Applications of Stem Cell-Derived Extracellular Vesicles in Nerve Regeneration.

Extracellular vesicles (EVs), including exosomes, microvesicles, and other lipid vesicles derived from cells, play a pivotal role in intercellular communication by transferring information between cells. EVs secreted by progenitor and stem cells have been associated with the therapeutic effects observed in cell-based therapies, and they also contribute to tissue regeneration following injury, such as in orthopaedic surgery cases. This review explores the involvement of EVs in nerve regeneration, their potential as drug carriers, and their significance in stem cell research and cell-free therapies. It underscores the importance of bioengineers comprehending and manipulating EV activity to optimize the efficacy of tissue engineering and regenerative therapies.

Marine Skeletons: Towards Hard Tissue Repair and Regeneration.

Musculoskeletal disorders in the elderly have significantly increased due to the increase in an ageing population. The treatment of these diseases necessitates surgical procedures, including total joint replacements such as hip and knee joints. Over the years a number of treatment options have been specifically established which are either permanent or use temporary natural materials such as marine skeletons that possess unique architectural structure and chemical composition for the repair and regeneration of bone tissue. This review paper will give an overview of presently used materials and marine structures for hard tissue repair and regeneration, drugs of marine origin and other marine products which show potential for musculoskeletal treatment.

Bioresorbable zinc hydroxyapatite guided bone regeneration membrane for bone regeneration.

OBJECTIVES: The aim of this study was to investigate the bone regenerative properties of a heat treated cross-linked GBR membrane with zinc hydroxyapatite powders in the rat calvarial defect model over a 6-week period. MATERIAL AND METHODS: In vitro physio-chemical characterization involved X-ray diffraction analysis, surface topology by scanning electron microscopy, and zinc release studies in physiological buffers. Bilateral rat calvarial defects were used to compare the Zn-HAp membranes against the commercially available collagen membranes and the unfilled defect group through radiological and histological evaluation. RESULTS: The synthesized Zn-MEM (100 µm thick) showed no zinc ions released in the phosphate buffer solution (PBS) buffer, but zinc was observed under acidic conditions. At 6 weeks, both the micro-CT and histological analyses revealed that the Zn-MEM group yielded significantly greater bone formation with 80 ± 2% of bone filled, as compared with 60 ± 5% in the collagen membrane and 40 ± 2% in the unfilled control group. CONCLUSION: This study demonstrated the use of heat treatment as an alternative method to cross-linking the Zn-MEM to be applied as a GBR membrane. Its synthesis and production are relatively simple to fabricate, and the membrane had rough surface features on one side, which might be beneficial for cellular activities. In a rat calvarial defect model, it was shown that new bone formation was accelerated in comparison with the collagen membrane and the unfilled defect groups. These results would suggest that Zn-MEM has the potential for further development in dental applications.

Marine structure derived calcium phosphate-polymer biocomposites for local antibiotic delivery.

Hydrothermally converted coralline hydroxyapatite (HAp) particles loaded with medically active substances were used to develop polylactic acid (PLA) thin film composites for slow drug delivery systems. The effects of HAp particles within PLA matrix on the gentamicin (GM) release and release kinetics were studied. The gentamicin release kinetics seemed to follow Power law Korsmeyer Peppas model with mainly diffusional process with a number of different drug transport mechanisms. Statistical analysis shows very significant difference on the release of gentamicin between GM containing PLA (PLAGM) and GM containing HAp microspheres within PLA matrix (PLAHApGM) devices, which PLAHApGM displays lower release rates. The use of HAp particles improved drug stabilization and higher drug encapsulation efficiency of the carrier. HAp is also the source of Ca2+ for the regeneration and repair of diseased bone tissue. The release profiles, exhibited a steady state release rate with significant antimicrobial activity against Staphylococcus aureus (S. aureus) (SH1000) even at high concentration of bacteria. The devices also indicated significant ability to control the growth of bacterial even after four weeks of drug release. Clinical release profiles can be easily tuned from drug-HAp physicochemical interactions and degradation kinetics of polymer matrix. The developed systems could be applied to prevent microbial adhesion to medical implant surfaces and to treat infections mainly caused by S. aureus in surgery.

Coral exoskeletons as a precursor material for the development of a calcium phosphate drug delivery system for bone tissue engineering.

With the global rise in aging of populations, the occurrence of osteoporosis will continue to increase. Biomaterial and pharmaceutical scientists continue to develop innovative strategies and materials to address this disease. In this article, we describe a new perspective and approach into the use of coral exoskeletons as a precursor material to synthesize a calcium phosphate-based drug delivery system. Studies detailing the methodology of the conversion methods and the strategies and approach for the development of these novel drug delivery systems are described. Furthermore, in vivo studies in osteoporotic mice using a drug loaded and chemically modified version of the biomimetic delivery system showed significant cortical and cancellous bone increases. These studies support the notion and the rationale for future research and development of the use of coral exoskeletons as materials for drug delivery applications.

Bone regeneration of rat tibial defect by zinc-tricalcium phosphate (Zn-TCP) synthesized from porous Foraminifera carbonate macrospheres.

Foraminifera carbonate exoskeleton was hydrothermally converted to biocompatible and biodegradable zinc-tricalcium phosphate (Zn-TCP) as an alternative biomimetic material for bone fracture repair. Zn-TCP samples implanted in a rat tibial defect model for eight weeks were compared with unfilled defect and beta-tricalcium phosphate showing accelerated bone regeneration compared with the control groups, with statistically significant bone mineral density and bone mineral content growth. CT images of the defect showed restoration of cancellous bone in Zn-TCP and only minimal growth in control group. Histological slices reveal bone in-growth within the pores and porous chamber of the material detailing good bone-material integration with the presence of blood vessels. These results exhibit the future potential of biomimetic Zn-TCP as bone grafts for bone fracture repair.

A therapeutic potential for marine skeletal proteins in bone regeneration.

A vital ingredient for engineering bone tissue, in the culture dish, is the use of recombinant matrix and growth proteins to help accelerate the growth of cultivated tissues into clinically acceptable quantities. The skeletal organic matrices of calcifying marine invertebrates are an untouched potential source of such growth inducing proteins. They have the advantage of being ready-made and retain the native state of the original protein. Striking evidence shows that skeleton building bone morphogenic protein-2/4 (BMP) and transforming growth factor beta (TGF-ß) exist within various marine invertebrates such as, corals. Best practice mariculture and the latest innovations in long-term marine invertebrate cell cultivation can be implemented to ensure that these proteins are produced sustainably and supplied continuously. This also guarantees that coral reef habitats are not damaged during the collection of specimens. Potential proteins for bone repair, either extracted from the skeleton or derived from cultivated tissues, can be identified, evaluated and retrieved using chromatography, cell assays and proteomic methods. Due to the current evidence for bone matrix protein analogues in marine invertebrates, together with the methods established for their production and retrieval there is a genuine prospect that they can be used to regenerate living bone for potential clinical use.

Effects of micromovement on the changes in stress distribution of partially stabilized zirconia (PS-ZrO2) dental implants and bridge during clenching: a three-dimensional finite element analysis.

OBJECTIVE: This investigation aims to evaluate the changes in stress magnitudes and distributions on Partially Stabilized Zirconia (PS-ZrO(2)) dental implants and bridges and on the mandible caused by fibrous encapsulations during clenching. MATERIALS AND METHODS: Four 3.26 mm diameter PS-ZrO(2) dental implants with lengths of 12 mm were modelled and placed in the second premolar and first molar region on both sides of the mandible model. A rigid zirconia bridge with a thickness of 0.5 mm connects the PS-ZrO(2) dental implants placed in the second premolar and first molar. Four periodontal ligament (PDL) case studies were examined: PDL in the second premolars; PDL in the first molars; PDL in both the second premolars and first molars; and no PDL present. RESULTS: The results reveal the magnitudes and distributions of stresses on the dental implants and connecting bridges were governed by the PDLs. A significant drop in stress levels were recorded when the PDL encapsulates the roots of the dental implants. Of the four PDL case studies, it was found that when the PDLs are present in both the second premolars and first molars the lowest stress magnitudes are generated. The analysis also revealed that, during the healing process after implant insertion and the result of fibrous encapsulation, the dental implant system will experience a varying amount of stress levels. CONCLUSION: This study was intended to produce more insight into the influence of the PDL on the changes in stress distribution on the dental implant system during clenching.

Finite element stress analysis of Ti-6Al-4V and partially stabilized zirconia dental implant during clenching.

OBJECTIVE: The purpose of this paper is to compare the differences in stress between Ti-6Al-4V and PS-ZrO(2) dental implant during clenching and whether these changes are clinically significant to limit the use of zirconia in oral implantology. MATERIALS AND METHODS: The model geometry was derived from position measurements taken from 28 diamond blade cut cross-sections of an average size human adult edentulous mandible and generated using a special sequencing method. Data on anatomical, structural, functional aspects and material properties were obtained from measurements and published data. Ti-6Al-4V and PS-ZrO(2) dental implants were modelled as cylindrical structure with a diameter of 3.26 mm and length of 12.00 mm was placed in the first molar region on the right hemimandible. RESULTS: The analysis revealed an increase of 2-3% in the averaged tensile and compressive stress and an increase of 8% in the averaged Von Mises stress were recorded in the bone-implant interface when PS-ZrO(2) dental implant was used instead of Ti-6Al-4V dental implant. The results also revealed only relatively low levels of stresses were transferred from the implant to the surrounding cortical and cancellous bone, with the majority of the stresses transferred to the cortical bone. CONCLUSION: Even though high magnitudes of tensile, compressive and Von Mises stresses were recorded on the Ti-6Al-4V and PS-ZrO(2) dental implants and in the surrounding osseous structures, the stresses may not be clinically critical since the mechanical properties of the implant material and the cortical and cancellous bone could withstand stress magnitudes far greater than those recorded in this analysis.

Exosome Structures Supported by Machine Learning Can Be Used as a Promising Diagnostic Tool.

Principal component analysis (PCA) as a machine-learning technique could serve in disease diagnosis and prognosis by evaluating the dynamic morphological features of exosomes via Cryo-TEM-imaging. This hypothesis was investigated after the crude isolation of similarly featured exosomes derived from the extracellular vehicles (EVs) of immature dendritic cells (IDCs) JAWSII. It is possible to identify functional molecular groups by FTIR, but the unique physical and morphological characteristics of exosomes can only be revealed by specialized imaging techniques such as cryo-TEM. On the other hand, PCA has the ability to examine the morphological features of each of these IDC-derived exosomes by considering software parameters such as various membrane projections and differences in Gaussians, Hessian, hue, and class to assess the 3D orientation, shape, size, and brightness of the isolated IDC-derived exosome structures. In addition, Brownian motions from nanoparticle tracking analysis of EV IDC-derived exosomes were also compared with EV IDC-derived exosome images collected by scanning electron microscopy and confocal microscopy. Sodium-Dodecyl-Sulphate-Polyacrylamide-Gel-Electrophoresis (SDS-PAGE) was performed to separate the protein content of the crude isolates showing that no considerable protein contamination occurred during the crude isolation technique of IDC-derived-exosomes. This is an important finding because no additional purification of these exosomes is required, making PCA analysis both valuable and novel.

In vitro testing and efficacy of poly-lactic acid coating incorporating antibiotic loaded coralline bioceramic on Ti6Al4V implant against Staphylococcus aureus.

Biofilm formation on an implant surface is most commonly caused by the human pathogenic bacteria Staphylococcus aureus, which can lead to implant related infections and failure. It is a major problem for both implantable orthopedic and maxillofacial devices. The current antibiotic treatments are typically delivered orally or in an injectable form. They are not highly effective in preventing or removing biofilms, and they increase the risk of antibiotic resistance of bacteria and have a dose-dependent negative biological effect on human cells. Our aim was to improve current treatments via a localized and controlled antibiotic delivery-based implant coating system to deliver the antibiotic, gentamicin (Gm). The coating contains coral skeleton derived hydroxyapatite powders (HAp) that act as antibiotic carrier particles and have a biodegradable poly-lactic acid (PLA) thin film matrix. The system is designed to prevent implant related infections while avoiding the deleterious effects of high concentration antibiotics in implants on local cells including primary human adipose derived stem cells (ADSCs). Testing undertaken in this study measured the rate of S. aureus biofilm formation and determined the growth rate and proliferation of ADSCs. After 24 h, S. aureus biofilm formation and the percentage of live cells found on the surfaces of all 5%-30% (w/w) PLA-Gm-(HAp-Gm) coated Ti6Al4V implants was lower than the control samples. Furthermore, Ti6Al4V implants coated with up to 10% (w/w) PLA-Gm-(HAp-Gm) did not have noticeable Gm related adverse effect on ADSCs, as assessed by morphological and surface attachment analyses. These results support the use and application of the antibacterial PLA-Gm-(HAp-Gm) thin film coating design for implants, as an antibiotic release control mechanism to prevent implant-related infections.

Bio-Activation of HA/ß-TCP Porous Scaffolds by High-Pressure CO2 Surface Remodeling: A Novel "Coating-from" Approach.

Biphasic macroporous Hydroxyapatite/ß-Tricalcium Phosphate (HA/ß-TCP) scaffolds (BCPs) are widely used for bone repair. However, the high-temperature HA and ß-TCP phases exhibit limited bioactivity (low solubility of HA, restricted surface area, low ion release). Strategies were developed to coat such BCPs with biomimetic apatite to enhance bioactivity. However, this can be associated with poor adhesion, and metastable solutions may prove difficult to handle at the industrial scale. Alternative strategies are thus desirable to generate a highly bioactive surface on commercial BCPs. In this work, we developed an innovative "coating from" approach for BCP surface remodeling via hydrothermal treatment under supercritical CO2, used as a reversible pH modifier and with industrial scalability. Based on a set of complementary tools including FEG-SEM, solid state NMR and ion exchange tests, we demonstrate the remodeling of macroporous BCP surface with the occurrence of dissolution-reprecipitation phenomena involving biomimetic CaP phases. The newly precipitated compounds are identified as bone-like nanocrystalline apatite and octacalcium phosphate (OCP), both known for their high bioactivity character, favoring bone healing. We also explored the effects of key process parameters, and showed the possibility to dope the remodeled BCPs with antibacterial Cu2+ ions to convey additional functionality to the scaffolds, which was confirmed by in vitro tests. This new process could enhance the bioactivity of commercial BCP scaffolds via a simple and biocompatible approach.

Synthetic tissue engineering with smart, cytomimetic protocells.

Synthetic protocells are rudimentary origin-of-life versions of natural cell counterparts. Protocells are widely engineered to advance efforts and useful accepted outcomes in synthetic biology, soft matter chemistry and bioinspired materials chemistry. Protocells in collective symbiosis generate synthetic proto-tissues that display unprecedented autonomy and yield advanced materials with desirable life-like features for smart multi-drug delivery, micro bioreactors, renewable fuel production, environmental clean-up, and medicine. Current levels of protocell and proto-tissue functionality and adaptivity are just sufficient to apply them in tissue engineering and regenerative medicine, where they animate biomaterials and increase therapeutic cell productivity. As of now, structural biomaterials for tissue engineering lack the properties of living biomaterials such as self-repair, stochasticity, cell synergy and the sequencing of molecular and cellular events. Future protocell-based biomaterials provide these core properties of living organisms, but excluding evolution. Most importantly, protocells are programmable for a broad array of cell functions and behaviors and collectively in consortia are tunable for multivariate functions. Inspired by upcoming designs of smart protocells, we review their developmental background and cover the most recently reported developments in this promising field of synthetic proto-biology. Our emphasis is on manufacturing proto-tissues for tissue engineering of organoids, stem cell niches and reprogramming and tissue formation through stages of embryonic development. We also highlight the exciting reported developments arising from fusing living cells and tissues, in a valuable hybrid symbiosis, with synthetic counterparts to bring about novel functions, and living tissue products for a new synthetic tissue engineering discipline.

Specifiable biomimetic microsponges for timed release of crystal entrapped biomolecules useful in bone repair.

Most marine materials, by nature, contain crystals of inorganic matter with specific structures that allow the loading, release, and delivery of biomolecules that can be utilized in clinical applications. These structures can be biomimetically synthesized. Aggregates of inorganic particles generated by biomimetic microsponges may provide surfaces and structures for cell attachment, organization, and promotion of matrix synthesis. Biomimetic microsponges have been developed with tunable release profiles differing by the rate (speed over distance), velocity (rate of change in direction), and the quantity discharged over time, according to biomolecular species. Specifically, the types of proteins involved guide and regulate cells in physical contact with the microsponges, for instance, reprogramming somatic cells, the switching phenotypes, or specifying stem cell differentiation. Applications for these microsponges include gene transfection of localized cells and promotion of bone matrix synthesis by the externalized display of RGD cell adhesive peptides and the release of crystal entrapped, occluded, adsorbed and infused rhBMP-2 and plasmid. A requirement for de novo bone formation is a solid structure to enable osteocytes to lay new bone tissue. In this study, biomimetic microsponges highlight tremendous potential as osteoconductive packing material in bone repair with parallel influence on regeneration. Majorly, microsponges offer pronounced osteoinductivity, unlike many other bone particulates, by solid-state integration of active regenerative biological molecules through the prism of the biomineral crystalline structure.

Comparative Analysis of NF-κB in the MyD88-Mediated Pathway After Implantation of Titanium Alloy and Stainless Steel and the Role of Regulatory T Cells.

OBJECTIVE: Development of immunologically smart implants, integrated to biological systems, is a key aim to minimize the inflammatory response of the host to biomaterial implants. METHODS: The aim of this study is to investigate the influence of titanium alloy and stainless steel implants on immunological responses in rats by comparative analysis of nuclear factor kappa B (NF-κB) profiles in the activation of inflammatory signaling pathways and the role of CD4+CD25+Foxp3+. RESULTS: Both Ti alloy and stainless steel alloy group implantation affect Toll-like receptors-4 pathways and CD4+CD25+ regulatory T cells in different ways. CONCLUSIONS: Results show that NF-κB/p65 and NF-κB1/p50 possess potential as a therapeutic target in the prevention of adverse reactions to metal, especially for controlling inflammation after the implantation.

Drug Delivery From Polymer-Based Nanopharmaceuticals-An Experimental Study Complemented by Simulations of Selected Diffusion Processes.

The success of medical therapy depends on the correct amount and the appropriate delivery of the required drugs for treatment. By using biodegradable polymers a drug delivery over a time span of weeks or even months is made possible. This opens up a variety of strategies for better medication. The drug is embedded in a biodegradable polymer (the "carrier") and injected in a particular position of the human body. As a consequence of the interplay between the diffusion process and the degrading polymer the drug is released in a controlled manner. In this work we study the controlled release of medication experimentally by measuring the delivered amount of drug within a cylindrical shell over a long time interval into the body fluid. Moreover, a simple continuum model of the Fickean type is initially proposed and solved in closed-form. It is used for simulating some of the observed release processes for this type of carrier and takes the geometry of the drug container explicitly into account. By comparing the measurement data and the model predictions diffusion coefficients are obtained. It turns out that within this simple model the coefficients change over time. This contradicts the idea that diffusion coefficients are constants independent of the considered geometry. The model is therefore extended by taking an additional absorption term into account leading to a concentration dependent diffusion coefficient. This could now be used for further predictions of drug release in carriers of different shape. For a better understanding of the complex diffusion and degradation phenomena the underlying physics is discussed in detail and even more sophisticated models involving different degradation and mass transport phenomena are proposed for future work and study.

Mechanical testing of antimicrobial biocomposite coating on metallic medical implants as drug delivery system.

Post-operative infection often occurs following orthopedic and dental implant placement requiring systemically administered antibiotics. However, this does not provide long-term protection. Over the last few decades, alternative methods involving slow drug delivery systems based on biodegradable poly-lactic acid and antibiotic loaded hydroxyapatite microspheres were developed to prevent post-operative infection. In this study, thermally anodised and untreated Ti6Al4V discs were coated with Poly-Lactic Acid (PLA) containing Gentamicin (Gm) antibiotic-loaded coralline Hydroxyapatite (HAp) are investigated. Following chemical characterization, mechanical properties of the coated samples were measured using nanoindentation and scratch tests to determine the elastic modulus, hardness and bonding adhesion between film and substrate. It was found that PLA biocomposite multilayered films were around 400nm thick and the influence and effect of the substrate were clearly observed during the nanoindentation studies with heavier loads. Scratch tests of PLA coated samples conducted at ~160nm depth showed the minimal difference in the measured friction between Gm and non Gm containing films. It is also observed that the hardness values of PLA film coated anodised samples ranged from 0.45 to 1.9GPa (dependent on the applied loads) against untreated coated samples which ranged from 0.28 to 0.8GPa.

Micro- and nano-indentation of a hydroxyapatite-carbon nanotube composite.

The mechanical properties of pure synthetic hydroxyapatite and hydroxyapatite-carbon nanotube composites were examined. Vickers microhardness and nanoindentation using a Berkovich tipped indenter were used to determine the hardness, fracture toughness and Young's modulus of the pure hydroxyapatite matrix and the composite materials. Microscopy showed that for the composites produced the carbon nanotubes were present as discrete clumps. These clumps induced a detrimental effect on the hardness of the materials, while the fracture toughness values were not affected. This would be undesirable in terms of using the material for biomedical implant applications. It should be noted that the carbon nanotubes used contained free graphite. As the properties of the composite materials studied were not greatly improved over the matrix, it is speculated that if the graphite phase were removed from the reagent, this could in-turn enhance the properties of the material.

Porous orbital implants in enucleation: a systematic review.

Orbital implants have been used for cosmesis following surgical removal of the eyeball, or enucleation, for over a century. Implant design has progressed significantly in recent years with the use of porous devices, with the theoretical advantages of reduced complications and improved cosmesis. However, in some cases the theoretical benefits have not fully translated into clinical results. In this article the use of orbital implants in enucleation, with a particular focus on the newer porous biomaterials that have gained prominence over the last 15 years, is reviewed. Specific factors identified as affecting the performance of porous orbital implants include the material used, pore size, and morphology. Mechanical factors have received little consideration in the past and may form a basis for the use of higher compliance porous materials in the future. Of the porous materials in use, current clinical evidence is not sufficient to suggest either that porous implants are superior to non-porous implants, or that one material is more suited to the application than another. Future developments in this field require randomized controlled clinical trials with extensive follow-up as complications may not become evident until over 5 years post-implantation.

Development and dissolution studies of bisphosphonate (clodronate)-containing hydroxyapatite-polylactic acid biocomposites for slow drug delivery.

An increase in clinical demand on the controlled release of bisphosphonates (BPs) due to complications associated with systemic administration, has been the current driving force on the development of BP drug-release systems. Bisphosphonates have the ability to bind to divalent metal ions, such as Ca2+ , in bone mineral and prevent bone resorption by influencing the apoptosis of osteoclasts. Localized delivery using biodegradable materials, such as polylactic acid (PLA) and hydroxyapatite (HAp), which are ideal in this approach, have been used in this study to investigate the dissolution of clodronate (non-nitrogen-containing bisphosphonate) in a new release system. The effects of coral structure-derived HAp and the release kinetics of the composites were evaluated. The release kinetics of clodronate from PLA-BP and PLA-HAp-BP systems seemed to follow the power law model described by Korsmeyer-Peppas. Drug release was quantified by 31 P-NMR with detection and quantification limits of 9.2 and 30.7 mM, respectively. The results suggest that these biocomposite systems could be tuned to release clodronate for both relatively short and prolonged period of time. In addition to drug delivery, the degradation of HAp supplies both Ca2+ and phosphate ions that can help in bone mineralization. Copyright © 2015 John Wiley & Sons, Ltd.