Influence of Fiber Diameter of Polycaprolactone Nanofibrous Materials on Biofilm Formation and Retention of Bacterial Cells.

To develop microbiologically safe nanofibrous materials, it is crucial to understand their interactions with microbial cells. Current research indicates that the morphology of nanofibers, particularly the diameter of the fibers, may play a significant role in biofilm formation and retention. However, it has not yet been determined how the fiber diameter of poly-ε-caprolactone (PCL), one of the most widely used biopolymers, affects these microbial interactions. In this study, two nanofibrous materials electrospun from PCL (PCL45 and PCL80) with different fiber diameter and characteristic distance δ between fibers were compared in terms of their ability to support or inhibit bacterial biofilm formation and retain bacterial cells. Strains of Escherichia coli (ATCC 25922 and ATCC 8739) and Staphylococcus aureus (ATCC 25923 and ATCC 6538) were used as model bacteria. Biofilm formation rate and retention varied significantly between the E. coli and S. aureus strains (p < 0.05) for the tested nanomaterials. In general, PCL showed a lower tendency to be colonized by the tested bacteria compared to the control material (polystyrene). Fiber diameter did not influence the biofilm formation rate of S. aureus strains and E. coli 25922 (p > 0.05), but it did significantly impact the biofilm formation rate of E. coli 8739 and biofilm morphology formed by all of the tested bacterial strains. In PCL45, thick uniform biofilm layers were formed preferably on the surface, while in PCL80 smaller clusters formed preferably inside the structure. Further, fiber diameter significantly influenced the retention of bacterial cells of all the tested strains (p < 0.001). PCL45, with thin fibers (average fiber diameter of 376 nm), retained up to 7 log (CFU mL-1) of staphylococcal cells (100% retention). The overall results indicate PCL45's potential for further research and highlight the nanofibers' morphology influence on bacterial interactions and differences in bacterial strains' behavior in the presence of nanomaterials.

The Impacts of the Sterilization Method and the Electrospinning Conditions of Nanofibrous Biodegradable Layers on Their Degradation and Hemocompatibility Behavior.

The use of electrospun polymeric biodegradable materials for medical applications is becoming increasingly widespread. One of the most important parameters regarding the functionality of nanofiber scaffolds during implantation and the subsequent regeneration of damaged tissues concerns their stability and degradation behavior, both of which are influenced by a wide range of factors (the properties of the polymer and the polymer solution, the technological processing approach, the sterilization method, etc.). This study monitored the degradation of nanofibrous materials fabricated from degradable polyesters as a result of the sterilization method applied (ethylene oxide and gamma irradiation) and the solvent system used to prepare the spun polymer solution. Aliphatic polyesters PCL and PLCL were chosen for this study and selected with respect to the applicability and handling in the surgical setting of these nanofibrous materials for vascular bandaging. The results revealed that the choice of solvent system exerts a significant impact on degradation during sterilization, especially at higher gamma irradiation values. The subsequent enzyme-catalyzed degradation of the materials following sterilization indicated that the choice of the sterilization method influenced the degradation behavior of the materials. Whereas wave-like degradation was evident concerning ethylene oxide sterilization, no such behavior was observed following gamma-irradiation sterilization. With concern for some of the tested materials, the results also indicated the potential for influencing the development of degradation within the bulk versus degradation from the surface of the material. Both the sterilization method and the choice of the spinning solvent system were found to impact degradation, which was observed to be most accelerated in the case of PLCL (L-lactide-co-caprolactone copolymer) electrospun from organic acids and subsequently sterilized using gamma irradiation. Since we planned to use these materials in cardiovascular applications, it was decided that their hemocompatibility would also be tested. The results of these tests revealed that changes in the structures of the materials initiated by sterilization may exert thrombogenic and anticoagulant impacts. Moreover, the microscopic analysis suggested that the solvent system used in the preparation of the materials potentially affects the behavior of erythrocytes; however, no indication of the occurrence of hemolysis was detected.

Current Approaches to Wound Repair in Burns: How far Have we Come From Cover to Close? A Narrative Review.

Burn injuries are a significant global health concern, with more than 11 million people requiring medical intervention each year and approximately 180,000 deaths annually. Despite progress in health and social care, burn injuries continue to result in socioeconomic burdens for victims and their families. The management of severe burn injuries involves preventing and treating burn shock and promoting skin repair through a two-step procedure of covering and closing the wound. Currently, split-thickness/full-thickness skin autografts are the gold standard for permanent skin substitution. However, deep burns treated with split-thickness skin autografts may contract, leading to functional and appearance issues. Conversely, defects treated with full-thickness skin autografts often result in more satisfactory function and appearance. The development of tissue-engineered dermal templates has further expanded the scope of wound repair, providing scar reductive and regenerative properties that have extended their use to reconstructive surgical interventions. Although their interactions with the wound microenvironment are not fully understood, these templates have shown potential in local infection control. This narrative review discusses the current state of wound repair in burn injuries, focusing on the progress made from wound cover to wound closure and local infection control. Advancements in technology and therapies hold promise for improving the outcomes for burn injury patients. Understanding the underlying mechanisms of wound repair and tissue regeneration may provide new insights for developing more effective treatments in the future.

A novel bifunctional multilayered nanofibrous membrane combining polycaprolactone and poly (vinyl alcohol) enriched with platelet lysate for skin wound healing.

Skin wound healing is a complex physiological process that involves various cell types, growth factors, cytokines, and other bioactive compounds. In this study, a novel dual-function multilayered nanofibrous membrane is developed for chronic wound application. The membrane is composed of five alternating layers of polycaprolactone (PCL) and poly (vinyl alcohol) (PVA) nanofibers (PCL-PVA) with a dual function: the PCL nanofibrous layers allow cell adhesion and growth, and the PVA layers enriched with incorporated platelet lysate (PCL-PVA + PL) serve as a drug delivery system for continuous release of bioactive compounds from PL into an aqueous environment. The material is produced using a needleless multi-jet electrospinning approach which can lead to homogeneous large-scale production. The bioactive PCL-PVA + PL membranes are cytocompatible and hemocompatible. A spatially compartmented co-culture of three cell types involved in wound healing - keratinocytes, fibroblasts and endothelial cells - is used for cytocompatibility studies. PCL-PVA + PL membranes enhance the proliferation of all cell types and increase the migration of both fibroblasts and endothelial cells. The membranes are also hemocompatible without any deleterious effect for thrombogenicity, hemolysis and coagulation. Thus, the beneficial effect of the PCL-PVA + PL membrane is demonstrated in vitro, making it a promising scaffold for the treatment of chronic wounds.

Assessment of thermally stabilized electrospun poly(vinyl alcohol) materials as cell permeable membranes for a novel blood salvage device.

The use of Intraoperative Cell Salvage (ICS) is currently limited in oncological surgeries, due to safety concerns associated with the ability of existing devices to successfully remove circulating tumour cells. In this work, we present the first stages towards the creation of an alternative platform to current cell savers, based on the extremely selective immunoaffinity membrane chromatography principle. Non-woven membranes were produced via electrospinning using poly(vinyl alcohol) (PVA), and further heat treated at 180 °C to prevent their dissolution in aqueous environments and preserve their fibrous morphology. The effects of the PVA degree of hydrolysis (DH) (98 % vs 99 %), method of electrospinning (needleless DC vs AC), and heat treatment duration (1-8 h) were investigated. All heat treated supports maintained their cytocompatibility, whilst tensile tests indicated that the 99 % hydrolysed DC electrospun mats were stronger compared to their 98 % DH counterparts. Although, and at the described conditions, AC electrospinning produced fibres with more than double the diameter compared to those from DC electrospinning, it was not chosen for subsequent experiments because it is still under development. Evidence of unimpeded passage of SY5Y neuroblastoma cells and undiluted defibrinated sheep's blood in flow-through filtration experiments confirmed the successful creation of 3D networks with minimum resistance to mass transfer and lack of non-specific cell binding to the base material, paving the way for the development of novel, highly selective ICS devices for tumour surgeries.

Novel lipophosphonoxin-loaded polycaprolactone electrospun nanofiber dressing reduces Staphylococcus aureus induced wound infection in mice.

Active wound dressings are attracting extensive attention in soft tissue repair and regeneration, including bacteria-infected skin wound healing. As the wide use of antibiotics leads to drug resistance we present here a new concept of wound dressings based on the polycaprolactone nanofiber scaffold (NANO) releasing second generation lipophosphonoxin (LPPO) as antibacterial agent. Firstly, we demonstrated in vitro that LPPO released from NANO exerted antibacterial activity while not impairing proliferation/differentiation of fibroblasts and keratinocytes. Secondly, using a mouse model we showed that NANO loaded with LPPO significantly reduced the Staphylococcus aureus counts in infected wounds as evaluated 7 days post-surgery. Furthermore, the rate of degradation and subsequent LPPO release in infected wounds was also facilitated by lytic enzymes secreted by inoculated bacteria. Finally, LPPO displayed negligible to no systemic absorption. In conclusion, the composite antibacterial NANO-LPPO-based dressing reduces the bacterial load and promotes skin repair, with the potential to treat wounds in clinical settings.

The Effect of the Controlled Release of Platelet Lysate from PVA Nanomats on Keratinocytes, Endothelial Cells and Fibroblasts.

Platelet lysate (PL) provides a natural source of growth factors and other bioactive molecules, and the local controlled release of these bioactive PL components is capable of improving the healing of chronic wounds. Therefore, we prepared composite nanofibrous meshes via the needleless electrospinning technique using poly(vinyl alcohol) (PVA) with a high molecular weight and with a high degree of hydrolysis with the incorporated PL (10% w/w). The morphology, wettability and protein release from the nanofibers was then assessed from the resulting composite PVA-PL nanomats. The bioactivity of the PVA-PL nanomats was proved in vitro using HaCaT keratinocytes, human saphenous endothelial cells (HSVECs) and 3T3 fibroblasts. The PVA-PL supported cell adhesion, proliferation, and viability. The improved phenotypic maturation of the HaCaT cells due to the PVA-PL was manifested via the formation of intermediate filaments positive for cytokeratin 10. The PVA-PL enhanced both the synthesis of the von Willebrand factor via HSVECs and HSVECs chemotaxis through membranes with 8 µm-sized pores. These results indicated the favorable effects of the PVA-PL nanomats on the three cell types involved in the wound healing process, and established PVA-PL nanomats as a promising candidate for further evaluation with respect to in vivo experiments.

The Effect of a Polyester Nanofibrous Membrane with a Fibrin-Platelet Lysate Coating on Keratinocytes and Endothelial Cells in a Co-Culture System.

Chronic wounds affect millions of patients worldwide, and it is estimated that this number will increase steadily in the future due to population ageing. The research of new therapeutic approaches to wound healing includes the development of nanofibrous meshes and the use of platelet lysate (PL) to stimulate skin regeneration. This study considers a combination of a degradable electrospun nanofibrous blend of poly(L-lactide-co-ε-caprolactone) and poly(ε-caprolactone) (PLCL/PCL) membranes (NF) and fibrin loaded with various concentrations of PL aimed at the development of bioactive skin wound healing dressings. The cytocompatibility of the NF membranes, as well as the effect of PL, was evaluated in both monocultures and co-cultures of human keratinocytes and human endothelial cells. We determined that the keratinocytes were able to adhere on all the membranes, and their increased proliferation and differentiation was observed on the membranes that contained fibrin with at least 50% of PL (Fbg + PL) after 14 days. With respect to the co-culture experiments, the membranes with fibrin with 20% of PL were observed to enhance the metabolic activity of endothelial cells and their migration, and the proliferation and differentiation of keratinocytes. The results suggest that the newly developed NF combined with fibrin and PL, described in the study, provides a promising dressing for chronic wound healing purposes.

Fabrication of dual-functional composite yarns with a nanofibrous envelope using high throughput AC needleless and collectorless electrospinning.

Nanotechnologies allow the production of yarns containing nanofibres for use in composites, membranes and biomedical materials. Composite yarns with a conventional thread core for mechanical strength and a nanofibrous envelope for functionality, e.g. biological, catalytic, have many advantages. Until now, the production of such yarns has been technologically difficult. Here, we show an approach to composite yarn production whereby a plume of nanofibers generated by high throughput AC needleless and collectorless electrospinning is wound around a classic thread. In the resulting yarn, nanofibres can form up to 80% of its weight. Our yarn production speed was 10 m/min; testing showed this can be increased to 60 m/min. After the yarn was embedded into knitwear, scanning electron microscope images revealed an intact nanofibrous envelope of the composite yarn. Our results indicate that this production method could lead to the widespread production and use of composite nanofibrous yarns on an industrial scale.

A cell-free nanofiber composite scaffold regenerated osteochondral defects in miniature pigs.

The aim of the study was to evaluate the effect of a cell-free hyaluronate/type I collagen/fibrin composite scaffold containing polyvinyl alcohol (PVA) nanofibers enriched with liposomes, basic fibroblast growth factor (bFGF) and insulin on the regeneration of osteochondral defects. A novel drug delivery system was developed on the basis of the intake effect of liposomes encapsulated in PVA nanofibers. Time-controlled release of insulin and bFGF improved MSC viability in vitro. Nanofibers functionalized with liposomes also improved the mechanical characteristics of the composite gel scaffold. In addition, time-controlled release of insulin and bFGF stimulated MSC recruitment from bone marrow in vivo. Cell-free composite scaffolds containing PVA nanofibers enriched with liposomes, bFGF, and insulin were implanted into seven osteochondral defects of miniature pigs. Control defects were left untreated. After 12 weeks, the composite scaffold had enhanced osteochondral regeneration towards hyaline cartilage and/or fibrocartilage compared with untreated defects that were filled predominantly with fibrous tissue. The cell-free composite scaffold containing PVA nanofibers, liposomes and growth factors enhanced migration of the cells into the defect, and their differentiation into chondrocytes; the scaffold was able to enhance the regeneration of osteochondral defects in minipigs.

A simple drug anchoring microfiber scaffold for chondrocyte seeding and proliferation.

The structural properties of microfiber meshes made from poly(2-hydroxyethyl methacrylate) (PHEMA) were found to significantly depend on the chemical composition and subsequent cross-linking and nebulization processes. PHEMA microfibres showed promise as scaffolds for chondrocyte seeding and proliferation. Moreover, the peak liposome adhesion to PHEMA microfiber scaffolds observed in our study resulted in the development of a simple drug anchoring system. Attached foetal bovine serum-loaded liposomes significantly improved both chondrocyte adhesion and proliferation. In conclusion, fibrous scaffolds from PHEMA are promising materials for tissue engineering and, in combination with liposomes, can serve as a simple drug delivery tool.

Analysis and three-dimensional visualization of collagen in artificial scaffolds using nonlinear microscopy techniques.

Extracellularly distributed collagen and chondrocytes seeded in gelatine and poly-ɛ-caprolactone scaffolds are visualized by two-photon excitation microscopy (TPEM) and second-harmonic generation (SHG) imaging in both forward and backward nondescanned modes. Joint application of TPEM and SHG imaging in combination with stereological measurements of collagen enables us not only to take high-resolution 3-D images, but also to quantitatively analyze the collagen volume and a spatial arrangement of cell-collagen-scaffold systems, which was previously impossible. This novel approach represents a powerful tool for the analysis of collagen-containing scaffolds with applications in cartilage tissue engineering.