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.

On the Biomechanical Performances of Duplex Stainless Steel Graded Scaffolds Produced by Laser Powder Bed Fusion for Tissue Engineering Applications.

This experimental study aims to extend the know-how on biomechanical performances of duplex stainless steel (DSS) for tissue engineering applications to a graded lattice geometry scaffold based on the F53 DSS (UNS S32750 according to ASTM A182) produced by laser powder bed fusion (LPBF). The same dense-out graded geometry based on rhombic dodecahedral elementary unit cells investigated in previous work on 316L stainless steel (SS) was adopted here for the manufacturing of the F53 DSS scaffold (SF53). Microstructural characterization and mechanical and biological tests were carried out on the SF53 scaffold, using the in vitro behavior of the 316L stainless steel scaffold (S316L) as a control. Results show that microstructure developed as a consequence of different volume energy density (VED) values is mainly responsible for the different mechanical behaviors of SF53 and S316L, both fabricated using the same LPBF manufacturing system. Specifically, the ultimate compressive strength (σUC) and elastic moduli (E) of SF53 are three times and seven times higher than S316L, respectively. Moreover, preliminary biological tests evidenced better cell viability in SF53 than in S316L already after seven days of culture, suggesting SF53 with dense-out graded geometry as a viable alternative to 316L SS for bone tissue engineering applications.

Influence of Trabecular Geometry on Scaffold Mechanical Behavior and MG-63 Cell Viability.

In a scaffold-based approach for bone tissue regeneration, the control over morphometry allows for balancing scaffold biomechanical performances. In this experimental work, trabecular geometry was obtained by a generative design process, and scaffolds were manufactured by vat photopolymerization with 60% (P60), 70% (P70) and 80% (P80) total porosity. The mechanical and biological performances of the produced scaffolds were investigated, and the results were correlated with morphometric parameters, aiming to investigate the influence of trabecular geometry on the elastic modulus, the ultimate compressive strength of scaffolds and MG-63 human osteosarcoma cell viability. The results showed that P60 trabecular geometry allows for matching the mechanical requirements of human mandibular trabecular bone. From the statistical analysis, a general trend can be inferred, suggesting strut thickness, the degree of anisotropy, connectivity density and specific surface as the main morphometric parameters influencing the biomechanical behavior of trabecular scaffolds, in the perspective of tissue engineering applications.

Altered type I collagen networking in osteoporotic human femoral head revealed by histomorphometric and Fourier transform infrared imaging correlated analyses.

Bone homeostasis is the equilibrium between organic and inorganic components of the extracellular matrix (ECM) and cells. Alteration of this balance has consequences on bone mass and architecture, resulting in conditions such as osteoporosis (OP). Given ECM protein mutual regulation and their effects on bone structure and mineralization, further insight into their expression is crucial to understanding bone biology under normal and pathological conditions. This study focused on Type I Collagen, which is mainly responsible for structural properties and mineralization of bone, and selected proteins implicated in matrix composition, mineral deposition, and cell-matrix interaction such as Decorin, Osteocalcin, Osteopontin, Bone Sialoprotein 2, Osteonectin and Transforming Growth Factor beta. We developed a novel multidisciplinary approach in order to assess bone matrix in healthy and OP conditions more comprehensively by exploiting the Fourier Transform Infrared Imaging (FTIRI) technique combined with histomorphometry, Sirius Red staining, immunohistochemistry, and Western Blotting. This innovatory procedure allowed for the analysis of superimposed tissue sections and revealed that the alterations in OP bone tissue architecture were associated with warped Type I Collagen structure and deposition but not with changes in the total protein amount. The detected changes in the expression and/or cooperative or antagonist role of Decorin, Osteocalcin, Osteopontin, and Bone Sialoprotein-2 indicate the deep impact of these NCPs on collagen features of OP bone. Overall, our strategy may represent a starting point for designing targeted clinical strategies aimed at bone mass preservation and sustain the FTIRI translational capability as upcoming support for traditional diagnostic methods.

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.

The multifaced role of HtrA1 in the development of joint and skeletal disorders.

HtrA1 (High temperature requirement A1) family proteins include four members, widely conserved from prokaryotes to eukaryotes, named HtrA1, HtrA2, HtrA3 and HtrA4. HtrA1 is a serine protease involved in a variety of biological functions regulating many signaling pathways degrading specific components and playing key roles in many human diseases such as neurodegenerative disorders, pregnancy complications and cancer. Due to its role in the breakdown of many ExtraCellular Matrix (ECM) components of articular cartilage such as fibronectin, decorin and aggrecan, HtrA1 encouraged many researches on studying its role in several skeletal diseases (SDs). These studies were further inspired by the fact that HtrA1 is able to regulate the signaling of one of the most important cytokines involved in SDs, the TGFß-1. This review aims to summarize the data currently available on the role of HtrA1 in skeletal diseases such as Osteoporosis, Rheumatoid Arthritis, Osteoarthritis and Intervertebral Disc Degeneration (IDD). The use of HtrA1 as a marker of frailty in geriatric medicine would represent a powerful tool for identifying older individuals at risk of developing skeletal disorders, evaluating an appropriate intervention to improve quality care in these people avoiding or improving age-related SDs in the elderly population.

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.

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.

Insights into oxidative stress in bone tissue and novel challenges for biomaterials.

The presence of Reactive Oxygen Species (ROS) in bone can influence resident cells behaviour as well as the extra-cellular matrix composition and the tissue architecture. Aging, in addition to excessive overloads, unbalanced diet, smoking, predisposing genetic factors, lead to an increase of ROS and, if it is accompanied with an inappropriate production of scavengers, promotes the generation of oxidative stress that encourages bone catabolism. Furthermore, bone injuries can be triggered by numerous events such as road and sports accidents or tumour resection. Although bone tissue possesses a well-known repair and regeneration capacity, these mechanisms are inefficient in repairing large size defects and bone grafts are often necessary. ROS play a fundamental role in response after the implant introduction and can influence its success. This review provides insights on the mechanisms of oxidative stress generated by an implant in vivo and suitable ways for its modulation. The local delivery of active molecules, such as polyphenols, enhanced bone biomaterial integration evidencing that the management of the oxidative stress is a target for the effectiveness of an implant. Polyphenols have been widely used in medicine for cardiovascular, neurodegenerative, bone disorders and cancer, thanks to their antioxidant and anti-inflammatory properties. In addition, the perspective of new smart biomaterials and molecular medicine for the oxidative stress modulation in a programmable way, by the use of ROS responsive materials or by the targeting of selective molecular pathways involved in ROS generation, will be analysed and discussed critically.

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.

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.

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.

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.

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.

Collagen and non-collagenous proteins molecular crosstalk in the pathophysiology of osteoporosis.

Collagenous and non-collagenous proteins (NCPs) in the extracellular matrix, as well as the coupling mechanisms between osteoclasts and osteoblasts, work together to ensure normal bone metabolism. Each protein plays one or more critical roles in bone metabolism, sometimes even contradictory, thus affecting the final mechanical, physical and chemical properties of bone tissue. Anomalies in the amount and structure of one or more of these proteins can cause abnormalities in bone formation and resorption, which consequently leads to malformations and defects, such as osteoporosis (OP). The connections between key proteins involved in matrix formation and resorption are far from being elucidated. In this review, we resume knowledge on the crosstalk between collagen type I and selected NCPs (Transforming Growth Factor-ß, Insulin-like Growth Factor-1, Decorin, Osteonectin, Osteopontin, Bone Sialoprotein and Osteocalcin) of bone matrix, focusing on their possible involvement and role in OP. The different elements of this network can be pharmacologically targeted or used for the design/development of innovative regenerative strategies to modulate a feedback loop in bone remodelling.

Newly-designed collagen/polyurethane bioartificial blend as coating on bioactive glass-ceramics for bone tissue engineering applications.

In the present work, a new combination of synthetic and natural biomaterials is proposed for bone tissue engineering (BTE) applications. In order to mimic the inorganic and organic phases of bone extracellular matrix (ECM), a bioactive glass-ceramic deriving from a SiO2-P2O5-CaO-MgO-Na2O-K2O parent glass, acting as a substrate in form of a slice, was surface-functionalised with a type I collagen-based coating. In particular, the collagen was blended with a water soluble polyurethane (PUR), synthesised from poly(ethylene glycol), 1,6-hexamethylene diisocyanate and N-BOC-serinol. The PUR was designed to expose amino groups on the polymeric chain, which can be exploited for the blend stabilisation through crosslinking. The newly synthesised PUR demonstrated to be non-cytotoxic, as assessed by a biological test with MG-63 osteoblast-like cells. The collagen/PUR blend showed good biocompatibility as well. The polymeric coating on the glass-ceramic samples was produced by surface-silanisation, followed by further chemical grafting of the blend, using genipin as a crosslinker. The glass-ceramic surface was characterised at each functionalisation step, demonstrating that the procedure allowed obtaining a covalent link between the blend and the substrate. Finally, biological tests performed using human periosteal derived precursor cells demonstrated that the proposed polymer-coated material was a good substrate for bone cell adhesion and growth, and a good candidate to mimic the composite nature of the bone ECM.

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.