In Vitro Characterization and Real-Time Label-Free Assessment of the Interaction of Chitosan-Coated Niosomes with Intestinal Cellular Monolayers

In vitro cell-based characterization methods of nanoparticles are generally static and require the use of secondary analysis techniques and labeling agents. In this study, bare niosomes and chitosan-coated niosomes (chitosomes) and their interactions with intestinal cells are studied under dynamic conditions and without fluorescent probes, using surface plasmon resonance (SPR)-based cell sensing. Niosomes and chitosomes were synthesized by using Tween 20 and cholesterol in a 15 mM:15 mM ratio and then characterized by dynamic light scattering (DLS). DLS analysis demonstrated that bare niosomes had average sizes of ∼125 nm, polydispersity index (PDI) below 0.2, and a negative zeta (ζ)-potential of −35.6 mV. In turn, chitosomes had increased sizes up to ∼180 nm, with a PDI of 0.2–0.3 and a highly positive ζ-potential of +57.9 mV. The viability of HT29-MTX, Caco-2, and Caco-2/HT29-MTX cocultured cells showed that both niosomes and chitosomes are cytocompatible up to concentrations of 31.6 μg/mL for at least 240 min. SPR analysis demonstrated that chitosomes interact more efficiently with HT29-MTX, Caco-2, and Caco-2/HT29-MTX cocultures compared to bare niosomes. The resulting SPR measurements were further supported by confocal microscopy and flow cytometry studies, which demonstrated that this method is a useful complementary or even alternative tool to directly characterize the interactions between niosomes and in vitro cell models in label-free and real-time conditions

The Involvement of PPARs in the Peculiar Energetic Metabolism of Tumor Cells

Energy homeostasis is crucial for cell fate, since all cellular activities are strongly dependent on the balance between catabolic and anabolic pathways. In particular, the modulation of metabolic and energetic pathways in cancer cells has been discussed in some reports, but subsequently has been neglected for a long time. Meanwhile, over the past 20 years, a recovery of the study regarding cancer metabolism has led to an increasing consideration of metabolic alterations in tumors. Cancer cells must adapt their metabolism to meet their energetic and biosynthetic demands, which are associated with the rapid growth of the primary tumor and colonization of distinct metastatic sites. Cancer cells are largely dependent on aerobic glycolysis for their energy production, but are also associated with increased fatty acid synthesis and increased rates of glutamine consumption. In fact, emerging evidence has shown that therapeutic resistance to cancer treatment may arise from the deregulation of glucose metabolism, fatty acid synthesis, and glutamine consumption. Cancer cells exhibit a series of metabolic alterations induced by mutations that lead to a gain-of-function of oncogenes, and a loss-of-function of tumor suppressor genes, including increased glucose consumption, reduced mitochondrial respiration, an increase of reactive oxygen species, and cell death resistance; all of these are responsible for cancer progression. Cholesterol metabolism is also altered in cancer cells and supports uncontrolled cell growth. In this context, we discuss the roles of peroxisome proliferator-activated receptors (PPARs), which are master regulators of cellular energetic metabolism in the deregulation of the energetic homeostasis, which is observed in cancer. We highlight the different roles of PPAR isotypes and the differential control of their transcription in various cancer cells

Minimal information for studies of extracellular vesicles (MISEV2023): From basic to advanced approaches

Extracellular vesicles (EVs), through their complex cargo, can reflect the state of their cell of origin and change the functions and phenotypes of other cells. These features indicate strong biomarker and therapeutic potential and have generated broad interest, as evidenced by the steady year-on-year increase in the numbers of scientific publications about EVs. Important advances have been made in EV metrology and in understanding and applying EV biology. However, hurdles remain to realising the potential of EVs in domains ranging from basic biology to clinical applications due to challenges in EV nomenclature, separation from non-vesicular extracellular particles, characterisation and functional studies. To address the challenges and opportunities in this rapidly evolving field, the International Society for Extracellular Vesicles (ISEV) updates its 'Minimal Information for Studies of Extracellular Vesicles', which was first published in 2014 and then in 2018 as MISEV2014 and MISEV2018, respectively. The goal of the current document, MISEV2023, is to provide researchers with an updated snapshot of available approaches and their advantages and limitations for production, separation and characterisation of EVs from multiple sources, including cell culture, body fluids and solid tissues. In addition to presenting the latest state of the art in basic principles of EV research, this document also covers advanced techniques and approaches that are currently expanding the boundaries of the field. MISEV2023 also includes new sections on EV release and uptake and a brief discussion of in vivo approaches to study EVs. Compiling feedback from ISEV expert task forces and more than 1000 researchers, this document conveys the current state of EV research to facilitate robust scientific discoveries and move the field forward even more rapidly

Development of hybrid nanocarrier library based biomaterials for cancer therapy

Introduction: The major challenges are requested to improve the properties of nanoparticles injected systemically: i) the increase of payloads inside nanoparticles, ii) the controlled release of payloads, iii) the decrease of interaction and uptake with mononuclear phagocyte system cells, iv) the selective targeting. Geometry and size of the therapeutic nanoparticles play a crucial role for their adhesion to tumor vasculature1. Discoidal non-spherical nanoparticles interact better with the fenestrated endothelium of tumor tissue with respect to spherical nanoparticles. Recently, discoidal nanoparticles have been developed to delivery payloads for anticancer therapy as well as cardiovascular diseases2. The aim of this project is of the synthesis of discoidal and hybrid nanocarrier (DHN) obtained by combining biomaterials, e.g. lipid/polymer, and extracellular vesicles for the treatment of Hepatoma and related-diseases. In particular, DHN shall allow to cure the liver diseases (i.e., cancer) and to stimulate the regeneration of tissue. Materials and Methods: DHN will be designed, synthesized and physicochemical characterize during different techniques such as DLS, NTA,TEM, FT-IR, LC ms/ms, and HPLC. In vitro and in vivo experiment will be carried out to test the toxicity and pre-clinic efficacy in suitable hepatic cellular lines and murine models. Conclusion: There resulting DHN may represent a new nanomedicine for the treatment of Hepatoma and related-diseases

Discoidal Nanoparticles: reaction environment-dependent Size Response

In recent years, the develop of new drug delivery system (DDS) was necessary to optimize the treatment efficiency, thus overcoming the limits of traditional teraphy such as targeting and drug half life. However, many treatmants cause immunogenicity. Several DDS was developed such as liposomes, niosomes and other tipe of nanoparticles, but they can have some limits like the recognition and elimination of the immune system or the tumour targeting. These drawbacks can be bypassed by modifing sizes and shapes of liposomes. Discoidal Nanopartcles (DNs) are obtained from liposomes by adding Styrene-Maleic Acid copolimer (SMA) (Fig. 1). The molecular ratio between styrene and maleic anhydride, pH and temperature of microenvironment reaction can affect the synthesis of DNs [1]. In this work we studied the use of SMA as copolymer, which is able to synthesis DNs by starting from spherical liposomes (DMPC) at different molar ratio of copolymers (2:1 and 4:1), pHs (range of pH from 3.5 to 11.5) and temperatures (4°C, 25°C, 37°C, 65°C). The SMA copolymer can form DNs, and their properties depend on the reaction enviroment. In fact, at different pHs, particle sizes are modified according to these physical parameters. The modifcation of temperature can influence the synthesis of DNs. Currently, the best condition is obtained at pH = 7.4 and 25°C by using 2:1 molar ratio of copolymer. The results are in agreement with previously reported data using SMA as a copolymer under different reaction conditions [1]. These properties could be affected the synthesis of DNs as well as drug delivery, but DNs could be a innovative DDS for anticancer therapy. Acknowledgment: This research is funded by the Italian Ministry of Instruction, University, and Research under the national project PON Ricerca e Innovazione 2014-2020

HYBRID NANOCARRIER LIBRARY BASED BIOMATERIALS FOR THERAPEUTIC APPLICATIONS.

Tumor targeting plays a pivot role in anticancer therapy. Tumor targeting can be carried out by conjugating specific targeting agents on the surface of nanoparticles or modifying their shape and size. To improve the selective targeting and accumulate therapeutic agents inside the tumor tissues, we designed new Discoidal Nanoparticles (DNs) having a non-spherical shape that better interact and cross through the fenestrated endothelium of tumor tissue compared to spherical nanoparticles. These nanoparticles are lipid based and are made up from the same lipids used to make liposomes and are obtained by changing the original spherical shape of liposomes in discoidal and decreasing liposomal native average sizes. Copolymers are basically used to modify spherical shape of liposomes and finally obtain discoidal form. SMAnh is a copolymer synthesized by reversible addition-fragmentation chain transfer polymerization (RAFT) between Styrene and Maleic Anhydride and can form nanodisks. Molecular ratio between styrene and maleic anhydride, pH and temperature of microenvironment reaction can however affect the synthesis of nanodiscks [1]. In this work we studied the use of SMAnh as copolymers capable to synthesis DNs starting from spherical liposomes at different molar ratio of copolymer, pH and temperature. MATERIALS AND METHODS Different liposomes were synthesized using thin layer evaporation method, freeze and thaw and extrusion. Liposomes were physicochemical characterized by using Dinamic Light Scattering (DLS) and UV-Vis spectrophotometer. The best selected formulations was DMPC liposomes that was used for further experiments. The DNs are synthesized by incubating the resulting liposomes with SMAnh (Styrene-Anhydride maleic molecular ratio 2:1) in Hepes 10 mM at pH = 7.4 and pH = 6.4. The resulting DNs are further physiochemically characterized by DLS and UV-Vis spectrophotometer. RESULT AND DISCUSSION SMAnh 2:1 molar ratio cannot make only DN at pH = 7.4. UV-Vis absorption of DN increased over time. This effect is directly related to the turbidity of nanoparticles. UV-Vis analysis agreed DLS data that show the presence of three different particle sizes at 5000 nm (80%), 200 nm (13%) and 13 nm (7%). Large particles depend on the presence of free SMAnh making aggregates in incubation medium. Narrow size distributed DN was obtained at pH = 6.4 using SMAnh 2:1 (molar ratio). DLS data shows two different peaks: 200 nm (10%) that represents bare liposomes and 13 nm (60%) that represents DN (Figure 1)