Sustainable Exploitation of Posidonia oceanica Sea Balls (Egagropili): A Review.

Posidonia oceanica (L.) Delile is the main seagrass plant in the Mediterranean basin that forms huge underwater meadows. Its leaves, when decomposed, are transported to the coasts, where they create huge banquettes that protect the beaches from sea erosion. Its roots and rhizome fragments, instead, aggregate into fibrous sea balls, called egagropili, that are shaped and accumulated by the waves along the shoreline. Their presence on the beach is generally disliked by tourists, and, thus, local communities commonly treat them as waste to remove and discard. Posidonia oceanica egagropili might represent a vegetable lignocellulose biomass to be valorized as a renewable substrate to produce added value molecules in biotechnological processes, as bio-absorbents in environmental decontamination, to prepare new bioplastics and biocomposites, or as insulating and reinforcement materials for construction and building. In this review, the structural characteristics, and the biological role of Posidonia oceanica egagropili are described, as well as their applications in different fields as reported in scientific papers published in recent years.

Self-esterified hyaluronan hydrogels: Advancements in the production with positive implications in tissue healing.

Hyaluronan-(HA) short half-life in vivo limits its benefits in tissue repair. Self-esterified-HA is of great interest because it progressively releases HA, promoting tissue-regeneration longer than the unmodified-polymer. Here, the 1-ethyl-3-(3-diethylaminopropyl)carbodiimide(EDC)-hydroxybenzotriazole(HOBt) carboxyl-activating-system was evaluated for self-esterifying HA in the solid state. The aim was to propose an alternative to the time-consuming, conventional reaction of quaternary-ammonium-salts of HA with hydrophobic activating-systems in organic media, and to the EDC-mediated reaction, limited by by-product formation. Additionally, we aimed to obtain derivatives releasing defined molecular-weight(MW)-HA that would be valuable for tissue renewal. A 250 kDa-HA(powder/sponge) was reacted with increasing EDC/HOBt amounts. HA-modification was investigated through Size-Exclusion-Chromatography-Triple-Detector-Array-analyses, FT-IR/1H NMR and the products(XHAs) extensively characterized. Compared to conventional protocols, the set procedure is more efficient, avoids side-reactions, allows for an easier processing to diverse clinically-usable 3D-forms, leads to products gradually releasing HA under physiological conditions with the possibility to tune the MW of the biopolymer-released. Finally, the XHAs exhibit sound stability to Bovine-Testicular-Hyaluronidase, hydration/mechanical properties suitable for wound-dressings, with improvements over available matrices, and prompt in vitro wound-regeneration, comparably to linear-HA. To the best of our knowledge, the procedure is the first valid alternative to conventional protocols for HA self-esterification with advances in the process itself and in product performance.

Enhanced Streptomyces roseochromogenes melanin production by using the marine renewable source Posidonia oceanica egagropili.

Since the possibility to biotechnologically produce melanin by Streptomycetes using plant biomass has been so far poorly investigated, Posidonia oceanica egagropili, a marine waste accumulating along the Mediterranean Sea coasts, was explored as a renewable source to enhance extracellular melanin production by Streptomyces roseochromogenes ATCC 13400. Therefore, different amounts of egagropili powder were added to a culture medium containing glucose, malt extract, and yeast extract, and their effect on the melanin biosynthesis was evaluated. A 2.5 g·L-1 supplementation in 120-h shake flask growths at 26 °C, at pH 6.0 and 250 rpm, was found to enhance the melanin production up to 3.94 ± 0.12 g·L-1, a value 7.4-fold higher than the control. Moreover, 2-L batches allowed to reach a concentration of 9.20 ± 0.12 g·L-1 in 96 h with a productivity of 0.098 g·L-1·h-1. Further studies also demonstrated that the melanin production enhancement was due to the synergistic effect of both the lignin carbohydrate complex and the holocellulose components of the egagropili. Finally, the pigment was purified from the broth supernatant by acidic precipitation and reversed-phase chromatography, characterized by UV absorbance and one- and two-dimensional NMR, and also tested for its chemical, antioxidant, and photo-protective properties. KEY POINTS: • S. roseochromogenes ATCC 13400 produces extracellular soluble melanin. • Egagropili added to the growth medium enhances melanin production and productivity. • Both the lignin carbohydrate complex and the holocellulose egagropili components influence the melanin biosynthesis.

Argan (Argania spinosa L.) Seed Oil Cake as a Potential Source of Protein-Based Film Matrix for Pharmaco-Cosmetic Applications.

Various different agri-food biomasses might be turned into renewable sources for producing biodegradable and edible plastics, potentially attractive for food, agricultural and cosmeceutical sectors. In this regard, different seeds utilized for edible and non-edible oil extraction give rise to high amounts of organic by-products, known as seed oil cakes (SOCs), potentially able to become protein-rich resources useful for the manufacturing of biodegradable films. This study reports the potential of SOC derived from Argania spinosa (argan), a well-known plant containing valuable non-refined oil suitable for food or cosmetic use, to be a promising valuable source for production of a protein-based matrix of biomaterials to be used in the pharmaco-cosmetic sector. Thus, glycerol-plasticized films were prepared by casting and drying using different amounts of argan seed protein concentrate, in the presence of increasing glycerol concentrations, and characterized for their morphological, mechanical, barrier, and hydrophilicity properties. In addition, their antioxidant activity and effects on cell viability and wound healing were investigated. The hydrophobic nature of the argan protein-based films, and their satisfying physicochemical and biological properties, suggest a biorefinery approach for the recycling of argan SOC as valuable raw material for manufacturing new products to be used in the cosmeceutical and food industries.

Polydatin Incorporated in Polycaprolactone Nanofibers Improves Osteogenic Differentiation.

Polycaprolactone nanofibers are used as scaffolds in the field of tissue engineering for tissue regeneration or drug delivery. Polycaprolactone (PCL) is a biodegradable hydrophobic polyester used to obtain implantable nanostructures, which are clinically applicable due to their biological safety. Polydatin (PD), a glycosidic precursor of resveratrol, is known for its antioxidant, antitumor, antiosteoporotic, and bone regeneration activities. We aimed to use the osteogenic capacity of polydatin to create a biomimetic innovative and patented scaffold consisting of PCL-PD for bone tissue engineering. Both osteosarcoma cells (Saos-2) and mesenchymal stem cells (MSCs) were used to test the in vitro cytocompatibility of the PD-PCL scaffold. Reverse-phase (RP) HPLC was used to evaluate the timing release of PD from the PCL-PD nanofibers and the MTT assay, scanning electron microscopy, and alkaline phosphatase (ALP) activity were used to evaluate the proliferation, adhesion, and cellular differentiation in both osteosarcoma and human mesenchymal stem cells (MSCs) seeded on PD-PCL nanofibers. The proliferation of osteosarcoma cells (Saos-2) on the PD-PCL scaffold decreased when compared to cells grown on PLC nanofibers, whereas the proliferation of MSCs was comparable in both PCL and PD-PCL nanofibers. Noteworthy, after 14 days, the ALP activity was higher in both Saos-2 cells and MSCs cultivated on PD-PCL than on empty scaffolds. Moreover, the same cells showed a spindle-shaped morphology after 14 days when grown on PD-PCL as shown by SEM. In conclusion, we provide evidence that nanofibers appropriately coated with PD support the adhesion and promote the osteogenic differentiation of both human osteosarcoma cells and MSCs.

Limosilactobacillus fermentum from buffalo milk is suitable for potential biotechnological process development and inhibits Helicobacter pylori in a gastric epithelial cell model.

Probiotics are living microorganisms that give beneficial health effects while consumed, and each strain possesses diverse and unique properties and also different technological characteristics that affect its ability to be produced at large scale. Limosilactobacillus fermentum is a widely studied member of probiotics, however, few data are available on the development of fermentation and downstream processes for the production of viable biomasses for potential industrial applications. In the present study a novel L. fermentum strain was isolated from buffalo milk and used as test example for biotechnological process development. The strain was able to produce up to 109 CFU/mL on a (glucose based) semi-defined medium deprived of animal-derived raw materials up to the pilot scale (150 L), demonstrating improved results compared to commonly used, although industrially not suitable, media rich of casein and beef extract. The study of strain behavior in batch experiments indicated that the highest concentration of viable cells was reached after only 8 h of growth, greatly shortening the process. Moreover, initial concentrations of glucose in the medium above 30 g/L, if not supported by higher nitrogen concentrations, reduced the yield of biomass and increased production of heterolactic fermentation by-products. Biomass concentration via microfiltration on hollow fibers, and subsequent spray-drying allowed to recover about 5.7 × 1010CFU/gpowder of viable cells, indicating strain resistance to harsh processing conditions. Overall, these data demonstrate the possibility to obtain and maintain adequate levels of viable L. fermentum cells by using a simple approach that is potentially suitable for industrial development. Moreover, since often exopolysaccharides produced by lactobacilli contribute to the strain's functionality, a partial characterization of the EPS produced by the newly identified L. fermentum strain was carried out. Finally, the effect of L. fermentum versus H. pylori in a gastric epithelial cell model was evaluated demonstrating its ability to stimulate the response of the immune system and displace the infective agent.

Niclosamide as a Repurposing Drug against Corynebacterium striatum Multidrug-Resistant Infections.

Corynebacterium striatum (C. striatum) is an emerging multidrug-resistant (MDR) pathogen associated with nosocomial infections. In this scenario, we screened the antimicrobial activity of the anthelmintic drugs doramectin, moxidectin, selamectin and niclosamide against 20 C. striatum MDR clinical isolates. Among these, niclosamide was the best performing drug against C. striatum. Niclosamide cytotoxicity was evaluated by a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay on immortalized human keratinocyte cells (HaCaT). After 20 h of treatment, the recorded 50% cytotoxic concentration (CC50) was 2.56 µg/mL. The antibacterial efficacy was determined via disc diffusion, broth microdilution method and time-killing. Against C. striatum, niclosamide induced a growth inhibitory area of 22 mm and the minimum inhibitory concentration that inhibits 90% of bacteria (MIC90) was 0.39 µg/mL, exhibiting bactericidal action. The biofilm biomass eradicating action was investigated through crystal violet (CV), MTT and confocal laser scanning microscopy (CLSM). Niclosamide affected the biofilm viability in a dose-dependent manner and degraded biomass by 55 and 49% at 0.39 µg/mL and 0.19 µg/mL. CLSM images confirmed the biofilm biomass degradation, showing a drastic reduction in cell viability. This study could promote the drug-repurposing of the anthelmintic FDA-approved niclosamide as a therapeutic agent to counteract the C. striatum MDR infections.

Potential of Biofermentative Unsulfated Chondroitin and Hyaluronic Acid in Dermal Repair.

Chondroitin obtained through biotechnological processes (BC) shares similarities with both chondroitin sulfate (CS), due to the dimeric repetitive unit, and hyaluronic acid (HA), as it is unsulfated. In the framework of this experimental research, formulations containing BC with an average molecular size of about 35 KDa and high molecular weight HA (HHA) were characterized with respect to their rheological behavior, stability to enzymatic hydrolysis and they were evaluated in different skin damage models. The rheological characterization of the HHA/BC formulation revealed a G' of 92 ± 3 Pa and a G″ of 116 ± 5 Pa and supported an easy injectability even at a concentration of 40 mg/mL. HA/BC preserved the HHA fraction better than HHA alone. BTH was active on BC alone only at high concentration. Assays on scratched keratinocytes (HaCaT) monolayers showed that all the glycosaminoglycan formulations accelerated cell migration, with HA/BC fastening healing 2-fold compared to the control. In addition, in 2D HaCaT cultures, as well as in a 3D skin tissue model HHA/BC efficiently modulated mRNA and protein levels of different types of collagens and elastin remarking a functional tissue physiology. Finally, immortalized human fibroblasts were challenged with TNF-α to obtain an in vitro model of inflammation. Upon HHA/BC addition, secreted IL-6 level was lower and efficient ECM biosynthesis was re-established. Finally, co-cultures of HaCaT and melanocytes were established, showing the ability of HHA/BC to modulate melanin release, suggesting a possible effect of this specific formulation on the reduction of stretch marks. Overall, besides demonstrating the safety of BC, the present study highlights the potential beneficial effect of HHA/BC formulation in different damage dermal models.

Evaluation of novel biomaterials for cartilage regeneration based on gelatin methacryloyl interpenetrated with extractive chondroitin sulfate or unsulfated biotechnological chondroitin.

Gelatin is widely proposed as scaffold for cartilage tissue regeneration due to its high similarities to the extracellular matrix. However, poor mechanical properties and high sensitivity to enzymatic degradation encouraged the scientific community to develop strategies to obtain better performing hydrogels. Gelatin networks, specifically gelatin-methacryloyl (GM), have been coupled to hyaluronan or chondroitin sulfate (CS). In this study, we evaluated the biophysical properties of an innovative photocross-linked hydrogel based on GM with the addition of CS or a new unsulfated biotechnological chondroitin (BC). Biophysical, mechanical, and biochemical characterization have been assessed to compare GM hydrogels to the chondroitin containing networks. Moreover, mesenchymal stem cells (MSCs) were seeded on these biomaterials in order to evaluate the differentiation toward the chondrocyte phenotype in 21 days. Rheological characterization showed that both CS and BC increased the stiffness (G' was about 2-fold), providing a stronger rigid matrix, with respect to GM alone. The biological tests confirmed the onset of MSCs differentiation process starting from 14 days of in vitro culture. In particular, the combination GM + BC resulted to be more effective than GM + CS in the up-regulation of key genes such as collagen type 2A1 (COLII), SOX-9, and aggrecan). In addition, the scanning microscope analyses revealed the cellular adhesion on materials and production of extracellular vesicles. Immunofluorescence staining confirmed an increase of COLII in presence of both chondroitins. Finally, the outcomes suggest that BC entangled within cross-linked GM matrix may represent a promising new biomaterial with potential applications in cartilage regeneration.

Hyaluronan-Based Gel Promotes Human Dental Pulp Stem Cells Bone Differentiation by Activating YAP/TAZ Pathway.

BACKGROUND: Hyaluronans exist in different forms, accordingly with molecular weight and degree of crosslinking. Here, we tested the capability to induce osteogenic differentiation in hDPSCs (human dental pulp stem cells) of three hyaluronans forms: linear pharmaceutical-grade hyaluronans at high and (HHA) low molecular weight (LHA) and hybrid cooperative complexes (HCC), containing both sizes. METHODS: hDPSCs were treated with HHA, LHA, HCC for 7, 14 and 21 days. The effects of hyaluronans on osteogenic differentiation were evaluated by qRT-PCR and WB of osteogenic markers and by Alizarin Red S staining. To identify the involved pathway, CD44 was analyzed by immunofluorescence, and YAP/TAZ expression was measured by qRT-PCR. Moreover, YAP/TAZ inhibitor-1 was used, and the loss of function of YAP/TAZ was evaluated by qRT-PCR, WB and immunofluorescence. RESULTS: We showed that all hyaluronans improves osteogenesis. Among these, HCC is the main inducer of osteogenesis, along with overexpression of bone related markers and upregulating CD44. We also found that this biological process is subordinate to the activation of YAP/TAZ pathway. CONCLUSIONS: We found that HA's molecular weight can have a relevant impact on HA performance for bone regeneration, and we unveil a new molecular mechanism by which HA acts on stem cells.

Hard-to-heal wound treated with Integra Flowable Wound Matrix: analysis and clinical observations.

Skin healing defects severely impair the quality of life of millions of people and burden healthcare systems globally. The therapeutic approach to these pathologies still represents a challenge. Novel scaffolds, used as dermal substitutes, possibly represent a promising strategy in complex wound management. Integra Flowable Wound Matrix (IFWM) is composed of a lyophilised, micronised form of collagen/chondroitin sulphate matrix, already used in regenerative medicine and endorsed in the therapy of diabetic foot lesions. In this paper, IFWM was applied to a tunnelling hard-to-heal skin lesion in order to restore tissue integrity. Although the different phases of skin wound healing are well established, the molecular mechanism underpinning IFWM-induced tissue repair are almost unknown. Here, we report, for the first time, the comparative analysis of molecular, histological and clinical observations of the healing process of a hard-to-heal tunnelling skin wound. The therapeutic success of this clinical case allowed us to recommend the use of IFWM as a tissue substitute in this rare type of hard-to-heal wound in which the high inflammatory status hampered the natural healing process.

Decreased expression of Malassezia furfur virulence factors after Q-switched Nd:YAG laser irradiation.

Malassezia spp. are lipophilic yeasts implicated in the pathogenesis of chronic skin diseases. Repeated therapies are often necessary due to the recurrence of this type of disease. Recently, laser and light-based devices used for the treatment of some skin diseases have shown good efficacy, few contraindications, and minimal side effects. The neodymium-doped yttrium aluminium garnet (Q-switched Nd:YAG) laser is one of the most commonly used lasers in dermatology. The aim of this study was to evaluate the effect of the Q-switched Nd:YAG laser (Medlite C6 laser, Conbio, USA) on the pathogenic mechanisms of M. furfur during skin infections. Following laser exposure, the ability of M. furfur to retain phospholipase activity, upregulate the aryl receptor and its associated pathway, and stimulate the immune response were tested. The Q-switched Nd:YAG laser was shown to attenuate the virulence of M. furfur. The Q-switched Nd:YAG laser should be considered as a valid therapeutic alternative for the treatment of Malassezia-associated infections.

Gelatin-biofermentative unsulfated glycosaminoglycans semi-interpenetrating hydrogels via microbial-transglutaminase crosslinking enhance osteogenic potential of dental pulp stem cells.

Gelatin hydrogels by microbial-transglutaminase crosslinking are being increasingly exploited for tissue engineering, and proved high potential in bone regeneration. This study aimed to evaluate, for the first time, the combination of enzymatically crosslinked gelatin with hyaluronan and the newly developed biotechnological chondroitin in enhancing osteogenic potential. Gelatin enzymatic crosslinking was carried out in the presence of hyaluronan or of a hyaluronan-chondroitin mixture, obtaining semi-interpenetrating gels. The latter proved lower swelling extent and improved stiffness compared to the gelatin matrix alone, whilst maintaining high stability. The heteropolysaccharides were retained for 30 days in the hydrogels, thus influencing cell response over this period. To evaluate the effect of hydrogel composition on bone regeneration, materials were seeded with human dental pulp stem cells and osteogenic differentiation was assessed. The expression of osteocalcin (OC) and osteopontin (OPN), both at gene and protein level, was evaluated at 7, 15 and 30 days of culture. Scanning electron microscopy (SEM) and two-photon microscope observations were performed to assess bone-like extracellular matrix (ECM) deposition and to observe the cell penetration depth. In the presence of the heteropolysaccharides, OC and OPN expression was upregulated and a higher degree of calcified matrix formation was observed. Combination with hyaluronan and chondroitin improved both the biophysical properties and the biological response of enzymatically crosslinked gelatin, fastening bone deposition.

Acellular Dermal Matrix Used in Diabetic Foot Ulcers: Clinical Outcomes Supported by Biochemical and Histological Analyses.

Diabetic foot ulcer (DFU) is a diabetes complication which greatly impacts the patient's quality of life, often leading to amputation of the affected limb unless there is a timely and adequate management of the patient. DFUs have a high economic impact for the national health system. Data have indeed shown that DFUs are a major cause of hospitalization for patients with diabetes. Based on that, DFUs represent a very important challenge for the national health system. Especially in developed countries diabetic patients are increasing at a very high rate and as expected, also the incidence of DFUs is increasing due to longevity of diabetic patients in the western population. Herein, the surgical approach focused on the targeted use of the acellular dermal matrix has been integrated with biochemical and morphological/histological analyses to obtain evidence-based information on the mechanisms underlying tissue regeneration. In this research report, the clinical results indicated decreased postoperative wound infection levels and a short healing time, with a sound regeneration of tissues. Here we demonstrate that the key biomarkers of wound healing process are activated at gene expression level and also synthesis of collagen I, collagen III and elastin is prompted and modulated within the 28-day period of observation. These analyses were run on five patients treated with Integra® sheet and five treated with the injectable matrix Integra® Flowable, for cavitary lesions. In fact, clinical evaluation of improved healing was, for the first time, supported by biochemical and histological analyses. For these reasons, the present work opens a new scenario in DFUs treatment and follow-up, laying the foundation for a tailored protocol towards complete healing in severe pathological conditions.

Unsulfated biotechnological chondroitin by itself as well as in combination with high molecular weight hyaluronan improves the inflammation profile in osteoarthritis in vitro model.

Several studies suggest that inflammation has a pivotal role during the progression of osteoarthritis (OA) and cytokines have been identified as the main process mediators. This study aimed to explore the ability to modulate the main OA pro-inflammatory biomarkers of novel gels (H-HA/BC) based on high molecular weight hyaluronan (H-HA) and unsulfated biotechnological chondroitin (BC). For the first time, BC was tested also in combination with H-HA on human primary cells isolated from pathological knee joints. Specifically, the experiments were performed using an OA in vitro model based on human chondrocytes and synoviocytes. To evaluate the anti-inflammatory effects of H-HA/BC in comparison with H-HA and BC single gels, NF-kB, COMP-2, MyD88, MMP-13 and a wide range of cytokines, known to be specific biomarkers in OA (e.g., IL-6, IL-8, and TNF-α), were evaluated. In addition, cell morphology and proliferation occurring in the presence of either H-HA/BC or single components were assessed using time-lapse video microscopy. It was shown that synovial fluids and cells isolated from OA suffering patients, presented a cytokine pattern respondent to an ongoing inflammation status. H-HA and BC significantly reduced the levels of 23 biomarkers associated with cartilage damage. However, H-HA/BC decreased significantly 24 biological mediators and downregulated 19 of them more efficiently than the single components. In synoviocytes cultures, cytokine analyses proved that H-HA/BC gels re-established an extracellular environment more similar to a healthy condition reducing considerably the concentration of 11 analytes. Instead, H-HA and BC significantly modulated 7 (5 only with a longer treatment) and 8 biological cytokines, respectively. Our results suggest that H-HA/BC beyond the viscosupplementation effect typical for HA-based gels, can improve the inflammation status in joints and thus could be introduced as a valid protective and anti-inflammatory intraarticular device in the field of Class III medical devices for OA treatments.

Streptomycetes as platform for biotechnological production processes of drugs.

Streptomyces is one of the most versatile genera for biotechnological applications, widely employed as platform in the production of drugs. Although streptomycetes have a complex life cycle and metabolism that would need multidisciplinary approaches, review papers have generally reported only studies on single aspects like the isolation of new strains and metabolites, morphology investigations, and genetic or metabolic studies. Besides, even if streptomycetes are extensively used in industry, very few review papers have focused their attention on the technical aspects of biotechnological processes of drug production and bioconversion and on the key parameters that have to be set up. This mini-review extensively illustrates the most innovative developments and progresses in biotechnological production and bioconversion processes of antibiotics, immunosuppressant, anticancer, steroidal drugs, and anthelmintic agents by streptomycetes, focusing on the process development aspects, describing the different approaches and technologies used in order to improve the production yields. The influence of nutrients and oxygen on streptomycetes metabolism, new fed-batch fermentation strategies, innovative precursor supplementation approaches, and specific bioreactor design as well as biotechnological strategies coupled with metabolic engineering and genetic tools for strain improvement is described. The use of whole, free, and immobilized cells on unusual supports was also reported for bioconversion processes of drugs. The most outstanding thirty investigations published in the last 8 years are here reported while future trends and perspectives of biotechnological research in the field have been illustrated. KEY POINTS: • Updated Streptomyces biotechnological processes for drug production are reported. • Innovative approaches for Streptomyces-based biotransformation of drugs are reviewed. • News about fermentation and genome systems to enhance secondary metabolite production.

The PI3K/AKT Pathway Is Activated by HGF in NT2D1 Non-Seminoma Cells and Has a Role in the Modulation of Their Malignant Behavior.

Overactivation of the c-MET/HGF system is a feature of many cancers. We previously reported that type II testicular germ cell tumor (TGCT) cells express the c-MET receptor, forming non-seminomatous lesions that are more positive compared with seminomatous ones. Notably, we also demonstrated that NT2D1 non-seminomatous cells (derived from an embryonal carcinoma lesion) increase their proliferation, migration, and invasion in response to HGF. Herein, we report that HGF immunoreactivity is more evident in the microenvironment of embryonal carcinoma biopsies with respect to seminomatous ones, indicating a tumor-dependent modulation of the testicular niche. PI3K/AKT is one of the signaling pathways triggered by HGF through the c-MET activation cascade. Herein, we demonstrated that phospho-AKT increases in NT2D1 cells after HGF stimulation. Moreover, we found that this pathway is involved in HGF-dependent NT2D1 cell proliferation, migration, and invasion, since the co-administration of the PI3K inhibitor LY294002 together with HGF abrogates these responses. Notably, the inhibition of endogenous PI3K affects collective cell migration but does not influence proliferation or chemotactic activity. Surprisingly, LY294002 administered without the co-administration of HGF increases cell invasion at levels comparable to the HGF-administered samples. This paradoxical result highlights the role of the testicular microenvironment in the modulation of cellular responses and stimulates the study of the testicular secretome in cancer lesions.

Biotechnological Transformation of Hydrocortisone into 16α-Hydroxyprednisolone by Coupling Arthrobacter simplex and Streptomyces roseochromogenes.

16α-Hydroxyprednisolone, an anti-inflammatory drug, could be potentially obtained from hydrocortisone bioconversion by combining a 1,2-dehydrogenation reaction performed by Arthrobacter simplexATCC31652 with a 16α-hydroxylation reaction by Streptomyces roseochromogenes ATCC13400. In this study we tested, for the first time, potential approaches to couple the two reactions using similar pH and temperature conditions for hydrocortisone bioconversion by the two strains. The A. simplex capability to 1,2-dehydrogenate the 16α-hydroxyhydrocortisone, the product of S. roseochromogenes transformation of hydrocortisone, and vice versa the capability of S. roseochromogenes to 16α-hydroxylate the prednisolone were assessed. Bioconversions were studied in shake flasks and strain morphology changes were observed by SEM. Whole cell experiments were set up to perform the two reactions in a sequential mode in alternate order or contemporarily at diverse temperature conditions. A. simplex catalyzed either the dehydrogenation of hydrocortisone into prednisolone efficiently or of 16α-hydroxyhydrocortisone into 16α-hydroxyprednisolone in 24 h (up to 93.9%). Surprisingly S. roseochromogenes partially converted prednisolone back to hydrocortisone. A 68.8% maximum of 16α-hydroxyprednisolone was obtained in 120-h bioconversion by coupling whole cells of the two strains at pH 6.0 and 26 °C. High bioconversion of hydrocortisone into 16α-hydroxyprednisolone was obtained for the first time by coupling A. simplex and S. roseochromogenes.

Annurca apple polyphenol extract promotes mesenchymal-to-epithelial transition and inhibits migration in triple-negative breast cancer cells through ROS/JNK signaling.

Aberrant activation of epithelial-to-mesenchymal transition has been shown to correlate with triple-negative breast cancer (TNBC) progression and metastasis. Thus, the induction of the reverse process might offer promising opportunities to restrain TNBC metastatic spreading and related mortality. Recently, the Annurca apple polyphenol extract (APE) has been highlighted as a multi-faceted agent that selectively kills TNBC cells by ROS generation and sustained JNK activation. Here, by qualitatively and quantitatively monitoring the real-time movements of live cells we provided the first evidence that APE inhibited the migration of MDA-MB-231 and MDA-MB-468 TNBC cells and downregulated metalloproteinase-2 and metalloproteinase-9. In MDA-MB-231 cells APE decreased SMAD-2/3 and p-SMAD-2/3 levels, increased E-cadherin/N-cadherin protein ratio, induced the switch from N-cadherin to E-cadherin expression and greatly reduced vimentin levels. Confocal and scanning electron microscopy imaging of APE-treated MDA-MB-231 cells evidenced a significant cytoskeletal vimentin and filamentous actin reorganization and revealed considerable changes in cell morphology highlighting an evident transition from the mesenchymal to epithelial phenotype with decreased migratory features. Notably, all these events were reverted by N-acetyl-L-cysteine and JNK inhibitor SP600125 furnishing evidence that APE exerted its effects through the activation of ROS/JNK signaling. The overall data highlighted APE as a potential preventing agent for TNBC metastasis.

Glutamic Acid as Repeating Building Block for Bio-Based Films.

Commercial inexpensive preparations of poly-γ-glutamic acid were used to obtain films made with a polypeptide constituted by a single repeating unit. The homopolymer was characterized by 1H-NMR spectroscopy and thermogravimetry, as well as by zeta potential and Z-average measurements. Manipulatable materials were obtained by casting film-forming solutions prepared at pH values between 3.0 and 4.0 and containing extensively dialyzed samples of the commercial product. The analysis of the mechanical properties highlighted a marked extensibility and plasticity of the films obtained without plasticizer, even though the addition of low amounts of glycerol (1-4%) was able to further increase these features. The characterization of poly-γ-glutamic acid molecular species, performed by membrane ultrafiltration and size-exclusion chromatography, coupled with triple-detection analysis of the obtained fractions, suggested that biopolymer chain length is responsible not only for its capacity to form film, but also for conferring to the films different features depending on the homopolymer molecular weight.