Histological assessment in peripheral nerve tissue engineering

The histological analysis of peripheral nerve regeneration is one of the most used methods to demonstrate the success of the regeneration through nerve conduits. Nowadays, it is possible to evaluate different parameters of nerve regeneration by using histological, histochemical, immunohistochemical and ultrastructural techniques. The histochemical methods are very sensible and are useful tools to evaluate the extracellular matrix remodeling and the myelin sheath, but they are poorly specific. In contrast, the immunohistochemical methods are highly specific and are frequently used for the identification of the regenerated axons, Sehwann cells and proteins associated to nerve regeneration or neural linage. The ultrastructural techniques offer the possibility to perform a high resolution morphological and quantitative analysis of the nerve regeneration. However, the use of a single histological method may not be enough to assess the degree of regeneration, and the combination of different histological techniques could be necessary.

Reduced graphene oxide-grafted bovine serum albumin/bredigite nanocomposites with high mechanical properties and excellent osteogenic bioactivity for bone tissue engineering

The optimization of the scaffolds to provide a suitable matrix and accelerate the regeneration process is vital for bone tissue engineering.However,poor mechanical and biological characteristics remain the primary challenges that must be addressed.For example,although bredigite(Br)has shown great potential for application in bone tissue engineering,it easily fails in replacement.In the present work,these challenges are addressed by reinforcing the Br matrix with nanosheets of graphene oxide(rGO)that have been reduced by bovine serum albumin(BSA)in order to enhance the mechanical properties and biological behavior.The reduction of graphene oxide by BSA improves the water stability of the nanosheets and provides an electrostatic interaction between theBSA-rGO nanosheets and theBr particles.The high thermal conductivity of theBSA-rGO nanosheets decreases the porosity of the Br by transferring heat to the core of the tablet.Furthermore,the addition of BSA-rGO nanosheets into the Br matrix enhances the adhesion of G-292 cells on the surface of the tablets.These findings suggest that the tablet consisting of BSA-rGO-reinforced Br has encouraging potential for application in bone tissue engineering.

Myelin histology:a key tool in nervous system research

The myelin sheath is a lipoprotein-rich,multilayered structure capable of increasing conduction velocity in central and peripheral myelinated nerve fibers.Due to the complex structure and composition of myelin,various histological techniques have been developed over the centuries to evaluate myelin under normal,pathological or experimental conditions.Today,methods to assess myelin integrity or content are key tools in both clinical diagnosis and neuroscience research.In this review,we provide an updated summary of the composition and structure of the myelin sheath and discuss some histological procedures,from tissue fixation and processing techniques to the most used and practical myelin histological staining methods.Considering the lipoprotein nature of myelin,the main features and technical details of the different available methods that can be used to evaluate the lipid or protein components of myelin are described,as well as the precise ultrastructural techniques.

Supposition of graphene stacks to estimate the contact resistance and conductivity of nanocomposites

In this study,the effects of stacked nanosheets and the surrounding interphase zone on the resistance of the contact region between nanosheets and the tunneling conductivity of samples are evaluated with developed equations superior to those previously reported.The contact resistance and nanocomposite conductivity are modeled by several influencing factors,including stack properties,interphase depth,tunneling size,and contact diameter.The developed model's accuracy is verified through numerous experimental measurements.To further validate the models and establish correlations between parameters,the effects of all the variables on contact resistance and nanocomposite conductivity are analyzed.Notably,the contact resistance is primarily dependent on the polymer tunnel resistivity,contact area,and tunneling size.The dimensions of the graphene nanosheets significantly influence the conductivity,which ranges from 0 S/m to90 S/m.An increased number of nanosheets in stacks and a larger gap between them enhance the nanocomposite's conductivity.Furthermore,the thicker interphase and smaller tunneling size can lead to higher sample conductivity due to their optimistic effects on the percolation threshold and network efficacy.

Simulating traumatic brain injury in vitro:developing high throughput models to test biomaterial based therapies

Traumatic brain injuries are serious clinical incidents associated with some of the poorest outcomes in neurological practice.Coupled with the limited regenerative capacity of the brain,this has significant implications for patients,carers,and healthcare systems,and the requirement for life-long care in some cases.Clinical treatment currently focuses on limiting the initial neural damage with longterm care/support from multidisciplinary teams.Therapies targeting neuroprotection and neural regeneration are not currently available but are the focus of intensive research.Biomaterial-based interventions are gaining popularity for a range of applications including biomolecule and drug delive ry,and to function as cellular scaffolds.Experimental investigations into the development of such novel therapeutics for traumatic brain injury will be critically underpinned by the availability of appropriate high thro ughput,facile,ethically viable,and pathomimetic biological model systems.This represents a significant challenge for researchers given the pathological complexity of traumatic brain injury.Specifically,there is a concerted post-injury response mounted by multiple neural cell types which includes microglial activation and astroglial scarring with the expression of a range of growth inhibito ry molecules and cytokines in the lesion environment.Here,we review common models used for the study of traumatic brain injury(ranging from live animal models to in vitro systems),focusing on penetrating traumatic brain injury models.We discuss their relative advantages and drawbacks for the developmental testing of biomaterial-based therapies.

Chitosan conduits enriched with fibrin-collagen hydrogel with or without adipose-derived mesenchymal stem cells for the repair of 15-mm-long sciatic nerve defect

Hollow conduits of natural or synthetic origins have shown acceptable regeneration results in short nerve gap repair;however,results are still not comparable with the current gold standard technique“autografts”.Hollow conduits do not provide a successful regeneration outcome when it comes to critical nerve gap repair.Enriching the lumen of conduits with different extracellular materials and cells could provide a better biomimicry of the natural nerve regenerating environment and is expected to ameliorate the conduit performance.In this study,we evaluated nerve regeneration in vivo using hollow chitosan conduits or conduits enriched with fibrin-collagen hydrogels alone or with the further addition of adipose-derived mesenchymal stem cells in a 15 mm rat sciatic nerve transection model.Unexpected changes in the hydrogel consistency and structural stability in vivo led to a failure of nerve regeneration after 15 weeks.Nevertheless,the molecular assessment in the early regeneration phase(7,14,and 28 days)has shown an upregulation of useful regenerative genes in hydrogel enriched conduits compared with the hollow ones.Hydrogels composed of fibrin-collagen were able to upregulate the expression of soluble NRG1,a growth factor that plays an important role in Schwann cell transdifferentiation.The further enrichment with adipose-derived mesenchymal stem cells has led to the upregulation of other important genes such as ErbB2,VEGF-A,BDNF,c-Jun,and ATF3.

Peripheral nerve regeneration through nerve conduits evokes differential expression of growth-associated protein-43 in the spinal cord

Growth-associated protein 43 plays a key role in neurite outgrowth through cytoskeleton remodeling.We have previously demonstrated that structural damage of peripheral nerves induces growth-associated protein 43 upregulation to promote growth cone formation.Conversely,the limited regenerative capacity of the central nervous system due to an inhibitory environment prevents major changes in neurite outgrowth and should be presumably associated with low levels of growth-associated protein 43 expression.However,central alterations due to peripheral nerve damage have never been assessed using the growthassociated protein 43 marker.In this study,we used the tubulization technique to repair 1 cm-long nerve gaps in the rat nerve injury/repair model and detected growth-associated protein 43 expression in the peripheral and central nervous systems.First,histological analysis of the regeneration process confirmed an active regeneration process of the nerve gaps through the conduit from 10 days onwards.The growth-associated protein 43 expression profile varied across regions and follow-up times,from a localized expression to an abundant and consistent expression throughout the regeneration tissue,confirming the presence of an active nerve regeneration process.Second,spinal cord changes were also histologically assessed,and no apparent changes in the structural and cellular organization were observed using routine staining methods.Surprisingly,remarkable differences and local changes appeared in growth-associated protein 43 expression at the spinal cord level,in particular at 20 days post-repair and beyond.Growth-associated protein 43 protein was first localized in the gracile fasciculus and was homogeneously distributed in the left posterior cord.These findings differed from the growth-associated protein 43 pattern observed in the healthy control,which did not express growth-associated protein 43 at these levels.Our results revealed a differential expression in growth-associated protein 43 protein not only in the regenerating nerve tissue but also in the spinal cord after peripheral nerve transection.These findings open the possibility of using this marker to monitor changes in the central nervous system after peripheral nerve injury.

掺锌羟基磷灰石-沸石/聚己内酯复合材料作为镁基体涂层提高其体外腐蚀和抗菌性能

开发掺锌羟基磷灰石-沸石(Zn HA-Zeo)和聚己内酯(PCL)复合涂层,以提高镁基体的耐腐蚀性能和抗菌性能。采用浸涂技术在室温下将Zn HA-Zeo/PCL涂覆在镁基体上,对样品进行场发射扫描电子显微镜(FESEM)、X射线衍射(XRD)、傅里叶变换红外光谱(FTIR)、能量色散X射线光谱(EDX)和抗菌电位测试。结果表明,复合涂层由羟基磷灰石(HA)、磷钙锌石(scholzite)、沸石和PCL相组成。EDX谱显示钙(Ca)、硅(Si)、铝(Al)、锌(Zn)、磷(P)和氧(O)元素的存在。复合涂层表面的显微形貌为球形,厚度为226~260μm。掺锌HA-Zeo复合涂层具有较高的耐蚀性,对镁基体表面提供足够的保护,使其不被电流腐蚀。与未掺杂的涂层相比,掺锌HA-Zeo涂层对大肠杆菌具有更好的抗菌活性。综合结果表明,Zn HA-Zeo涂层不仅具有良好的耐腐蚀性能,而且具有良好的抗菌性能,可作为种植体应用的理想材料。

Mathematical modeling of drug release from biodegradable polymeric microneedles

Transdermal drug delivery systems have overcome many limitations of other drug administration routes,such as injection pain and first-pass metabolism following oral route,although transdermal drug delivery systems are limited to drugs with low molecular weight.Hence,new emerging technology allowing high molecular weight drug delivery across the skin—known as‘microneedles’—has been developed,which creates microchannels that facilitate drug delivery.In this report,drug-loaded degradable conic microneedles are modeled to characterize the degradation rate and drug release profile.Since a lot of data are available for polylactic acid-co-glycolic acid(PLGA)degradation in the literature,PLGA of various molecular weights-as a biodegradable polymer in the polyester family-is used for modeling and verification of the drug delivery in themicroneedles.The main reaction occurring during polyester degradation is hydrolysis of steric bonds,leading to molecular weight reduction.The acid produced in the degradation has a catalytic effect on the reaction.Changes in water,acid and steric bond concentrations over time and for different radii of microneedles are investigated.To solve the partial and ordinary differential equations simultaneously,finite difference and Runge–Kutta methods are employed,respectively,with the aid of MATLAB.Correlation of the polymer degradation rate with its molecular weight and molecular weight changes versus time are illustrated.Also,drug diffusivity is related to matrix molecular weight.The molecular weight reduction and accumulative drug release within the system are predicted.In order to validate and assess the proposed model,data series of the hydrolytic degradation of aspirin(180.16 Da)-and albumin(66,000 Da)-loaded PLGA(1:1 molar ratio)are used for comparison.The proposed model is in good agreement with experimental data from the literature.Considering diffusion as themain phenomena and autocatalytic effects in the reaction,the drug release profile is predicted.Based on our results for a microneedle containing drug,we are able to estimate drug release rates before fabrication.

Fluid Flow and Sub-Bactericidal Release of Silver from Organic Nanocomposite Coatings Enhance <i>ica</i>Operon Expression in <i>Staphylococcus epidermidis</i>

The present study investigates the effect of a silver (Ag)-containing nanocomposite coating on Staphylococcus epidermidis adhesion and icaA gene expression. Bacterial interactions with organic coatings with and without Ag nanoclusters were assessed through a combination of both conventional phenotypic analysis, using microscopy, and genotypic analysis, using the relative reverse transcription Real-Time Polymerase Chain Reaction (RT-PCR). The results suggest that the incorporation of Ag in organic coatings can significantly decrease bacterial adhesion and viability with time, in comparison to the organic coating alone. The initial Ag release though at concentrations lower than the bactericidal, significantly increased icaA gene expression for the bacteria interacting with the Ag containing coating two hours post adhesion, especially under the higher shear rate. Stress-inducing conditions such as sub-bactericidal concentrations of Ag and high shear rate can therefore increase icaA expression, indicating that analysis of gene expression can not only refine our knowledge of bacterial-material interactions, but also yield novel biomarkers for potential use in assessing biomaterials antimicrobial performance.

Cellular interplay to 3D in vitro microphysiological disease model:cell patterning microbiota-gut-brain axis

The microbiota-gut-brain axis(MGBA)has emerged as a key prospect in the bidirectional communication between two major organ systems:the brain and the gut.Homeostasis between the two organ systems allows the body to function without disease,whereas dysbiosis has long-standing evidence of etiopathological conditions.The most common communication paths are the microbial release of metabolites,soluble neurotransmitters,and immune cells.However,each pathway is intertwined with a complex one.With the emergence of in vitro models and the popularity of three-dimensional(3D)cultures and Transwells,engineering has become easier for the scientific understanding of neurodegenerative diseases.This paper briefly retraces the possible communication pathways between the gut microbiome and the brain.It further elaborates on three major diseases:autism spectrum disorder,Parkinson’s disease,and Alzheimer’s disease,which are prevalent in children and the elderly.These diseases also decrease patients’quality of life.Hence,understanding them more deeply with respect to current advances in in vitro modeling is crucial for understanding the diseases.Remodeling of MGBA in the laboratory uses many molecular technologies and biomaterial advances.Spheroids and organoids provide a more realistic picture of the cell and tissue structure than monolayers.Combining them with the Transwell system offers the advantage of compartmentalizing the two systems(apical and basal)while allowing physical and chemical cues between them.Cutting-edge technologies,such as bioprinting and microfluidic chips,might be the future of in vitro modeling,as they provide dynamicity.

Important parameters in plasma jets for the production of RONS in liquids for plasma medicine: A brief review

Reactive oxygen and nitrogen species (RONS) are among the key factors in plasma medicine. They are generated by atmospheric plasmas in biological fluids, living tissues and in a variety of liquids. This ability of plasmas to create a delicate mix of RONS in liquids has been used to design remote or indirect treatments for oncological therapy by treating biological fluids by plasmas and putting them in contact with the tumour;Documented effects include selective cancer cell toxicity, even though the exact mechanisms involved are still under investigation. However, the "right" dose for suitable therapeutical activity is crucial and still under debate. The wide variety of plasma sources hampers comparisons. This review focuses on atmospheric pressure plasma jets as the most studied plasma devices in plasma medicine and compiles the conditions employed to generate RONS in relevant liquids and the concentration ranges obtained. The concentrations of H2O2, NO2^-, NO3^- and short-lived oxygen species are compared critically to provide a useful overview for the reader.

Powder metallurgy with space holder for porous titanium implants:A review

One of the biggest challenges in the biocompatibility of implantable metals is the prevention of the stress shielding effect,which is related to the coupling of the bone-metal mechanical properties.This stress shielding phenomenon provokes bone resorption and the consequent adverse effects on prosthesis fixation.However,it can be inhibited by adapting the stiffness of the implant material.Since the use of titanium(Ti)porous structures is a great alternative not only to inhibit this effect but also to improve the osteointegration of orthopedic and dental implants,a brief description of the techniques used for their manufacturing and a review of the current commercialized implants produced from porous Ti assemblies are compiled in this work.As powder metallurgy(PM)with space holder(SH)is a powerful technology used to produce porous Ti structures,it is here discussed its potential for the fabrication of medical devices from the perspectives of both design and manufacture.The most important parameters of the technique such as the size and shape of the initial metallic particles,the SH and binder type of materials,the compaction pressure of the green form,and in the sintering stage,the temperature,atmosphere,and time are reviewed according to the bibliography reported.Furthermore,the importance of the porosity and its types together with the influence of the mentioned parameters in the final porosity and,consequently,in the ultimate mechanical properties of the structure are discussed.Finally,a few examples of the PM-SH application for the manufacturing of orthopedic implants are presented.

Bioactive Materials:A Comprehensive Review on Interactions with Biological Microenvironment Based on the Immune Response

Application of“bioactive materials”,as a modified version of biomaterials,can optimize the response of the biological system due to their surface reactivity and formation of strong interactions with the adjacent tissue upon implantation.However,choosing an appropriate bioactive material that suits to the application and provides the desired mechanical,physical,chemical and biological functionality,as well as understanding the aspects of biological reaction to the biomaterial,in particular immune response,it plays a key role in successful integration of the implant.In this review,we will discuss different bioactive materials including bioactive ceramics,polymers and composites and their applications in drug delivery and scaffold preparation in order to provide an adequate introduction to the recent studies.Considering the necessity of regulation of implant fate for higher biocompatibility,the comprehensive overview to the immune response will be reviewed with the focus on representing the cell-biomaterial interactions and more importantly,the inflammatory responses.Ultimately,we will also discuss about different approaches namely as immunomodulation to elicit the desired physiochemical properties and mimicking native cellular response using bioactive compounds,functionalizing the implant surface with active molecules and alteration of the surface morphology.With better understanding of bioactive materials and their interactions with body,more novel biomaterials representing desired properties can be designed.

Transplantation of engineered exosomes derived from bone marrow mesenchymal stromal cells ameliorate diabetic peripheral neuropathy under electrical stimulation

Diabetic peripheral neuropathy(DPN)is a long-term complication associated with nerve dysfunction and uncontrolled hyperglycemia.In spite of new drug discoveries,development of effective therapy is much needed to cure DPN.Here,we have developed a combinatorial approach to provide biochemical and electrical cues,considered to be important for nerve regeneration.Exosomes derived from bone marrow mesenchymal stromal cells(BMSCs)were fused with polypyrrole nanoparticles(PpyNps)containing liposomes to deliver both the cues in a single delivery vehicle.We developed DPN rat model and injected intramuscularly the fused exosomal system to understand its long-term therapeutic effect.We found that the fused system along with electrical stimulation normalized the nerve conduction velocity(57.60±0.45 m/s)and compound muscle action potential(16.96±0.73 mV)similar to healthy control(58.53±1.10 m/s;18.19±1.45 mV).Gastrocnemius muscle morphology,muscle mass,and integrity were recovered after treatment.Interestingly,we also observed paracrine effect of delivered exosomes in controlling hyperglycemia and loss in body weight and also showed attenuation of damage to the tissues such as the pancreas,kidney,and liver.This work provides a promising effective treatment and also contribute cutting edge therapeutic approach for the treatment of DPN.

Zn-Mg and Zn-Cu alloys for stenting applications:From nanoscale mechanical characterization to in vitro degradation and biocompatibility

In the recent decades,zinc(Zn)and its alloys have been drawing attention as promising candidates for bioresorbable cardiovascular stents due to its degradation rate more suitable than magnesium(Mg)and iron(Fe)alloys.However,its mechanical properties need to be improved in order to meet the criteria for vascular stents.This work investigates the mechanical properties,biodegradability and biocompatibility of Zn-Mg and Zn-Cu alloys in order to determine a proper alloy composition for optimal stent performance.Nanoindentation measurements are performed to characterize the mechanical properties at the nanoscale as a function of the Zn microstructure variations induced by alloying.The biodegradation mechanisms are discussed and correlated to microstructure,mechanical performance and bacterial/cell response.Addition of Mg or Cu alloying elements refined the microstructure of Zn and enhanced yield strength(YS)and ultimate tensile strength(UTS)proportional to the volume fraction of secondary phases.Zn-1Mg showed the higher YS and UTS and better performance in terms of degradation stability in Hanks’solution.Zn-Cu alloys presented an antibacterial effect for S.aureus controlled by diffusion mechanisms and by contact.Biocompatibility was dependent on the degradation rate and the nature of the corrosion products.

Novel synthesis method combining a foaming agent with freeze-drying to obtain hybrid highly macroporous bone scaffolds

Three-dimensional macroporous scaffolds are commonly used in bone tissue engineering applications since they provide sufficient space for cell migration and proliferation, facilitating bone ingrowth and implant vascularisation. The aim of this work was to combine two simple methods, freeze-drying and gas-foaming, in order to fabricate highly macroporous bone scaffolds made of chitosan/agarose matrix reinforced with nanohydroxyapatite. The secondary goal of this research was to comprehensively assess biomedical potential of developed biomaterials. In this work, it was demonstrated that simultaneous application of freeze-drying and gas-foaming technique allows to obtain hybrid(as proven by ATR-FTIR)macroporous bone scaffolds(pore diameter > 50 um) characterized by high open(70%) and interconnected porosity. Novel scaffolds were non-toxic, favoured osteoblasts adhesion and growth and induced apatite formation on their surfaces, indicating their high bioactivity that is essential for good implant osseointegration. Biomaterials were also prone to enzymatic degradation, degradation in acidified microenvironment(e.g. osteoclast-mediated), and slow degradation under physiological p H of 7.4.Moreover, the scaffolds revealed microstructure(70% open porosity, SSA approx. 30 m2/g, high share of macropores with diameter in the range 100-410 um) and compressive strength(1–1.4 MPa) comparable to cancellous bone, indicating that they are promising implants for cancellous bone regeneration.