157 research outputs found
Prevention and treatment of dermatological diseases: optimizing the protective action and efficacy of phenylpropanoid glycosides delivered through the skin by inclusion in biodegradable nanovesicles
No full textThe skin, the human body’s largest organ, serves critical roles in defense and physiological regulation. Maintaining its integrity is essential, yet it is constantly challenged by environmental factors such as ultraviolet radiation (UVR), extreme temperatures, and chemical pollutants. As a "complete carcinogen," UVR has the unique capacity to both initiate and promote tumor development, making it a significant contributor to skin cancer, with melanoma being the most aggressive and lethal form. The global incidence of melanoma continues to rise. Although targeted therapies and immunotherapies have advanced, clinical outcomes remain suboptimal due to tumor resistance and adverse effects, underscoring the need for innovative treatment strategies. Natural antioxidants, including Phenylpropanoid Glycosides (PPGs) and Hyperforin (HPF), show promising antitumor potential among emerging thera peutic agents. PPGs, a group of over 200 plant-derived polyphenols, and HPF, an acylphloroglucinol extracted from Hypericum perforatum, exhibit comple mentary pharmacological properties, such as antioxidant, anti-inflammatory, and antitumor activities. This thesis examines the dual role of the PPGs, particularly Echinacoside and Forsythoside B, in preventing skin tumors and treating melanoma and other skin cancers. To explore the protective properties of PPGs against UVR, normal hu man epidermal melanocytes (NHEM) and normal human dermal fibroblasts (NHDF) were pre-treated with these compounds. Solar UV radiation was then simulated by exposing cultured cells to a UV lamp. The protective effects of PPGs pre-treatment were analyzed by assessing cell viability and the activation of multiple stress signaling pathways. Our findings suggest that PPGs exert a mild UV protective effect in normal cells, although statistical significance was not consistently observed. To improve their stability and bioavailability under UV and temperature exposure, PPGs were nanoencapsulated. In collaboration with the company “Nanomnia”, a natural, biodegradable polymer-based process was employed, eliminating microplastics and enhancing targeted delivery and controlled release. This approach can also mitigate the environmental impact of conven tional sunscreen components, which are associated with marine ecosystem disruption. Preliminary studies revealed a mild UV protective effect in normal cells from encapsulated PPGs. Particular attention is given to the antitumor properties of Echinacoside and Forsythoside B against BRAF-mutated melanoma cell line (FO-1) and a human skin tumor cell line (A-431). The results reveal that these compounds selectively induce cytotoxicity in malignant cells without affecting normal f ibroblasts, keratinocytes, or melanocytes, even at concentrations four times higher than their EC50 values in cancer cells. Within 48 h of in vitro administration, PPGs promote apoptosis by cleaving Poly (ADP-ribose) Polymerase-1 (PARP-1), activating Ataxia-Telangiectasia Mutated (ATM) kinase, and increasing phosphorylated H2A histone family member X (γH2AX) levels, indicating DNA damage. Additionally, the phosphorylation of p38 mitogen-activated protein kinases (MAPK) suggests that stress response signaling pathways are involved. These findings demonstrate a slight protective effect on normal skin cells following UV stress, as well as the selective cytotoxicity and apoptotic potential of Echinacoside and Forsythoside B in skin cancer cell lines, highlighting their therapeutic promise in melanoma and other skin cancer. Encapsulated PPGs were evaluated to assess their protective activity on NHDF exposed to UV-A radiation. Preliminary results indicate that lower concentrations of encapsulated PPGs revealed a remarkable protective response through morphological analysis. However, quantitative analysis need to be carried out. Furthermore, Hyperforin (HPF), an acylphloroglucinol derived from Hy pericum perforatum, exhibits a variety of pharmacological activities, including antidepressant, anti-inflammatory, antimicrobial, and antitumor effects. Our f indings demonstrate that HPF exerts potent antitumor activity against BRAF-mutated melanoma cell lines (A375, FO-1, SK-Mel-28), including inhibition of proliferation, motility, colony formation, and induction of apop tosis. Its cytotoxic mechanisms involve lipid peroxidation and disruption of iron homeostasis through downregulation of glutathione peroxidase-4 (GPX 4), cystine transporter SLC7A11, and ferritin, thereby increasing cellular susceptibility to oxidative damage. Concurrently, HPF upregulates heme oxygenase-1 (HO-1), which promotes the release of free iron. Cytofluorimetric analysis confirmed an increase in lipid peroxidation, as evidenced by enhanced BODIPY-C11 fluorescence. HPF also induces autophagy, as indicated by elevated LC3B expression. Moreover, HPF exerts cytostatic effects by modulating key cell cycle regulators, including downregulation of cyclins D1 and A2, CDK4, and phos phorylated Rb, and upregulation of P21/waf1 and activated P53. Apoptotic effects are further enhanced by reductions in Bcl-2 and Bcl-xL levels and increased cleaved PARP-1. Furthermore, HPF alters the metabolism of melanoma cells inhibiting mitochondrial enzymes expression and compro mising the mitochondrial membrane potential. Although HPF modulates TRPC6 channels and elevates cytosolic Ca2+ and Zn2+ levels, its cytostatic effects appear largely independent of these pathways. Furthermore, HPF suppresses survival signaling by reducing the activity of transcription factors nuclear factor-kappa B (NF-κB) P65 and Signal Transducer and Activator of Transcription 3 (STAT3). HPF’s antimetastatic properties further reinforce its therapeutic potential. It downregulates many invasion markers, consistent with observed reductions in cell migration and colony formation. Collectively, these findings underscore HPF’s pleiotropic antitumor properties, includ ing cytostatic, cytotoxic, and antimetastatic effects, thereby supporting its potential as a therapeutic agent in melanoma management. These findings collectively highlight the therapeutic potential of PPGs and HPF. While PPGs demonstrate a slight UV-protective effect on normal f ibroblasts and melanocytes and a selective cytotoxic effect on malignant skin cells, HPF exerts broader cytostatic, cytotoxic, and antimetastatic activities on melanoma cell lines, providing a strong foundation for future translational research and therapeutic applications
Orthogonal-Array based Design Methodology for Complex, Coupled Space Systems
No full textThe process of designing a complex system, formed by many elements and sub-elements interacting between each other, is usually completed at a system level and in the preliminary phases in two major steps: design-space exploration and optimization. In a classical approach, especially in a company environment, the two steps are usually performed together, by experts of the field inferring on major phenomena, making assumptions and doing some trial-and-error runs on the available mathematical models. To support designers and decision makers during the design phases of this kind of complex systems, and to enable early discovery of emergent behaviours arising from interactions between the various elements being designed, the authors implemented a parametric methodology for the design-space exploration and optimization. The parametric technique is based on the utilization of a particular type of matrix design of experiments, the orthogonal arrays. Through successive design iterations with orthogonal arrays, the optimal solution is reached with a reduced effort if compared to more computationally-intense techniques, providing sensitivity and robustness information. The paper describes the design methodology in detail providing an application example that is the design of a human mission to support a lunar base
Effect of environmental enrichment and group size on behaviour and live weight in growing rabbits
No full text[EN] The aim of this research was to study the effects of group size and environmental enrichment on behaviour and growth of 108 hybrid growing rabbits (Oryctolagus cuniculus). We compared the behaviour (time budget and reactions to specific behavioural tests: "tonic immobility" and "emergence test") and live weight of growing rabbits housed in cages with a different number of rabbits per cage (2, 3 and 4; same density:14 rabbits/m2). Half of the cages were enriched with a wooden stick (Robinia Pseudoacacia, length: 20 cm ¿ diameter: 6 cm, cylindrical) hanging from the ceiling of the cage. The stick and number of animals per cage had no effect on weight gain or on behavioural tests responses. Interaction with the stick was significantly higher at the beginning of the growing period. Principal component analysis performed on the data for the whole period showed significant differences according to the treatments: increasing the number of rabbits per cage and introducing a wooden stick seemed to affect locomotor activity frequency and social interactions. Rabbits housed 3 and 4 per cage showed less lying behaviour and higher locomotor activity and sitting. The larger functional space allowance enabled rabbits to perform more natural behaviours compared to smaller cages (2 rabbits/cage). Environmental enrichment seems to be related to higher allogrooming behaviour frequency, which could indicate a social behaviour related to pheromonal olfactory stimulation and mutual recognition.Zucca, D.; Marelli, S.; Redaelli, V.; Heinzl, E.; Cardile, H.; Ricci, C.; Verga, M.... (2012). Effect of environmental enrichment and group size on behaviour and live weight in growing rabbits. World Rabbit Science. 20(2):89-95. https://doi.org/10.4995/wrs.2012.1082SWORD899520
DEVELOPMENT OF MEASUREMENT METHODOLOGIES IN METROLOGY FOR CELL BIOLOGY AND REGENERATIVE MEDICINE
No full textAim of this thesis is the development of measurement methodologies in metrology for cell biology and regenerative medicine. Regenerative medicine is a novel branch of medicine based on the use of autologous stem cells and biocompatible medical devices to regenerate and repair damaged tissues of patients, i.e. by using three-dimensional scaffolds to implant stem cells into the tissue to be regenerated. Stakeholders of metrology for regenerative medicine are: health care providers who require safe, reliable and cost effective treatments, supported by evidence and approved by regulators; regulators who require standard materials and traceable data demonstrating the safety and efficacy of new products and treatments; medical products companies who require advanced and traceable techniques to develop new products and need methods to measure processes, such as cell growth on scaffolds, to ensure quality and efficiency of the medical products implanted into the patients. Consequently, regenerative medicine has the important requisite of a real time monitoring and not invasiveness neither destructiveness processes to measure the cell-scaffold interactions, in order to preserve the samples from any contamination or modification. Thus non-invasive measurement methodologies need to be developed for analysing the 3D cell culture on scaffolds and, in order to evaluate the uncertainty, highly reproducible measurement procedures are strongly required to minimize the type A uncertainties and to define the type B uncertainties. The non-invasive and non-destructive measurement of cell-scaffold interactions (i.e. stem cell proliferation and differentiation on scaffolds) is one of the most effective methodology to answer the need of testing the efficacy of the design, production/manufacturing, development and performances of stem cell-scaffold products. To satisfy the requirements and the needs for metrology in regenerative medicine, for this thesis it has been chosen to develop a measurement methodology for cellular activity (proliferation and differentiation) on 3D Biocoral® scaffolds and to conduce a metrological study to evaluate the uncertainty of the methodology. This thesis has been developed in the Bioscience group of the Italian National Metrological Institute (Istituto Nazionale di Ricerca Metrologica - INRIM). The main important contributes of this thesis to the metrology in biosciences have been: • to lay the foundations for a metrological approach to cell biology and particularly to regenerative medicine research and applications; • to address the filling of the lack of traceability in the metrology for cell biology metabolic methodologies used to evaluate cellular activities in living sample with non-invasive procedures. The main results and originalities achieved during this PhD work are: • a metabolic assay, the resazurin/resorufin assay, for the first time, has been metrologically characterized and the uncertainty of the measurement has been evaluated; • the resazurin/resorufin assay has been for the first time tailored for a 3D cell culture on Biocoral® scaffolds and the uncertainty of the measurement has been evaluated; • it was demonstrated that Biocoral® induces osteodifferentiation of stem cells and for the first time it was demonstrated on human mesenchymal stem cells; • it was demonstrated, for the first time, that the resazurin/resorufin metabolic assay can be a methodology to detect not only the proliferation but also the differentiation of stem cells on Biocoral® scaffolds; A description of the METREGEN regional project, which this thesis is part of, will follow in the introduction. The chapter 1 will give an overview on regenerative medicine field and its application with scaffolds, particularly referring to the Biocoral® scaffold. The resazurin/resorufin methodology will be deeply described in chapter 2 with a uncertainty budget evaluation and discussion. Chapter 3 will present in details a series of experiments made to establish and characterize a hMSCs in vitro 2D culture, establish a hMSCs in vitro 3D culture on Biocoral, tailor the resazurin/resorufin assay for 3D cell culture on Biocoral and evaluate the hMSCs osteodifferentiation induced by Biocoral scaffolds. All the results have been analysed with a metrological approach to evaluate the uncertainty. Finally, the conclusion will give a recapitulation and some interesting perspective of employment for the resazurin/resorufin methodology to final users, such as the cell factorie
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