Mimicking natural electrical environment with cellulose acetate scaffolds enhances collagen formation of osteoblasts.

The medical field is continuously seeking new solutions and materials, where cellulose materials due to their high biocompatibility have great potential. Here we investigate the applicability of cellulose acetate (CA) electrospun fibers for bone tissue regeneration. For the first time we show the piezoelectric properties of electrospun CA fibers via high voltage switching spectroscopy piezoresponse force microscopy (HVSS-PFM) tests, which are followed by surface potential studies using Kelvin probe force microscopy (KPFM) and zeta potential measurements. Piezoelectric coefficient for CA fibers of 6.68 ± 1.70 pmV-1 along with high surface (718 mV) and zeta (-12.2 mV) potentials allowed us to mimic natural electrical environment favoring bone cell attachment and growth. Importantly, the synergy between increased surface potential and highly developed structure of the fibrous scaffold led to the formation of a vast 3D network of collagen produced by osteoblasts only after 7 days of in vitro culture. We clearly show the advantages of CA scaffolds as a bone replacement material, when long-lasting structural support is needed.

Enhanced Cells Anchoring to Electrospun Hybrid Scaffolds With PHBV and HA Particles for Bone Tissue Regeneration.

Hybrid materials combining organic and inorganic compounds used as scaffolds are highly beneficial in bone regeneration. In this study, we successfully produced by blend electrospinning poly(3-hydroxybutyric acid-co-3-hydrovaleric acid) (PHBV) scaffolds enriched with hydroxyapatite (HA) particles to biomimic bone tissue for improved and faster regeneration processes. The morphology, fiber diameters, and composition of the scaffolds were investigated by scanning electron microscopy (SEM) techniques followed by focused ion beam (FIB) sectioning to verify HA particles integration with PHBV fibers. In vitro cell culture was performed for 7 days and followed with the cell proliferation test (CellTiter-Blue® Assay). Additionally, cell integration with the scaffold was visualized by confocal and SEM imaging. We developed a simple way of obtaining hybrid scaffolds by electrospinning PHBV solution with HA particles without any post-processing. The PHBV + HA scaffold enhanced cell proliferation and filopodia formation responsible for cell anchoring within the created 3D environment. The obtained results show the great potential in the development of hybrid scaffolds stimulating bone tissue regeneration.

Enhanced osteoblasts adhesion and collagen formation on biomimetic polyvinylidene fluoride (PVDF) films for bone regeneration.

Bone tissue engineering can be utilized to study the early events of osteoconduction. Fundamental research in cell adhesion to various geometries and proliferation has shown the potential of extending it to implantable devices for regenerative medicine. Following this concept in our studies, first, we developed well-controlled processing of polyvinylidene fluoride (PVDF) film to obtain a surface biomimicking ECM. We optimized the manufacturing dependent on humidity and temperature during spin-coating of a polymer solution. The mixture of solvents such as dimethylacetamide and acetone together with high humidity conditions led to a biomimetic, highly porous and rough surface, while with lower humidity and high temperatures drying allowed us to obtain a smooth and flat PVDF film. The roughness of the PVDF film was biofabricated and compared to smooth films in cell culture studies for adhesion and proliferation of osteoblasts. The bioinspired roughness of our films enhanced the osteoblast adhesion by over 44%, and there was collagen formation already after 7 days of cell culturing that was proved via scanning electron microscopy observation, light microscopy imaging after Sirius Red staining, and proliferation test such as MTS. Cell development, via extended filopodia, formed profoundly on the rough PVDF surface, demonstrated the potential of the structural design of biomimetic surfaces to enhance further bone tissue regeneration.

Cell Integration with Electrospun PMMA Nanofibers, Microfibers, Ribbons, and Films: A Microscopy Study.

Tissue engineering requires properly selected geometry and surface properties of the scaffold, to promote in vitro tissue growth. In this study, we obtained three types of electrospun poly(methyl methacrylate) (PMMA) scaffolds-nanofibers, microfibers, and ribbons, as well as spin-coated films. Their morphology was imaged by scanning electron microscopy (SEM) and characterized by average surface roughness and water contact angle. PMMA films had a smooth surface with roughness, Ra below 0.3 µm and hydrophilic properties, whereas for the fibers and the ribbons, we observed increased hydrophobicity, with higher surface roughness and fiber diameter. For microfibers, we obtained the highest roughness of 7 µm, therefore, the contact angle was 140°. All PMMA samples were used for the in vitro cell culture study, to verify the cells integration with various designs of scaffolds. The detailed microscopy study revealed that higher surface roughness enhanced cells' attachment and their filopodia length. The 3D structure of PMMA microfibers with an average fiber diameter above 3.5 µm, exhibited the most favorable geometry for cells' ingrowth, whereas, for other structures we observed cells growth only on the surface. The study showed that electrospinning of various scaffolds geometry is able to control cells development that can be adjusted according to the tissue needs in the regeneration processes.

Thermal insulation design bioinspired by microstructure study of penguin feather and polar bear hair.

Nature is an amazing source of inspiration for the design of thermal insulation strategies, which are key for saving energy. In nature, thermal insulation structures, such as penguin feather and polar bear hair, are well developed; enabling the animals' survival in frigid waters. The detailed microscopy investigations conducted in this study, allowed us to perform microstructural analysis of these thermally insulating materials, including statistical measurements of keratin fiber and pore dimensions directly from high resolution Scanning Electron Microscope (SEM) images. The microscopy study revealed many similarities in both materials, and showed the importance of their hierarchically-organized porous structure. Finally, we propose the schematic configuration of a thermally-insulating structure, based on the penguin feather and polar bear hair. These optimized thermal-insulator systems indicate the road maps for future development, and new approaches in the design of material properties. STATEMENT OF SIGNIFICANCE: We present the first detailed comparison of microstructures of penguin feather and polar bear hair for designing optimum thermal insulation properties. This unique study involves the measurement of the sizes of pores and fibers of these two keratin-based materials, including the investigation of their 3D arrangements. We revealed porosity interconnection, especially in polar bear hair, which is one of the key designs exhibited by thermal insulation materials.

Surface-Potential-Controlled Cell Proliferation and Collagen Mineralization on Electrospun Polyvinylidene Fluoride (PVDF) Fiber Scaffolds for Bone Regeneration.

This study represents the unique analysis of the electrospun scaffolds with the controlled and stable surface potential without any additional biochemical modifications for bone tissue regeneration. We controlled surface potential of polyvinylidene fluoride (PVDF) fibers with applied positive and negative voltage polarities during electrospinning, to obtain two types of scaffolds PVDF(+) and, PVDF(-). The cells' attachments to PVDF scaffolds were imaged in great details with advanced scanning electron microscopy (SEM) and 3D tomography based on focus ion beam (FIB-SEM). We presented the distinct variations in cells shapes and in filopodia and lamellipodia formation according to the surface potential of PVDF fibers that was verified with Kelvin probe force microscopy (KPFM). Notable, cells usually reach their maximum spread area through increased proliferation, suggesting the stronger adhesion, which was indeed double for PVDF(-) scaffolds having surface potential of -95 mV. Moreover, by tuning the surface potential of PVDF fibers, we were able to enhance collagen mineralization for possible use in bone regeneration. The scaffolds built of PVDF(-) fibers demonstrated the greater potential for bone regeneration than PVDF(+), showing after 7 days in osteoblasts culture produce well-mineralized osteoid required for bone nodules. The collagen mineralization was confirmed with energy dispersive X-ray spectroscopy (EDX) and Sirius Red staining, additionally the cells proliferation with fluorescence microscopy and Alamar Blue assays. The scaffolds made of PVDF fibers with the similar surface potential to the cell membranes promoting bone growth for next-generation tissue scaffolds, which are on a high demand in bone regenerative medicine.