Sílvia Santos Pedrosa

Biocompatibility of a self-assembled glycol chitosan nanogel

The research of chitosan-based nanogel for biomedical applications has grown exponentially in the last years; however, its biocompatibility is still insufficiently reported. Hence, the present work provides a thorough study of the biocompatibility of a glycol chitosan (GC) nanogel. The obtained results showed that GC nanogel induced slight decrease on metabolic activity of RAW, 3T3 and HMEC cell cultures, although no effect on cell membrane integrity was verified. The nanogel does not promote cell death by apoptosis and/or necrosis, exception made for the HMEC cell line challenged with the higher GC nanogel concentration. Cell cycle arrest on G1 phase was observed only in the case of RAW cells. Remarkably, the nanogel is poorly internalized by bone marrow derived macrophages and does not trigger the activation of the complement system. GC nanogel blood compatibility was confirmed through haemolysis and whole blood clotting time assays. Overall, the results demonstrated the safety of the use of the GC nanogel as drug delivery system.

A novel crosslinked hyaluronic acid nanogel for drug delivery.

An amphiphilic hyaluronic acid conjugate is successfully developed based on grafting a thiolated hydrophobic molecule to the polysaccharide backbone. The engineered conjugate is capable of assembling into nanostructures once dispersed in water, with average diameter of 80.2 ± 0.4 nm (n = 5), stable up to 6 months. The thiolated HyA conjugate is reticulated by dissulfide bond with a homofunctional crosslinker-1,4-Bis(3-[2-pyridyldithio]propionamido)butane (DPDPB). The drug loading efficiency of the reticulated and non-reticulated nanogel is accessed with two hydrophobic drugs, curcumin and simvastatin. Results suggest that crosslinked nanogel exhibit higher stability upon dilution and drug loading efficiency and proves to be a redox sensitive material. The nanogels hold great potential as stealth carriers of lipophilic drugs.

siRNA inhibition of endocytic pathways to characterize the cellular uptake mechanisms of folate functionalized glycol chitosan nanogels

Glycol chitosan nanogels have been widely used in gene, drug, and contrast agent delivery in an effort to improve disease diagnosis and treatment. Herein, we evaluate the internalization mechanisms and intracellular fate of previously described glycol chitosan nanogels decorated with folate to target the folate receptor. Uptake of the folate-decorated nanogel was impaired by free folate, suggesting competitive inhibition and shared internalization mechanisms via the folate receptor. Nanogel uptake was shown to occur mainly through flotillin-1 and Cdc42-dependent endocytosis. This was determined by inhibition of uptake reduction observed upon siRNA depletion of these two proteins and the pathways that they regulate. The data also suggest the involvement of the actin cytoskeleton in nanogel uptake via macropinocytosis. After 7 h of incubation with HeLa cells, approximately half of the nanogel population was localized in endolysosomal compartments, whereas the remaining 50% of the material was in undefined regions of the cytoplasm. Glycol chitosan nanogels may thus have potential as drug delivery vectors for targeting different intracellular compartments.

Antiproliferative Activity of Fucan Nanogel

Sulfated fucans comprise families of polydisperse natural polysaccharides based on sulfated L-fucose. Our aim was to investigate whether fucan nanogel induces cell-specific responses. To that end, a non toxic fucan extracted from Spatoglossum schröederi was chemically modified by grafting hexadecylamine to the polymer hydrophilic backbone. The resulting modified material (SNFuc) formed nanosized particles. The degree of substitution with hydrophobic chains was close to 100%, as estimated by elemental analysis. SNFfuc in aqueous media had a mean diameter of 123 nm and zeta potential of −38.3 ± 0.74 mV, as measured by dynamic light scattering. Nanoparticles conserved their size for up to 70 days. SNFuc cytotoxicity was determined using the MTT assay after culturing different cell lines for 24 h. Tumor-cell (HepG2, 786, H-S5) proliferation was inhibited by 2.0%–43.7% at nanogel concentrations of 0.05–0.5 mg/mL and rabbit aorta endothelial cells (RAEC) non-tumor cell line proliferation displayed inhibition of 8.0%–22.0%. On the other hand, nanogel improved Chinese hamster ovary (CHO) and monocyte macrophage cell (RAW) non-tumor cell line proliferation in the same concentration range. The antiproliferative effect against tumor cells was also confirmed using the BrdU test. Flow cytometric analysis revealed that the fucan nanogel inhibited 786 cell proliferation through caspase and caspase-independent mechanisms. In addition, SNFuc blocks 786 cell passages in the S and G2-M phases of the cell cycle.

The Nasal Cavity of the Rat and Mouse-Source of Mesenchymal Stem Cells for Treatment of Peripheral Nerve Injury: THE NASAL CAVITY OF THE RAT AND MOUSE

The nasal cavity performs several crucial functions in mammals, including rodents, being involved in respiration, behavior, reproduction, and olfaction. Its anatomical structure is complex and divided into several regions, including the olfactory recess where the olfactory mucosa (OM) is located and where the capture and interaction with the environmental odorants occurs. Among the cells of this region are the OM mesenchymal stem cells (MSCs), whose location raises the possibility that these cells could be involved in the peculiar ability of the olfactory nerve to regenerate continuously throughout life, although this relationship has not yet been confirmed. These cells, like all MSCs, present functional characteristics that make them candidates in new therapies associated with regenerative medicine, namely to promote the regeneration of the peripheral nerve after injury. The availability of stem cells to be therapeutically applied essentially depends on their collection in the tissue of origin. In the case of mice and rats OM‐MSCs, knowledge about the anatomy and histology of their nasal cavity is essential in establishing effective collection protocols. The present article describes the morphological characteristics of rodents OM and establishes an alternative protocol for access to the olfactory recess and collection of the OM. Anat Rec, 301:1678–1689, 2018.

Targetability of hyaluronic acid nanogel to cancer cells: In vitro and in vivo studies

Abstract We have, in previous work developed, characterized and evaluated the biocompatibility of an engineered hyaluronic acid nanogel. Here we assess the targetability of a hyaluronic acid nanogel towards CD44 overexpressing cells, in vitro and in vivo. Results obtained by flow cytometry and confocal fluorescence microscopy shows that nanogel is greatly internalized by non‐small cancer lung cells (A549 cells), that overexpress CD44 receptors. The biodistribution and tumor targetability of the nanogel labelled with a near‐infrared (NIR) probe were performed, in mice, through a non‐invasive imaging system. Results revealed nanogel high targetability towards an induced subcutaneous A549 tumor. Nanogels pharmacokinetics was evaluated also in healthy animals, and Alexa Fluor 680 labelled nanogel exhibited higher accumulation in liver, kidneys and skin. Also, a comparative biodistribution study was performed, using two NIR imaging probes, Cy5.5 and Alexa Fluor 680. Graphical abstract Figure. No Caption available.

Peripheral nerve injury and axonotmesis: State of the art and recent advances

Abstract Peripheral nerve lesions are frequent occurrences in both human and animal patients, leading to important physiological and labor complications that affect the quality of life of those who suffer the injury. More severe injuries are often associated with poor nerve regeneration and inadequate functional recovery, even with early medical and surgical interventions. Peripheral nerve crush lesions are frequent and, therefore, an experimental lesion paradigm widely used in researches involving nerve injury models and therapies for its resolution. In recent years, many studies have focused on innovative approaches to peripheral nerve treatment after crush injuries with more or less success. This review addresses the theme of peripheral nerve injury, with a special focus on the axonotmesis lesion, its etiology, pathophysiological mechanisms, methods of functional evaluation and regenerative processes, therapeutic options and corresponding recent advances.