Optimizing Platelet and Leucocyte-Rich Plasma as Biomaterials for Ophthalmic Applications: Impact of Centrifugation Speed.

Purpose: To assess the impact of varying centrifugation speeds on platelet and leucocyte-rich plasma (L-PRP) in liquid and gel form cellularity and growth factor concentrations for potential use against ocular surface disorders. Patients and Methods: L-PRP was collected from 16 healthy subjects using three different centrifugation speeds: 580, 1000, and 2000 g, each for 8 min at 25°C. Platelet and leukocyte counts were automatically evaluated. The concentrations of vascular endothelial growth factor (VEGF), platelet-derived growth factor (PDGF), and transforming growth factor beta 1 (TGF-B1) were measured using enzyme-linked immunosorbent assays. L-PRP gel cellularity was assessed through hematoxylin-eosin and Masson's trichrome staining, categorized as moderate or abundant, and statistically analyzed. L-PRP gel membrane's chemical composition was analyzed using Fourier-transform infrared spectroscopy (FTIR), crystallization was investigated with X-ray diffraction (XRD), and ultrastructure was assessed using surface electron microscopy (SEM). Additionally, membrane degradation was evaluated over a 7-day period. Results: No significant differences in cellularity and growth factor concentrations among centrifugation speeds (p > 0.05) were found. Moderate cellularity predominated at 580 g and 2000 g, while abundant cellularity was observed at 1000 g. No significant differences were found techniques (p = 0.16). Masson's trichrome staining suggested the existence of abundant fibrin at 1000 g but without significant differences (p = 0.07). FTIR analysis exhibited the characteristic fibrin bands at all speeds, and XRD indicated a keratin-like pattern. SEM revealed greater porosity at 580 g and fibrin membrane degradation was lower at this speed (p = 0.0001). Conclusion: Centrifugation speed did not significantly affect growth factor concentration or cellularity in both liquid and gel L-PRP. Further studies should explore the impact of different separation techniques for L-PRP used in ophthalmic applications.

Customizable Collagen Vitrigel Membranes and Preliminary Results in Corneal Engineering.

Corneal opacities are a leading cause of visual impairment that affect 4.2 million people annually. The current treatment is corneal transplantation, which is limited by tissue donor shortages. Corneal engineering aims to develop membranes that function as scaffolds in corneal cell transplantation. Here, we describe a method for producing transplantable corneal constructs based on a collagen vitrigel (CVM) membrane and corneal endothelial cells (CECs). The CVMs were produced using increasing volumes of collagen type I: 1X (2.8 µL/mm2), 2X, and 3X. The vitrification process was performed at 40% relative humidity (RH) and 40 °C using a matryoshka-like system consisting of a shaking-oven harboring a desiccator with a saturated K2CO3 solution. The CVMs were characterized via SEM microscopy, cell adherence, FTIR, and manipulation in an ex vivo model. A pilot transplantation of the CECs/CVM construct in rabbits was also carried out. The thickness of the CVMs was 3.65-7.2 µm. The transparency was superior to a human cornea (92.6% = 1X; 94% = 2X; 89.21% = 3X). SEM microscopy showed a homogenous surface and laminar organization. The cell concentration seeded over the CVM increased threefold with no significant difference between 1X, 2X, and 3X (p = 0.323). The 2X-CVM was suitable for surgical manipulation in the ex vivo model. Constructs using the CECs/2X-CVM promoted corneal transparency restoration.

Reinforcement of Alginate-Gelatin Hydrogels with Bioceramics for Biomedical Applications: A Comparative Study.

This study states the preparation of novel ink with potential use for bone and cartilage tissue restoration. 3Dprint manufacturing allows customizing prostheses and complex morphologies of any traumatism. The quest for bioinks that increase the restoration rate based on printable polymers is a need. This study is focused on main steps, the synthesis of two bioceramic materials as WO3 and Na2Ti6O13, its integration into a biopolymeric-base matrix of Alginate and Gelatin to support the particles in a complete scaffold to trigger the potential nucleation of crystals of calcium phosphates, and its comparative study with independent systems of formulations with bioceramic particles as Al2O3, TiO2, and ZrO2. FT-IR and SEM studies result in hydroxyapatite's potential nucleation, which can generate bone or cartilage tissue regeneration systems with low or null cytotoxicity. These composites were tested by cell culture techniques to assess their biocompatibility. Moreover, the reinforcement was compared individually by mechanical tests with higher results on synthesized materials Na2Ti6O13 with 35 kPa and WO3 with 63 kPa. Finally, the integration of these composite materials formulated by Alginate/Gelatin and bioceramic has been characterized as functional for further manufacturing with the aid of novel biofabrication techniques such as 3D printing.

Hydroxyapatite Nanoparticles in Drug Delivery: Physicochemistry and Applications.

Hydroxyapatite (HAP) has been the gold standard in the biomedical field due to its composition and similarity to human bone. Properties such as shape, size, morphology, and ionic substitution can be tailored through the use of different synthesis techniques and compounds. Regardless of the ability to determine its physicochemical properties, a conclusion for the correlation with the biological response it is yet to be found. Hence, a special focus on the most desirable properties for an appropriate biological response needs to be addressed. This review provides an overview of the fundamental properties of hydroxyapatite nanoparticles and the characterization of physicochemical properties involved in their biological response and role as a drug delivery system. A summary of the main chemical properties and applications of hydroxyapatite, the advantages of using nanoparticles, and the influence of shape, size, functional group, morphology, and crystalline phase in the biological response is presented. A special emphasis was placed on the analysis of chemical and physical interactions of the nanoparticles and the cargo, which was explained through the use of spectroscopic and physical techniques such as FTIR, Raman, XRD, SEM, DLS, and BET. We discuss the properties tailored for hydroxyapatite nanoparticles for a specific biomolecule based on the compilation of studies performed on proteins, peptides, drugs, and genetic material.

Alginate/Gelatin Hydrogels Reinforced with TiO2 and ß-TCP Fabricated by Microextrusion-based Printing for Tissue Regeneration.

Three-dimensional (3D) printing technologies have become an attractive manufacturing process to fabricate scaffolds in tissue engineering. Recent research has focused on the fabrication of alginate complex shaped structures that closely mimic biological organs or tissues. Alginates can be effectively manufactured into porous three-dimensional networks for tissue engineering applications. However, the structure, mechanical properties, and shape fidelity of 3D-printed alginate hydrogels used for preparing tissue-engineered scaffolds is difficult to control. In this work, the use of alginate/gelatin hydrogels reinforced with TiO2 and ß-tricalcium phosphate was studied to tailor the mechanical properties of 3D-printed hydrogels. The hydrogels reinforced with TiO2 and ß-TCP showed enhanced mechanical properties up to 20 MPa of elastic modulus. Furthermore, the pores of the crosslinked printed structures were measured with an average pore size of 200 µm. Additionally, it was found that as more layers of the design were printed, there was an increase of the line width of the bottom layers due to its viscous deformation. Shrinkage of the design when the hydrogel is crosslinked and freeze dried was also measured and found to be up to 27% from the printed design. Overall, the proposed approach enabled fabrication of 3D-printed alginate scaffolds with adequate physical properties for tissue engineering applications.

Electrospun Polycaprolactone Fibrous Membranes Containing Ag, TiO2 and Na2Ti6O13 Particles for Potential Use in Bone Regeneration.

Biocompatible and biodegradable membrane treatments for regeneration of bone are nowadays a promising solution in the medical field. Bioresorbable polymers are extensively used in membrane elaboration, where polycaprolactone (PCL) is used as base polymer. The goal of this work was to improve electrospun membranes' biocompatibility and antibacterial properties by adding micro- and nanoparticles such as Ag, TiO2 and Na2Ti6O13. Micro/nanofiber morphologies of the obtained membranes were characterized by X-ray diffraction, Fourier-transform infrared spectroscopy, differential scanning calorimetry, scanning electron microscopy, energy-dispersive X-ray spectroscopy and a tensile test. Also, for this study optical microscopy was used to observe DAPI-stained cells. Membranes of the different systems were electrospun to an average diameter of 1.02⁻1.76 µm. To evaluate the biological properties, cell viability was studied by growing NIH/3T3 cells on the microfibers. PCL/TiO2 strength was enhanced from 0.6 MPa to 6.3 MPa in comparison with PCL without particles. Antibacterial activity was observed in PCL/TiO2 and PCL/Na2Ti6O13 electrospun membranes using Staphylococcus aureus bacteria. Bioactivity of the membranes was confirmed with simulated body fluid (SBF) treatment. From this study, the ceramic particles TiO2 and Na2Ti6O13, combined with a PCL matrix with micro/nanoparticles, enhanced cell proliferation, adhesion and antibacterial properties. The electrospun composite with Na2Ti6O13 can be considered viable for tissue regenerative processes.

On the Rule of Mixtures for Predicting Stress-Softening and Residual Strain Effects in Biological Tissues and Biocompatible Materials.

In this work, we use the rule of mixtures to develop an equivalent material model in which the total strain energy density is split into the isotropic part related to the matrix component and the anisotropic energy contribution related to the fiber effects. For the isotropic energy part, we select the amended non-Gaussian strain energy density model, while the energy fiber effects are added by considering the equivalent anisotropic volumetric fraction contribution, as well as the isotropized representation form of the eight-chain energy model that accounts for the material anisotropic effects. Furthermore, our proposed material model uses a phenomenological non-monotonous softening function that predicts stress softening effects and has an energy term, derived from the pseudo-elasticity theory, that accounts for residual strain deformations. The model's theoretical predictions are compared with experimental data collected from human vaginal tissues, mice skin, poly(glycolide-co-caprolactone) (PGC25 3-0) and polypropylene suture materials and tracheal and brain human tissues. In all cases examined here, our equivalent material model closely follows stress-softening and residual strain effects exhibited by experimental data.