Mesenchymal stem cells derived-exosomes enhanced amniotic membrane extract promotes corneal keratocyte proliferation.

Amniotic membrane extract (AME) and Wharton's jelly mesenchymal stem cells derived-exosomes (WJ-MSC-Exos) are promising therapeutic solutions explored for their potential in tissue engineering and regenerative medicine, particularly in skin and corneal wound healing applications. AME is an extract form of human amniotic membrane and known to contain a plethora of cytokines and growth factors, making it a highly attractive option for topical applications. Similarly, WJ-MSC-Exos have garnered significant interest for their wound healing properties. Although WJ-MSC-Exos and AME have been used separately for wound healing research, their combined synergistic effects have not been studied extensively. In this study, we evaluated the effects of both AME and WJ-MSC-Exos, individually and together, on the proliferation of corneal keratocytes as well as their ability to promote in vitro cell migration, wound healing, and their impact on cellular morphology. Our findings indicated that the presence of both exosomes (3 × 105 Exo/mL) and AME (50 µg/mL) synergistically enhance the proliferation of corneal keratocytes. Combined use of these solutions (3 × 105 Exo/mL + 50 µg/mL) increased cell proliferation compared to only 50 µg/mL AME treatment on day 3 (**** p < 0.0001). This mixture treatment (3 × 105 Exo/mL + 50 µg/mL) increased wound closure rate compared to isolated WJ-MSC-Exo treatment (3 × 105 Exo/mL) (*p < 0.05). Overall, corneal keratocytes treated with AME and WJ-MSC-Exo (3 × 105 Exo/mL + 50 µg/mL) mixture resulted in enhanced proliferation and wound healing tendency. Utilization of combined use of AME and WJ-MSC-Exo can pave the way for a promising foundation for corneal repair research.

Development of photoreactive demineralized bone matrix 3D printing colloidal inks for bone tissue engineering.

Demineralized bone matrix (DBM) has been widely used clinically for dental, craniofacial and skeletal bone repair, as an osteoinductive and osteoconductive material. 3D printing (3DP) enables the creation of bone tissue engineering scaffolds with complex geometries and porosity. Photoreactive methacryloylated gelatin nanoparticles (GNP-MAs) 3DP inks have been developed, which display gel-like behavior for high print fidelity and are capable of post-printing photocrosslinking for control of scaffold swelling and degradation. Here, novel DBM nanoparticles (DBM-NPs, ∼400 nm) were fabricated and characterized prior to incorporation in 3DP inks. The objectives of this study were to determine how these DBM-NPs would influence the printability of composite colloidal 3DP inks, assess the impact of ultraviolet (UV) crosslinking on 3DP scaffold swelling and degradation and evaluate the osteogenic potential of DBM-NP-containing composite colloidal scaffolds. The addition of methacryloylated DBM-NPs (DBM-NP-MAs) to composite colloidal inks (100:0, 95:5 and 75:25 GNP-MA:DBM-NP-MA) did not significantly impact the rheological properties associated with printability, such as viscosity and shear recovery or photocrosslinking. UV crosslinking with a UV dosage of 3 J/cm2 directly impacted the rate of 3DP scaffold swelling for all GNP-MA:DBM-NP-MA ratios with an ∼40% greater increase in scaffold area and pore area in uncrosslinked versus photocrosslinked scaffolds over 21 days in phosphate-buffered saline (PBS). Likewise, degradation (hydrolytic and enzymatic) over 21 days for all DBM-NP-MA content groups was significantly decreased, ∼45% less in PBS and collagenase-containing PBS, in UV-crosslinked versus uncrosslinked groups. The incorporation of DBM-NP-MAs into scaffolds decreased mass loss compared to GNP-MA-only scaffolds during collagenase degradation. An in vitro osteogenic study with bone marrow-derived mesenchymal stem cells demonstrated osteoconductive properties of 3DP scaffolds for the DBM-NP-MA contents examined. The creation of photoreactive DBM-NP-MAs and their application in 3DP provide a platform for the development of ECM-derived colloidal materials and tailored control of biochemical cue presentation with broad tissue engineering applications.

Multifunctional Silk Fibroin/Carbon Nanofiber Scaffolds for In Vitro Cardiomyogenic Differentiation of Induced Pluripotent Stem Cells and Energy Harvesting from Simulated Cardiac Motion.

In this proof-of-concept study, cardiomyogenic differentiation of induced pluripotent stem cells (iPSCs) is combined with energy harvesting from simulated cardiac motion in vitro. To achieve this, silk fibroin (SF)-based porous scaffolds are designed to mimic the mechanical and physical properties of cardiac tissue and used as triboelectric nanogenerator (TENG) electrodes. The load-carrying mechanism, ß-sheet content, degradation characteristics, and iPSC interactions of the scaffolds are observed to be interrelated and regulated by their pore architecture. The SF scaffolds with a pore size of 379 ± 34 µm, a porosity of 79 ± 1%, and a pore interconnectivity of 67 ± 1% upregulated the expression of cardiac-specific gene markers TNNT2 and NKX2.5 from iPSCs. Incorporating carbon nanofibers (CNFs) enhances the elastic modulus of the scaffolds to 45 ± 3 kPa and results in an electrical conductivity of 0.021 ± 0.006 S/cm. The SF and SF/CNF scaffolds are used as conjugate TENG electrodes and generate a maximum power output of 0.37 × 10-3 mW/m2, with an open-circuit voltage and a short circuit current of 0.46 V and 4.5 nA, respectively, under simulated cardiac motion. A novel approach is demonstrated for fabricating scaffold-based cardiac patches that can serve as tissue scaffolds and simultaneously allow energy harvesting.

Adsorption behavior of serum proteins on anodized titanium is driven by surface nanomorphology.

Protein adsorption behavior can play a critical role in defining the outcome of a material by affecting the subsequent in vivo response to it. To date, the effect of surface properties on protein adsorption behavior has been mainly focused on surface chemistry, but research on the effect of nanoscale surface topography remains limited. In this study, the adsorption behavior of human serum albumin, immunoglobulin G, and fibrinogen in terms of the adsorbed amount and conformational changes were investigated on bare and anodized titanium (Ti) samples (40 and 60 V applied voltages). While the surface chemistry, RMS surface roughness, and arithmetic surface roughness of the anodized samples were similar, they had distinctly different nanomorphologies identified by atomic force microscopy and scanning electron microscopy, and the surface statistical parameters, surface skewness Ssk and kurtosis Sku. The Feret pore size distribution was more uniform on the 60 V sample, and surface nanostructures were more symmetrical with higher peaks and deeper pores. On the other hand, the 40 V sample surface presented a nonuniform pore size distribution and asymmetrical surface nanostructures with lower peaks and shallower pores. The amount of surface-adsorbed protein increased on the sample surfaces in the order of Ti < 40 V < 60 V with the predominant factor affecting the amount of surface-adsorbed protein being the increased surface area attained by pore formation. The secondary structure of all adsorbed proteins deviated from that of their native counterparts. While comparing the secondary structure components of proteins on anodized surfaces, it was observed that all three proteins retained more of their secondary structure composition on the surface with more uniform and symmetrical nanofeatures than the surface having asymmetrical nanostructures. Our results suggest that the nanomorphology of the peaks and outer walls of the nanotubes can significantly influence the conformation of adsorbed serum proteins, even for surfaces having similar roughness values.

Development of electrically conductive porous silk fibroin/carbon nanofiber scaffolds.

Tissue engineering applications typically require three-dimensional scaffolds which provide the requisite surface area for cellular functions, while allowing transport of nutrients, waste and oxygen to and from the surrounding tissues. Scaffolds need to ensure sufficient mechanical properties to provide mechanically stable frameworks under physiologically relevant stress levels. Meanwhile, electrically conductive platforms are also desirable for the regeneration of specific tissues, where electrical impulses are transmitted throughout the tissue for proper physiological functioning. Towards this goal, carbon nanofibers (CNFs) were incorporated into silk fibroin (SF) scaffolds whose pore size and porosity were controlled during a salt leaching process. In our methodology, CNFs were dispersed in SF due to the hydrogen bond-forming ability of hexafluoro-2-propanol, a fluoroalcohol used as a solvent for SF. Results showed enhanced electrical conductivity and mechanical properties upon the incorporation of CNFs into the SF scaffolds, while the metabolic activities of cells cultured on SF/CNF nanocomposite scaffolds were significantly improved by optimizing the CNF content, porosity and pore size range of the scaffolds. Specifically, SF/CNF nanocomposite scaffolds with electrical conductivities as high as 0.023 S cm-1, tangent modulus values of 260 ± 30 kPa, a porosity as high as 78% and a pore size of 376 ± 53 µm were fabricated for the first time in the literature. Furthermore, an increase of about 34% in the wettability of SF was achieved by the incorporation of 10% CNF, which provided enhanced fibroblast spreading on scaffold surfaces.

Fabrication and cellular interactions of nanoporous tantalum oxide.

Tantalum possesses remarkable chemical and mechanical properties, and thus it is considered to be one of the next generation implant materials. However, the biological properties of tantalum remain to be improved for its use in tissue engineering applications. To enhance its cellular interactions, implants made of tantalum could be modified to obtain nanofeatured surfaces via the electrochemical anodization process. In this study, anodization parameters were adjusted to obtain a nanoporous surface morphology on tantalum surfaces and systematically altered to control the pore sizes from 25 to 65 nm using an aqueous HF:H2 SO4 electrolyte. Results indicated the formation of Ta2 O5 -based nanoporous surface layers, which had up to 28% more surface area and increased nanophase roughness (more than twofolds) compared to nonporous tantalum upon the anodization. It was observed that the nanoporous tantalum oxide surfaces promoted nearly 25% more fibroblast proliferation at 5 days in vitro and 15.5% more cellular spreading. Thus, nanoporous tantalum oxide surfaces can be used to increase biological interactions of the cells and provide a means of improving bioactivity of tantalum for biomaterial applications.