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Chitosan nanoparticles (CS NPs) showed promising results in drug, vaccine and gene delivery for the treatment of various diseases. The considerable attention towards CS was owning to its outstanding biological properties, however, the main challenge in the application of CS NPs was faced during their size-controlled synthesis. Herein, ionic gelation reaction between CS and sodium tripolyphosphate (TPP), a widely used and safe CS cross-linker for biomedical application, was exploited by a microfluidic approach based on a staggered herringbone micromixer (SHM) for the synthesis of TPP cross-linked CS NPs (CS/TPP NPs). Screening design of experiments was applied to systematically evaluate the main process and formulative factors affecting CS/TPP NPs physical properties (mean size and size distribution). Effectiveness of the SHM-assisted manufacturing process was confirmed by the preliminary evaluation of the biological performance of the optimized CS/TPP NPs that were internalized in the cytosol of human mesenchymal stem cells through clathrin-mediated mechanism. Curcumin, selected as a challenging model drug, was successfully loaded into CS/TPP NPs (EE% > 70%) and slowly released up to 48 h via the diffusion mechanism. Finally, the comparison with the conventional bulk mixing method corroborated the efficacy of the microfluidics-assisted method due to the precise control of mixing at microscales., Competing Interests: The authors declare no conflict of interest.

Adipose tissue comprises both adipose and non-adipose cells such as mesenchymal stem cells. These cells show a surface antigenic profile similar to that of bone-marrow-derived MSC. The cells derived from the dedifferentiation of mature adipocytes (DFAT) are another cell population with characteristics of stemness. The aim of this study is to provide evidence of the stemness, proliferation, and differentiation of human adipose stem cells (hASC) and DFAT obtained from human subcutaneous AT and evaluate their potential use in regenerative medicine. Cell populations were studied by histochemical and molecular biology techniques. Both hASC and DFAT were positive for MSC markers. Their proliferative capacity was similar and both populations were able to differentiate into osteogenic, chondrogenic, and adipogenic lineages. DFAT were able to accumulate lipids and their lipoprotein lipase and adiponectin gene expression were high. Alkaline phosphatase and RUNX2 gene expression were greater in hASC than in DFAT at 14 days but became similar after three weeks. Both cell populations were able to differentiate into chondrocytes, showing positive staining with Alcian Blue and gene expression of SOX9 and ACAN . In conclusion, both hASC and DFAT populations derived from AT have a high differentiation capacity and thus may have applications in regenerative medicine., Competing Interests: The authors declare no conflict of interest.

To characterize different cell populations in the human ovary, morphological and functional characteristics of cell populations collected during routine IVF procedures were studied. Cells obtained from follicular fluid grew in vitro under minimal medium conditions, without growth factor, including leukaemia-inhibiting factor. Morphological analysis revealed a heterogeneous cell population, with cells displaying a fibroblast-like, epithelial-like and also neuron-like features. Morpho-functional characteristics of fibroblast-like cells were similar to mesenchymal stem cells, and, in particular, were positive for mesenchymal stemness markers, including CD90, CD44, CD105, CD73, but negative for epithelial proteins, such as cytokeratins, CD34 and CD45 antigens. Cell proliferation activity at different times and colony-forming unit capability were evaluated, and multipotency of a subset of granulosa cells was established by in-vitro differentiation studies (e.g. osteogenic, chondrogenic and adipogenic differentiation). This study suggests that cells provided by mesenchymal plasticity can be easily isolated by waste follicular fluid, avoiding scraping of human ovaries, and cultivated in minimal conditions. Successful growth of such progenitor cells on three-dimensional cryogel scaffold provides the basis for future developments in tissue engineering. This culture system may be regarded as an experimental model in which biological behaviour is not influenced by specific growth factors., (Copyright © 2014 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.)

Recent studies have reported that stem cells (human follicular fluid mesenchymal stem cells or hFF-MSCs) are present in ovarian follicular fluid (hFF) and that they have a proliferative and differentiative potential which is similar to that of MSCs derived from other adult tissue. These mesenchymal stem cells, isolated from human follicular fluid waste matter discarded after retrieval of oocytes during the IVF process, constitute another, as yet unutilized, source of stem cell materials. There has been little work on the compatibility of these hFF-MSCs with scaffolds useful for bone tissue engineering applications and the aim of this study was to evaluate the osteogenic capacity of hFF-MSCs seeded on bioglass 58S-coated titanium and to provide an assessment of their suitability for bone tissue engineering purposes. Following a chemical and morphological characterization with scanning electron microscopy (SEM) and energy dispersive spectroscopy (EDS), cell viability, morphology and expression of specific osteogenic markers were examined after 7 and 21 days of culture. The hFF-MSCs seeded on bioglass and cultured with osteogenic factors, when compared with those seeded on tissue culture plate or on uncoated titanium, exhibited enhanced cell viability and osteogenic differentiation, as reflected by increased calcium deposition and increased ALP activity with expression and production of bone-related proteins. Taken together, these results demonstrate that MSCs from human follicular fluid waste materials can be easily cultured in titanium scaffolds coated with bioglass, having osteoinductive properties. This process has significant potential for regenerative medicine applications and indicates that hFF-MSCs may be a valid alternative to hBM-MSC cells in experimental models in bone tissue engineering. [ABSTRACT FROM AUTHOR]

The article discusses a study conducted to investigate the biological effects of three distinct nanosized formulations with similar physical features, each composed of a precise Hyaluronic Acid molecular weight, in human bone marrow mesenchymal stem cells CD44+. Cytocompatibility and cell proliferation were assessed by MTT assay and Bromodeoxyuridine assay.

Nanotechnology offers advanced biomedical tools for diagnosis and drug delivery, stressing the value of investigating the mechanisms by which nanocarriers interact with the biological environment. Herein, the cellular response to CD44-targeted nanoparticles (NPs) was investigated. CD44, the main hyaluronic acid (HA) receptor, is widely exploited as a target for therapeutic purposes. HA NPs were produced by microfluidic platform starting from HA with different molecular weights (Mw, 280, 540, 820 kDa) by polyelectrolyte complexation with chitosan (CS). Thanks to microfluidic technology, HA/CS NPs with the same physical features were produced, and only the effects of HA Mw on CD44-overexpressing cells (human mesenchymal stem cells, hMSCs) were studied. This work provides evidence of the HA/CS NPs biocompatibility regardless the HA Mw and reveals the effect of low Mw HA in improving the cell proliferation. Special attention was paid to the endocytic mechanisms used by HA/CS NPs to enter hMSCs. The results show the notable role of CD44 and the pronounced effect of HA Mw in the NPs' internalization. HA/CS NPs uptake occurs via different endocytic pathways simultaneously, and most notably, NPs with 280 kDa HA were internalized by clathrin-mediated endocytosis. Instead, NPs with 820 kDa HA revealed a greater contribution of caveolae and cytoskeleton components. [ABSTRACT FROM AUTHOR]

In this study, an innovative microfluidics-based method was developed for one-step synthesis of hyaluronic acid (HA)-based nanoparticles (NPs), by exploiting polyelectrolytic interactions between HA and chitosan (CS), in order to improve reliability, reproducibility and possible scale-up of the NPs preparation. The on-chip synthesis, using a staggered herringbone micromixer, allowed to produce HA/CS NPs with tailored-made size and suitable for both parenteral (117.50 ± 4.51 nm) and loco-regional (349.15 ± 38.09 nm) administration, mainly composed by HA (more than 85% wt) with high negative surface charge (< −20 mV). HA/CS NPs were successfully loaded with a challenging water-insoluble molecule, Everolimus (EVE), an FDA- and EMA-approved anticancer drug able to lead to cell cycle arrest, reduced angiogenesis and promotion of apoptosis. HA/CS NPs resulted to be massively internalized in CD44+ human mesenchymal stem cells via CD44 receptor-mediated endocytosis. HA/CS NPs selectiveness towards CD44 was highlighted by blocking CD44 receptor by anti-CD44 primary antibody and by comparison to CS-based NPs cellular uptake. Eventually, high effectiveness in inhibiting cell proliferation was demonstrated on-chip synthetized EVE loaded HA/CS NPs by tracking in vitro DNA synthesis. [ABSTRACT FROM AUTHOR]