Novel magnetic iron oxide-dextran sulphate nanocomposites as potential anticoagulants: Investigating interactions with blood components and assessing cytotoxicity.

Examining the critical role of anticoagulants in medical practice, particularly their central function in preventing abnormal blood clotting, is of the utmost importance. However, the study of interactions between blood proteins and alternative anticoagulant nano-surfaces is still understood poorly. In this study, novel approach involving direct functionalisation of magnetic iron oxide nanoparticles (MNPs) as carriers with sulphated dextran (s-dext) is presented, with the aim of evaluating the potential of magnetically-responsive MNPs@s-dext as anticoagulants. The physicochemical characterisation of the synthesised MNPs@s-dext includes crystal structure analysis, morphology study, surface and electrokinetic properties, thermogravimetric analysis and magnetic properties` evaluation, which confirms the successful preparation of the nanocomposite with sulfonate groups. The anticoagulant potential of MNPs@s-dext was investigated using a standardised activated partial thromboplastin time (APTT) test and a modified APTT test with a quartz crystal microbalance with dissipation (QCM-D) which confirmed the anticoagulant effect. Time-resolved solid-liquid interactions between the MNPs@s-dext and model blood proteins bovine serum albumin and fibrinogen were also investigated, to gain insight into their hemocompatibility, and revealed protein-repellence of MNPs@s-dext against blood proteins. The study also addressed comprehensive cytotoxicity studies of prepared nanocomposites, and provided valuable insights into potential applicability of MNPs@s-dext as a promising magnetic anticoagulant in biomedical contexts.

Clinical Properties and Non-Clinical Testing of Mineralocorticoid Receptor Antagonists in In Vitro Cell Models.

Mineralocorticoid receptor antagonists (MRAs) are one of the renin-angiotensin-aldosterone system inhibitors widely used in clinical practice. While spironolactone and eplerenone have a long-standing profile in clinical medicine, finerenone is a novel agent within the MRA class. It has a higher specificity for mineralocorticoid receptors, eliciting less pronounced adverse effects. Although approved for clinical use in patients with chronic kidney disease and heart failure, intensive non-clinical research aims to further elucidate its mechanism of action, including dose-related selectivity. Within the field, animal models remain the gold standard for non-clinical testing of drug pharmacological and toxicological properties. Their role, however, has been challenged by recent advances in in vitro models, mainly through sophisticated analytical tools and developments in data analysis. Currently, in vitro models are gaining momentum as possible platforms for advanced pharmacological and pathophysiological studies. This article focuses on past, current, and possibly future in vitro cell models research with clinically relevant MRAs.

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.

Editorial on the Special Issue Entitled "Recent Advances in Aerogels".

Aerogels are unique solid materials that consist mainly of air and have an extremely low density, large open pores, and a large internal surface area [...].

Evaluation of ethanol-induced chitosan aerogels with human osteoblast cells.

The following article provides an insight into the production of chitosan aerogels as potential materials for tissue engineering. Chitosan aerogels were prepared following two different protocols: formation in ethanol and formation in sodium hydroxide in an ethanol solution. The main objective was to apply a new route to obtain chitosan aerogels with no external cross-linkers and compare the mentioned preparation approaches. Forming chitosan aerogels in ethanol implies a simple, environmentally friendly, and efficient method. The prepared materials showed specific surface areas of up to 450 m2/g, highly porous networks and great mechanical properties. In vitro degradation studies revealed high stability for up to 10 weeks. The differences between the samples were significant. While the chitosan aerogels prepared in ethanol showed superior textural, morphological and mechanical properties, the chitosan aerogels prepared in the sodium hydroxide solution proved that a considerable influence on end properties could be made simply by adjusting the ageing medium. In vitro cell analysis with primary human osteoblasts showed good biocompatibility and pointed towards the potential use of these aerogels for orthopedic applications. This testing showed further that adjustments in structural properties by sodium hydroxide also come with a cost regarding their suitability to host bone cells.

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.

Renal Proximal Tubular Epithelial Cells: From Harvesting to Use in Studies.

The kidneys are the body's main excretion organ with several additional functions, and the nephron represents their central structural unit. It is comprised of endothelial, mesangial, glomerular, and tubular epithelial cells, as well as podocytes. Treatment of acute kidney injury or chronic kidney disease (CKD) is complex due to broad etiopathogenic mechanisms and limited regeneration potential as kidney cells finish their differentiation after 34 weeks of gestation. Despite the ever-increasing prevalence of CKD, very limited treatment modalities are available. The medical community should therefore strive to improve existing treatments and develop new ones. Furthermore, polypharmacy is present in most CKD patients, while current pharmacologic study designs lack effectiveness in predicting potential drug-drug interactions and the resulting clinically relevant complications. An opportunity for addressing these issues lies in developing in vitro cell models based on patient-derived renal cells. Currently, several protocols have been described for isolating desired kidney cells, of which the most isolated are the proximal tubular epithelial cells. These play a significant role in water homeostasis, acid-base control, reabsorption of compounds, and secretion of xenobiotics and endogenous metabolites. When developing a protocol for the isolation and culture of such cells, one must focus on several steps. These include harvesting cells from biopsy specimens or after nephrectomies, using different digestion enzymes and culture mediums to facilitate the selective growth of only the desired cells. The literature reports several existing models, from simple 2D in vitro cultures to more complex ones created with bioengineering methods, such as kidney-on-a-chip models. While their creation and use depend on the target research, one should consider factors such as equipment, cost, and, even more importantly, source tissue quality and availability.

Efficient and Green Isolation of Keratin from Poultry Feathers by Subcritical Water.

The isolation of keratin from poultry feathers using subcritical water was studied in a batch reactor at temperatures (120-250 °C) and reaction times (5-75 min). The hydrolyzed product was characterized by FTIR and elemental analysis, while the molecular weight of the isolated product was determined by SDS-PAGE electrophoresis. To determine whether disulfide bond cleavage was followed by depolymerization of protein molecules to amino acids, the concentration of 27 amino acids in the hydrolysate was analyzed by GC/MS. The optimal operating parameters for obtaining a high molecular weight protein hydrolysate from poultry feathers were 180 °C and 60 min. The molecular weight of the protein hydrolysate obtained under optimal conditions ranged from 4.5 to 12 kDa, and the content of amino acids in the dried product was low (2.53% w/w). Elemental and FTIR analyses of unprocessed feathers and dried hydrolysate obtained under optimal conditions showed no significant differences in protein content and structure. Obtained hydrolysate is a colloidal solution with a tendency for particle agglomeration. Finally, a positive influence on skin fibroblast viability was observed for the hydrolysate obtained under optimal processing conditions for concentrations below 6.25 mg/mL, which makes the product interesting for various biomedical applications.

Review of Potential Drug-Eluting Contact Lens Technologies.

The field of ophthalmology is expanding exponentially, both in terms of diagnostic and therapeutic capabilities, as well as the worldwide increasing incidence of eye-related diseases. Due to an ageing population and climate change, the number of ophthalmic patients will continue to increase, overwhelming healthcare systems and likely leading to under-treatment of chronic eye diseases. Since drops are the mainstay of therapy, clinicians have long emphasised the unmet need for ocular drug delivery. Alternative methods, i.e., with better compliance, stability and longevity of drug delivery, would be preferred. Several approaches and materials are being studied and used to overcome these drawbacks. We believe that drug-loaded contact lenses are among the most promising and are a real step toward dropless ocular therapy, potentially leading to a transformation in clinical ophthalmic practice. In this review, we outline the current role of contact lenses in ocular drug delivery, focusing on materials, drug binding and preparation, concluding with a look at future developments.

Single-Strain Probiotic Lactobacilli for the Treatment of Atopic Dermatitis in Children: A Systematic Review and Meta-Analysis.

Probiotics are known for their positive effects on the gut microbiota. There is growing evidence that the infant gut and skin colonization have a role in the development of the immune system, which may be helpful in the prevention and treatment of atopic dermatitis. This systematic review focused on evaluating the effect of single-strain probiotic lactobacilli consumption on treating children's atopic dermatitis. Seventeen randomized placebo-controlled trials with the primary outcome of the Scoring Atopic Dermatitis (SCORAD) index were included in the systematic review. Clinical trials using single-strain lactobacilli were included. The search was conducted until October 2022 using PubMed, ScienceDirect, Web of Science, Cochrane library and manual searches. The Joanna Briggs Institute appraisal tool was used to assess the quality of the included studies. Meta-analyses and sub meta-analyses were performed using Cochrane Collaboration methodology. Due to different methods of reporting the SCORAD index, only 14 clinical trials with 1124 children were included in the meta-analysis (574 in the single-strain probiotic lactobacilli group and 550 in the placebo group) and showed that single-strain probiotic lactobacilli statistically significantly reduced the SCORAD index compared to the placebo in children with atopic dermatitis (mean difference [MD]: -4.50; 95% confidence interval [CI]: -7.50 to -1.49; Z = 2.93; p = 0.003; heterogeneity I2 = 90%). The subgroup meta-analysis showed that strains of Limosilactobacillus fermentum were significantly more effective than strains of Lactiplantibacillus plantarum, Lacticaseibacillus paracasei or Lacticaseibacillus rhamnosus. A longer treatment time and younger treatment age statistically significantly reduced symptoms of atopic dermatitis. The result of this systematic review and meta-analysis shows that certain single-strain probiotic lactobacilli are more successful than others in reducing atopic dermatitis severity in children. Therefore, careful consideration to strain selection, treatment time and the age of the treated patients are important factors in enhancing the effectiveness of reducing atopic dermatitis in children when choosing probiotic single-strain lactobacilli.

Screen-printing of chitosan and cationised cellulose nanofibril coatings for integration into functional face masks with potential antiviral activity.

Masks proved to be necessary protective measure during the COVID-19 pandemic, but they provided a physical barrier rather than inactivating viruses, increasing the risk of cross-infection. In this study, high-molecular weight chitosan and cationised cellulose nanofibrils were screen-printed individually or as a mixture onto the inner surface of the first polypropylene (PP) layer. First, biopolymers were evaluated by various physicochemical methods for their suitability for screen-printing and antiviral activity. Second, the effect of the coatings was evaluated by analysing the morphology, surface chemistry, charge of the modified PP layer, air permeability, water-vapour retention, add-on, contact angle, antiviral activity against the model virus phi6 and cytotoxicity. Finally, the functional PP layers were integrated into face masks, and resulting masks were tested for wettability, air permeability, and viral filtration efficiency (VFE). Air permeability was reduced for modified PP layers (43 % reduction for kat-CNF) and face masks (52 % reduction of kat-CNF layer). The antiviral potential of the modified PP layers against phi6 showed inhibition of 0.08 to 0.97 log (pH 7.5) and cytotoxicity assay showed cell viability above 70 %. VFE of the masks remained the same (~99.9 %), even after applying the biopolymers, confirming that these masks provided high level of protection against viruses.

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%).

Practical Use of Quartz Crystal Microbalance Monitoring in Cartilage Tissue Engineering.

Quartz crystal microbalance (QCM) is a real-time, nanogram-accurate technique for analyzing various processes on biomaterial surfaces. QCM has proven to be an excellent tool in tissue engineering as it can monitor key parameters in developing cellular scaffolds. This review focuses on the use of QCM in the tissue engineering of cartilage. It begins with a brief discussion of biomaterials and the current state of the art in scaffold development for cartilage tissue engineering, followed by a summary of the potential uses of QCM in cartilage tissue engineering. This includes monitoring interactions with extracellular matrix components, adsorption of proteins onto biomaterials, and biomaterial-cell interactions. In the last part of the review, the material selection problem in tissue engineering is highlighted, emphasizing the importance of surface nanotopography, the role of nanofilms, and utilization of QCM as a "screening" tool to improve the material selection process. A step-by-step process for scaffold design is proposed, as well as the fabrication of thin nanofilms in a layer-by-layer manner using QCM. Finally, future trends of QCM application as a "screening" method for 3D printing of cellular scaffolds are envisioned.

A Promising Method for the Determination of Cell Viability: The Membrane Potential Cell Viability Assay.

Determining the viability of cells is fraught with many uncertainties. It is often difficult to determine whether a cell is still alive, approaching the point of no return, or dead. Today, there are many methods for determining cell viability. Most rely on an indirect determination of cell death (metabolism, molecular transport, and leakage, to name a few). In contrast, we have developed a promising novel method for a "direct" determination of cell viability. The potential method assesses cell membrane integrity (which is essential for all viable cells) by measuring the electrical potential of the cell membrane. To test the assay, we chose two different cell types, blood macrophages (TLT) and breast cancer epithelial cells (MCF 7). We exposed them to seven different toxic scenarios (arsenic (V), UV light, hydrogen peroxide, nutrient starvation, Tetrabromobisphenol A, fatty acids, and 5-fluorouracil) to induce different cell death pathways. Under controlled test conditions, the assay showed good accuracy when comparing the toxicity assessment with well-established methods. Moreover, the method showed compatibility with live cell imaging. Although we know that further studies are needed to confirm the performance of the assay in other situations, the results obtained are promising for their wider application in the future.

Organic acid cross-linked 3D printed cellulose nanocomposite bioscaffolds with controlled porosity, mechanical strength, and biocompatibility.

Herein, we fabricated chemically cross-linked polysaccharide-based three-dimensional (3D) porous scaffolds using an ink composed of nanofibrillated cellulose, carboxymethyl cellulose, and citric acid (CA), featuring strong shear thinning behavior and adequate printability. Scaffolds were produced by combining direct-ink-writing 3D printing, freeze-drying, and dehydrothermal heat-assisted cross-linking techniques. The last step induces a reaction of CA. Degree of cross-linking was controlled by varying the CA concentration (2.5-10.0 wt.%) to tune the structure, swelling, degradation, and surface properties (pores: 100-450 µm, porosity: 86%) of the scaffolds in the dry and hydrated states. Compressive strength, elastic modulus, and shape recovery of the cross-linked scaffolds increased significantly with increasing cross-linker concentration. Cross-linked scaffolds promoted clustered cell adhesion and showed no cytotoxic effects as determined by the viability assay and live/dead staining with human osteoblast cells. The proposed method can be extended to all polysaccharide-based materials to develop cell-friendly scaffolds suitable for tissue engineering applications.

The Endplate Role in Degenerative Disc Disease Research: The Isolation of Human Chondrocytes from Vertebral Endplate-An Optimised Protocol.

BACKGROUND: Degenerative disc disease is a progressive and chronic disorder with many open questions regarding its pathomorphological mechanisms. In related studies, in vitro organ culture systems are becoming increasingly essential as a replacement option for laboratory animals. Live disc cells are highly appealing to study the possible mechanisms of intervertebral disc (IVD) degeneration. To study the degenerative processes of the endplate chondrocytes in vitro, we established a relatively quick and easy protocol for isolating human chondrocytes from the vertebral endplates. METHODS: The fragments of human lumbar endplates following lumbar fusion were collected, cut, ground and partially digested with collagenase I in Advanced DMEM/F12 with 5% foetal bovine serum. The sediment was harvested, and cells were seeded in suspension, supplemented with special media containing high nutrient levels. Morphology was determined with phalloidin staining and the characterisation for collagen I, collagen II and aggrecan with immunostaining. RESULTS: The isolated cells retained viability in appropriate laboratory conditions and proliferated quickly. The confluent culture was obtained after 14 days. Six to 8 h after seeding, attachments were observed, and proliferation of the isolated cells followed after 12 h. The cartilaginous endplate chondrocytes were stable with a viability of up to 95%. Pheno- and geno-typic analysis showed chondrocyte-specific expression, which decreased with passages. CONCLUSIONS: The reported cell isolation process is simple, economical and quick, allowing establishment of a viable long-term cell culture. The availability of a vertebral endplate cell model will permit the study of cell properties, biochemical aspects, the potential of therapeutic candidates for the treatment of disc degeneration, and toxicology studies in a well-controlled environment.

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.