Design of Carboxymethylcellulose/Poloxamer-Based Bioformulation Embedding Trichoderma afroharzianum for Agricultural Applications.

Microbial biological control agents are believed to be a potential alternative to classical fertilizers to increase the sustainability of agriculture. In this work, the formulation of Trichoderma afroharzianum T22 (T22) spores with carboxymethyl cellulose (CMC) and Pluronic F-127 (PF-127) solutions was investigated. Rheological and microscopical analysis were performed on T22-based systems at three different CMC/PF-127 concentrations, showing that polymer aggregates tend to surround T22 spores, without viscosity, and the viscoelastic properties of the formulations were affected. Contact angle measurements showed the ability of PF-127 to increase the wettability of the systems, and the effect of the formulations on the viability of the spores was evaluated. The viability of the spores was higher over 21 days in all the formulations, compared to the control in water, at 4 and 25 °C. Finally, the effectiveness of the formulations on sweet basil was estimated by greenhouse tests. The results revealed a beneficial effect of the CMC/PF-127 mixture, but none on the formulation with T22. The data show the potential of CMC/PF-127 mixtures for the future design of microorganism-based formulations.

Effect of Hyaluronic Acid and Mesenchymal Stem Cells Secretome Combination in Promoting Alveolar Regeneration.

Pharmacological therapies in lung diseases are nowadays useful in reducing the symptomatology of lung injury. However, they have not yet been translated to effective treatment options able to restore the lung tissue damage. Cell-therapy based on Mesenchymal Stem Cells (MSCs) is an attractive, as well as new therapeutic approach, although some limitations can be ascribed for therapeutic use, such as tumorigenicity and immune rejection. However, MSCs have the capacity to secrete multiple paracrine factors, namely secretome, capable of regulating endothelial and epithelial permeability, decrease inflammation, enhancing tissue repair, and inhibiting bacterial growth. Furthermore, Hyaluronic acid (HA) has been demonstrated to have particularly efficacy in promoting the differentiation of MSCs in Alveolar type II (ATII) cells. In this frame, the combination of HA and secretome to achieve the lung tissue regeneration has been investigated for the first time in this work. Overall results showed how the combination of HA (low and medium molecular weight HA) plus secretome could enhance MSCs differentiation in ATII cells (SPC marker expression of about 5 ng/mL) compared to the only HA or secretome solutions alone (SPC about 3 ng/mL, respectively). Likewise, cell viability and cell rate of migration were reported to be improved for HA and secretome blends, indicating an interesting potentiality of such systems for lung tissue repair. Moreover, an anti-inflammatory profile has been revealed when dealing with HA and secretome mixtures. Therefore, these promising results can allow important advance in the accomplishment of the future therapeutic approach in respiratory diseases, up to date still missing.

How should incidental NEN of the pancreas and gastrointestinal tract be followed?

Neuroendocrine gastro-entero-pancreatic neoplasms (GEP-NENs) constitute a heterogeneous group of tumors, whose incidence has increased over the years. The most frequent site for primary disease is the stomach followed by small and large intestine, and pancreas. In the last decade, a dramatic growing in the incidence of small, incidental GEP-NENs has been recorded. In parallel, an increasing attitude toward more conservative approaches instead of surgical management has being widely spreading. This is particularly true for small, asymptomatic, pancreatic NEN as for these tumor forms an active surveillance has proven to be safe and feasible. Primary site and biological features of the neoplasms lead to different strategies and indications for surveillance and follow-up. This review focuses on the current evidence on modality and timing of surveillance and conservative treatment of incidentally discovered lesions.

Collagen-Mesenchymal Stem Cell Microspheres Embedded in Hyaluronic Acid Solutions as Biphasic Stem Niche Delivery Systems for Pulmonary Differentiation.

Cell therapy has the potential to become a feasible solution for several diseases, such as those related to the lungs and airways, considering the more beneficial intratracheal administration route. However, in lung diseases, an impaired pulmonary extracellular matrix (ECM) precludes injury resolution with a faulty engraftment of mesenchymal stem cells (MSCs) at the lung level. Furthermore, a shielding strategy to avoid cell damage as well as cell loss due to backflow through the injection path is required. Here, an approach to deliver cells encapsulated in a biomimetic stem niche is used, in which the interplay between cells and physiological lung ECM constituents, such as collagen and hyaluronic acid (HA), can occur. To this aim, a biphasic delivery system based on MSCs encapsulated in collagen microspheres (mCOLLs) without chemical modification and embedded in an injectable HA solution has been developed. Such biphasic delivery systems can both increase the mucoadhesive properties at the site of interest and improve cell viability and pulmonary differentiation. Rheological results showed a similar viscosity at high shear rates compared to the MSC suspension used in intratracheal administration. The size of the mCOLLs can be controlled, resulting in a lower value of 200 µm, suitable for delivery in alveolar sacs. Biological results showed that mCOLLs maintained good cell viability, and when they were suspended in lung medium implemented with low molecular weight HA, the differentiation ability of the MSCs was further enhanced compared to their differentiation ability in only lung medium. Overall, the results showed that this strategy has the potential to improve the delivery and viability of MSCs, along with their differentiation ability, in the pulmonary lineage.

A Covalently Cross-Linked Hyaluronic Acid/Carboxymethyl Cellulose Composite Hydrogel as a Potential Filler for Soft Tissue Augmentation.

The use of fillers for soft tissue augmentation is an approach to restore the structure in surgically or traumatically created tissue voids. Hyaluronic acid (HA), is one of the main components of the extracellular matrix, and it is widely employed in the design of materials with features similar to human tissues. HA-based fillers already find extensive use in soft tissue applications, but are burdened with inherent drawbacks, such as poor thermal stability. A well-known strategy to improve the HA properties is to reticulate it with 1,4-Butanediol diglycidyl ether (BDDE). The aim of this work was to improve the design of HA hydrogels as fillers, by developing a crosslinking HA method with carboxymethyl cellulose (CMC) by means of BDDE. CMC is a water soluble cellulose ether, whose insertion into the hydrogel can lead to increased thermal stability. HA/CMC hydrogels at different ratios were prepared, and their rheological properties and thermal stability were investigated. The hydrogel with an HA/CMC ratio of 1/1 resulted in the highest values of viscoelastic moduli before and after thermal treatment. The morphology of the hydrogel was examined via SEM. Biocompatibility response, performed with the Alamar blue assay on fibroblast cells, showed a safety percentage of around 90% until 72 h.

Correction: Pomegranate extract-loaded sphingosomes for the treatment of cancer: Phytochemical investigations, formulation, and antitumor activity evaluation.

[This corrects the article DOI: 10.1371/journal.pone.0293115.].

Fermentation-Derived Albumin-Based Hydrogels for Tissue Adhesion Applications.

Currently, most of the clinically available surgical glues and sealants lack elasticity, good adhesion and biocompatibility properties. Hydrogels as tissue adhesives have received extensive attention for their tissue-mimicking features. Here, a novel surgical glue hydrogel based on a fermentation-derived human albumin (rAlb) and biocompatible crosslinker for tissue-sealant applications has been developed. In order to reduce the risks of viral transmission diseases and an immune response, Animal-Free Recombinant Human Albumin from the saccharomyces yeast strain was used. A more biocompatible crosslinking agent, 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC), was used and compared with glutaraldehyde (GA). The design of crosslinked albumin-based adhesive gels was optimized by varying the albumin concentration, the mass ratio between albumin and the crosslinking agent as well as the crosslinker type. Tissue sealants were characterized in terms of mechanical (tensile and shear), adhesive and in vitro biocompatibility properties. The results indicated that the mechanical and adhesive properties improved as the albumin concentration increased and the mass ratio between albumin and crosslinker decreased. Moreover, the EDC-crosslinked albumin gels have better biocompatibility properties than GA-crosslinked glues.

Effect of Composition of Lung Biomimetic Niche on the Mesenchymal Stem Cell Differentiation toward Alveolar Type II Pneumocytes.

Pulmonary niche dynamically orchestrates the signals, such as proliferation or differentiation of mesenchymal stem cells (MSCs), which allows inducing tissue repair. Lung niche includes extracellular matrix (ECM), comprising hyaluronic acid (HA) and collagen (COLL), and several types of MSCs. Impaired ECM, in lung pathologies, makes the promising therapies based on MSCs ineffective, as it results in a reduced attachment and homing of MSCs, precluding their differentiation and viability. To overcome this problem, in this study a pulmonary biomimetic niche based on HA and COLL hydrogel is developed, with the specific aim to elucidate the role of COLL and HA/COLL semi-interpenetrating polymer networks (SIPNs) in directing the differentiation of MSCs into Alveolar Type II (ATII) cells. The effect of low (L), medium (M), and high (H) molecular weight (MW) HA is investigated, both like structural component of the SIPNs hydrogel and like trophic factor in cell culture media solution. HA in the culture media significantly improves surfactant protein (SP)-C expression (≈2 ng mL-1 ), without showing difference in the MW tested, compared to control only (≈1 ng mL-1 ). Furthermore, LMWHA/COLL hydrogel promotes the SPC expression (approximately two times) compared to COLL, MMWHA/COLL, and HMWHA/COLL hydrogels.

Design of functional nanoparticles by microfluidic platforms as advanced drug delivery systems for cancer therapy.

Nanoparticle systems are functional carriers that can be used in the cancer therapy field for the delivery of a variety of hydrophobic and/or hydrophilic drugs. Recently, the advent of microfluidic platforms represents an advanced approach to the development of new nanoparticle-based drug delivery systems. Particularly, microfluidics can simplify the design of new nanoparticle-based systems with tunable physicochemical properties such as size, size distribution and morphology, ensuring high batch-to-batch reproducibility and consequently, an enhanced therapeutic effect in vitro and in vivo. In this perspective, we present accurate state-of-the-art microfluidic platforms focusing on the fabrication of polymer-based, lipid-based, lipid/polymer-based, inorganic-based and metal-based nanoparticles for biomedical applications.

Ultrasmall Solid-Lipid Nanoparticles via the Polysorbate Sorbitan Phase-Inversion Temperature Technique: A Promising Vehicle for Antioxidant Delivery into the Skin.

Solid lipid nanoparticles promote skin hydration via stratum corneum occlusion, which prevents water loss by evaporation, and via the reinforcement of the skin's lipid-film barrier, which occurs through the adhesion of the nanoparticles to the stratum corneum. The efficacy of both phenomena correlates with lower nanoparticle size and the increased skin permeation of loaded compounds. The so-called Polysorbate Sorbitan Phase-Inversion Temperature method has, therefore, been optimized in this experimental work, in order to engineer ultrasmall solid-lipid nanoparticles that were then loaded with α-tocopherol, as the anti-age ingredient for cosmetic application. Ultrasmall solid-lipid nanoparticles have been proven to be able to favor the skin absorption of loaded compounds via the aforementioned mechanisms.

A Hyaluronic Acid-Based Formulation with Simultaneous Local Drug Delivery and Antioxidant Ability for Active Viscosupplementation.

Hyaluronic acid (HA) and its derivatives are widely used for intra-articular injection to augment compromised viscoelastic properties of damaged synovial fluid. Combining HA-based devices with anti-inflammatory drugs or bioactive principles in order to provide an additional benefit to the viscosupplementation is emerging as a new promising approach to improve the clinical outcome. Here, we aim to design a novel active viscosupplementation agent that can load and release hydrophobic drugs and at the same time possessing antioxidant properties. Optimized ternary systems named HCV based on HA, (2-hydroxypropyl)-ß-cyclodextrin (CD), and vitamin E (VE), without being engaged in formal chemical bonding with each other, showed the best viscoelastic and lubrication properties along with antioxidant capabilities, able to solubilize and release DF. The physical-chemical characterization suggested that the HCV system displayed rheological synergism and higher thermal stability because of the presence of VE and its antioxidant activity, and the loading of hydrophobic drugs was improved by the presence of CD and VE. Cell morphology and viability tests on L929 cells exhibited high biocompatibility of the HCV system with higher level expression of anti-inflammatory interleukin-10.

Hyaluronan-coated nanoparticles for active tumor targeting: Influence of polysaccharide molecular weight on cell uptake.

Here we aimed to correlate different molecular weights of hyaluronic acid (HA), 200, 800 and 1437 kDa, used to decorate poly(lactic-co-glycolic acid) (PLGA)-based nanoparticles (NPs), to their cell uptakes. NP internalization kinetics in CD44-overexpressing breast carcinoma cells were quantified, using healthy fibroblast cells as reference. Actually, NP uptake and selectivity by tumor cells were maximized for NPs HA 800 kDa, while being minimum for NPs HA1400 kDa. This unexpected result could be explained considering that the interaction between NPs and tumor cells is dictated by rearrangement and conformation of that segment of HA chain that actually protrudes from the NPs. Overall, results obtained in this work point at how HA molecular weight, is pivotal project parameter in NP formulation to promote active targeting in the CD44 overexpressing cancer cells.

Advances in Hyaluronic-Acid-Based (Nano)Devices for Cancer Therapy.

Cancer is the main cause of fatality all over the world with a considerable growth rate. Many biologically active nanoplatforms are exploited for tumor treatment. Of nanodevices, hyaluronic acid (HA)-based systems have shown to be promising candidates for cancer therapy due to their high biocompatibility and cell internalization. Herein, surface functionalization of different nanoparticles (NPs), e.g., organic- and inorganic-based NPs, is highlighted. Subsequently, HA-based nanostructures and their applications in cancer therapy are presented.

Shall We Tune? From Core-Shell to Cloud Type Nanostructures in Heparin/Silica Hybrids.

Heparin plays multiple biological roles depending on the availability of active sites strongly influenced by the conformation and the structure of polysaccharide chains. Combining different components at the molecular scale offers an extraordinary chance to easily tune the structural organization of heparin required for exploring new potential applications. In fact, the combination of different material types leads to challenges that cannot be achieved by each single component. In this study, hybrid heparin/silica nanoparticles were synthesized, and the role of silica as a templating agent for heparin supramolecular organization was investigated. The effect of synthesis parameters on particles compositions was deeply investigated by Fourier Transform Infrared Spectroscopy (FTIR) and Thermogravimetric Analysis (TGA). Transmission Electron Microscopy (TEM) reveals a different supramolecular organization of both components, leading to amazing organic-inorganic nanoparticles with different behavior in drug encapsulation and release. Furthermore, favorable biocompatibility for healthy human dermal fibroblasts (HDF) and tumor HS578T cells has been assessed, and a different biological behavior was observed, ascribed to different surface charge and morphology of synthesized nanoparticles.

Effect of Hyaluronic Acid on the Differentiation of Mesenchymal Stem Cells into Mature Type II Pneumocytes.

Hyaluronic acid (HA) is an essential component of the extracellular matrix (ECM) of the healthy lung, playing an important role in the structure of the alveolar surface stabilizing the surfactant proteins. Alveolar type II (ATII) cells are the fundamental element of the alveolus, specializing in surfactant production. ATII cells represent the main target of lung external lesion and a cornerstone in the repair process of pulmonary damage. In this context, knowledge of the factors influencing mesenchymal stem cell (MSC) differentiation in ATII cells is pivotal in fulfilling therapeutic strategies based on MSCs in lung regenerative medicine. To achieve this goal, the role of HA in promoting the differentiation of MSCs in mature Type II pneumocytes capable of secreting pulmonary surfactant was evaluated. Results demonstrated that HA, at a specific molecular weight can greatly increase the expression of lung surfactant protein, indicating the ability of HA to influence MSC differentiation in ATII cells.

Mechanical behavior of bioactive poly(ethylene glycol) diacrylate matrices for biomedical application.

The biomedical applications of physically entangled polymeric hydrogels are generally limited due to their weak mechanical properties, rapid swelling and dissolution in physiologically relevant environment. Chemical crosslinking helps stabilizing hydrogel structure and enhancing mechanical properties, thereby allowing a higher stability in phisiological environment. In this context, it is known that the mechanical properties of the hydrogel are affected by both the molecular weight (MW) of the starting polymer and the concentration of the crosslinker. Here, our aim was to assess the influence of polymer MW and concentration in the precursor solution on the mechanical features of the final hydrogel and their influence on cells-material interaction. In detail, 3D synthetic matrices based on poly(ethylene glycol) diacrylate (PEGDA) at two molecular weights (PEG 700 and PEG 3400) and at three different concentrations (10, 20, 40 w/v %), which were photopolymerized using darocour as an initiator, were studied. Then, infrared and swelling analyses, along with a comprehensive mechanical characterization of the obtained hydrogels (i.e. oscillatory shear and confined compression tests), were performed. Finally, to evaluate the influence of the mechanical features on the biological behaviour, the hydrogels were characterized in terms of cell adhesion percentage and cell viability after functionalizing the substrates with RGD peptide at three different concentrations. Results have demonstrated that both the Young's modulus (E) in compression and the elastic modulus (G') in shear of the hydrogels increase with increasing polymer precursor concentration. E decreased as MW increased, and the differences are more relevant for more concentrated hydrogels. On the contrary, G' appears to increase with increasing PEGDA MW and in particular for the lowest polymer precursor concentration. The biological results have demonstrated that cells cultured for longer times seem to prefer PEG 3400 hydrogels with a larger mesh size structure that posses higher viscoelastic properties in shear.

Natural Formulations Provide Antioxidant Complement to Hyaluronic Acid-Based Topical Applications Used in Wound Healing.

Hyaluronic acid (HA) promotes wound healing, and, accordingly, formulations based on HA have been widely used in regenerative medicine. In addition, naturally derived compounds, e.g., plant-based extracts and vitamin E, have exhibited antioxidant activity. In this study, a formulation containing hyaluronic acid, vitamin E, raspberry extract, and green tea was developed for potential topical applications, targeting wound healing. Rheological analysis was performed along with antioxidant and biological studies. The rheological characterization showed that the HA-based formulation is a thixotropic platform and possesses higher mechanical properties than the control formulation. To evaluate the wound healing potential of the formulation, an in vitro "wound healing" assay was carried out using human derived fibroblasts (HDF) with a cell-free gap on the tissue culture dish. The formulation showed better wound healing ability than the control formulation.

Hyaluronic acid/corn silk extract based injectable nanocomposite: A biomimetic antibacterial scaffold for bone tissue regeneration.

Injectable hydrogels have revealed the great potential for use as scaffolds in cartilage and bone tissue engineering. Here, thermosensitive and injectable hydrogels containing ß-tricalcium phosphate, hyaluronic acid, and corn silk extract-nanosilver (CSE-Ag NPs) were synthesized for their potential use in bone tissue regeneration applications. Spherical nanoparticles of silver were biosynthesized through microwave-assisted green approach using CSE in organic solvent-free medium. Rheological experiments demonstrated that the thermosensitive hydrogels have gelification temperature (Tgel) close to body temperature. The samples containing Ag NPs showed antibacterial activity toward gram-positive (Bacillus Subtilis, Staphylococcus Aureus) and gram-negative (Pseudomonas Aeruginosa, Escherichia Coli) bacteria along without cytotoxicity after 24 h. Mesenchymal stem cells seeded in the nanocomposite exhibited high bone differentiation which indicate that thay could be a good candidate as a potential scaffold for bone tissue regeneration.

Biosynthesis and characterization of antibacterial thermosensitive hydrogels based on corn silk extract, hyaluronic acid and nanosilver for potential wound healing.

The wounds closure after physical injury or surgery is of significant clinical and research importance. In this study, thermosensitive and injectable hydrogels based on hyaluronic acid (HA), corn silk extract (CSE) and nanosilver were prepared and their potential use as a wound care material was investigated. Silver nanoparticles (Ag NPs) were biosynthesized by a microwave-assisted green technique using corn silk extract in an organic solvent-free medium. Rheological analysis demonstrated that the nanocomposites have good mechanical properties with gelation temperature close to the body temperature; hence, they can be easily administrated locally on wounded skins. The samples exhibited antibacterial activity toward gram-positive and gram-negative bacteria. Cytotoxicity assay showed that the hydrogels have good biocompatibility. Interestingly, an in-vitro model of wound healing revealed that the nanocomposites allow faster wound closure and repair, compared to the control. The obtained results highlight the potential application of these novel injectable hydrogels as wound dressing.

Dynamics of nanoparticle diffusion and uptake in three-dimensional cell cultures.

This study aims at elucidating the effect of three-dimensional (3D) extracellular matrix on cell behaviour and nanoparticle (NP) diffusion and its consequences on NP cellular uptake mechansims. For this purpose, human dermal fibroblasts (HDF) and human fibrosarcoma (HT1080) cell lines were grown within a 3D collagen gel and exposed to model polystyrene (PS) NPs of controlled size (44 and 100nm). Results indicate that, in 3D, cell morphology dramatically changes compared to standard 2D cultures and NP diffusion within the matrix is hampered by the interaction with the collagen fibres. As a consequence, NP cellular uptake, modeled with equations describing the stoichiometric exchange between NPs and cell membrane, is significantly slowed down in 3D and in the case of 100 nm NPs, in part due to the hampered diffusion of NPs in collagen gel compared to their transport in standard cell culture medium. Furthermore, our outcomes point at a significant contribution of the cytoskeleton assembly, in particular actin microfilaments, in governing the uptake of PS NPs in a 3D environment, and also that the macropinocytosis process is preserved and is mainly involved in the internalization of PS NPs in a 3D environment. However, depending on cell type and nanoparticle size, other endocytic pathways are also implicated when moving from 2D to 3D culture systems. This work highlights the importance of studying the nano-bio interaction in experimental models that resembles in vivo conditions in order to better predict the therapeutic efficacy of drug delivery systems.