We review recently developed advanced nanotechnologies for overcoming antimicrobial resistance and tackling of biofilm infections.

We discuss the recent progress in the development of scaffold-free techniques for the fabrication of cell spheroids and their biomedical applications.

Microbial biofilms are a major concern in wound care, implant devices, and organ infections. Biofilms allow higher tolerance to antimicrobial drugs, can impair wound healing, and potentially lead to sepsis. There has been a recent focus on developing novel nanocarrier-based delivery vehicles to enhance the biofilm penetration of traditional antibacterial drugs. However, a feasible in vitro human skin model to mimic the biofilm formation and its treatment for clearance have not yet been reported. This study describes the benefits of using an innovative bacterial biofilm-infected keratinocyte clusteroid model for the first time. It paves a new way for testing innovative nanomedicine delivery systems in a rapid and reproducible way on a realistic human cell-based platform, free of any animal testing. Herein, we have developed a novel composite 3D biofilm/human keratinocyte clusteroid coculture platform, which was used to measure biofilm clearance efficiency of nanoparticle (NP)-based therapeutics. We tested this model by treating the biofilm-infected 3D coculture layers by a ciprofloxacin-loaded Carbopol nanogel particles, surface-functionalized by the cationic protease Alcalase. We measured the antibacterial efficiency of the NP treatment on clearing Staphylococcus aureus and Pseudomonas aeruginosa biofilms on the 3D keratinocyte clusteroid/biofilm coculture model. Our experiments showed that these bacteria can infect the 3D layer of keratinocyte clusteroids and produce a stable biofilm. The biofilms were efficiently cleared by treatment with a formulation of 0.0032 wt % ciprofloxacin-loaded in 0.2 wt % Carbopol NPs surface-functionalized with 0.2 wt % Alcalase. Taken together, these promising results demonstrate that our coculture model can be exploited as a novel platform for testing the biofilm-eliminating efficiency of various NP formulations emulating skin and wound infections and could have wider applicability to replace animal models in similar experiments. This 3D cell culture-based platform could help in developing and testing of more effective antibacterial agents for clinical applications of antiplaque dental treatments, implants, infection control, and wound dressings.

We report a novel 3D urothelial clusteroid/fungal biofilm co-culture model for the advanced testing of antifungal nanotechnologies.

A massive amount of chalaza with nearly 400 metric tons is produced annually as waste in the liquid-egg industry. The present study aimed to look for ways to utilize chalaza as a natural emulsifier for high internal phase emulsions (HIPEs) at the optimal production conditions to expand the utilization of such abundant material. To the author’s knowledge, for the first time, we report the usage of hen egg chalaza particles as particulate emulsifiers for Pickering (HIPEs) development. The chalaza particles with partial wettability were fabricated at different pH or ionic strengths by freeze-drying. The surface electricity of the chalaza particles was neutralized when the pH was adjusted to 4, where the chalaza contained a particle size around 1500 nm and held the best capability to stabilize the emulsions. Similarly, the chalaza reaches proper electrical charging (−6 mv) and size (700 nm) after the ionic strength was modified to 0.6 M. Following the characterization of chalaza particles, we successfully generated stable Pickering HIPEs with up to 86% internal phase at proper particle concentrations (0.5–2%). The emulsion contained significant stability against coalescence and flocculation during long term storage due to the electrical hindrance raised by the chalaza particles which absorbed on the oil–water interfaces. Different rheological models were tested on the formed HIPEs, indicating the outstanding stability of such emulsions. Concomitantly, a percolating 3D-network was formed in the Pickering HIPES stabilized by chalaza which provided the emulsions with viscoelastic and self-standing features. Moreover, the current study provides an attractive strategy to convert liquid oils to viscoelastic soft solids without artificial trans fats.

BACKGROUND: The egg yolk is complex, which makes it difficult to understand why mayonnaise can be stabilized into a high internal-phase emulsion. This study aimed to assess the possibility of developing oil-in-water emulsions through unmodified natural egg-yolk granules (EYGs) at various pH levels, to further understand the precise mechanism of mayonnaise. RESULTS: Egg-yolk granules were obtained from hen egg yolk by centrifugation. The sizes of the EYGs were characterized using dynamic light scattering (DLS). Zeta potential of EYGs was detected by DLS and its microstructure was observed by microscope and scanning electron microscope (SEM). Oil / water emulsions were made with EYGs and the size distribution and creaming index of those emulsions were measured at different storage times (1 h and 14 days). The interfacial morphology of EYGs was observed using the emulsion polymerization method. Our results suggested that the prepared EYGs were mainly in an aggregated state but individual EYGs displayed spherical shapes, with a size of 1.0 ± 0.2 m. The emulsion stabilized by EYGs displayed better stability against creaming at acidic pH (<4.0). At the same time, the interfacial morphology and microscopic observation of the emulsions strongly demonstrated that the emulsions were of the Pickering type. CONCLUSION: The above results are of great importance for an understanding of the mechanism by which mayonnaise is stabilized by egg, together with the applications of egg in food formulations.

We report a novel inexpensive and up-scalable fabrication technique for viable hepatocyte clusteroids and demonstrate that they grow faster than individual cells in tissue engineering applications.

Tumor cell spheroids and 3D cell culture have generated a lot of interest in the past decade due to their relative ease of production and biomedical research applications. To date, the frontier in tumor 3D models has been pushed to the level of personalized cancer treatment and customized tissue engineering applications. However, without vascularization, the central parts of these artificial constructs cannot survive without an adequate oxygen and nutrient supply. The formation of a necrotic core into in vitro 3D cell models still serves as the major obstacle in their wider practical application. Here, we propose a rapid formation protocol based on using a water-in-water (w/w) Pickering emulsion template to generate phenotypically endothelial/hepatic (ECV304/Hep-G2) coculture cell clusteroids with angiogenic capability. The w/w Pickering emulsion template was based on a dextran/poly­(ethylene oxide) aqueous two-phase system stabilized by whey protein particles. The initial cell proportion in the coculture clusteroids can easily be manipulated for optimal performance. The cocultured pattern of the endothelial/hepatic cells could significantly promote the production of angiogenesis-related proteins. Our study confirmed that cocultured clusteroids can stimulate cell sprouting without the addition of vascular endothelial growth factor (VEGF) or other angiogenesis inducers at a 1:2 ratio of Hep-G2/ECV304. Angiogenesis gene production in the coculture clusteroids was enhanced with VEGF, urea, and insulin-like growth factor-binding protein along with angiogenesis-related marker CD34 levels, also indicating angiogenesis progress. Our aqueous two-phase Pickering emulsion templates provided a convenient approach to vascularize a target cell type in 3D cell coculture without additional stimulating factors, which could potentially apply to either cell lines or biopsy tissues, expanding the clusteroids downstream applications.