attraverso alcuni pathways di segnalazione cellulare e che può anche essere un buon biomarker per la diagnosi tumorale. Infine, mostriamo che l’irisina può regolare lipolisi e termogenesi in vitro. Complessivamente, questi risultati indicano che il tessuto adiposo è un medium eccellente per interventi farmacologici o biomedicali per il trattamento di disordini metabolici.

Adipose Tissue Metabolism in Response to Food Intake

The quality and quantity of the food we consume have a major impact on our general health and longevity [...

Development of an HPLC-DAD Method for the Extraction and Quantification of 5-Fluorouracil, Uracil, and 5-Fluorodeoxyuridin Monophosphate in Cells and Culture Media of Lactococcus lactis

The drug 5-fluorouracil (5-FU) is a common cancer chemotherapeutic, presenting toxicity. Mild toxicity is treated with administration of probiotics. The interaction of these probiotics with the drug may have a crucial effect on its therapeutic efficacy. In the present work, a method for the quantification of uracil, 5-FU, and its active metabolite 5-fluorodeoxyuridin monophosphate in cells and culture medium of the probiotic L. lactis is presented. Extraction using H2O containing 0.05% v/v formic acid (1:5 v/v) was followed by ammonium sulphate protein precipitation and SPE. Analysis was conducted in a Nucleosil column using a gradient of water, formic acid, and acetonitrile. Calibration curves were constructed for 5-FU (5&ndash;100 &mu;g/mL), uracil (5&ndash;20 &mu;g/mL), and 5-fluorodeoxyuridin monophosphate (5&ndash;20 &mu;g/mL) using 5-bromouracil as the internal standard (R2 &ge; 0.999). The photodegradation of 5-FU amounted to 36.2% at 96 h. An administration experiment in the dark revealed a decline in 5-FU concentration in the culture media (88.3%) and uptake by the cells, while the uracil and FdUMP levels increased in the cells. The inactive metabolite 5,6 dihydrofluorouracil was detected in the medium. Our results demonstrate that uptake and metabolism of 5-FU in L. lactis cells leads to a decline in the drug levels and in the formation of both the active and the inactive metabolites of the drug

Development of an HPLC-DAD Method for the Extraction and Quantification of 5-Fluorouracil, Uracil, and 5-Fluorodeoxyuridin Monophosphate in Cells and Culture Media of <i>Lactococcus lactis</i>

The drug 5-fluorouracil (5-FU) is a common cancer chemotherapeutic, presenting toxicity. Mild toxicity is treated with administration of probiotics. The interaction of these probiotics with the drug may have a crucial effect on its therapeutic efficacy. In the present work, a method for the quantification of uracil, 5-FU, and its active metabolite 5-fluorodeoxyuridin monophosphate in cells and culture medium of the probiotic L. lactis is presented. Extraction using H2O containing 0.05% v/v formic acid (1:5 v/v) was followed by ammonium sulphate protein precipitation and SPE. Analysis was conducted in a Nucleosil column using a gradient of water, formic acid, and acetonitrile. Calibration curves were constructed for 5-FU (5–100 μg/mL), uracil (5–20 μg/mL), and 5-fluorodeoxyuridin monophosphate (5–20 μg/mL) using 5-bromouracil as the internal standard (R2 ≥ 0.999). The photodegradation of 5-FU amounted to 36.2% at 96 h. An administration experiment in the dark revealed a decline in 5-FU concentration in the culture media (88.3%) and uptake by the cells, while the uracil and FdUMP levels increased in the cells. The inactive metabolite 5,6 dihydrofluorouracil was detected in the medium. Our results demonstrate that uptake and metabolism of 5-FU in L. lactis cells leads to a decline in the drug levels and in the formation of both the active and the inactive metabolites of the drug

Characteristics of the Protocols Used in Electrical Pulse Stimulation of Cultured Cells for Mimicking In Vivo Exercise: A Systematic Review, Meta-Analysis, and Meta-Regression

While exercise benefits a wide spectrum of diseases and affects most tissues and organs, many aspects of its underlying mechanistic effects remain unsolved. In vitro exercise, mimicking neuronal signals leading to muscle contraction in vitro, can be a valuable tool to address this issue. Following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines for this systematic review and meta-analysis, we searched EMBASE and PubMed (from database inception to 4 February 2022) for relevant studies assessing in vitro exercise using electrical pulse stimulation to mimic exercise. Meta-analyses of mean differences and meta-regression analyses were conducted. Of 985 reports identified, 41 were eligible for analysis. We observed variability among existing protocols of in vitro exercise and heterogeneity among protocols of the same type of exercise. Our analyses showed that AMPK, Akt, IL-6, and PGC1a levels and glucose uptake increased in stimulated compared to non-stimulated cells, following the patterns of in vivo exercise, and that these effects correlated with the duration of stimulation. We conclude that in vitro exercise follows motifs of exercise in humans, allowing biological parameters, such as the aforementioned, to be valuable tools in defining the types of in vitro exercise. It might be useful in transferring obtained knowledge to human research

The impact of adipokines on vascular networks in adipose tissue

Adipose tissue (AT) is a highly active and plastic endocrine organ. It secretes numerous soluble molecules known as adipokines, which act locally to AT control the remodel and homeostasis or exert pleiotropic functions in different peripheral organs. Aberrant production or loss of certain adipokines contributes to AT dysfunction associated with metabolic disorders, including obesity. The AT plasticity is strictly related to tissue vascularization. Angiogenesis supports the AT expansion, while regression of blood vessels is associated with AT hypoxia, which in turn mediates tissue inflammation, fibrosis and metabolic dysfunction. Several adipokines can regulate endothelial cell functions and are endowed with either pro- or anti-angiogenic properties. Here we address the role of adipokines in the regulation of angiogenesis. A better understanding of the link between adipokines and angiogenesis will open the way for novel therapeutic approaches to treat obesity and metabolic diseases

The contribution of obesogenic behavior on global warming

Background: obesogenic behaviors, unbalanced diet and sedentary behaviors are continuously rising in our society, posing serious threats to public health (1). Several studies analyzed the effects of how the obesogenic behavior exacerbating metabolic and pathological alterations supports pathological conditions (2,3,4) while little is known about the association between obesogenic behaviors and global warming. To address this relation, we performed a systematic review. Material and methods: obesogenic behavior, non-obesogenic behavior, obesity, GHG emissions, and adverse climate change were used to develop the research algorithm which was applied to four databases: including PubMed, Scopus, Web of Science and EBSCO. The algorithm was refined and its sensitivity was tested on a pre-defined Golden Library. Results: The search yielded more than 36,000 results. Exploiting a pre-defined Golden Library, the refined search algorithm showed a sensitivity of approx 80%. Papers were retrieved in PubMed (4687), Scopus (9503), Web of Science (3416) and EBSCO (18898). The databases were explored without applying any limiting timeframe. Results were filtered to include only peer-reviewed results published in english. Only studies on humans were considered. The results have been screened to remove duplicates and preliminary assessment suggests that an association between obesogenic habits and global warming may exist. These results will be analyzed further to confirm this relation. Conclusions: our study will provide possible evidence of how preventing obesogenic habits may mitigate adverse climate change

Natural Histogel-Based Bio-Scaffolds for Sustaining Angiogenesis in Beige Adipose Tissue

In the treatment of obesity and its related disorders, one of the measures adopted is weight reduction by controlling nutrition and increasing physical activity. A valid alternative to restore the physiological function of the human body could be the increase of energy consumption by inducing the browning of adipose tissue. To this purpose, we tested the ability of Histogel, a natural mixture of glycosaminoglycans isolated from animal Wharton jelly, to sustain the differentiation of adipose derived mesenchymal cells (ADSCs) into brown-like cells expressing UCP-1. Differentiated cells show a higher energy metabolism compared to undifferentiated mesenchymal cells. Furthermore, Histogel acts as a pro-angiogenic matrix, induces endothelial cell proliferation and sprouting in a three-dimensional gel in vitro, and stimulates neovascularization when applied in vivo on top of the chicken embryo chorioallantoic membrane or injected subcutaneously in mice. In addition to the pro-angiogenic activity of Histogel, also the ADSC derived beige cells contribute to activating endothelial cells. These data led us to propose Histogel as a promising scaffold for the modulation of the thermogenic behavior of adipose tissue. Indeed, Histogel simultaneously supports the acquisition of brown tissue markers and activates the vasculature process necessary for the correct function of the thermogenic tissue. Thus, Histogel represents a valid candidate for the development of bioscaffolds to increase the amount of brown adipose tissue in patients with metabolic disorders