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The recovery of injured nervous tissue, one of the main goals for regenerative therapeutic approaches, is often hindered by the limited axonal regeneration ability of the central nervous system (CNS). In this regard, the identification of scaffolds that support the reconstruction of functional neuronal tissues and guide the alignment of regenerating neurons is a major challenge in tissue engineering. Ideally, the usage of such scaffolds would promote and guide the axonal growth, a crucial phase for the restoration of neuronal connections and, consequently, the nerve function. Among the materials proposed as scaffolds for CNS regeneration, silk has been used to exploit its outstanding features as a biomaterial to promote axonal regeneration. In this study, we explore, for the first time, the possibility of using high-performance regenerated silk fibers obtained by straining flow spinning (SFS) to serve as scaffolds for inducing and guiding the axonal growth. It is shown that SFS fibers promote the spontaneous organization of dissociated cortical primary cells into highly interconnected cellular spheroid-like tissue formations. Neuronal projections (i.e., axons) from these cellular spheroids span hundreds of microns along the SFS fibers that act as guides and allow the connection of distant spheroids. In addition, it is also shown that SFS fibers serve as scaffolds for neuronal migration covering short and long distances. As a consequence, the usage of high-performance SFS fibers appears as a promising basis for the development of novel therapies, leading to directed axonal regeneration., Comunidad de Madrid (S2017/BMD-3760), Ministerio de Ciencia e Innovación (MAT2016-79832-R), Depto. de Biodiversidad, Ecología y Evolución, Centro de Vigilancia Sanitaria Veterinaria (VISAVET), TRUE, pub

The true stress-true strain curves of 11 Australian spider species from the Entelegynae lineage were tensile tested and classified based on the values of the alignment parameter, alpha*, in the framework of the Spider Silk Standardization Initiative (S3I). The application of the S3I methodology allowed the determination of the alignment parameter in all cases, and were found to range between alpha* = 0.03 and alpha* = 0.65. These data, in combination with previous results on other species included in the Initiative, were exploited to illustrate the potential of this approach by testing two simple hypotheses on the distribution of the alignment parameter throughout the lineage: (1) whether a uniform distribution may be compatible with the values obtained from the studied species, and (2) whether any trend may be established between the distribution of the alpha* parameter and phylogeny. In this regard, the lowest values of the alpha* parameter are found in some representatives of the Araneidae group, and larger values seem to be found as the evolutionary distance from this group increases. However, a significant number of outliers to this apparent general trend in terms of the values of the alpha* parameter are described.

Titanium (Ti-6Al-4V) substrates were functionalized through the covalent binding of fibronectin, and the effect of the existence of this extracellular matrix protein on the surface of the material was assessed by employing mesenchymal stem cell (MSC) cultures. The functionalization process comprised the usage of the activation vapor silanization (AVS) technique to deposit a thin film with a high surface density of amine groups on the material, followed by the covalent binding of fibronectin to the amine groups using the N-(3-dimethylaminopropyl)-N & PRIME;-ethylcarbodiimide hydrochloride/N-hydroxysuccinimide (EDC/NHS) crosslinking chemistry. The biological effect of the fibronectin on murine MSCs was assessed in vitro. It was found that functionalized samples not only showed enhanced initial cell adhesion compared with bare titanium, but also a three-fold increase in the cell area, reaching values comparable to those found on the polystyrene controls. These results provide compelling evidence of the potential to modulate the response of the organism to an implant through the covalent binding of extracellular matrix proteins on the prosthesis.

The adhesion forces of cells to peptide-coated functionalized materials were assessed through the Single Cell Force Spectroscopy (SCFS) technique in order to develop a methodology that allows the fast selection of peptide motifs that favor the interaction between cells and the biomaterial. Borosilicate glasses were functionalized using the activated vapor silanization process (AVS) and subsequently decorated with an RGD- containing peptide using the EDC/NHS crosslinking chemistry.It is shown that the RGD-coated glass induces larger attachment forces on mesenchymal stem cell cultures (MSCs), compared to the bare glass substrates. These higher forces correlate well with the enhanced adhesion of the MSCs observed on RGD-coated substrates through conventional adhesion cell cultures and inverse centrifugation tests. The methodology based on the SCFS technique presented in this work constitutes a fast procedure for the screening of new peptides or their combinations to select candidates that may enhance the response of the organism to the implant of the functionalized biomaterials.

After an injury, the limited regenerative capacity of the central nervous system makes the reconnection and functional recovery of the affected nervous tissue almost impossible. To address this problem, biomaterials appear as a promising option for the design of scaffolds that promote and guide this regenerative process. Based on previous seminal works on the ability of regenerated silk fibroin fibers spun through the straining flow spinning (SFS) technique, this study is intended to show that the usage of functionalized SFS fibers allows an enhancement of the guidance ability of the material when compared with the control (nonfunctionalized) fibers. It is shown that the axons of the neurons not only tend to follow the path marked by the fibers, in contrast to the isotropic growth observed on conventional culture plates, but also that this guidance can be further modulated through the biofunctionalization of the material with adhesion peptides. Establishing the guidance ability of these fibers opens the possibility of their use as implants for spinal cord injuries, so that they may represent the core of a therapy that would allow the reconnection of the injured ends of the spinal cord.

Central nervous system (CNS) diseases represent an extreme burden with significant social and economic costs. A common link in most brain pathologies is the appearance of inflammatory components that can jeopardize the stability of the implanted biomaterials and the effectiveness of therapies. Different silk fibroin scaffolds have been used in applications related to CNS disorders. Although some studies have analyzed the degradability of silk fibroin in non-cerebral tissues (almost exclusively upon non-inflammatory conditions), the stability of silk hydrogel scaffolds in the inflammatory nervous system has not been studied in depth. In this study, the stability of silk fibroin hydrogels exposed to different neuroinflammatory contexts has been explored using an in vitro microglial cell culture and two in vivo pathological models of cerebral stroke and Alzheimer's disease. This biomaterial was relatively stable and did not show signs of extensive degradation across time after implantation and during two weeks of in vivo analysis. This finding contrasted with the rapid degradation observed under the same in vivo conditions for other natural materials such as collagen. Our results support the suitability of silk fibroin hydrogels for intracerebral applications and highlight the potentiality of this vehicle for the release of molecules and cells for acute and chronic treatments in cerebral pathologies.

Great effort has been devoted to the synthesis of novel multi-target directed tacrine derivatives in the search of new treatments for Alzheimer’s disease (AD). Herein we describe the proof of concept of MBA121, a compound designed as a tacrine–ferulic acid hybrid, and its potential use in the therapy of AD. MBA121 shows good β-amyloid (Aβ) anti-aggregation properties, selective inhibition of human butyrylcholinesterase, good neuroprotection against toxic insults, such as Aβ1–40, Aβ1–42, and H2O2, and promising ADMET properties that support translational developments. A passive avoidance task in mice with experimentally induced amnesia was carried out, MBA121 being able to significantly decrease scopolamine-induced learning deficits. In addition, MBA121 reduced the Aβ plaque burden in the cerebral cortex and hippocampus in APPswe/PS1ΔE9 transgenic male mice. Our in vivo results relate its bioavailability with the therapeutic response, demonstrating that MBA121 is a promising agent to treat the cognitive decline and neurodegeneration underlying AD.

Objective: Ischemic stroke is one of the main causes of death and disability worldwide and currently has limited treatment options. Electroencephalography (EEG) signals are significantly affected in stroke patients during the acute stage. In this study, we preclinically characterized the brain electrical rhythms and seizure activity during the hyperacute and late acute phases in a hemispheric stroke model with no reperfusion. Methods: EEG signals and seizures were studied in a model of hemispheric infarction induced by permanent occlusion of the middle cerebral artery (pMCAO), which mimics the clinical condition of stroke patients with permanent ischemia. Electrical brain activity was also examined using a photothrombotic (PT) stroke model. In the PT model, we induced a similar (PT group-1) or smaller (PT group-2) cortical lesion than in the pMCAO model. For all models, we used a nonconsanguineous mouse strain that mimics human diversity and genetic variation. Results: The pMCAO hemispheric stroke model exhibited thalamic-origin nonconvulsive seizures during the hyperacute stage that propagated to the thalamus and cortex. The seizures were also accompanied by progressive slowing of the EEG signal during the acute phase, with elevated delta/theta, delta/alpha, and delta/beta ratios. Cortical seizures were also confirmed in the PT stroke model of similar lesions as in the pMCAO model, but not in the PT model of smaller injuries. Significance: In the clinically relevant pMCAO model, poststroke seizures and EEG abnormalities were inferred from recordings of the contralateral hemisphere (noninfarcted hemisphere), emphasizing the reciprocity of interhemispheric connections and that injuries affecting one hemisphere had consequences for the other. Our results recapitulate many of the EEG signal hallmarks seen in stroke patients, thereby validating this specific mouse model for the examination of the mechanistic aspects of brain function and for the exploration of the rever

Brain stroke is a highly prevalent pathology and a main cause of disability among older adults. If not promptly treated with recanalization therapies, primary and secondary mechanisms of injury contribute to an increase in the lesion, enhancing neurological deficits. Targeting excitotoxicity and oxidative stress are very promising approaches, but only a few compounds have reached the clinic with relatively good positive outcomes. The exploration of novel targets might overcome the lack of clinical translation of previous efficient preclinical neuroprotective treatments. In this study, we examined the neuroprotective properties of 2-aminoethoxydiphenyl borate (2-APB), a molecule that interferes with intracellular calcium dynamics by the antagonization of several channels and receptors. In a permanent model of cerebral ischemia, we showed that 2-APB reduces the extent of the damage and preserves the functionality of the cortical territory, as evaluated by somatosensory evoked potentials (SSEPs). While in this permanent ischemia model, the neuroprotective effect exerted by the antioxidant scavenger cholesteronitrone F2 was associated with a reduction in reactive oxygen species (ROS) and better neuronal survival in the penumbra, 2-APB did not modify the inflammatory response or decrease the content of ROS and was mostly associated with a shortening of peri-infarct depolarizations, which translated into better cerebral blood perfusion in the penumbra. Our study highlights the potential of 2-APB to target spreading depolarization events and their associated inverse hemodynamic changes, which mainly contribute to extension of the area of lesion in cerebrovascular pathologies.

High-performance regenerated silkworm (Bombyx mori) silk fibers can be produced efficiently through the straining flow spinning (SFS) technique. In addition to an enhanced biocompatibility that results from the removal of contaminants during the processing of the material, regenerated silk fibers may be functionalized conveniently by using a range of different strategies. In this work, the possibility of implementing various functionalization techniques is explored, including the production of fluorescent fibers that may be tracked when implanted, the combination of the fibers with enzymes to yield fibers with catalytic properties, and the functionalization of the fibers with cell-adhesion motifs to modulate the adherence of different cell lineages to the material. When considered globally, all these techniques are a strong indication not only of the high versatility offered by the functionalization of regenerated fibers in terms of the different chemistries that can be employed, but also on the wide range of applications that can be covered with these functionalized fibers.

Titanium implants are widely used in traumatology and various orthopedic fields. Titanium and other metallic-based implants have limited structural and functional integration into the body, which translates into progressive prosthesis instability and the need for new surgical interventions that have enormous social and economic impacts. To enhance the biocompatibility of titanium implants, numerous biofunctionalization strategies have been developed. However, the problem persists, as more than 70% of implant failures are due to aseptic loosening. In this study we addressed the problem of improving the physiological engraftability and acceptability of titanium-based implants by applying a robust and versatile functionalization method based on the covalent immobilization of extracellular matrix (ECM)derived oligopeptides on Ti-6Al-4V surfaces treated by activated vapor silanization (AVS). The feasibility of this technique was evaluated with two oligopeptides of different structures and compositions. These oligopeptides were immobilized on Ti-6Al-4V substrates by a combination of AVS and N-(3-dimethylaminopropyl)-N???-ethylcarbodiimide hydrochloride/N-hydroxysuccinimide (EDC/NHS) crosslinking chemistry. The immobilization was shown to be stable and resistant to chemical denaturing upon sodium dodecyl sulfate treatment. On Ti-6Al-4V surfaces both peptides increased the attachment, spreading, rearrangement and directional growth of mesenchymal stem and progenitor cells (MSC) with chondro- and osteo-regenerative capacities. We also found that this biofunctionalization method (AVS-EDC/NHS) increased the attachment capacity of an immortalized cell line of neural origin with poor adhesive properties, highlighting the versatility and robustness of this method in terms of potential oligopeptides that may be used, and cell lineages whose anchorage to the biomaterial may be enhanced. Collectively, this novel functionalization strategy can accelerate the development of advanced p

Silk gut fibers were produced from the silkworm Samia cynthia ricini silk glands by the usual procedure of immersion in a mildly acidic solution and subsequent stretching. The morphology of the silk guts was assessed by scanning electron microscopy, and their microstructure was assessed by infrared spectroscopy and X-ray diffraction. It was found that both naturally spun and Samia silk guts share a common semicrystalline microstructure. The mechanical characterization of the silk guts revealed that these fibers show an elastomeric behavior when tested in water, and exhibit a genuine ground state to which the fiber may revert independently of its previous loading history. In spite of its large cross-sectional area compared with naturally spun silk fibers, Samia silk guts show values of work to fracture up to 160 MJ m(-3), much larger than those of most of their natural counterparts, and establish a new record value for this parameter in silk guts.

Nitrones are encouraging drug candidates for the treatment of oxidative stress-driven diseases such as acute ischemic stroke (AIS). In a previous study, we found a promising quinolylnitrone, QN23, which exerted a neuroprotective effect in neuronal cell cultures subjected to oxygen–glucose deprivation and in experimental models of cerebral ischemia. In this paper, we update the biological and pharmacological characterization of QN23. We describe the suitability of intravenous administration of QN23 to induce neuroprotection in transitory four-vessel occlusion (4VO) and middle cerebral artery occlusion (tMCAO) experimental models of brain ischemia by assessing neuronal death, apoptosis induction, and infarct area, as well as neurofunctional outcomes. QN23 significantly decreased the neuronal death and apoptosis induced by the ischemic episode in a dose-dependent manner and showed a therapeutic effect when administered up to 3 h after post-ischemic reperfusion onset, effects that remained 11 weeks after the ischemic episode. In addition, QN23 significantly reduced infarct volume, thus recovering the motor function in a tMCAO model. Remarkably, we assessed the antioxidant activity of QN23 in vivo using dihydroethidium as a molecular probe for radical species. Finally, we describe QN23 pharmacokinetic parameters. All these results pointing to QN23 as an interesting and promising preclinical candidate for the treatment of AIS.

Silk gut fibers were produced from the silkworm Samia cynthia ricini silk glands by the usual procedure of immersion in a mildly acidic solution and subsequent stretching. The morphology of the silk guts was assessed by scanning electron microscopy, and their microstructure was assessed by infrared spectroscopy and X-ray diffraction. It was found that both naturally spun and Samia silk guts share a common semicrystalline microstructure. The mechanical characterization of the silk guts revealed that these fibers show an elastomeric behavior when tested in water, and exhibit a genuine ground state to which the fiber may revert independently of its previous loading history. In spite of its large cross-sectional area compared with naturally spun silk fibers, Samia silk guts show values of work to fracture up to 160 MJ m, much larger than those of most of their natural counterparts, and establish a new record value for this parameter in silk guts.

Brain stroke is a highly prevalent pathology and a main cause of disability among older adults. If not promptly treated with recanalization therapies, primary and secondary mechanisms of injury contribute to an increase in the lesion, enhancing neurological deficits. Targeting excitotox-icity and oxidative stress are very promising approaches, but only a few compounds have reached the clinic with relatively good positive outcomes. The exploration of novel targets might overcome the lack of clinical translation of previous efficient preclinical neuroprotective treatments. In this study, we examined the neuroprotective properties of 2-aminoethoxydiphenyl borate (2-APB), a molecule that interferes with intracellular calcium dynamics by the antagonization of several channels and receptors. In a permanent model of cerebral ischemia, we showed that 2-APB reduces the extent of the damage and preserves the functionality of the cortical territory, as evaluated by soma-tosensory evoked potentials (SSEPs). While in this permanent ischemia model, the neuroprotective effect exerted by the antioxidant scavenger cholesteronitrone F2 was associated with a reduction in reactive oxygen species (ROS) and better neuronal survival in the penumbra, 2-APB did not modify the inflammatory response or decrease the content of ROS and was mostly associated with a shortening of peri-infarct depolarizations, which translated into better cerebral blood perfusion in the penumbra. Our study highlights the potential of 2-APB to target spreading depolarization events and their associated inverse hemodynamic changes, which mainly contribute to extension of the area of lesion in cerebrovascular pathologies.

Silk refers to a family of natural fibers spun by several species of invertebrates such as spiders and silkworms. In particular, silkworm silk, the silk spun by Bombyx mori larvae, has been primarily used in the textile industry and in clinical settings as a main component of sutures for tissue repairing and wound ligation. The biocompatibility, remarkable mechanical performance, controllable degradation, and the possibility of producing silk-based materials in several formats, have laid the basic principles that have triggered and extended the use of this material in regenerative medicine. The field of neural soft tissue engineering is not an exception, as it has taken advantage of the properties of silk to promote neuronal growth and nerve guidance. In addition, silk has notable intrinsic properties and the by-products derived from its degradation show anti-inflammatory and antioxidant properties. Finally, this material can be employed for the controlled release of factors and drugs, as well as for the encapsulation and implantation of exogenous stem and progenitor cells with therapeutic capacity. In this article, we review the state of the art on manufacturing methodologies and properties of fiber-based and non-fiber-based formats, as well as the application of silk-based biomaterials to neuroprotect and regenerate the damaged nervous system. We review previous studies that strategically have used silk to enhance therapeutics dealing with highly prevalent central and peripheral disorders such as stroke, Alzheimer's disease, Parkinson's disease, and peripheral trauma. Finally, we discuss previous research focused on the modification of this biomaterial, through biofunctionalization techniques and/or the creation of novel composite formulations, that aim to transform silk, beyond its natural performance, into more efficient silk-based-polymers towards the clinical arena of neuroprotection and regeneration in nervous system diseases.

Age-related macular degeneration is an incurable chronic neurodegenerative disease, causing progressive loss of the central vision and even blindness. Up-to-date therapeutic approaches can only slow down he progression of the disease. Objective. Feasibility study for a multilayered, silk fibroin-based, 3D biohybrid retina. Approach. Fabrication of silk fibroin-based biofilms; culture of different types of cells: retinal pigment epithelium, retinal neurons, Muller and mesenchymal stem cells ; creation of a layered structure glued with silk fibroin hydrogel. Main results. In vitro evidence for the feasibility of layered 3D biohybrid retinas; primary culture neurons grow and develop neurites on silk fibroin biofilms, either alone or in presence of other cells cultivated on the same biomaterial; cell organization and cellular phenotypes are maintained in vitro for the seven days of the experiment. Significance. 3D biohybrid retina can be built using silk silkworm fibroin films and hydrogels to be used in cell replacement therapy for AMD and similar retinal neurodegenerative diseases.

Age-related Macular Degeneration (AMD) is an up-to-date untreatable chronic neurodegenerative eye disease of multifactorial origin, and the main causes of blindness in over 65 years old people. It is characterized by a slow progression and the presence of a multitude of factors, highlighting those related to diet, genetic heritage and environmental conditions, present throughout each of the stages of the illness. Current therapeutic approaches, mainly consisting of intraocular drug delivery, are only used for symptoms relief and/or to decelerate the progression of the disease. Furthermore, they are overly simplistic and ignore the complexity of the disease and the enormous differences in the symptomatology between patients. Due to the wide impact of the AMD and the up-to-date absence of clinical solutions, the development of biomaterials-based approaches for a personalized and controlled delivery of therapeutic drugs and biomolecules represents the main challenge for the defeat of this neurodegenerative disease. Here we present a critical review of the available and under development AMD therapeutic approaches, from a biomaterials and biotechnological point of view. We highlight benefits and limitations and we forecast forthcoming alternatives based on novel biomaterials and biotechnology methods. In the first part we expose the physiological and clinical aspects of the disease, focusing on the multiple factors that give origin to the disorder and highlighting the contribution of these factors to the triggering of each step of the disease. Then we analyze available and under development biomaterials-based drug-delivery devices (DDD), taking into account the anatomical and functional characteristics of the healthy and ill retinal tissue.

Age-related Macular Degeneration (AMD) is an up-to-date untreatable chronic neurodegenerative eye disease of multifactorial origin, and the main causes of blindness in over 65 y.o. people. It is characterized by a slow progression and the presence of a multitude of factors, highlighting those related to diet, genetic heritage and environmental conditions, present throughout each of the stages of the illness. Current therapeutic approaches, mainly consisting on intraocular drug delivery, are only used for symptoms relief and/or to decelerate the progression of the disease. Furthermore, they are overly simplistic and ignore the complexity of the disease and the enormous differences in the symptomatology between patients. Due to the wide impact of the AMD and the up-to-date absence of clinical solutions, Due to the wide impact of the AMD and the up-to-date absence of clinical solutions, different treatment options have to be considered. Cell therapy is a very promising alternative to drug-based approaches for AMD treatment. Cells delivered to the affected tissue as a suspension have shown poor retention and low survival rate. A solution to these inconveniences has been the encapsulation of these cells on biomaterials, which contrive to their protection, gives them support, and favor their retention of the desired area. We offer a two-papers critical review of the available and under development AMD therapeutic approaches, from a biomaterials and biotechnological point of view. We highlight benefits and limitations and we forecast forthcoming alternatives based on novel biomaterials and biotechnology methods. In this second part we review the preclinical and clinical cell-replacement approaches aiming at the development of efficient AMD-therapies, the employed cell types, as well as the cell-encapsulation and cell-implant systems. We discuss their advantages and disadvantages and how they could improve the survival and integration of the implanted cells.

Ischemic stroke represents one of the most prevalent pathologies in humans and is a leading cause of death and disability. Anti-thrombolytic therapy with tissue plasminogen activator (t-PA) and surgical thrombectomy are the primary treatments to recanalize occluded vessels and normalize the blood flow in ischemic and peri-ischemic regions. A large majority of stroke patients are refractory to treatment or are not eligible due to the narrow time window of therapeutic efficacy. In recent decades, we have significantly increased our knowledge of the molecular and cellular mechanisms that inexorably lead to progressive damage in infarcted and peri-lesional brain areas. As a result, promising neuroprotective targets have been identified and exploited in several stroke models. However, these considerable advances have been unsuccessful in clinical contexts. This lack of clinical translatability and the emerging use of biomaterials in different biomedical disciplines have contributed to developing a new class of biomaterial-based systems for the better control of drug delivery in cerebral disorders. These systems are based on specific polymer formulations structured in nanoparticles and hydrogels that can be administered through different routes and, in general, bring the concentrations of drugs to therapeutic levels for prolonged times. In this review, we first provide the general context of the molecular and cellular mechanisms impaired by cerebral ischemia, highlighting the role of excitotoxicity, inflammation, oxidative stress, and depolarization waves as the main pathways and targets to promote neuroprotection avoiding neuronal dysfunction. In the second part, we discuss the versatile role played by distinct biomaterials and formats to support the sustained administration of particular compounds to neuroprotect the cerebral tissue at risk of damage.

The regenerative capacity of the peripheral nervous system after an injury is limited, and a complete function is not recovered, mainly due to the loss of nerve tissue after the injury that causes a separation between the nerve ends and to the disorganized and intermingled growth of sensory and motor nerve fibers that cause erroneous reinnervations. Even though the development of biomaterials is a very promising field, today no significant results have been achieved. In this work, we study not only the characteristics that should have the support that will allow the growth of nerve fibers, but also the molecular profile necessary for a specific guidance. To do this, we carried out an exhaustive study of the molecular profile present during the regeneration of the sensory and motor fibers separately, as well as of the effect obtained by the administration and inhibition of different factors involved in the regeneration. In addition, we offer a complete design of the ideal characteristics of a biomaterial, which allows the growth of the sensory and motor neurons in a differentiated way, indicating (1) size and characteristics of the material; (2) necessity to act at the microlevel, on small groups of neurons; (3) combination of molecules and specific substrates; and (4) temporal profile of those molecules expression throughout the regeneration process. The importance of the design we offer is that it respects the complexity and characteristics of the regeneration process; it indicates the appropriate temporal conditions of molecular expression, in order to obtain a synergistic effect; it takes into account the importance of considering the process at the group of neuron level; and it gives an answer to the main limitations in the current studies.

The recovery of injured nervous tissue, one of the main goals for regenerative therapeutic approaches, is often hindered by the limited axonal regeneration ability of the central nervous system (CNS). In this regard, the identification of scaffolds that support the reconstruction of functional neuronal tissues and guide the alignment of regenerating neurons is a major challenge in tissue engineering. Ideally, the usage of such scaffolds would promote and guide the axonal growth, a crucial phase for the restoration of neuronal connections and, consequently, the nerve function. Among the materials proposed as scaffolds for CNS regeneration, silk has been used to exploit its outstanding features as a biomaterial to promote axonal regeneration. In this study, we explore, for the first time, the possibility of using high-performance regenerated silk fibers obtained by straining flow spinning (SFS) to serve as scaffolds for inducing and guiding the axonal growth. It is shown that SFS fibers promote the spontaneous organization of dissociated cortical primary cells into highly interconnected cellular spheroid-like tissue formations. Neuronal projections (i.e., axons) from these cellular spheroids span hundreds of microns along the SFS fibers that act as guides and allow the connection of distant spheroids. In addition, it is also shown that SFS fibers serve as scaffolds for neuronal migration covering short and long distances. As a consequence, the usage of high-performance SFS fibers appears as a promising basis for the development of novel therapies, leading to directed axonal regeneration.

Ischemic stroke represents one of the most prevalent pathologies in humans and is a leading cause of death and disability. Anti-thrombolytic therapy with tissue plasminogen activator (t-PA) and surgical thrombectomy are the primary treatments to recanalize occluded vessels and normalize the blood flow in ischemic and peri-ischemic regions. A large majority of stroke patients are refractory to treatment or are not eligible due to the narrow time window of therapeutic efficacy. In recent decades, we have significantly increased our knowledge of the molecular and cellular mechanisms that inexorably lead to progressive damage in infarcted and peri-lesional brain areas. As a result, promising neuroprotective targets have been identified and exploited in several stroke models. However, these considerable advances have been unsuccessful in clinical contexts. This lack of clinical translatability and the emerging use of biomaterials in different biomedical disciplines have contributed to developing a new class of biomaterial-based systems for the better control of drug delivery in cerebral disorders. These systems are based on specific polymer formulations structured in nanoparticles and hydrogels that can be administered through different routes and, in general, bring the concentrations of drugs to therapeutic levels for prolonged times. In this review, we first provide the general context of the molecular and cellular mechanisms impaired by cerebral ischemia, highlighting the role of excitotoxicity, inflammation, oxidative stress, and depolarization waves as the main pathways and targets to promote neuroprotection avoiding neuronal dysfunction. In the second part, we discuss the versatile role played by distinct biomaterials and formats to support the sustained administration of particular compounds to neuroprotect the cerebral tissue at risk of damage.

Physical and cognitive disabilities are hallmarks of a variety of neurological diseases. Stem cell-based therapies are promising solutions to neuroprotect and repair the injured brain and overcome the limited capacity of the central nervous system to recover from damage. It is widely accepted that most benefits of different exogenously transplanted stem cells rely on the secretion of different factors and biomolecules that modulate inflammation, cell death and repair processes in the damaged host tissue. However, few cells survive in cerebral tissue after transplantation, diminishing the therapeutic efficacy. As general rule, cell encapsulation in natural and artificial polymers increases the in vivo engraftment of the transplanted cells. However, we have ignored the consequences of such encapsulation on the secretory activity of these cells. In this study, we investigated the biological compatibility between silk fibroin hydrogels and stem cells of mesenchymal origin, a cell population that has gained increasing attention and popularity in regenerative medicine. Although the survival of mesenchymal stem cells was not affected inside hydrogels, this biomaterial format caused adhesion and proliferation deficits and impaired secretion of several angiogenic, chemoattractant and neurogenic factors while concurrently potentiating the anti-inflammatory capacity of this cell population through a massive release of TGF-Beta-1. Our results set a milestone for the exploration of engineering polymers to modulate the secretory activity of stem cell-based therapies for neurological disorders.

This work summarizes the main principles and some of the most significant results of straining flow spinning (SFS), a technology developed originally by the authors of this work. The principles on which the technology is based, inspired by the natural spinning system of silkworms and spiders, are presented, as well as some of the main achievements of the technique. Among these achievements, spinning under environmentally friendly conditions, obtaining high-performance fibers, and imparting the fibers with emerging properties such as supercontraction are discussed. Consequently, SFS appears as an efficient process that may represent one of the first realizations of a biomimetic technology with a significant impact at the production level.

Straining Flow Spinning is a versatile and robust spinning technique for the production of regeneratedsilkworm silkfibers using mild chemistries. However, reaching high values of tensile strength and strainat breaking requires a step ofwet-stretchingin water, which limits scalability and the practical usage ofthe technique. Here, we show that adding apost-spinning drawingstep to the procedure improves theperformance of thefibers, and allows the development of a scalable process. It is also shown that theproperties of thefiber can be tuned by varying the parameters of the post-spinning step. Finally,equivalence is established between the discrete wet-stretching process and the continuous post-spinning drawing step.

This study presents the development of an efficient procedure for covalently immobilizing collagen molecules on AVS-functionalized Ti-6Al-4V samples, and the assessment of the survival and proliferation of cells cultured on these substrates. Activated Vapor Silanization (AVS) is a versatile functionalization technique that allows obtaining a high density of active amine groups on the surface. A procedure is presented to covalently bind collagen to the functional layer using EDC/NHS as cross-linker. The covalently bound collagen proteins are characterized by fluorescence microscopy and atomic force microscopy and their stability is tested. The effect of the cross-linker concentration on the process is assessed. The concentration of the cross-linker is optimized and a reliable cleaning protocol is developed for the removal of the excess of carbodiimide from the samples. The results demonstrate that the covalent immobilization of collagen type I on Ti-6Al-4V substrates, using the optimized protocol, increases the number of viable cells present on the material. Consequently, AVS in combination with the carbodiimide chemistry appears as a robust method for the immobilization of proteins and, for the first time, it is shown that it can be used to enhance the biological response to the material.

This work summarizes the main principles and some of the most significant results of straining flow spinning (SFS), a technology developed originally by the authors of this work. The principles on which the technology is based, inspired by the natural spinning system of silkworms and spiders, are presented, as well as some of the main achievements of the technique. Among these achievements, spinning under environmentally friendly conditions, obtaining high-performance fibers, and imparting the fibers with emerging properties such as supercontraction are discussed. Consequently, SFS appears as an efficient process that may represent one of the first realizations of a biomimetic technology with a significant impact at the production level.

The use of advanced biomaterials as a structural and functional support for stem cells-based therapeutic implants has boosted the development of tissue engineering applications in multiple clinical fields. In relation to neurological disorders, we are still far from the clinical reality of restoring normal brain function in neurodegenerative diseases and cerebrovascular disorders. Hydrogel polymers show unique mechanical stiffness properties in the range of living soft tissues such as nervous tissue. Furthermore, the use of these polymers drastically enhances the engraftment of stem cells as well as their capacity to produce and deliver neuroprotective and neuroregenerative factors in the host tissue. Along this article, we review past and current trends in experimental and translational research to understand the opportunities, benefits, and types of tentative hydrogel-based applications for the treatment of cerebral disorders. Although the use of hydrogels for brain disorders has been restricted to the experimental area, the current level of knowledge anticipates an intense development of this field to reach clinics in forthcoming years.

The restitution of damaged circuitry and functional remodeling of peri-injured areas constitute two main mechanisms for sustaining recovery of the brain after stroke. In this study, a silk fibroin-based biomaterial efficiently supports the survival of intracerebrally implanted mesenchymal stem cells (mSCs) and increases functional outcomes over time in a model of cortical stroke that affects the forepaw sensory and motor representations. We show that the functional mechanisms underlying recovery are related to a substantial preservation of cortical tissue in the first days after mSCs-polymer implantation, followed by delayed cortical plasticity that involved a progressive functional disconnection between the forepaw sensory (FLs1) and caudal motor (cFLm1) representations and an emergent sensory activity in peri-lesional areas belonging to cFLm1. Our results provide evidence that mSCs integrated into silk fibroin hydrogels attenuate the cerebral damage after brain infarction inducing a delayed cortical plasticity in the peri-lesional tissue, this later a functional change described during spontaneous or training rehabilitation-induced recovery. This study shows that brain remapping and sustained recovery were experimentally favored using a stem cell-biomaterial-based approach.