Targeting the Complexity of In Vitro Skin Models: A Review of Cutting-Edge Developments.

Skin in vitro models offer much promise for research, testing drugs, cosmetics, and medical devices, reducing animal testing and extensive clinical trials. There are several in vitro approaches to mimicking human skin behavior, ranging from simple cell monolayer to complex organotypic and bioengineered 3-dimensional models. Some have been approved for preclinical studies in cosmetics, pharmaceuticals, and chemicals. However, development of physiologically reliable in vitro human skin models remains in its infancy. This review reports on advances in in vitro complex skin models to study skin homeostasis, aging, and skin disease.

Novel 3D printed polysaccharide-based materials with prebiotic activity for potential treatment of diaper rash.

Diaper rash, mainly occurring as erythema and itching in the diaper area, causes considerable distress to infants and toddlers. Increasing evidence suggests that an unequal distribution of microorganisms on the skin contributes to the development of diaper dermatitis. Probiotic bacteria, like Staphylococcus epidermidis, are crucial for maintaining a healthy balance in the skin's microbiome, among others, through their fermentative metabolites, such as short-chain fatty acids. Using a defined prebiotic as a carbon source (e.g., as part of the diaper formulation) can selectively trigger the fermentation of probiotic bacteria. A proper material choice can reduce diaper rash incidence by diminishing the skin exposure to wetness and faeces. Using 3D printing, we fabricated carbon-rich materials for the top sheet layer of baby diapers that enhance the probiotic activity of S. epidermidis. The developed materials' printability, chemical composition, swelling ability, and degradation rate were analysed. In addition, microbiological tests evaluated their potential as a source of in situ short-chain fatty acid production. Finally, biocompatibility testing with skin cells evaluated their safety for potential use as part of diapers. The results demonstrate a cost-effective approach for producing novel materials that can tailor the ecological balance of the skin microflora and help treat diaper rash.

Development of a novel NiCu nanoparticle-loaded polysaccharide-based hydrogel for 3D printing of customizable dressings with promising cytotoxicity against melanoma cells.

Polysaccharide hydrogels and metal alloy nanoparticles have already found use in a range of biomedical applications. Nickel-copper nanoparticles (NiCu NPs) are particularly promising due to their tunable properties, such as ferromagnetism, biocompatibility, and antimicrobial activity. At the same time, polysaccharide hydrogels made of polymer mixtures such as alginate and methylcellulose with incorporated metal alloy nanoparticles are reported in the scientific literature. In view of this, in this work, NiCu NPs are combined with polysaccharide hydrogels and 3D printed to construct geometrically customizable dressings with tailorable properties for melanoma treatment. This novel combination exploits the intrinsic magnetic properties of NiCu NPs and the same time builds on their less known properties to improve the mechanic stability of 3D printed materials, both contributing to a previously not reported application as potent cytotoxic dressing against melanoma cells. The dressings were evaluated in terms of their physico-chemical characteristics, and their potential application, namely melanoma cell cytotoxicity. While all dressings exhibited similar degradation profiles regardless of composition, the addition of NiCu NPs had an effect on the hydrophilicity, swelling rates, and topographical properties of the dressings. Compression results showed that the presence of NPs increased the stiffness of the dressings, while the ultimate tensile strength was highest at 0.31 MPa for the dressings with 0.5 wt% NPs. We show that although the base formulation of the dressings is biocompatible with skin-derived cells, dressings loaded with NPs exhibit promising antimelanoma activity. Extracts obtained from dressings containing 0.5 wt% NPs reduced melanoma cell viability to 61% ± 11% and 40% ± 2% after 24 h and 72 h of soaking, respectively. Furthermore, extracts of dressings with 1 wt% NPs reduced melanoma cell viability to less than 15% within the first 24 h. By adjusting the NP content, the mechanical properties, surface roughness, and wettability can be tuned so that the dressings can be functionally customized. In addition, by using 3D printing as a fabrication process, the shape and composition of the dressings can be tailored to the patient's needs. The dressings also remained intact after soaking in simulated physiological solution for 14 days, indicating their suitability for long-term topical application.

Cell Electrospinning: A Mini-Review of the Critical Processing Parameters and Its Use in Biomedical Applications.

Functional tissue engineering is a widely studied area of research with increasing importance in regenerative medicine, as well as in the development of in vitro models used for drug discovery and mimicking diseased tissues, among other applications. Electrospinning (ES) is one of the most widely used methods in these fields. It has attracted considerable interest because it can produce materials resembling the extracellular matrix of native tissues. The micro/nanofibers produced by this method provide a cell-friendly environment that promotes cellular activities. Cell electrospinning (C-ES) is based on the fundamental ES process and enables the encapsulation of viable cells in a micro/nanofibrous mesh. In this review, the process of C-ES and the materials used in this process are discussed. This work also discusses the applications of C-ES in tissue engineering, focusing on recent advances in this field.

Multilayer Methacrylate-Based Wound Dressing as a Therapeutic Tool for Targeted Pain Relief.

This study presents an innovative wound dressing system that offers a highly effective therapeutic solution for treating painful wounds. By incorporating the widely used non-steroidal anti-inflammatory drug diclofenac, we have created an active wound dressing that can provide targeted pain relief with ease. The drug was embedded within a biocompatible matrix composed of polyhydroxyethyl methacrylate and polyhydroxypropyl methacrylate. The multilayer structure of the dressing, which allows for sustained drug release and an exact application, was achieved through the layer-by-layer coating technique and the inclusion of superparamagnetic iron platinum nanoparticles. The multilayered dressings' physicochemical, structural, and morphological properties were characterised using various methods. The synergistic effect of the incorporated drug molecules and superparamagnetic nanoparticles on the surface roughness and release kinetics resulted in controlled drug release. In addition, the proposed multilayer wound dressings were found to be biocompatible with human skin fibroblasts. Our findings suggest that the developed wound dressing system can contribute to tailored therapeutic strategies for local pain relief.

The development of an electropolymerized, molecularly imprinted polymer (MIP) sensor for insulin determination using single-drop analysis.

An electrochemical sensor for the detection of insulin in a single drop (50 µL) was developed based on the concept of molecularly imprinted polymers (MIP). The synthetic MIP receptors were assembled on a screen-printed carbon electrode (SPCE) by the electropolymerization of pyrrole (Py) in the presence of insulin (the protein template) using cyclic voltammetry. After electropolymerization, insulin was removed from the formed polypyrrole (Ppy) matrix to create imprinting cavities for the subsequent analysis of the insulin analyte in test samples. The surface characterization, before and after each electrosynthesis step of the MIP sensors, was performed using atomic force microscopy, scanning electron microscopy, and energy-dispersive X-ray spectroscopy. The performance of the developed MIP-SPCE sensor was evaluated using a single drop of solution containing K3Fe(CN)6 and the square-wave voltammetry technique. The MIP-SPCE showed a linear concentration range of 20.0-70.0 pM (R2 = 0.9991), a limit of detection of 1.9 pM, and a limit of quantification of 6.2 pM. The rapid response time to the protein target and the portability of the developed sensor, which is considered a disposable MIP-based system, make this MIP-SPCE sensor a promising candidate for point-of-care applications. In addition, the MIP-SPCE sensor was successfully used to detect insulin in a pharmaceutical sample. The sensor was deemed to be accurate (the average recovery was 108.46%) and precise (the relative standard deviation was 7.23%).

Novel Methacrylate-Based Multilayer Nanofilms with Incorporated FePt-Based Nanoparticles and the Anticancer Drug 5-Fluorouracil for Skin Cancer Treatment.

Despite medical advances, skin-associated disorders continue to pose a unique challenge to physicians worldwide. Skin cancer is one of the most common forms of cancer, with more than one million new cases reported each year. Currently, surgical excision is its primary treatment; however, this can be impractical or even contradictory in certain situations. An interesting potential alternative could lie in topical treatment solutions. The goal of our study was to develop novel multilayer nanofilms consisting of a combination of polyhydroxyethyl methacrylate (PHEMA), polyhydroxypropyl methacrylate (PHPMA), sodium deoxycholate (NaDOC) with incorporated superparamagnetic iron-platinum nanoparticles (FePt NPs), and the potent anticancer drug (5-fluorouracil), for theranostic skin cancer treatment. All multilayer systems were prepared by spin-coating and characterised by atomic force microscopy, infrared spectroscopy, and contact angle measurement. The magnetic properties of the incorporated FePt NPs were evaluated using magnetisation measurement, while their size was determined using transmission electron microscopy (TEM). Drug release performance was tested in vitro, and formulation safety was evaluated on human-skin-derived fibroblasts. Finally, the efficacy for skin cancer treatment was tested on our own basal-cell carcinoma cell line.

Artificial Biomimetic Electrochemical Assemblies.

Rapid, selective, and cost-effective detection and determination of clinically relevant biomolecule analytes for a better understanding of biological and physiological functions are becoming increasingly prominent. In this regard, biosensors represent a powerful tool to meet these requirements. Recent decades have seen biosensors gaining popularity due to their ability to design sensor platforms that are selective to determine target analytes. Naturally generated receptor units have a high affinity for their targets, which provides the selectivity of a device. However, such receptors are subject to instability under harsh environmental conditions and have consequently low durability. By applying principles of supramolecular chemistry, molecularly imprinted polymers (MIPs) can successfully replace natural receptors to circumvent these shortcomings. This review summarizes the recent achievements and analytical applications of electrosynthesized MIPs, in particular, for the detection of protein-based biomarkers. The scope of this review also includes the background behind electrochemical readouts and the origin of the gate effect in MIP-based biosensors.

Neurosurgical Approaches to Brain Tissue Harvesting for the Establishment of Cell Cultures in Neural Experimental Cell Models.

In recent decades, cell biology has made rapid progress. Cell isolation and cultivation techniques, supported by modern laboratory procedures and experimental capabilities, provide a wide range of opportunities for in vitro research to study physiological and pathophysiological processes in health and disease. They can also be used very efficiently for the analysis of biomaterials. Before a new biomaterial is ready for implantation into tissues and widespread use in clinical practice, it must be extensively tested. Experimental cell models, which are a suitable testing ground and the first line of empirical exploration of new biomaterials, must contain suitable cells that form the basis of biomaterial testing. To isolate a stable and suitable cell culture, many steps are required. The first and one of the most important steps is the collection of donor tissue, usually during a surgical procedure. Thus, the collection is the foundation for the success of cell isolation. This article explains the sources and neurosurgical procedures for obtaining brain tissue samples for cell isolation techniques, which are essential for biomaterial testing procedures.

Advanced Bio-Based Polymers for Astrocyte Cell Models.

The development of in vitro neural tissue analogs is of great interest for many biomedical engineering applications, including the tissue engineering of neural interfaces, treatment of neurodegenerative diseases, and in vitro evaluation of cell-material interactions. Since astrocytes play a crucial role in the regenerative processes of the central nervous system, the development of biomaterials that interact favorably with astrocytes is of great research interest. The sources of human astrocytes, suitable natural biomaterials, guidance scaffolds, and ligand patterned surfaces are discussed in the article. New findings in this field are essential for the future treatment of spinal cord and brain injuries.

Hybrid 3D Printing of Advanced Hydrogel-Based Wound Dressings with Tailorable Properties.

Despite the extensive utilization of polysaccharide hydrogels in regenerative medicine, current fabrication methods fail to produce mechanically stable scaffolds using only hydrogels. The recently developed hybrid extrusion-based bioprinting process promises to resolve these current issues by facilitating the simultaneous printing of stiff thermoplastic polymers and softer hydrogels at different temperatures. Using layer-by-layer deposition, mechanically advantageous scaffolds can be produced by integrating the softer hydrogel matrix into a stiffer synthetic framework. This work demonstrates the fabrication of hybrid hydrogel-thermoplastic polymer scaffolds with tunable structural and chemical properties for applications in tissue engineering and regenerative medicine. Through an alternating deposition of polycaprolactone and alginate/carboxymethylcellulose gel strands, scaffolds with the desired architecture (e.g., filament thickness, pore size, macro-/microporosity), and rheological characteristics (e.g., swelling capacity, degradation rate, and wettability) were prepared. The hybrid fabrication approach allows the fine-tuning of wettability (approx. 50-75°), swelling (approx. 0-20× increased mass), degradability (approx. 2-30+ days), and mechanical strength (approx. 0.2-11 MPa) in the range between pure hydrogels and pure thermoplastic polymers, while providing a gradient of surface properties and good biocompatibility. The controlled degradability and permeability of the hydrogel component may also enable controlled drug delivery. Our work shows that the novel hybrid hydrogel-thermoplastic scaffolds with adjustable characteristics have immense potential for tissue engineering and can serve as templates for developing novel wound dressings.

Clindamycin-Based 3D-Printed and Electrospun Coatings for Treatment of Implant-Related Infections.

This study presents the development and characterisation of two novel bioactive coatings deposited on TiAlV and AISI 316LVM substrates. The coatings were prepared using 3D printing and electrospinning. The 3D-printed coating consisted of the cellulose nanofibril suspension, alginate, and carboxymethylcellulose (CMC), while CMC and polyethylene oxide were used to prepare the electrospun coating. Both coatings were loaded with the antibiotic clindamycin (CLIN), which is a bacteriostatic lincosamide known for its activity against streptococci, staphylococci, pneumococci, Bacteroides species, and other anaerobes. Initial characterisation of the coatings was performed by attenuated total reflectance Fourier transform infrared spectroscopy, field emission scanning electron microscopy, and atomic force microscopy. Furthermore, the contact angle measurements, swelling rate, and biodegradability of the coatings were investigated. The released concentration of CLIN in PBS (pH = 7.4 at 25 °C) was determined by UV-VIS spectrophotometry. The coatings' biocompatibility was determined using an MTT (3(4,5 dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide) assay using an osteoblast cell culture (hFOB 1.19, ATCC CRL 11372).

Polysaccharide-Based Bioink Formulation for 3D Bioprinting of an In Vitro Model of the Human Dermis.

Limitations in wound management have prompted scientists to introduce bioprinting techniques for creating constructs that can address clinical problems. The bioprinting approach is renowned for its ability to spatially control the three-dimensional (3D) placement of cells, molecules, and biomaterials. These features provide new possibilities to enhance homology to native skin and improve functional outcomes. However, for the clinical value, the development of hydrogel bioink with refined printability and bioactive properties is needed. In this study, we combined the outstanding viscoelastic behavior of nanofibrillated cellulose (NFC) with the fast cross-linking ability of alginate (ALG), carboxymethyl cellulose (CMC), and encapsulated human-derived skin fibroblasts (hSF) to create a bioink for the 3D bioprinting of a dermis layer. The shear thinning behavior of hSF-laden bioink enables construction of 3D scaffolds with high cell density and homogeneous cell distribution. The obtained results demonstrated that hSF-laden bioink supports cellular activity of hSF (up to 29 days) while offering proper printability in a biologically relevant 3D environment, making it a promising tool for skin tissue engineering and drug testing applications.

Polysaccharide Thin Solid Films for Analgesic Drug Delivery and Growth of Human Skin Cells.

Chronic wounds not only lower the quality of patient's life significantly, but also present a huge financial burden for the healthcare systems around the world. Treatment of larger wounds often requires the use of more complex materials, which can ensure a successful renewal or replacement of damaged or destroyed tissues. Despite a range of advanced wound dressings that can facilitate wound healing, there are still no clinically used dressings for effective local pain management. Herein, alginate (ALG) and carboxymethyl cellulose (CMC), two of the most commonly used materials in the field of chronic wound care, and combination of ALG-CMC were used to create a model wound dressing system in the form of multi-layered thin solid films using the spin-assisted layer-by-layer (LBL) coating technique. The latter multi-layer system was used to incorporate and study the release kinetics of analgesic drugs such as diclofenac and lidocaine at physiological conditions. The wettability, morphology, physicochemical and surface properties of the coated films were evaluated using different surface sensitive analytical tools. The influence of in situ incorporated drug molecules on the surface properties (e.g., roughness) and on the proliferation of human skin cells (keratinocytes and skin fibroblasts) was further evaluated. The results obtained from this preliminary study should be considered as the basis for the development "real" wound dressing materials and for 3D bio-printing applications.

Systematic Evaluation of a Diclofenac-Loaded Carboxymethyl Cellulose-Based Wound Dressing and Its Release Performance with Changing pH and Temperature.

Development of drug-loaded wound dressings is often performed without systematic consideration of the changing wound environment that can influence such materials' performance. Among the crucial changes are the wound pH and temperature, which have an immense effect on the drug release. Detailed release studies based on the consideration of these changing properties provide an important aspect of the in vitro performance testing of novel wound dressing materials. A sodium carboxymethyl cellulose-based wound dressing, with the incorporated non-steroidal anti-inflammatory drug diclofenac, was developed and characterised in regard to its physico-chemical, structural and morphological properties. Further, the influence of pH and temperature were studied on the drug release. Finally, the biocompatibility of the wound dressing towards human skin cells was tested. Incorporation of diclofenac did not alter important properties (water retention value, air permeability) of the host material. Changes in the pH and temperature were shown to influence the release performance and have to be accounted for in the evaluation of such dressings. Furthermore, the knowledge about the potential changes of these parameters in the wound bed could be used potentially to predict, and potentially even to control the drug release from the developed wound dressing. The prepared wound dressing was also proven biocompatible towards human skin cells, making it interesting for potential future use in the clinics.

Reusability of SPE and Sb-modified SPE Sensors for Trace Pb(II) Determination.

In this work, unmodified screen-printed electrode (bare SPE) and Sb-film modified SPE (SbFSPE) sensors were employed for the analysis of trace amounts of Pb(II) in non-deaerated water solutions. The modified electrode was performed in situ in 0.5 mg/L Sb(III) and 0.01 M HCl. The methodology was validated for an accumulation potential of ⁻1.1 V vs. Ag/AgCl and an accumulation time of 60 s. A comparative analysis of bare SPE and SbFSPE showed that the detection and quantification limits decrease for the bare SPE. The method with the bare SPE showed a linear response in the 69.8⁻368.4 µg/L concentration range, whereas linearity for the SbFSPE was in the 24.0⁻319.1 µg/L concentration range. This work also reports the reason why the multiple standard addition method instead of a linear calibration curve for Pb(II) analysis should be employed. Furthermore, the analytical method employing SbFSPE was found to be more accurate and precise compared to the use of bare SPE when sensors were employed for the first time, however this performance changed significantly when these sensors were reused in the same manner. Furthermore, electrochemical impedance spectroscopy was used for the first time to analyse the electrochemical response of sensors after being used for multiple successive analyses. Surface characterisation before and after multiple successive uses of bare SPE and SbFSPE sensors, with atomic force microscopy and field emission scanning electron microscopy, showed sensor degradation. The interference effect of Cd(II), Zn(II), As(III), Fe(II), Na(I), K(I), Ca(II), Mg(II), NO3⁻, Bi(III), Cu(II), Sn(II), and Hg(II) on the Pb(II) stripping signal was also studied. Finally, the application of SbFSPE was tested on a real water sample (from a local river), which showed high precision (RSD = 8.1%, n = 5) and accurate results.

Advanced therapies of skin injuries.

The loss of tissue is still one of the most challenging problems in healthcare. Efficient laboratory expansion of skin tissue to reproduce the skins barrier function can make the difference between life and death for patients with extensive full-thickness burns, chronic wounds, or genetic disorders such as bullous conditions. This engineering has been initiated based on the acute need in the 1980s and today, tissue-engineered skin is the reality. The human skin equivalents are available not only as models for permeation and toxicity screening, but are frequently applied in vivo as clinical skin substitutes. This review aims to introduce the most important recent development in the extensive field of tissue engineering and to describe already approved, commercially available skin substitutes in clinical use.

A review of herbal medicines in wound healing.

Herbs have been integral to both traditional and non-traditional forms of medicine dating back at least 5000 years. The enduring popularity of herbal medicines may be explained by the perception that herbs cause minimal unwanted side effects. More recently, scientists increasingly rely on modern scientific methods and evidence-based medicine to prove efficacy of herbal medicines and focus on better understanding of mechanisms of their action. However, information concerning quantitative human health benefits of herbal medicines is still rare or dispersed, limiting their proper valuation. Preparations from traditional medicinal plants are often used for wound healing purposes covering a broad area of different skin-related diseases. Herbal medicines in wound management involve disinfection, debridement, and provision of a suitable environment for aiding the natural course of healing. Here we report on 22 plants used as wound healing agents in traditional medicine around the world. The aim of this review is therefore to review herbal medicines, which pose great potential for effective treatment of minor wounds.

Novel cellulose based materials for safe and efficient wound treatment.

The present study aims at achieving effects of improved hydrophilicity and microorganism inhibition, which are rarely simultaneously present in wound dressings. Viscose fibers in their non-woven form were modified using two different pathways. Effects of a two-step procedure, i.e. alkaline or oxygen plasma treatment followed by the attachment of silver chloride nanoparticles were compared to a one-step procedure, i.e. ammonium plasma treatment, which results in both desired material characteristics simultaneously. The surface properties of untreated and differently modified cellulose samples were analyzed by X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM), in vitro silver release, and hydrophilicity measurements. The treatment effect on antimicrobial activity was determined by the AATCC 100-1999 standard test. In light of the introduced wound dressing preparation procedures and the desired wound dressing characteristics, the effectiveness of the used procedures was evaluated. Antimicrobial activity was proven against all Gram negative bacteria, while the Gram positive bacteria survive the as-prepared samples. Hydrophilicity was proven to be excellent using both preparation procedures. The mentioned results prove the potential of the used procedures and encourage future developments toward the clinical proof of concept.