Oleuropein-Rich Gellan Gum/Alginate Films as Innovative Treatments against Photo-Induced Skin Aging.

Olea europaea L. leaf extracts (OLEs) represent highly value-added agro-industrial byproducts, being promising sources of significant antioxidant compounds, such as their main component, oleuropein. In this work, hydrogel films based on low-acyl gellan gum (GG) blended with sodium alginate (NaALG) were loaded with OLE and crosslinked with tartaric acid (TA). The films' ability to act as an antioxidant and photoprotectant against UVA-induced photoaging, thanks to their capability to convey oleuropein to the skin, were examined with the aim of a potential application as facial masks. Biological in vitro performances of the proposed materials were tested on normal human dermal fibroblasts (NhDFs), both under normal conditions and after aging-induced UVA treatment. Overall, our results clearly show the intriguing properties of the proposed hydrogels as effective and fully naturally formulated anti-photoaging smart materials for potential use as facial masks.

Oxidative stress in retinal pigment epithelium impairs stem cells: a vicious cycle in age-related macular degeneration.

Aging, chronic oxidative stress, and inflammation are major pathogenic factors in the development and progression of age-related macular degeneration (AMD) with the loss of retinal pigment epithelium (RPE). The human RPE contains a subpopulation of progenitors (i.e., RPE stem cells-RPESCs) whose role in the RPE homeostasis is under investigation. We evaluated the paracrine effects of mature RPE cells exposed to oxidative stress (H2O2) on RPESCs behavior through co-cultural, morphofunctional, and bioinformatic approaches. RPESCs showed a decline in proliferation, an increase of the senescence-associated ß-galactosidase activity, the acquisition of a senescent-like secretory phenotype (SASP), and the reduction of their stemness and differentiation competencies. IL-6 and Superoxide Dismutase 2 (SOD2) seem to be key molecules in RPESCs response to oxidative stress. Our results get insight into stress-induced senescent-associated molecular mechanisms implicated in AMD pathogenesis. The presence of chronic oxidative stress in the microenvironment reduces the RPESCs abilities, inducing and/or maintaining a pro-inflammatory retinal milieu that in turn could affect AMD onset and progression.

Tissue Biomarkers Predicting Lymph Node Status in Cutaneous Melanoma.

Cutaneous melanoma is a severe neoplasm that shows early invasiveness of the lymph nodes draining the primary site, with increased risk of distant metastases and recurrence. The tissue biomarker identification could be a new frontier to predict the risk of early lymph node invasiveness, especially in cases considered by current guidelines to be at low risk of lymph node involvement and not requiring evaluation of the sentinel lymph node (SLN). For this reason, we present a narrative review of the literature, seeking to provide an overview of current tissue biomarkers, particularly vascular endothelium growth factors (VEGF), Tetraspanin CD9, lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1), D2-40, and gene expression profile test (31-GEP). Among these, 31-GEP seems to be able to provide a distinction between low or high risk for positive SLN classes. VEGF receptor-3 and CD9 expression may be independent predictors of positive SLN. Lastly, LYVE-1 and D2-40 allow an easier assessment of lymph vascular invasion, which can be considered a good predictor of SLN status. In conclusion, biomarkers to assess the lymph node status of cutaneous melanoma patients may play an important role in those cases where the clinician is in doubt whether or not to perform SLN biopsy.

Tetraspanin CD9 Expression Predicts Sentinel Node Status in Patients with Cutaneous Melanoma.

The tetraspanin CD9 is considered a metastasis suppressor in many cancers, however its role is highly debated. Currently, little is known about CD9 prognostic value in cutaneous melanoma. Our aim was to analyse CD9 expression in melanocytic nevi and primary cutaneous melanomas through immunohistochemistry and immunofluorescence approaches to determine its correlation with invasiveness and metastatic potential. CD9 displayed homogeneous staining in all melanocytic nevi. In contrast, it showed a complete loss of reactivity in all thin melanomas. Interestingly, CD9 was re-expressed in 46% of intermediate and thick melanomas in small tumor clusters predominantly located at sites of invasion near or inside the blood or lymphatic vessels. The most notable finding is that all CD9 stained melanomas presented sentinel node positivity. Additionally, a direct association between CD9 expression and presence of distant metastasis was reported. Finally, we confirm that CD9 expression is consistent with an early protective role against tumorigenesis, however, our data endorse in melanoma a specific function of CD9 in vascular dissemination during late tumor progression. The presence of CD9 hotspots could be essential for melanoma cell invasion in lymphatic and endothelial vessels. CD9 could be a valid prognostic factor for lymph node metastasis risk.

Paraoxonase-2: A potential biomarker for skin cancer aggressiveness.

BACKGROUND: Cutaneous neoplasms include melanoma and non-melanoma skin cancers (NMSCs). Among NMSCs, basal cell carcinoma (BCC) represents the most common lesion. On the contrary, although accounting for less than 5% of all skin cancers, melanoma is responsible for most of cutaneous malignancy-related deaths. Paraoxonase-2 (PON2) is an intracellular enzyme exerting a protective role against production of reactive oxygen species within mitochondrial respiratory chain. Recently, a growing attention has been focused on exploring the role of PON2 in cancer. The aim of this study was to investigate the diagnostic and prognostic role of PON2 in skin neoplasms. MATERIALS AND METHODS: 36 cases of BCC, distinguished between nodular and infiltrative lesions, as well as 29 melanoma samples were analysed by immunohistochemistry to evaluate PON2 protein expression. Subsequent statistical analyses were carried out to explore the existence of correlations between intratumour enzyme levels and clinicopathological features. RESULTS: Results obtained showed PON2 overexpression in BCCs compared with controls. In particular, distinguishing between less and more aggressive tumour forms, we found no significant differences in enzyme levels between nodular BCCs and controls. Conversely, PON2 expression was significantly higher in infiltrative BCCs compared with controls. Moreover, the enzyme was strongly upregulated in melanoma samples with respect to controls. Interestingly, PON2 levels were positively correlated with Breslow thickness, Clark level, regression, mitoses, lymph node metastases, primary tumour (pT) parameter and pathological stage. CONCLUSIONS: Reported findings seem to suggest that PON2 expression levels could be positively related with tumour aggressiveness of both BCC and melanoma.

Pro-inflammatory cytokines and microRNAs in male infertility.

BACKGROUND: Male infertility is a problem that affects 10-15% of men of reproductive age. In particular, gametogenesis is a complex process in which inflammation may play a central role through the secretion of cytokines and the expression of microRNAs. We assessed the potential role of proinflammatory cytokines (TNF-α, IL-6 and IL-1α) and microRNAs (miR-146a-5p, miR-34a-5p and miR-23a-3p) in the seminal plasma of infertile men compared to controls, evaluating their correlation with seminal and biochemical parameters. METHODS AND RESULTS: Expression of cytokines and microRNAs was analyzed by ELISA and q-PCR. Our data shows that IL-1α was significantly increased in the azoospermic group compared to controls, TNF-α mRNA was more expressed in the oligozoospermic group than controls. There were no significant differences in miRNAs expression among the three groups. The correlations between sperm parameters and inflammatory markers were evaluated, however no significance was highlighted. CONCLUSIONS: The determination of each inflammatory marker separately in the seminal plasma of subfertile men, despite some significant differences, does not have a diagnostic value in male infertility even if an assay of selective pro-inflammatory cytokines and microRNAs in the semen may improve the diagnosis of male infertility.

Innovative Eco-Friendly Hydrogel Film for Berberine Delivery in Skin Applications.

Hydrogel formulations (masks or patches, without tissue support) represent the new frontier for customizable skin beauty and health. The employment of these materials is becoming popular in wound dressing, to speed up the healing process while protecting the affected area, as well as to provide a moisturizing reservoir, control the inflammatory process and the onset of bacterial development. Most of these hydrogels are acrylic-based at present, not biodegradable and potentially toxic, due to acrylic monomers residues. In this work, we selected a new class of cellulose-derived and biodegradable hydrogel films to incorporate and convey an active compound for dermatological issues. Films were obtained from a combination of different polysaccharides and clays, and berberine hydrochloride, a polyphenolic molecule showing anti-inflammatory, immunomodulatory, antibacterial and antioxidant properties, was chosen and then embedded in the hydrogel films. These innovative hydrogel-based systems were characterized in terms of water uptake profile, in vitro cytocompatibility and skin permeation kinetics by Franz diffusion cell. Berberine permeation fitted well to Korsmeyer-Peppas kinetic model and achieved a release higher than 100 µg/cm2 within 24 h. The latter study, exploiting a reliable skin model membrane, together with the biological assessment, gained insights into the most promising formulation for future investigations.

Traditional Chinese Medicine in Oncotherapy: The Research Status.

In China, Traditional Chinese Medicine (TCM) plays a vital role in the comprehensive treatment of cancer. As an auxiliary and supplement of major treatment modalities for cancer such as surgery, chemotherapy, and radiotherapy, both clinical observations and biomolecular research have confirmed the therapeutic efficacy of TCM in cancer.

Pectin-GPTMS-Based Biomaterial: toward a Sustainable Bioprinting of 3D scaffolds for Tissue Engineering Application.

Developing green and nontoxic biomaterials, derived from renewable sources and processable through 3D bioprinting technologies, is an emerging challenge of sustainable tissue engineering. Here, pectin from citrus peels was cross-linked for the first time with (3-glycidyloxypropyl)trimethoxysilane (GPTMS) through a one-pot procedure. Freeze-dried porous pectin sponges, with tunable properties in terms of porosity, water uptake, and compressive modulus, were obtained by controlling GPTMS content. Cell experiments showed that GPTMS did not affect the cytocompatibility of pectin. The addition of GPTMS improved the printability of pectin due to an increase of viscosity and yield stress. Three-dimensional woodpile and complex anatomical-shaped scaffolds with interconnected micro- and macropores were, therefore, bioprinted without the use of any additional support material. These results show the great potential of using pectin cross-linked with GPTMS as biomaterial ink to fabricate patient-specific scaffolds, which could be used to promote tissue regeneration in vivo.

How the Pathological Microenvironment Affects the Behavior of Mesenchymal Stem Cells in the Idiopathic Pulmonary Fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a chronic disease characterized by fibroblasts activation, ECM accumulation, and diffused alveolar inflammation. The role of inflammation in IPF is still controversial and its involvement may follow nontraditional mechanisms. It is seen that a pathological microenvironment may affect cells, in particular mesenchymal stem cells (MSCs) that may be able to sustain the inflamed microenvironment and influence the surrounding cells. Here MSCs have been isolated from fibrotic (IPF-MSCs) and control (C-MSCs) lung tissue; first cells were characterized and compared by the expression of molecules related to ECM, inflammation, and other interdependent pathways such as hypoxia and oxidative stress. Subsequently, MSCs were co-cultured between them and with NHLF to test the effects of the cellular crosstalk. Results showed that pathological microenvironment modified the features of MSCs: IPF-MSCs, compared to C-MSCs, express higher level of molecules related to ECM, inflammation, oxidative stress, and hypoxia; notably, when co-cultured with C-MSCs and NHLF, IPF-MSCs are able to induce a pathological phenotype on the surrounding cell types. In conclusion, in IPF the pathological microenvironment affects MSCs that in turn can modulate the behavior of other cell types favoring the progression of IPF.

Pre-eclampsia onset and SPARC: A possible involvement in placenta development.

Pre-eclampsia (PE) is a multisystem disorder commonly diagnosed in the latter half of pregnancy and it is a leading cause of intrauterine fetal growth retardation (IUGR). The aim of this study was to investigate the localization and the role of SPARC, secreted protein acidic, and rich in cysteine, in PE and PE-IUGR placentas in comparison with normal placentas. SPARC was mainly expressed in the villous and extravillous cytotrophoblastic cells in first trimester, whereas in PE, PE-IUGR and at term placentas, SPARC immunostaining was visible in both cytotrophoblastic cells and syncytiotrophoblast. SPARC expression significantly decreased in normal placenta from first to third trimester and a further significant reduction was demonstrated in PE and PE-IUGR. The latter downregulation of SPARC depends on hypoxic condition as shown by in vitro models. In conclusion, SPARC can play a pivotal role in PE and PE-IUGR onset and it should be considered as a key molecule for future investigations in such pathologies.

Dental pulp stem cells senescence and regenerative potential relationship.

Uncomplicated treatments for pulpitis and periodontitis continues to be challenging and regenerative approaches could meet this contingency. Dental pulp stem cells (DPSCs) represent a good candidate for oral recovering therapies. Here, we investigated changes in morphology, proliferation, and in vitro differentiation toward mesenchymal and neuronal phenotypes of human DPSCs harvested from differently aged donors. Aging is a physiologic phenomenon occurring with time that hamper body's capability to maintain homeostasis also affecting the functional reserve. Cytofluorimetric, immunohistochemical, quantitative reverse-transcription polymerase chain reaction (qRT-PCR), and western blot analyses were performed to gain insight for successful regenerative strategies in elderly. We observed a decline in DPSCs proliferation and differentiation potential with age. Interestingly, these cells behaved differently under osteogenic or odontogenic stimuli, showing different age-related mineralization capabilities. Similarly, neurogenic differentiation decreased with age. In conclusion, our observations represent a valid tool for the development of tailored regenerative strategies in an aging society.

Synovium-derived stromal cell-induced osteoclastogenesis: a potential osteoarthritis trigger.

Purpose: To shed light on the idea that mesenchymal stem/stromal cells (MSCs) recruited in synovium (SM) (i.e. Synovium-Derived Stromal Cells, SDSCs) could be involved in Osteoarthritis (OA) pathophysiology. Attention was also paid to a further stromal cell type with a peculiar ultrastructure called telocytes (TCs), whose role is far from clarified. Methods: In the present in vitro study, we compared SDSCs isolated from healthy and OA subjects in terms of phenotype, morphology and differentiation potential as well as in their capability to activate normal Peripheral Blood Mononuclear Cells (PBMCs). Histological, immunohistochemical and ultrastructural analyses were integrated by qRT-PCR and functional resorbing assays. Results: Our data demonstrated that both SDSC populations stimulated the formation of osteoclasts from PBMCs: the osteoclast-like cells generated by healthy-SDSCs via transwell co-cultures were inactive, while OA-derived SDSCs have a much greater effectiveness. Moreover, the presence of TCs was more evident in cultures obtained from OA subjects and suggests a possible involvement of these cells in OA. Conclusions: Osteoclastogenic differentiation capability of PBMCs from OA subjects, also induced by B synoviocytes has been already documented. Here we hypothesized that SDSCs, generally considered for their regenerative potential in cartilage lesions, have also a role in the onset/maintenance of OA. Clinical relevance: Our observations may represent an interesting opportunity for the development of a holistic approach for OA treatment, that considers the multifaceted capability of MSCs in relation to the environment.

The use of cell conditioned medium for musculoskeletal tissue regeneration.

Tissue regenerative medicine combines the use of cells, scaffolds, and molecules to repair damaged tissues. Different cell types are employed for musculoskeletal diseases, both differentiated and mesenchymal stromal cells (MSCs). In recent years, the hypothesis that cell-based therapy is guided principally by cell-secreted factors has become increasingly popular. The aim of the present literature review was to evaluate preclinical and clinical studies that used conditioned medium (CM), rich in cell-factors, for musculoskeletal regeneration. Thirty-one were in vitro, 12 in vivo studies, 1 was a clinical study, and 2 regarded extracellular vesicles. Both differentiated cells and MSCs produce CM that induces reduction in inflammation and increases synthetic activity. MSC recruitment and differentiation, endothelial cell recruitment and angiogenesis have also been observed. In vivo studies were performed with CM in bone and periodontal defects, arthritis and muscle dystrophy pathologies. The only clinical study was performed with CM from MSCs in patients needing alveolar bone regeneration, showing bone formation and no systemic or local complications. Platelet derived growth factor receptor ß, C3a, vascular endothelial growth factor, monocyte chemoattractant protein-1 and -3, interleukin 3 and 6, insulin-like growth factor-I were identified as responsible of cell migration, proliferation, osteogenic differentiation, and angiogenesis. The use of CM could represent a new regenerative treatment in several musculoskeletal pathologies because it overcomes problems associated with the use of cells and avoids the use of exogenous GFs or gene delivery systems. However, some issues remain to be clarified.

Detecting senescent fate in mesenchymal stem cells: a combined cytofluorimetric and ultrastructural approach.

Senescence can impair the therapeutic potential of stem cells. In this study, senescence-associated morphofunctional changes in periosteum-derived progenitor cells (PDPCs) from old and young individuals were investigated by combining cytofluorimetry, immunohistochemistry, and transmission electron microscopy. Cell cycle analysis demonstrated a large number of G0/G1 phase cells in PDPCs from old subjects and a progressive accumulation of G0/G1 cells during passaging in cultures from young subjects. Cytofluorimetry documented significant changes in light scattering parameters and closely correlated with the ultrastructural features, especially changes in mitochondrial shape and autophagy, which are consistent with the mitochondrial-lysosomal axis theory of ageing. The combined morphological, biofunctional, and ultrastructural approach enhanced the flow cytometric study of PDPC ageing. We speculate that impaired autophagy, documented in replicative senescent and old PDPCs, reflect a switch from quiescence to senescence. Its demonstration in a tissue with limited turnover-like the cambium layer of the periosteum, where reversible quiescence is the normal stem cell state throughout life-adds a new piece to the regenerative medicine jigsaw in an ageing society.

Silver-loaded chitosan coating as an integrated approach to face titanium implant-associated infections: analytical characterization and biological activity.

The present work focuses on the idea to prevent and/or inhibit the colonization of implant surfaces by microbial pathogens responsible for post-operative infections, adjusting antimicrobial properties of the implant surface prior to its insertion. An antibacterial coating based on chitosan and silver was developed by electrodeposition techniques on poly(acrylic acid)-coated titanium substrates. When a silver salt was added during the chitosan deposition step, a stable and scalable silver incorporation was achieved. The physico-chemical composition of the coating was studied by X-ray photoelectron spectroscopy (XPS), while atomic force microscopy in intermittent contact mode (ICAFM) was used to explore the coating morphology. The amount of silver released from the coating up to 21 days was evaluated by inductively coupled plasma mass spectrometry (ICP-MS). The capability of the proposed coating to interact in vitro with the biological environment in terms of compatibility and antibacterial properties was assessed using MG-63 osteoblast-like cell line and S. aureus and P. aeruginosa strains, respectively. These studies revealed that a coating showing a silver surface atomic percentage equal to 0.3% can be effectively used as antibacterial system, while providing good viability of osteoblast-like cells after 7 days. The antibacterial effectiveness of the prepared coating is mainly driven by a contact killing mechanism, although the low concentration of silver released (below 0.1 ppm up to 21 days) is enough to inhibit bacterial growth, advantaging MG-63 cells in the race for the surface.

Evidence Supporting a Paracrine Effect of IGF-1/VEGF on Human Mesenchymal Stromal Cell Commitment.

Healing of skeletal defects is strictly dependent on osteogenesis and efficient vascularization of engineered scaffolds. Insulin-like growth factor-1 (IGF-1) and vascular endothelial growth factor (VEGF) are both involved in these processes. The in vitro administration of IGF-1 in association with VEGF is able to modulate the osteoblastic or endothelial commitment of mesenchymal stromal cells (MSCs) of different origins (e.g. periosteum and skin). In the present study, in order to deepen a possible paracrine effect of IGF-1 and VEGF on periosteum-derived progenitor cells (PDPCs) and skin-derived MSCs (S-MSCs), a Transwell coculture approach was used. We explored the genes involved in endothelial and osteoblastic differentiation, those modulating mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3'-kinase (PI3K)-AKT signaling pathways as well as genes implicated in stemness (i.e. Sox2, Oct4, and Nanog). Periosteal cells, which are typically committed toward osteoblastogenesis, are driven in the direction of endothelial gene expression when influenced by S-MSCs. The latter, once influenced by PDPCs, lose their endothelial commitment and increase the expression of osteoblast-associated genes. PI3K/AKT and MAPK signaling pathways seem to be markedly involved in this behavior. Our results evidence that paracrine signals between MSCs may differently modulate their commitment in a bone microenvironment, opening stimulating viewpoints for skeletal tissue engineering strategies coupling angiogenesis and osteogenesis processes.

mRNAs and miRNAs profiling of mesenchymal stem cells derived from amniotic fluid and skin: the double face of the coin.

Mesenchymal stem cells (MSCs) can be isolated from different adult sources and, even if the minimal criteria for defining MSCs have been reported, the scientific question about the potential distinctions among MSCs derived from different sources is still open. In particular, it is debated whether MSCs of different origin have the same grade of stemness or whether the source affects their undifferentiated status. Here, we report not only the isolation and the traditional characterization of MSCs derived from amniotic fluid (AF-MSCs) and skin (S-MSCs) but also a molecular characterization based on mRNAs and miRNAs profiling. Our results show that, even if both AF- and S-MSCs are mostly regulated by the same pathways (such as Wnt, MAPK and TGF-ß), there are some important differences at the molecular level that directly affect important cellular features, such as the ability to differentiate into adipocytes. In conclusion, even if further studies are necessary to improve the knowledge about the role of each dysregulated miRNAs gene, these differences may actually strengthen the question about the importance of tissue origin.

Emerging biomedical applications of nano-chitins and nano-chitosans obtained via advanced eco-friendly technologies from marine resources.

The present review article is intended to direct attention to the technological advances made in the 2010-2014 quinquennium for the isolation and manufacture of nanofibrillar chitin and chitosan. Otherwise called nanocrystals or whiskers, n-chitin and n-chitosan are obtained either by mechanical chitin disassembly and fibrillation optionally assisted by sonication, or by e-spinning of solutions of polysaccharides often accompanied by poly(ethylene oxide) or poly(caprolactone). The biomedical areas where n-chitin may find applications include hemostasis and wound healing, regeneration of tissues such as joints and bones, cell culture, antimicrobial agents, and dermal protection. The biomedical applications of n-chitosan include epithelial tissue regeneration, bone and dental tissue regeneration, as well as protection against bacteria, fungi and viruses. It has been found that the nano size enhances the performances of chitins and chitosans in all cases considered, with no exceptions. Biotechnological approaches will boost the applications of the said safe, eco-friendly and benign nanomaterials not only in these fields, but also for biosensors and in targeted drug delivery areas.

Early Biofilm Formation on the Drain Tip after Total Knee Arthroplasty Is Not Associated with Prosthetic Joint Infection: A Pilot Prospective Case Series Study of a Single Center.

BACKGROUND: Periprosthetic joint infection (PJI) is a devastating complication of arthroplasties that could occur during the surgery. The purpose of this study was to analyze the biofilm formation through microbiological culture tests and scanning electron microscopy (SEM) on the tip of surgical drainage removed 24 h after arthroplasty surgery. METHODS: A total of 50 consecutive patients were included in the present prospective observational study. Drains were removed under total aseptic conditions twenty-four hours after surgery. The drain tip was cut in three equal parts of approximately 2-3 cm in length and sent for culture, culture after sonication, and SEM analysis. The degree of biofilm formation was determined using a SEM semi-quantitative scale. RESULTS: From the microbiological analysis, the cultures of four samples were positive. The semi-quantitative SEM analysis showed that no patient had grade 4 of biofilm formation. A total of 8 patients (16%) had grade 3, and 14 patients (28%) had grade 2. Grade 1, scattered cocci with immature biofilm, was contemplated in 16 patients (32%). Finally, 12 patients (24%) had grade 0 with a total absence of bacteria. During the follow-up (up to 36 months), no patient showed short- or long-term infectious complications. CONCLUSIONS: Most of the patients who underwent primary total knee arthroplasty (TKA) showed biofilm formation on the tip of surgical drain 24 h after surgery even though none showed a mature biofilm formation (grade 4). Furthermore, 8% of patients were characterized by a positivity of culture analysis. However, none of the patients included in the study showed signs of PJI up to 3 years of follow-up.