No full-text available
To read the full-text of this research,
you can request a copy directly from the authors.
Early peripheral nerve healing in collagen and silicone tube implants: Myofibroblasts and the cellular response
Abstract
Injuries to peripheral nerves innervating a limb cause paralysis, and can necessitate amputation. The inability of the nerves to regenerate spontaneously and the limitations of autograft procedures led to the development of treatments involving insertion of the nerve ends into prosthetic tubular devices. Previous work showed that 'entubulation' of the nerve ends in a silicone tube containing a specific porous, resorbable collagen-GAG (CG) copolymer, serving as an analog of extracellular matrix, improved regeneration compared to an empty silicone tube. However, long-term treatment with silicone tubes produced constriction that caused partial degradation of the regenerated axons; for this and other reasons, implementation of a nondegradable tube may require a second surgical procedure for removal. In this study the silicone tube was replaced with porous and non-porous collagen tubes in order to produce fully degradable devices. CG-filled collagen tubes and controls (CG-filled silicone tubes and empty collagen and silicone tubes) were implanted in a 10-mm gap in the rat sciatic nerve, with three rats in each group. The regeneration was evaluated after six weeks using light microscope images of cross sections of the nerve that were digitized and analyzed. Histograms of the diameters of the axons were generated and compared. The cellular response to the implanted biomaterials was assessed histologically, and immunohistochemistry was performed using an antibody to alpha-smooth muscle actin in order to determine the presence of myofibroblasts (contractile cells). Axonal regrowth was comparable in porous collagen, non-porous collagen, and silicone tubes filled with a CG matrix. These results support the implementation of a degradable collagen tube in place of a silicone device. Confirming earlier work, regeneration through the silicone and collagen tubes was enhanced by the CG copolymer, compared to empty tubes. A notable finding was a continuous layer of myofibroblasts on the surfaces of all of the six silicone tube prostheses, but on the inner surface of only one of six collagen tubes (Fisher's exact tests; P < 0.01). This is the first report of contractile capsules around silicone tubes, and supports the use of degradable collagen tubes in peripheral nerve regeneration. Macrophages were found bordering both the silicone and collagen tubes, and in the case of the collagen tubes, appeared to be participating in the regulation of the tubes.
... If distal and proximal aspects of regeneration process are not connected, Schwann cells from distal stump and axons from proximal stump, after short migration (about 4-5 mm), usually stop to migrate. Schwann cells create cup-like structures covering distal stump [10,15,21,25,26]. Myelinization process between 14 -36 day and advances from proximal to distal stump [20,22]. ...
... Regenerated nerve fibers are thinner; both axons and myelin sheaths [10,15,20,21,22,25,26,28]. During tissue maturation number of axons decreases while diameter of myelin sheahs increases [22,29,30]. ...
... Je¿eli nie dochodzi do po³¹czenia siê obu procesów komórki Schwanna w kikucie obwodowym, aksony w kikucie proksymalnym, po przejciu pewnego odcinka (oko³o 4-5 mm) przestaj¹ migrowaae. Komórki Schwanna tworz¹ strukturê na kszta³t "czapeczki", która pokrywa kikut dystalny [10,15,21,25,26]. Pocz¹tek mielinizacji postêpuj¹cy od kikuta proksymalnego w kierunku dystalnym obserwuje siê miêdzy 14-36 dniem [20,22]. ...
... 28,29 Several approaches utilize directional freezing and freeze-drying to form aligned scaffolds from natural polymers such as silk and collagen. [30][31][32][33][34][35][36] Wray et al. used gradient freezing of a silk solution in a mold of arrayed wires to fabricate silk-based scaffolds with tunable porosity, channel architecture, mechanical properties, and degradation rates. 30 Stoppel et al. then demonstrated that these aligned silk-based scaffolds could be combined with cardiac tissuederived ECM to greatly enhance cellular infiltration and vascularization of the scaffold constructs when implanted subcutaneously in rats. ...
... Yannas et al. have prepared collagen and glycosaminoglycan sponges with axially aligned pores through unidirectional freezing and subsequent lyophilization of a collagen-chondroitin sulfate suspension aimed at scaffolds for nerve regeneration. [32][33][34][35] Unlike the methods described herein, this approach does not start with a continuous, entangled network of selfassembled, fibrillar collagen, but rather a mixture of soluble collagen and insoluble collagen fragments. More recent work from the same laboratory has demonstrated that collagen-only scaffolds with axially aligned pores can be fabricated through gradient freezing of an acidified collagen suspension. ...
... However, the characterization of these scaffolds was largely limited to the interior pore structure, and little evidence was presented to characterize the scaffold surface. [32][33][34][35][36] In our approach, the highly aligned surface topography is dependent on the continuous fibrillar hydrogel network prior to freezing and the high aspect ratio conduit used for fabrication. SEM images of cross sections revealed that the aligned scaffolds fabricated from self-assembled gels cast in cylindrical conduits had a highly porous interior. ...
Matrix and cellular alignment are critical factors in the native function of many tissues, including muscle, nerve, and ligaments. Collagen is frequently a component of these aligned tissues, and collagen biomaterials are widely used in tissue engineering applications. However, the generation of aligned collagen scaffolds that maintain the native architecture of collagen fibrils has not been straightforward, with many methods requiring specialized equipment or technical procedures, extensive incubation times, or denaturing of the collagen. Herein, we present a simple, rapid method for fabrication of highly aligned collagen scaffolds. Collagen was assembled to form a fibrillar hydrogel in a cylindrical conduit with high aspect ratio and then frozen and lyophilized. The resulting collagen scaffolds demonstrated highly aligned topographical features along the scaffold surface. This presence of an initial fibrillar network and the high-aspect ratio vessel were both required to generate alignment. The diameter of fabricated scaffolds was found to vary significantly with both the collagen concentration of the hydrogel suspension and the diameter of conduits used for fabrication. Additionally, the size of individual aligned topographical features was significantly dependent on the conduit diameter and the freezing temperature. When cultured on aligned collagen scaffolds, both rat dermal fibroblasts and axons emerging from chick dorsal root ganglia explants demonstrated elongated, aligned morphology and growth on the aligned topographical features. Overall, this method presents a simple means for generating aligned collagen scaffolds that can be applied to a wide variety of tissue types, particularly those where such alignment is critical to native function.
... The use of tubular conduit is capable of enhance the regeneration process, guiding axonal growth, isolating, protecting and maintaining trophic factors in the injury site (Chamberlain et al., 1998;Jang et al., 2016;Safa and Buncke, 2016). Among the several types, biodegradable tubes have shown major application in basic and clinical research, because its nature provides biocompatibility, and decrease the probability of an immune response (Muheremu and Ao, 2015). ...
... The tubulization technique is another promising strategy that can enhance axonal regeneration. The nerve conduits are described by their ability to guide axonal growth, isolate, protect and maintain trophic factors at the site of injury (Chamberlain et al., 1998). Among the various types of tubes, biodegradable ones have shown great application in basic and clinical research, because its nature provides biocompatibility, and decreases the probability of an increase in the immune response. ...
The regenerative potential of the peripheral nervous system (PNS) is widely known, but functional recovery, particularly in humans, is seldom complete. Therefore, it is necessary to resort to strategies that induce or potentiate the PNS regeneration. Our main objective was to test the effectiveness of Olfactory Ensheathing Cells (OEC) transplantation into a biodegradable conduit as a therapeutic strategy to improve the repair outcome after nerve injury. Sciatic nerve transection was performed in C57BL/6 mice; proximal and distal stumps of the nerve were sutured into the collagen conduit. Two groups were analyzed: DMEM (acellular grafts) and OEC (1×105/2μL). Locomotor function was assessed weekly by Sciatic Function Index (SFI) and Global Mobility Test (GMT). After eight weeks the sciatic nerve was dissected for morphological analysis. Our results showed that the OEC group exhibited many clusters of regenerated nerve fibers, a higher number of myelinated fibers and myelin area compared to DMEM group. The G-ratio analysis of the OEC group showed significantly more fibers on the most suitable sciatic nerve G-ratio index. Motor recovery was accelerated in the OEC group. These data provide evidence that the OEC therapy can improve sciatic nerve functional and morphological recovery and can be potentially translated to the clinical setting.
... In a previous study, collagen-GAG-filled collagen tubes were assessed in vivo. The results demonstrated a continuous layer of myofibroblasts, and axonal regrowth was more effective [48]. Furthermore, chitosan was used to form nanofiber mesh tubes in another investigation. ...
Chitosan (Chi) is a natural polymer that has been demonstrated to have potential as a promoter of neural regeneration. In this study, Chi was prepared with various amounts (25, 50, and 100 ppm) of gold (Au) nanoparticles for use in in vitro and in vivo assessments. Each as-prepared material was first characterized by UV-visible spectroscopy (UV-Vis), Fourier-transform infrared spectroscopy (FTIR), atomic force microscopy (AFM), scanning electron microscopy (SEM), and Dynamic Light Scattering (DLS). Through the in vitro experiments, Chi combined with 50 ppm of Au nanoparticles demonstrated better biocompatibility. The platelet activation, monocyte conversion, and intracellular ROS generation was remarkably decreased by Chi–Au 50 pm treatment. Furthermore, Chi–Au 50 ppm could facilitate colony formation and strengthen matrix metalloproteinase (MMP) activation in mesenchymal stem cells (MSCs). The lower expression of CD44 in Chi–Au 50 ppm treatment demonstrated that the nanocomposites could enhance the MSCs undergoing differentiation. Chi–Au 50 ppm was discovered to significantly induce the expression of GFAP, β-Tubulin, and nestin protein in MSCs for neural differentiation, which was verified by real-time PCR analysis and immunostaining assays. Additionally, a rat model involving subcutaneous implantation was used to evaluate the superior anti-inflammatory and endothelialization abilities of a Chi–Au 50 ppm treatment. Capsule formation and collagen deposition were decreased. The CD86 expression (M1 macrophage polarization) and leukocyte filtration (CD45) were remarkably reduced as well. In summary, a Chi polymer combined with 50 ppm of Au nanoparticles was proven to enhance the neural differentiation of MSCs and showed potential as a biosafe nanomaterial for neural tissue engineering.
... Overall, predictions of the computational model indicate that a porous sheath has both advantages and drawbacks. This ambiguity is also present in experimental studies; some report increased neuronal regeneration [75][76][77][78] and offer the hypothesis that this could be a result of increased oxygen diffusion and infiltration of supportive cells, whereas others report poorer results when using porous materials [62,79] and suggest this is caused by loss of growth factors and infiltration of host immune cells. We point out however that the model developed in this work is limited to archetypal porous structures and offers only a simplified description of the initial steps of the nerve repair process. ...
Millions of people worldwide are affected by peripheral nerve injuries (PNI), involving billions of dollars in healthcare costs. Common outcomes for patients include paralysis and loss of sensation, often leading to lifelong pain and disability. Engineered Neural Tissue (EngNT) is being developed as an alternative to the current treatments for large-gap PNIs that show underwhelming functional recovery in many cases. EngNT repair constructs are composed of a stabilised hydrogel cylinder, surrounded by a sheath of material, to mimic the properties of nerve tissue. The technology also enables the spatial seeding of therapeutic cells in the hydrogel to promote nerve regeneration. The identification of mechanisms leading to maximal nerve regeneration and to functional recovery is a central challenge in the design of EngNT repair constructs. Using in vivo experiments in isolation is costly and time-consuming, offering a limited insight on the mechanisms underlying the performance of a given repair construct. To bridge this gap, we derive a cell-solute model and apply it to the case of EngNT repair constructs seeded with therapeutic cells which produce vascular endothelial growth factor (VEGF) under low oxygen conditions to promote vascularisation in the construct. The model comprises a set of coupled non-linear diffusion-reaction equations describing the evolving cell population along with its interactions with oxygen and VEGF fields during the first 24h after transplant into the nerve injury site. This model allows us to evaluate a wide range of repair construct designs (e.g. cell-seeding strategy, sheath material, culture conditions), the idea being that designs performing well over a short timescale could be shortlisted for in vivo trials. In particular, our results suggest that seeding cells beyond a certain density threshold is detrimental regardless of the situation considered, opening new avenues for future nerve tissue engineering.
... Different cell types have varying pore size requirements. For example, while 100 µm pore diameter is optimal for cardiomyocyte function [261,262], nerve cells require far smaller pores between 5 and 30 µm [263,264]. It has been further shown that graded pore sizes can be favourable for bone regeneration and interface tissue [261,[265][266][267]. Similarly, specific regenerative functions are influenced by pore size [4]. ...
A range of biomaterials and fabrication methods have been explored to produce biomimetic scaffolds to facilitate cardiac tissue regeneration. Ice-templated collagen scaffolds have demonstrated translational success in other clinical applications. The ice templating technique utilizes phase separation dynamics during solidification and subsequent sublimation of ice to produce scaffolds with interconnected porosity. Although composition has been found to be key to determining cellular response, both nano-scale and micro-scale surface features of ice templated collagen scaffolds have also been found to encourage cellular ingrowth and attachment. Previous research has introduced techniques to control pore size and anisotropy. To date, however, ice-templating has not been shown to allow control of architecture to the extent that it is possible to replicate the structure of more complex tissue morphologies such as the myocardium. In this thesis, the underpinning physics of ice formation is leveraged to determine the final architecture of ice-templated collagen scaffolds. A controllable directional freezing apparatus was designed and built to enable fine control of the thermal environment during solidification. A relationship between the set thermal parameters and final pore architecture was established. This relationship enabled the production of structures with controlled pore alignment and size. The direct control and monitoring capabilities of the freezing apparatus enabled observations of intrinsic freezing kinetics. This insight allowed the solidification processes of anisotropic and isotropic ice-templating to be compared, and a link between the previously distinct fields was hypothesized. A novel thermal control technique was developed that dictated ice growth directions and achieved complex lamellar orientation of ice-templated collagen scaffolds. A new mould design was produced, with a heat-sink at the base and heat sources in the mould walls, which afforded three-dimensional thermal control during the solidification process. Ultimately, this created complex lamellar orientation of ice-templated collagen scaffolds. The technique is presented alongside a finite element model, developed as a predictive tool for the design of final lamellar orientation. Heat source moulds were used to introduce controlled thermal gradients during the solidification phase of the ice-templating process. Various heat source profiles were implemented and simulated. It was found that by introducing controlled complex thermal gradients during solidification, scaffolds with multidirectional pore orientations were produced, and the finite element simulation was found to accurately predict lamellar orientation. Taken together, the model and heat source freezing technique provide the opportunity for design and production of regenerative collagen scaffolds with tailored architectural morphologies. After establishing a control protocol for producing structures with tailored local lamellar architecture, patches were tested by observing the effects of scaffold architecture on cellular behaviour and mechanical conformation to the dynamic movements of the heart. Cardiomyocytes (H9 hESCs) were seeded onto scaffolds with aligned and isotropic pore structures and the cell signalling patterns were then compared. It was determined that the biomimetic accuracy of the aligned scaffold improved the uniformity of calcium signalling in cardiomyocytes when compared with those on isotropic structures. These results indicate that myocardial function is enhanced by defined scaffold orientation. The application of an ex vivo ovine cardiac perfusion model enabled direct observation of the native myocardial movement during the cardiac cycle. Through direct optical imaging and digital image correlation, collagen scaffolds were tested to assess their response to native myocardial deformation patterns. The strain dynamics of aligned and isotropic scaffold architectures were compared, and the efficacy of both glue and suture fixation methods were explored. It was determined that aligned scaffolds adhered with suture fixation complied with the native physio-mechanical environment. Similarly adhered isotropic scaffolds and patches adhered with glue, however, resulted in reduced deformation relative to the native myocardium. The work in this thesis has established a novel freeze casting technique to afford specific three dimensional control of collagen scaffold alignment. The resulting scaffolds with directionally aligned pore architectures were found to enhance cellular and mechanical dynamics to better replicate the native behaviour of myocardial tissue.
... It should be noted that a silicone conduit was used as an outer sheath during transplantation in this study. Silicone conduits have been used extensively in animal models to investigate peripheral nerve regeneration [46][47][48] and also used in a 5-year follow-up clinical study by Lundborg et al. [49] . Silicone is not an ideal conduit material for clinical nerve repair due to its mechanical mismatch and non-biodegradability, which can cause nerve compression syndrome requiring a second operation to remove the silicone tube [50 , 51] . ...
Vascularisation is important in nerve tissue engineering to provide blood supply and nutrients for long-term survival of implanted cells. Furthermore, blood vessels in regenerating nerves have been shown to serve as tracks for Schwann cells to migrate along and thus form Bands of Büngner which promote axonal regeneration. In this study, we have developed tissue-engineered constructs containing aligned endothelial cells, or co-cultures of both endothelial cells and Schwann cells to test whether these structures could promote regeneration across peripheral nerve gaps. Type I rat tail collagen gels containing HUVECs (Human Umbilical Vein Endothelial Cells, 4 × 10^6 cells/ml) were cast in perforated tethering silicone conduits to facilitate cellular self-alignment and tube formation for 4 days of culture. For co-culture constructs, optimal tube formation and cellular alignment was achieved with a ratio of 4:0.5 × 10^6 cells/ml (HUVECs:Schwann cells). An in vivo test of the engineered constructs to bridge a 10 mm gap in rat sciatic nerves for 4 weeks revealed that constructs containing only HUVECs significantly promoted axonal regeneration and vascularisation across the gap, as compared to conventional aligned Schwann cell constructs and those containing co-cultured HUVECs and Schwann cells. Our results suggest that tissue-engineered constructs containing aligned endothelial cells within collagen matrix could be good candidates to treat peripheral nerve injury.
... It should be noted that a silicone conduit was used as an outer sheath during transplantation in this study. Silicone conduits have been used extensively in animal models to investigate peripheral nerve regeneration [46][47][48] and also used in a 5-year follow-up clinical study by Lundborg et al. [49] . Silicone is not an ideal conduit material for clinical nerve repair due to its mechanical mismatch and non-biodegradability, which can cause nerve compression syndrome requiring a second operation to remove the silicone tube [50 , 51] . ...
Vascularisation is important in nerve tissue engineering to provide blood supply and nutrients for long-term survival of implanted cells. Furthermore, blood vessels in regenerating nerves have been shown to serve as tracks for Schwann cells to migrate along and thus form Bands of Büngner which promote axonal regeneration. In this study, we have developed tissue-engineered constructs containing aligned endothelial cells, or co-cultures of both endothelial cells and Schwann cells to test whether these structures could promote regeneration across peripheral nerve gaps. Type I rat tail collagen gels containing HUVECs (Human Umbilical Vein Endothelial Cells, 4×10⁶ cells/ml) were cast in perforated tethering silicone tubes to facilitate cellular self-alignment and tube formation for 4 days of culture. For co-culture constructs, optimal tube formation and cellular alignment was achieved with a ratio of 4:0.5×10⁶ cells/ml (HUVECs:Schwann cells). An in vivo test of the engineered constructs to bridge a 10 mm gap in rat sciatic nerves for 4 weeks revealed that constructs containing only HUVECs significantly promoted axonal regeneration and vascularisation across the gap, as compared to conventional aligned Schwann cell constructs and those containing co-cultured HUVECs and Schwann cells. Our results suggest that tissue-engineered constructs containing aligned endothelial cells within collagen matrix could be good candidates to treat peripheral nerve injury.
Statement of Significance
Nerve tissue engineering provides a potential way to overcome the limitations associated with current clinical grafting techniques for the repair of severe peripheral nerve injuries. However, the therapeutic cells within engineered nerve tissue require effective vascularisation in order to survive. This work therefore aimed to develop engineered nerve constructs containing aligned tube-like structures made from endothelial cells. Not only did this provide a method to improve vascularisation, it demonstrated for the first time that aligned endothelial cells can outperform Schwann cells in promoting nerve regeneration in the rat sciatic nerve model. This has introduced the concept of developing pre-vascularised engineered nerve tissues, and indicated the potential usefulness of endothelial cell structures in tissue engineering for peripheral nerve repair.
... This condition is essential to permit application of the collagen tube in nerve gaps of different length. 19 Chamberlain et al. 21 used silicon tubes filled with collagen and show improved regeneration over a 10mm rat sciatic nerve gap compared to empty silicone controls. Our results demonstrate that both collagen and autograft repairs promote nerve regeneration. ...
... This thesis mainly focused on developing artificial tissues to mimic the inner part of the nerve, which includes endoneurium and perineurium layers. The silicone conduit was used as an outer sheath of the construct during transplantation, and has been used extensively in animal models to investigate peripheral nerve regeneration (Chamberlain et al., 1998;Kim et al., 2007;Wu et al., 2008). It was also used in a 5-year follow-up clinical study by Lundborg et al. (2004) (Lundborg et al., 2004). ...
Peripheral nerve injury can be debilitating and may result in loss of sensory or motor function. Nerve autograft remains the gold standard to repair damage that results in long gaps. The efficacy of nerve grafts, however, can be limited by necrosis at the central region. This is also a limitation in the effectiveness of cellular biomaterials developed as tissue engineered repair approaches. Although vascularised nerve grafts were introduced, some limitations such as availability of nerves and donor site morbidity need to be overcome. Therefore, there is a need for vascularised tissue-engineered nerve constructs.
This study established an anisotropic nerve construct which contains self-aligned human umbilical cord vein endothelial cells (HUVECs) that form tube-like structures with and without aligned Schwann cells within a tethered collagen matrix. In an in vitro model, aligned tube-like structures supported and enhanced Schwann cell migration when compared to aligned Schwann cell-only constructs. Additionally, their efficacy to promote axonal regeneration in vitro was comparable to that of aligned Schwann cell constructs. In a rat sciatic nerve injury model, the aligned HUVEC tube-like structure constructs supported robust neuronal regeneration. HUVEC-only constructs also showed significantly improved vascularisation and Schwann cell migration at the repair site.
In addition, the gel aspiration-ejection (GAE) technique offers a rapid and robust approach to produce stable anisotropic hydrogel scaffolds. This work optimised the GAE technique to generate aligned Schwann cells in collagen scaffolds. These scaffolds were stable and exhibited similar linear viscoelastic behaviours to rat sciatic nerves. They supported and promoted axonal regeneration in vivo when compared to the empty conduit groups. Together, this study has developed for the first time pre-vascularised tissue-engineered nerve constructs and shown their potential in promoting vascularisation and axonal regeneration. A novel GAE technique has also been shown to be useful in producing aligned Schwann cell-containing hydrogel constructs. In the future, the GAE system can be potentially integrated with the pre-vascularisation concept to create vascularised engineered-nerve conduits.
... Next-generation nerve conduits made use of biologically derived materials (e.g., collagen) as conduits and hydrogel substrates [53,54]. Conduits containing collagen hydrogels were found to perform better than hollow or salinefilled conduits, and performed the same as autografts when used to bridge short nerve gaps [55]. ...
Tissue engineering is the use of engineering methods to replace, replicate, or improve biological tissues. Neural tissue engineering involves the integrated use of biomaterials, cellular engineering, and drug delivery technologies with the purpose of protecting, repairing, or regenerating cells and tissues of the nervous system. Through the introduction of biochemical, topographic, immunomodulatory, and other types of cues, tissues can be therapeutically controlled to direct growth and tissue function in order to overcome biological constraints on tissue repair and regeneration. These strategies can be applied when injury or disease occurs in the brain, spinal cord, for damaged peripheral nerves, or to improve chronic functionality of implantable neural interfaces. In this chapter, we present an overview of neural tissue engineering using examples of therapeutic systems including nerve conduits, implantable hydrogels, delivery of neurotrophic factors and stem cells, genetic approaches to tissue engineering, immunomodulation, and electrical stimulation.
... oxygen and nutrients is crucial for the survival of seeded cells and the success of neuronal regeneration, and this can be facilitated through the use of porous guidance constructs. Careful design of NRC sheaths could also contribute towards greater control over spatial distributions of growth factors and cells via the use of asymmetric or spatially varying porosity;it is possible that the sheath design could be tailored to complement the chosen distribution or density of seeded cells.However, the exact effect of introducing porosity appears to differ according to the scenario, with some studies reporting better results with impermeable sheaths or tubular NRCs[66,107]. These contrasting results could be due to the positive effects of restricting the migration of inflammatory cells into the construct and the loss of important growth factors by diffusion out into the surround tissue. ...
Currently, the design of tissue engineered constructs for peripheral nerve repair is informed predominantly by experiments. However, translation to the clinical setting is slow, and engineered tissues have not surpassed the outcomes achieved by nerve grafts. Therapeutic cell survival and vascularisation are important for the assimilation of engineered tissue, and vascularisation provides vital directional cues for regenerating nerves. In this thesis, mathematical modelling informed by experimental data is used to investigate the impact of different therapeutic cell seeding strategies on cell survival and vascularisation in engineered tissue nerve repair constructs. A mathematical model of interactions between cells, oxygen and vascular endothelial growth factor (VEGF), consisting of three partial differential equations, is developed and parameterised against in vitro data. Key cell type-specific parameter values are derived, and the model is then used to simulate cell-solute interactions in a nerve repair construct over the first five days post-implantation in vivo. Simulations using uniform seeding cell densities of 88 and 13 × 10⁶ cells/ml result in the highest mean viable cell densities across the construct after 1 and 5 days respectively. However, simulations using seeding densities in the range of 200 – 300 ×10⁶ cells/ml result in steeper VEGF gradients and higher total VEGF concentrations across the construct, which could be beneficial for vascularisation. Simulations incorporating a porous construct sheath result in higher viable cell density predictions, but also lower total VEGF concentrations, than those run using an impermeable sheath. Subsequently, the cell-solute model is combined with a discrete model of angiogenesis that simulates vascular growth in response to gradients of VEGF. Simulation results suggest that different cell seeding strategies could influence the density, rate and morphology of vascularisation. The predictions generated in this work demonstrate how mathematical modelling as part of a wider multidisciplinary approach can provide direction for future experimental work.
... However, there are several criteria that NGTs must demonstrate prior to clinical deployment. First, NGTs should be porous to allow for waste and nutrient diffusion and exhibit a pore size in the 5-30 µm range to minimize excessive fibrosis and inflammatory cell infiltration [18][19][20]. The impact of pore size is increasingly important in clinically-relevant long gap nerve injury. ...
Promising biomaterials should be tested in appropriate large animal models that recapitulate human inflammatory and regenerative responses. Previous studies have shown tyrosine-derived polycarbonates (TyrPC) are versatile biomaterials with a wide range of applications across multiple disciplines. The library of TyrPC has been well studied and consists of thousands of polymer compositions with tunable mechanical characteristics and degradation and resorption rates that are useful for nerve guidance tubes (NGTs). NGTs made of different TyrPCs have been used in segmental nerve defect models in small animals. The current study is an extension of this work and evaluates NGTs made using two different TyrPC compositions in a 1 cm porcine peripheral nerve repair model. We first evaluated a nondegradable TyrPC formulation, demonstrating proof-of-concept chronic regenerative efficacy up to 6 months with similar nerve/muscle electrophysiology and morphometry to the autograft repair control. Next, we characterized the acute regenerative response using a degradable TyrPC formulation. After 2 weeks in vivo , TyrPC NGT promoted greater deposition of pro-regenerative extracellular matrix (ECM) constituents (in particular collagen I, collagen III, collagen IV, laminin and fibronectin) compared to commercially available collagen-based NGTs. This corresponded with dense Schwann cell infiltration and axon extension across the lumen. These findings confirmed results reported previously in a mouse model and reveal that TyrPC NGTs were well tolerated in swine and facilitated host axon regeneration and Schwann cell infiltration in the acute phase across segmental defects - likely by eliciting a favorable neurotrophic ECM milieu. This regenerative response ultimately can contribute to functional recovery.
... Tissue engineering (or regenerative medicine) is defined as the scientific principles applied in the production of living tissues through a use of bioreactors, cells, scaffolds, growth factors, or a combination of them (4). The scaffold can be implanted single-handedly to prompt host cell colonization to the wounded position and tissue restoration, or it can be seeded with cells and/or growth factors and serve to control the release and targeting of these treatments (5)(6)(7). Bone tissue engineering shows promise as an alternative to repair critical bone defects. Accordingly, bone tissue engineering was developed in both scope and significance in biomedical engineering. ...
Issues of safety are very crucial with biomaterials and medical devices. Sixteen male New Zealand White rabbits equally into four groups: Group A, rabbits had part of their radial bone (2 cm, mid shaft) and left empty as a control. Group B, rabbits were implanted with scaffold 5211. Group C, rabbits were implanted with scaffold 5211GTA+Alginate. Group D, rabbits were implanted with 5211PLA. All scaffolds were prepared by freeze-drying method. Blood samples were collected at day 0 and 1 st , 2 nd , 3 rd , 4 th and 8 th week after implantation. The blood examination included complete hemogram and certain serum biochemical parameters. The results showed that there was no significant difference (P>0.05) among each treatment group in comparison with control group (day 0). However, red blood cells, hemoglobin, packed cell volume, mean corpuscular hemoglobin concentration, monocyte, plasma protein, inorganic phosphate, sodium, chloride and urea were significantly increased (P<0.05) among treatment groups at week 8. An abnormal architecture of viscera was observed in all animals, thus indicating a form of toxicity related to the degrading scaffold materials. The severity of histopathological lesions in viscera was not coated polymers dependent nor development materials. In conclusion, implantation of 5211 scaffold with or without coated framework has a significant impact on histopathological and certain hematological and biochemical parameters.
... Tissue engineering (or regenerative medicine) is defined as the scientific principles applied in the production of living tissues through a use of bioreactors, cells, scaffolds, growth factors, or a combination of them (4). The scaffold can be implanted single-handedly to prompt host cell colonization to the wounded position and tissue restoration, or it can be seeded with cells and/or growth factors and serve to control the release and targeting of these treatments (5)(6)(7). Bone tissue engineering shows promise as an alternative to repair critical bone defects. Accordingly, bone tissue engineering was developed in both scope and significance in biomedical engineering. ...
Issues of safety are very crucial with biomaterials and medical devices. Sixteen male New Zealand White rabbits equally into four groups: Group A, rabbits had part of their radial bone (2 cm, mid shaft) and left empty as a control. Group B, rabbits were implanted with scaffold 5211. Group C, rabbits were implanted with scaffold 5211GTA+Alginate. Group D, rabbits were implanted with 5211PLA. All scaffolds were prepared by freeze-drying method. Blood samples were collected at day 0 and 1 st , 2 nd , 3 rd , 4 th and 8 th week after implantation. The blood examination included complete hemogram and certain serum biochemical parameters. The results showed that there was no significant difference (P>0.05) among each treatment group in comparison with control group (day 0). However, red blood cells, hemoglobin, packed cell volume, mean corpuscular hemoglobin concentration, monocyte, plasma protein, inorganic phosphate, sodium, chloride and urea were significantly increased (P<0.05) among treatment groups at week 8. An abnormal architecture of viscera was observed in all animals, thus indicating a form of toxicity related to the degrading scaffold materials. The severity of histopathological lesions in viscera was not coated polymers dependent nor development materials. In conclusion, implantation of 5211 scaffold with or without coated framework has a significant impact on histopathological and certain hematological and biochemical parameters. سفان ه محم خضر و د 1a,b ، محمد ز و زكريا بكر ابو كي 1a,2* انتان ، بنت سميحة الرزاق عبد 1a يوسف محمد لقمان ، 3 ، بكر ابو عبد ادامو 3c محم خضر زيد ، و د 4 م و حمد التب اب بن قيوم 1a 1a فرع ،البيطرية السريرية قبل ما العلوم 2 ،البيولوجية العلوم معهد ،الجزيئي الحيوي الطب مختبر 3 فرع الحيوان والجراحة الطب ية ،المصاحبة 4 فرع ماليزيا بوترا جامعة ،البيطري الطب كلية ،البيطرية السريرية الدراسات ، سيردانغ ،ماليزيا ،إحسان دارول سيالنجور ، 1b فرع ،العراق ،الموصل ،الموصل جامعة ،البيطري الطب كلية ،البيطري التشريح c فرع أوسم جامعة ،واألشعة البيطرية الجراحة ،دانفوديو انو نيجيريا ،سوكوتو الخالصة مهمة السالمة امور تعتبر جدا استخدام عند الطبية واألجهزة الحيوية المواد. تقسيم تم اربعة الى نيوزيلنديا ابيضا ارنبا عشر ستة أ المجموعة :مجاميع ، قطع تم حيث 2 ب المجموعة .سيطرة كمجموعة فارغة وتركها الكعبرة عظم جسم من سم ، زراعة تم عظم الكعبرة
... The effects of pore sizes on regeneration should be carefully considered to prevent fibrotic tissue infiltration. Early studies of PNI repair with hollow conduits suggest that pore sizes of~5-10 μm seem to be optimal to facilitate nutrients and waste exchange while minimizing fibrotic tissue infiltration in vivo (Chamberlain et al., 1998;Jenq et al., 1987;Vleggeert-Lankamp et al., 2007). It is also important to consider the material properties when considering the effects of pore size. ...
Nerve injuries can be life-long debilitating traumas that severely impact patients' quality of life. While many acellular neural scaffolds have been developed to aid the process of nerve regeneration, complete functional recovery is still very difficult to achieve, especially for long-gap peripheral nerve injury and most cases of spinal cord injury. Cell-based therapies have shown many promising results for improving nerve regeneration. With recent advances in neural tissue engineering, the integration of biomaterial scaffolds and cell transplantation are emerging as a more promising approach to enhance nerve regeneration. This review provides an overview of important considerations for designing cell-carrier biomaterial scaffolds. It also discusses current biomaterials used for scaffolds that provide permissive and instructive microenvironments for improved cell transplantation.
... In this sense, the use of biodegradable polymers for constructing nerve guide channels is ideal because it eliminates the need of a second surgery to remove the nerve guide channels from the body to avoid chronic tissue responses or nerve compression. In particular, poly (phosphoester) [16,17], collagen [18,19], polyglycolide [20], collagen and poly-glycolide [21], poly (L-lactide-co-glycolide) (PLGA) [22,23], and poly-L-lactic acid/caprolactone [24] are among the most common biodegradable polymers used for this purpose. ...
The human nervous system lacks an inherent ability to regenerate its components upon damage or diseased conditions. During the last decade, this has motivated the development of a number of strategies for nerve regeneration. However, most of those approaches have not been used in clinical applications till today. For instance, although biomaterial-based scaffolds have been extensively used for nerve reparation, the lack of more customized structures have hampered their use in vivo. This highlight focuses mainly on how 3D bioprinting technology, using polymeric hydrogels as bio-inks, can be used for the development of new nerve guidance channels or devices for peripheral nerve cell regeneration. In this concise contribution, some of the most recent and representative examples are highlighted to discuss the challenges involved in various aspects of 3D bioprinting for nerve cell regeneration, specifically when using polymeric hydrogels.
... 6 In this context, freeze-casting, a technique initially developed for the structuration of colloidal suspensions for the elaboration of ceramic materials, 7 has been adapted to produce scaffolds based on collagen and other biopolymers for tissue engineering purpose. [8][9][10] This technique enables to create oriented macroporous scaffolds from colloidal suspensions or polymer solutions by means of directional freezing followed by ice sublimation. 11 Upon anisotropic cooling, an oriented segregation occurs between the newly-formed ice and the solutes and/or particles initially present in the solution/suspension. ...
Type I collagen is the main component of the extra-cellular matrix (ECM). In vitro, under a narrow window of physico-chemical conditions, type I collagen self-assembles to form complex supramolecular architectures reminiscent of those found in native ECM. Presently, a major challenge in collagen-based biomaterials is to couple the delicate collagen fibrillogenesis events with a controlled shaping process in non-denaturating conditions. In this work an ice-templating approach promoting the structuration of collagen into macroporous monoliths is used. Instead of common solvent removal procedures, a new topotactic conversion approach yielding self-assembled ordered fibrous materials is implemented. These collagen-only, non-cross-linked scaffolds exhibit uncommon mechanical properties in the wet state. With the help of the ice-patterned micro-ridge features, Normal Human Dermal Fibroblasts and C2C12 murine myoblasts successfully migrate and form highly-aligned populations within the resulting 3D biomimetic collagen scaffolds. These results open a new pathway to the development of new tissue engineering scaffolds ordered across various organization levels from the molecule to the macropore, and are of particular interest for biomedical applications where large scale 3D cell alignment is needed such as for muscular or nerve reconstruction.
... 52,53 Collagen is a major component of ECM and has widespread use as a biological material including peripheral nerve repair. [54][55][56][57][58][59][60][61] When purified, collagen is only weakly antigenic, and its antigenicity can be further reduced via enzymatic removal of the non-helical telopeptide regions of the molecule, or by crosslinking. 62 Diffusion processes through collagen matrices are facilitated by its smooth microgeometry and transmurale permeability ($100,000 D). 60,61 A molecular weight cut-off of approximately 50,000 D has been reported to facilitate diffusional transportation of nutrients and other molecules, whilst preventing cells from entering the conduit structure. ...
... In addion, NGF inhibits Schwann cell apoptosis by activating the P13k/Akt/GSK3ß and ERK1/2 pathways [30]. Presence of Schwann cells is necessary for advancement of sxons [31]. ...
Objective
To determine the effects of chicken embryo brain extract (BE) on transects sciatic nerve in male rats.
Methods
Thirty adult male Sprague-Dawley rats weighing 200 to 250 g, were randomized into three groups treated with (1) sham surgery, (2) normal saline (NS), and (3) brain extract (BE). The BE was taken from incubating chick embryos at day 8. The sciatic nerve was exposed and sharply transected at the mid thigh level. Immediate epineurial repair was then performed. The BE treated animals were given 400 µl/kg of the chick embryo BE intraperitoneal, once daily, for 2 weeks. All animals were evaluated by sciatic functional index (SFI), electrophysiology, histology, and immunohistochemistry at days 28, 90 after surgery.
Results
The mean SFI difference between BE and NS groups at days 28, 60 and 90 after surgery was statistically significant (p=0.086). The mean number of myelinated fibers in the BE group was significantly greater than that of the NS group on days 28 and 90 after surgery (p=0.034). At days 28 and 90 after surgery, the mean nerve conduction velocity (NCV) in the BE group was significantly faster than that of the NS group (p=0.041).
Conclusion
These results indicate for the first time that chick embryo brain extract can enhance peripheral nerve regeneration in rat.
... [118] With a rat sciatic nerve model, Chamberlain et al. found 6-weeks post-operation a significant increase in the number of axons in the middle of 10-mm-long collagen-glycosaminoglycan scaffolds relative to saline-filled controls of the same length. [119] Because of collagen's structural role, addition of native microarchitectural features to collagen could improve cell binding and alignment. Ceballos et al. used a magnetic field to align collagen fibers and showed that peripheral nerve regeneration in mice implanted with aligned collagen grafts was significantly higher than for unaligned collagen grafts, after 60 days post-operation. ...
In article number 1701713, Kevin J. Otto, Jack W. Judy, Christine E. Schmidt and co‐workers review the latest in peripheral nerve interfaces, peripheral nerve tissue engineering, and the intersection of these fields to create regenerative peripheral nerve interfaces. With these devices, axons regenerate into a hydrogel‐like environment that allows them to intimately interface with electrodes. This technology could ultimately be used to control robotic prostheses.
... Initial attempts to define the biological changes that occur during surgical repair of the injured nerve have focused on regeneration within silicone and elastomer tubes. [6][7][8][9][10] These efforts have documented the genomic and proteomic changes in a crush injury (a different classification of nerve injury undergoing altered mechanisms of repair), [11][12][13][14][15][16] across short transection injuries, or have only considered a limited range of proteins using proteomic microarrays (15 proteins analyzed). [17] The complex orchestrated series of events needs to be further characterized to achieve an optimal treatment strategy. ...
The regenerative environment within different biomaterial nerve conduits remains poorly understood. In article number 1702170, Abhay Pandit and co‐workers highlight the proteomic changes that occur spatially throughout both natural and synthetic biomaterial conduits compared to autografts. The study explores how different materials activate different biological functions during the early stages of regeneration and where they can be supplemented to enhance repair.
... Initial attempts to define the biological changes that occur during surgical repair of the injured nerve have focused on regeneration within silicone and elastomer tubes. [6][7][8][9][10] These efforts have documented the genomic and proteomic changes in a crush injury (a different classification of nerve injury undergoing altered mechanisms of repair), [11][12][13][14][15][16] across short transection injuries, or have only considered a limited range of proteins using proteomic microarrays (15 proteins analyzed). [17] The complex orchestrated series of events needs to be further characterized to achieve an optimal treatment strategy. ...
The treatment of peripheral nerve injuries remains a major problem worldwide despite the availability of a number of Food and Drug Administration (FDA) approved devices which fail to match the efficacy of autografts. Different strategies are used to improve regeneration and functional recovery using biomaterial nerve conduits. However, there is little investigation of the transcriptomic and proteomic changes which occur as a result of these interventions, particularly regarding transection injuries. This study explores differences between autograft-mediated repair and conduit-material-mediated repair of peripheral nerve injuries to understand fundamental differences in their repair mechanisms at the proteomics level at the proximal, middle, and distal components in the early stages of repair. Pathway analysis demonstrates that each material selectively activates different regenerative pathways and alters different biological functions spatially throughout the biomaterial conduits. The analysis highlights some of the deficiencies in conduit-mediated repair in comparison to autograft (e.g., recycling of myelin and cholesterol, reduction in reactive oxygen species, and higher expression of regenerative proteins). These findings thus suggest that by supplementing the expression of these proteins on the biomaterial of choice, this study can potentially attain regeneration equivalent to autograft. This approach paves the way for incorporating future biomaterial-specific functionalities in nerve guidance conduits.
... [118] With a rat sciatic nerve model, Chamberlain et al. found 6-weeks post-operation a significant increase in the number of axons in the middle of 10-mm-long collagen-glycosaminoglycan scaffolds relative to saline-filled controls of the same length. [119] Because of collagen's structural role, addition of native microarchitectural features to collagen could improve cell binding and alignment. Ceballos et al. used a magnetic field to align collagen fibers and showed that peripheral nerve regeneration in mice implanted with aligned collagen grafts was significantly higher than for unaligned collagen grafts, after 60 days post-operation. ...
Research on neural interfaces has historically concentrated on development of systems for the brain; however, there is increasing interest in peripheral nerve interfaces (PNIs) that could provide benefit when peripheral nerve function is compromised, such as for amputees. Efforts focus on designing scalable and high-performance sensory and motor peripheral nervous system interfaces. Current PNIs face several design challenges such as undersampling of signals from the thousands of axons, nerve-fiber selectivity, and device-tissue integration. To improve PNIs, several researchers have turned to tissue engineering. Peripheral nerve tissue engineering has focused on designing regeneration scaffolds that mimic normal nerve extracellular matrix composition, provide advanced microarchitecture to stimulate cell migration, and have mechanical properties like the native nerve. By combining PNIs with tissue engineering, the goal is to promote natural axon regeneration into the devices to facilitate close contact with electrodes; in contrast, traditional PNIs rely on insertion or placement of electrodes into or around existing nerves, or do not utilize materials to actively facilitate axon regeneration. This review presents the state-of-the-art of PNIs and nerve tissue engineering, highlights recent approaches to combine neural-interface technology and tissue engineering, and addresses the remaining challenges with foreign-body response.
... It is known that the conduit wall porosity can impact nerve re-growth [7,8]. Numerous studies have investigated conduit porosity in the context of peripheral nerve regeneration and reported contradictory results [9][10][11]. Increased porosity in the conduit walls may enhance permeability to nutrients, and this may potentially augment nerve regeneration [3]. ...
Porous conduits provide a protected pathway for nerve regeneration, while still allowing exchange of nutrients and wastes. However, pore sizes >30 µm may permit fibrous tissue infiltration into the conduit, which may impede axonal regeneration. Coating the conduit with Fibrin Glue (FG) is one option for controlling the conduit’s porosity. FG is extensively used in clinical peripheral nerve repair, as a tissue sealant, filler and drug-delivery matrix. Here, we compared the performance of FG to an alternative, hyaluronic acid (HA) as a coating for porous conduits, using uncoated porous conduits and reverse autografts as control groups. The uncoated conduit walls had pores with a diameter of 60 to 70 µm that were uniformly covered by either FG or HA coatings. In vitro, FG coatings degraded twice as fast as HA coatings. In vivo studies in a 1 cm rat sciatic nerve model showed FG coating resulted in poor axonal density (993 ± 854 #/mm²), negligible fascicular area (0.03 ± 0.04 mm²), minimal percent wet muscle mass recovery (16 ± 1 in gastrocnemius and 15 ± 5 in tibialis anterior) and G-ratio (0.73 ± 0.01). Histology of FG-coated conduits showed excessive fibrous tissue infiltration inside the lumen, and fibrin capsule formation around the conduit. Although FG has been shown to promote nerve regeneration in non-porous conduits, we found that as a coating for porous conduits in vivo, FG encourages scar tissue infiltration that impedes nerve regeneration. This is a significant finding considering the widespread use of FG in peripheral nerve repair.
Graphical Abstract
... The macrophages were found on rough surfaces as opposed to smooth surfaces both in vitro and in vivo (28,29) with evident signs of phagocytosis of the biomaterial. The macrophages can secrete a variety of growth factors that can enhance healing, as well as promoters and regulators of inflammation and tissue degradation (30). ...
Background: Peripheral nerves may be damaged during an injury and its current standard treatment is using an autologous nerve.
Objectives: The purpose of this experimental study is to evaluate and compare the histological results of nerve regeneration after using the eggshell membrane (ESM) guidance channel with autograft.
Materials and Methods: Thirty adult male rats were divided into three experimental groups: ESM guidance channel, autograft, and sham surgery. The decalcifying membrane of egg rotated over the Teflon mandrel and dried at 37°C. A 10 mm nerve segment of left sciatic nerve was cut and removed. In ESM group, the ends of the sciatic nerve were telescoped into the nerve guides. In autograft group, the nerve segment was reversed and used as an autologous nerve graft. At 90 days after surgery, all animals were evaluated by histological and immunohistochemical assessment.
Results: The diameters of regenerated myelinated fibers were 5.24±2.14 µm for the ESM group, and 5.89±2.99 µm for the autograft group. The number of myelinated axons regenerated in the ESM group (9824±218 nerve fibers) was significantly greater than autograft group (7865±314 nerve fibers) (p<0.05).
Conclusion: These findings demonstrate that ESM effectively enhances nerve regeneration in injured rat sciatic nerve.
... In one previous study, undegradable materials were used to prepare the NGC, especially the silicone tube, which was used in the clinical. However, the silicone NGC cannot be absorbed by the body, and long-term retention in the human body will compress the neurals, therefore reoperation was needed to remove it again [27]. With the development of science and technology, degradable materials were used to prepare NGC, especially synthetic materials [28], such as PCL, PLA, Poly (l-lactic acid-coε-caprolactone) (P(LLA-CL)) and so on, which can provide sufficient mechanical support, and it were easy to be processed into nanofiber scaffold by electrospinning. ...
Neural guidance conduits (NGCs) have emerged as a potential alternative to autologous neural grafts, the gold standard for peripheral neural repair. A number of NGCs have been used experimentally to bridge peripheral neural defects in various animal tests, where the outcome is neural regeneration and functional recovery. In this chapter, we describe the latest advances of nanofiber composite NGC application in the field of neural tissue engineering (NTE) for peripheral neural regeneration. Firstly, application of synthetic and natural biomaterials used to fabricate nanofiber NGC is discussed. Then we outline new approaches to developing a regulated structural design of NGC for peripheral neural regeneration, including uniaxial aligned nanofiber membrane, nanoyarn inner-filler NGCs, microtube array sheet inner-filler NGCs, and multichannel NGCs. Furthermore, neurotrophic factors incorporated into nanofiber NGCs to promote Schwann cells proliferation and axonal regeneration are discussed. We also discuss the significance of conductive nanofiber NGCs and electrical stimulation in the field of NTE. Finally, we concluded that composite nanofibers offer good potential for NTE applications.
... Next generation nerve conduits saw the advent of conduits and hydrogel fillers made from biological materials such as collagen [69,70]. Collagen hydrogel ä Fig. 19.3 (continued) spinal cords treated with CA-Rac1, the axons extended a significantly further distance within the inhibitory region than the untreated and agarose controls. ...
Injury to the nervous system leads to several debilitating long-term disabilities that can severely impair quality of life. Regenerative failure following injury is the primary cause of disability and is mainly attributed to the localized upregulation of nerve inhibitory molecules in the case of central nervous system (CNS) injuries, and the presence of inhibitory molecules along with the absence of structural support in the case of peripheral nervous system (PNS) injuries. While treatments using autografts and allografts do result in appreciable nerve regeneration in the case of peripheral nerve gaps, the same is not true of CNS injuries which are difficult to treat.
This chapter discusses innate differences and challenges in treating CNS and PNS injuries, and the current methodologies being employed to enhance the endogenous regenerative potential and plasticity. The state-of-the-art in facilitating repair and rehabilitation by means of biochemical and cellular therapies as well as by electrical stimulation of neuromuscular tissue are also discussed.
... 20 Effects of conduit pore size on the outcome of nerve regeneration have been investigated and, although results and interpretations of this critical aspect vary widely, the optimal pore size for conduits was reported to be in the 5-30 mm range to enable nutrient and waste diffusion. [21][22][23][24] Considering that non-neural cells can easily penetrate through pores of these sizes, it is of interest to determine how infiltration of non-neural cells into porous conduits affects nerve regeneration when combined with a cellfriendly filler matrix. ...
Nerve conduits pre-filled with hydrogels are frequently explored in an attempt to promote nerve regeneration. This study examines the interplay between the porosity of the conduit wall and the level of bioactivity of the hydrogel used to fill the conduit. Nerve regeneration in porous (P) or non-porous (NP) conduits that were filled with either collagen-only or collagen enhanced with a covalently attached neurite-promoting peptide mimic of the glycan Human Natural Killer Cell Antigen-1 (m-HNK) were compared in a 5 mm critical size defect in the mouse femoral nerve repair model. While collagen is a cell-friendly matrix that does not differentiate between neural and non-neural cells, the m-HNK-enhanced collagen specifically promotes axon growth and appropriate motor neuron targeting. In this study, animals treated with non-porous (NP) conduits filled with collagen grafted with m-HNK (CollagenHNK) had the best functional recovery, commensurate with a substantial improvement in all tested histomorphometric parameters (number of axons, degree of raw tissue area, percentage of myelinated nerve fibers, cross-sectional area of myelinated nerve fibers) relative to all other conduit conditions. Our data indicate that under some conditions, the use of generally cell friendly fillers such as collagen, may limit nerve regeneration. This finding is significant, considering the frequent use of collagen based hydrogels as fillers of nerve conduits.
... Studies of the effect of conduit pore size on the outcome of nerve regeneration have shown the optimal pore size for conduits to be in the 5-30 µm range to enable nutrient and waste diffusion and minimize fibrotic and inflammatory cell infiltration. [29][30][31] However, considerable variation in outcomes has been reported in studies using conduits that differ in composition, method of fabrication, and the animal model used to test the conduits. It is important to note that most studies in vivo are performed in mice or rats where the conduit length is usually around 10 mm only. ...
Here, we report on the design of braided peripheral nerve conduits with barrier coatings. Braiding of extruded polymer fibers generates nerve conduits with excellent mechanical properties, high flexibility, and significant kink-resistance. However, braiding also results in variable levels of porosity in the conduit wall, which can lead to the infiltration of fibrous tissue into the interior of the conduit. This problem can be controlled by the application of secondary barrier coatings. Using a critical size defect in a rat sciatic nerve model, the importance of controlling the porosity of the nerve conduit walls was explored. Braided conduits without barrier coatings allowed cellular infiltration that limited nerve recovery. Several types of secondary barrier coatings were tested in animal studies, including (1) electrospinning a layer of polymer fibers onto the surface of the conduit and (2) coating the conduit with a cross-linked hyaluronic acid-based hydrogel. Sixteen weeks after implantation, hyaluronic acid-coated conduits had higher axonal density, displayed higher muscle weight, and better electrophysiological signal recovery than uncoated conduits or conduits having an electrospun layer of polymer fibers. This study indicates that braiding is a promising method of fabrication to improve the mechanical properties of peripheral nerve conduits and demonstrates the need to control the porosity of the conduit wall to optimize functional nerve recovery.
In response to severe injury to the skin or peripheral nerves, adult mammals typically undergo an irreversible repair process that results in contraction and the formation of non-physiologic scar tissue. However, recent advancements with induced regeneration using biologically active scaffolds have demonstrated that it is possible to intervene during the healing process to partially or near-completely restore the physiologic function of damaged skin or peripheral nerves. The aim of these scaffolds is to promote regeneration and minimize the contraction and scar formation mediated by stromal fibroblasts. For instance, some scaffolds appear to downregulate TGF-β signaling, a key inductor of myofibroblasts which promote contraction and scar formation. Two collagen-based and three synthetic-based regenerative devices have been approved by the Food and Drug Administration (FDA), two for the regeneration of the skin and three for the regeneration of peripheral nerves. Increasingly, these devices are establishing themselves as a viable alternative to autografting.
Functional regeneration of anisotropically aligned tissues such as ligaments, microvascular networks, myocardium, or skeletal muscle requires a temporal and spatial series of biochemical and biophysical cues to direct cell functions that promote native tissue regeneration. When these cues are lost during traumatic injuries such as volumetric muscle loss (VML), scar formation occurs, limiting the regenerative capacity of the tissue. Currently, autologous tissue transfer is the gold standard for treating injuries such as VML but can result in adverse outcomes including graft failure, donor site morbidity, and excessive scarring. Tissue-engineered scaffolds composed of biomaterials, cells, or both have been investigated to promote functional tissue regeneration but are still limited by inadequate tissue ingrowth. These scaffolds should provide precisely tuned topographies and stiffnesses using proregenerative materials to encourage tissue-specific functions such as myoblast orientation, followed by aligned myotube formation and recovery of functional contraction. In this study, we describe the design and characterization of novel porous fibrin scaffolds with anisotropic microarchitectural features that recapitulate the native tissue microenvironment and offer a promising approach for regeneration of aligned tissues. We used directional freeze-casting with varied fibrin concentrations and freezing temperatures to produce scaffolds with tunable degrees of anisotropy and strut widths. Nanoindentation analyses showed that the moduli of our fibrin scaffolds varied as a function of fibrin concentration and were consistent with native skeletal muscle tissue. Quantitative morphometric analyses of myoblast cytoskeletons on scaffold microarchitectures demonstrated enhanced cell alignment as a function of microarchitectural morphology. The ability to precisely control the anisotropic features of fibrin scaffolds promises to provide a powerful tool for directing aligned tissue ingrowth and enhance functional regeneration of tissues such as skeletal muscle.
This paper presents a systematic review of a key sector of the much promising and rapidly evolving field of biomedical engineering, specifically on the fabrication of three-dimensional open, porous collagen-based medical devices, using the prominent freeze-drying process. Collagen and its derivatives are the most popular biopolymers in this field, as they constitute the main components of the extracellular matrix, and therefore exhibit desirable properties, such as biocompatibility and biodegradability, for in vivo applications. For this reason, freeze-dried collagen-based sponges with a wide variety of attributes can be produced and have already led to a wide range of successful commercial medical devices, chiefly for dental, orthopedic, hemostatic, and neuronal applications. However, collagen sponges display some vulnerabilities in other key properties, such as low mechanical strength and poor control of their internal architecture, and therefore many studies focus on the settlement of these defects, either by tampering with the steps of the freeze-drying process or by combining collagen with other additives. Furthermore, freeze drying is still considered a high-cost and time-consuming process that is often used in a non-optimized manner. By applying an interdisciplinary approach and combining advances in other technological fields, such as in statistical analysis, implementing the Design of Experiments, and Artificial Intelligence, the opportunity arises to further evolve this process in a sustainable and strategic manner, and optimize the resulting products as well as create new opportunities in this field.
Freeze-drying is a well-established process in biomedical engineering for the fabrication of three-dimensional open-porous medical devices, especially those based on biopolymers. One of the most used biopolymers in this field is collagen, the most abundant protein in the human body and the main component of the extracellular-matrix, as well as its derivatives. Freeze-dried collagen-based sponges with a wide variety of attributes can be produced by design and have led to a wide range of successful commercial medical devices, foremost for dental, orthopedic, hemostatic and neuronal applications. However, this is still considered a high-cost and time-consuming process that is often used in a non-optimized manner. By combining advances in other technological fields, the opportunity arises to further evolve this process in a sustainable manner, and optimize the resulting products as well as create new opportunities in this field.
Peripheral nerve injury causes severe loss of motor and sensory functions, consequently increasing morbidity in affected patients. An autogenous nerve graft is considered the current gold standard for reconstructing nerve defects and recovering lost neurological functions; however, there are certain limitations to this method, such as limited donor nerve supply. With advances in regenerative medicine, recent research has focused on the fabrication of tissue-engineered nerve grafts as promising alternatives to the autogenous nerve grafts. In this study, we designed a nerve guidance conduit using an electrospun poly(lactide-co-ε-caprolactone) (PLCL) membrane with a visible light-crosslinked gelatin hydrogel. The PLCL nanoporous membrane with permeability served as a flexible and non-collapsible epineurium for the nerve conduit; the inner-aligned gelatin hydrogel paths were fabricated via 3D printing and a photocrosslinking system. The resultant gelatin hydrogel with microgrooved surface pattern was established as a conducting guidance path for the effective regeneration of axons and served as a reservoir that can incorporate and release bioactive molecules. From in vivo performance tests using a rat sciatic nerve defect model, our PLCL/gelatin conduit demonstrated successful axonal regeneration, remyelination capacities and facilitated functional recovery. Hence, the PLCL/gelatin conduit developed in this study is a promising substitute for autogenous nerve grafts.
Statement of Significance
Nerve guidance conduits(NGCs) are developed as promising recovery techniques for bridging peripheral nerve defects. However, there are still technological limitations including differences in the structures and components between natural peripheral nerve and NGCs. In this study, we designed a NGC composed of an electrospun poly(lactide-co-ε-caprolactone) (PLCL) membrane and 3D printed inner gelatin hydrogel to serve as a flexible and non-collapsible epineurium and a conducting guidance path, respectively, to mimic the fascicular structure of the peripheral nerve. In particular, in vitro cell tests clearly showed that gelatin hydrogel could guide the cells and function as a reservoir that incorporate and release nerve growth factor. From in vivo performance tests, our regenerative conduit successfully led to axonal regeneration with effective functional recovery.
Commercial nerve guidance conduits (NGCs) for repair of peripheral nerve discontinuities are of little use in gaps larger than 30 mm, and for smaller gaps they often fail to compete with the autografts that they are designed to replace. While recent research to develop new technologies for use in NGCs has produced many advanced designs with seemingly positive functional outcomes in animal models, these advances have not been translated into viable clinical products. While there have been many detailed reviews of the technologies available for creating NGCs, none of these have focussed on the requirements of the commercialisation process which are vital to ensure the translation of a technology from bench to clinic. Consideration of the factors essential for commercial viability, including regulatory clearance, reimbursement processes, manufacturability and scale up, and quality management early in the design process is vital in giving new technologies the best chance at achieving real-world impact. Here we have attempted to summarise the major components to consider during the development of emerging NGC technologies as a guide for those looking to develop new technology in this domain. We also examine a selection of the latest academic developments from the viewpoint of clinical translation, and discuss areas where we believe further work would be most likely to bring new NGC technologies to the clinic.
Statement of Significance
: NGCs for peripheral nerve repairs represent an adaptable foundation with potential to incorporate modifications to improve nerve regeneration outcomes. In this review we outline the regulatory processes that functionally distinct NGCs may need to address and explore new modifications and the complications that may need to be addressed during the translation process from bench to clinic.
Peripheral nerves have complex and precise structures that differ from other types of tissues and intrinsic regeneration abilities after injury. Spontaneous recovery is possible for neuropraxia and axonotmesis, while surgical treatment is required for neurotmesis. It remains a challenge to repair nerve gaps, a series of severe neurotmesis. It seems that 3 cm is the upper limit distance for primate peripheral nerves to regenerate spontaneously. Nerve autografts are the gold standard treatment for bridging nerve gaps. In the present review, current biomaterials for repairing gaps after peripheral nerve injury are briefly summarized. Moreover, the microstructure of the peripheral nerve, classifications of peripheral nerve injury, and the Wallerian degeneration are reviewed in the biological view and clinical practice. The failure of nerve regeneration in nerve conduits bridging longer than 3 cm gaps may be contributing to the insufficient vascularization of nerve conduit materials. Future researchers could focus on advanced biomaterials that promoting the angiogenesis of nerve conduits.
The pursuit of tissue engineering and regenerative medicine is to regenerate functional tissues and organs to replace damaged or diseased tissue. As a basic element of tissue engineering, scaffolds provide three-dimensional structure for cell growth while maintaining cell function, so the selection of scaffold materials is of great importance. Natural extracellular matrix is a potential therapeutic material because it is a naturally structured non-cellular tissue component with low immunogenicity, high biological activity, and can provide a unique microenvironment for cells. In addition, acellular materials can not only provide chemical and mechanical signals that determine the fate of cells, but also act as components of newly formed tissues, promoting damage repair. This chapter will focus on the repair ability of acellular materials in different tissues, and provide reference for the experimental research and development of different tissue repair materials and their clinical application.
Promising biomaterials should be tested in appropriate large animal models that recapitulate human inflammatory and regenerative responses. Previous studies have shown tyrosine-derived polycarbonates (TyrPC) are versatile biomaterials with a wide range of applications across multiple disciplines. The library of TyrPC has been well studied and consists of thousands of polymer compositions with tunable mechanical characteristics and degradation and resorption rates that are useful for nerve guidance tubes (NGTs). NGTs made of different TyrPCs have been used in segmental nerve defect models in small animals. The current study is an extension of this work and evaluates NGTs made using two different TyrPC compositions in a 1 cm porcine peripheral nerve repair model. We first evaluated a nondegradableTyrPC formulation, demonstrating proof-of-concept chronic regenerative efficacy up to 6 months with similar nerve/muscle electrophysiology and morphometry to the autograft repair control. Next, we characterized the acute regenerative response using a degradable TyrPC formulation. After 2 weeks in vivo, TyrPC NGT promoted greater deposition of pro-regenerative extracellular matrix (ECM) constituents (in particular collagen I, collagen III, collagen IV, laminin and fibronectin) compared to commercially available collagen-based NGTs. This corresponded with dense Schwann cell infiltration and axon extension across the lumen. These findings confirmed results reported previously in a mouse model and reveal that TyrPC NGTs were well tolerated in swine and facilitated host axon regeneration and Schwann cell infiltration in the acute phase across segmental defects - likely by eliciting a favorable neurotrophic ECM milieu. This regenerative response ultimately can contribute to functional recovery. This article is protected by copyright. All rights reserved.
Type I collagen is the main component of the extra-cellular matrix (ECM). In vitro, under a narrow window of physico-chemical conditions, type I collagen self-assembles to form complex supramolecular architectures reminiscent of those found in native ECM. Presently, a major challenge in collagen-based biomaterials is to couple the delicate collagen fibrillogenesis events with a controlled shaping process in non-denaturating conditions. In this work an ice-templating approach promoting the structuration of collagen into macroporous monoliths is used. Instead of common solvent removal procedures, a new topotactic conversion approach yielding self-assembled ordered fibrous materials is implemented. These collagen-only, non-cross-linked scaffolds exhibit uncommon mechanical properties in the wet state. With the help of the ice-patterned micro-ridge features, Normal Human Dermal Fibroblasts and C2C12 murine myoblasts successfully migrate and form highly-aligned populations within the resulting 3D biomimetic collagen scaffolds. These results open a new pathway to the development of new tissue engineering scaffolds ordered across various organization levels from the molecule to the macropore, and are of particular interest for biomedical applications where large scale 3D cell alignment is needed such as for muscular or nerve reconstruction.
Injury to the mammalian fetus is reversible during early stages of gestation and the spontaneous wound response is capable of restoring the structure and function of the original organ, a process called regeneration . By contrast, the unimpaired response to severe injury in adult mammals is an irreversible repair process leading to closure of the injured site by contraction and formation of scar, a nonphysiological tissue. The consequences of irreversible healing at the organ scale are far-reaching: they typically result in an essentially nonfunctional organ. Numerous approaches have been investigated to restore the loss of organ function in adults following irreversible injury. These strategies include transplantation, autografting, implantation of permanent prostheses, the use of stem cells, in vitro synthesis of the organ, and regenerative medicine (Yannas, Tissue and organ regeneration in adults. Springer; 2001). The last of these strategies is also referred to as induced organ regeneration , or the recovery of physiological structure and function of nonregenerative tissues in an organ (also known as de novo synthesis) by use of elementary reactants, such as biologically active scaffolds, either unseeded or seeded with cells. There is accumulating evidence that the spontaneous healing process of an injured organ in the adult mammal can be modified to yield a partially or completely regenerated organ. Regenerative medicine is an emerging field of study involving the implantation of biomaterials to facilitate formation (regeneration) of tissue in vivo. This field is undergoing rapid growth at this time, as evidenced by observation of regeneration or reported progress in on-going research efforts in a wide range of organs including skin (Butler and Orgill, Adv Biochem Eng Biotechnol 94:23-41, 2005), conjunctiva (Hatton and Rubin, Adv Biochem Eng Biotechnol 94:125-140, 2005), peripheral nerves (Zhang and Yannas, Adv Biochem Eng Biotechnol 94:67-89, 2005), bone (Mistry and Mikos, Adv Biochem Eng Biotechnol 94: 1-22, 2005), heart valves (Rabkin-Aikawa et al., Adv Biochem Eng Biotechnol 94:141-178, 2005), liver (Takimoto et al., Cell Transplant 12(4):413-421, 2003) articular cartilage (Kinner et al., Adv Biochem Eng Biotechnol 94:91-123, 2005), urological organs (Atala, Adv Biochem Eng Biotechnol 94:179-208, 2005), and the spinal cord (Verma and Fawcett, Adv Biochem Eng Biotechnol. 94:43-66, 2005). © Springer Science+Business Media, LLC 2012. All rights reserved.
Tissue engineering scaffolds are used extensively as three-dimensional analogs of the extracellular matrix (ECM). As a native component of the ECM, collagen-based biomaterials have a history of use in the field of tissue engineering. Collagen-glycosaminoglycan (CG) scaffolds in particular have long been utilized in vivo as ECM analogs for the regeneration of skin and peripheral nerves, and are currently being considered for the regeneration of a range of mineralized and soft tissues. Experimental characterization and theoretical modeling techniques have been used to describe a number of properties of CG scaffolds, notably pore microstructure, specific surface area, tensile and compressive mechanical properties, cell attachment, and permeability. Here we describe the fabrication, and characterization, and modeling of a series of CG and mineralized CG scaffolds. We will discuss their use in vitro as standardized 3D materials to study the influence of material parameters on cell phenotype and behaviors such as motility, contraction, differentiation, and matrix synthesis. We will conclude with a discussion of their use in vivo to induce tissue regeneration following injury for a range of soft and hard tissues, notably skin, peripheral nerves, brain, lung, cartilage, bone, fibrocartilage disks, and the conjunctiva.
Die Überbrückung kurzer Defekte von Nerven durch Röhrchen (Entubulation) ist eine einfache und elegante Lösung für ein komplexes Problem. Diese Technik erbringt eine praktische Lösung für viele mechanische-, zell- und biochemischen Forderungen, die überwunden werden müssen, um einen durchtrennten Nerven effektiv wieder zu vereinen, wobei dies nicht nur durch die klassische mikrochirurgische Neurorrhaphie gelöst wird.
Traumatic spinal cord injury (SCI) is a damage to the spinal cord that results in loss or impaired motor and/or sensory function. SCI is a sudden and unexpected event characterized by high morbidity and mortality rate during both acute and chronic stages, and it can be devastating in human, social and economical terms. Despite significant progresses in the clinical management of SCI, there remain no effective treatments to improve neurological outcomes. Among experimental strategies, bioengineered scaffolds have the potential to support and guide injured axons contributing to neural repair. The major aim of this study was to investigate a novel composite type I collagen scaffold with micropatterned porosity in a rodent model of severe spinal cord injury. After segment resection of the thoracic spinal cord we implanted the scaffold in female Sprague-Dawley rats. Controls were injured without receiving implantation. Behavioral analysis of the locomotor performance was monitored up to 55 days postinjury. Two months after injury histopathological analysis were performed to evaluate the extent of scar and demyelination, the presence of connective tissue and axonal regrowth through the scaffold and to evaluate inflammatory cell infiltration at the injured site. We provided evidence that the new collagen scaffold was well integrated with the host tissue, slightly ameliorated locomotor function, and limited the robust recruitment of the inflammatory cells at the injury site during both the acute and chronic stage in spinal cord injured rats. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2016.
A synthetic biodegradable nerve guide was constructed of two polymeric layers: an inner microporous layer prepared from a copolymer of L-lactide and epsilon-caprolactone (pore size range 0.5-1 micron) and an outer microporous layer prepared from a polyurethane/poly(L-lactide) mixture (pore size range 30-70 microns). This nerve guide was used to bridge a 7 mm gap in the right sciatic nerve of rats. It enabled the sciatic nerve to regenerate across the gap, forming a new, well-defined nerve that effectively re-established the contact between the proximial and distal nerve end, as effective as an autograft.
This communication presents an 11-center prospective randomized trial using the artificial dermis invented by Burke and Yannas. Patients with life-threatening burns who underwent primary excision and grafting within 7 days of injury had comparable sites randomized to receive either the artificial dermis (study site) or the investigator's usual skin grafting material (control site). Control materials were autograft, allograft, xenograft, or a synthetic dressing. Epidermal grafts were applied to the study site during a second operation, and surviving patients were followed for 1 year after grafting. One hundred thirty-nine sites on 106 patients were studied. Mean burn size was 46.5 +/- 15% mean total body surface (TBSA). Overall mortality was 13%, and mean hospital stay was 68 +/- 45 days. Median artificial dermis take was 80% compared with 95% for all comparative sites, but the take was equivalent to that of all nonautograft control materials. Results with the artificial dermis improved slightly as the investigators became more familiar with the material. Donor site thickness for the study site averaged .006'' +/- .002'' compared to .013'' +/- .018'' for control (p less than .0001) and the epidermal donor site healed an average of 4 days sooner (10 +/- 6 vs. 14 +/- 8 days) (p less than .0001). As the wounds matured during the first year, both patients and surgeons felt that both sites became more comparable in appearance and function. At the completion of the study, there was less hypertrophic scarring of the artificial dermis, and more patients preferred the artificial dermis to the control graft. Artificial dermis with an epidermal graft provides a permanent cover that is at least as satisfactory as currently available skin grafting techniques, and uses donor grafts that are thinner and donor sites that heal faster.
A monoclonal antibody (anti-alpha sm-1) recognizing exclusively alpha-smooth muscle actin was selected and characterized after immunization of BALB/c mice with the NH2-terminal synthetic decapeptide of alpha-smooth muscle actin coupled to keyhole limpet hemocyanin. Anti-alpha sm-1 helped in distinguishing smooth muscle cells from fibroblasts in mixed cultures such as rat dermal fibroblasts and chicken embryo fibroblasts. In the aortic media, it recognized a hitherto unknown population of cells negative for alpha-smooth muscle actin and for desmin. In 5-d-old rats, this population is about half of the medial cells and becomes only 8 +/- 5% in 6-wk-old animals. In cultures of rat aortic media SMCs, there is a progressive increase of this cell population together with a progressive decrease in the number of alpha-smooth muscle actin-containing stress fibers per cell. Double immunofluorescent studies carried out with anti-alpha sm-1 and anti-desmin antibodies in several organs revealed a heterogeneity of stromal cells. Desmin-negative, alpha-smooth muscle actin-positive cells were found in the rat intestinal muscularis mucosae and in the dermis around hair follicles. Moreover, desmin-positive, alpha-smooth muscle actin-negative cells were identified in the intestinal submucosa, rat testis interstitium, and uterine stroma. alpha-Smooth muscle actin was also found in myoepithelial cells of mammary and salivary glands, which are known to express cytokeratins. Finally, alpha-smooth muscle actin is present in stromal cells of mammary carcinomas, previously considered fibroblastic in nature. Thus, anti-alpha sm-1 antibody appears to be a powerful probe in the study of smooth muscle differentiation in normal and pathological conditions.
Résumé Au cours de la contraction du tissu de granulation, de nombreux fibroblastes acquièrent des caractéristiques ultrastructurelles qui les rendent semblables à des cellules musculaires lisses. Il est probable que ces éléments modifiés jouent un rôle dans le processus de contraction des plaies.
The process of wound repair is of vital importance for animals as well as for plants (Shigo, 1985), since a wound perturbs body homeostasis and may result in infection by microorganisms. A wound may occur without or with tissue loss (Robbins et al, 1984). In both cases, but more clearly in the second case, wound healing consists schematically of acute inflammation followed by formation of granulation tissue, a transitional tissue able to retract the wound space, and finally scar formation.
A porous biodegradable collagen-glycosaminoglycan bridge induced regeneration of myelinated and unmyelinated axons over large distances (15 mm) between the severed ends of the adult rat sciatic nerve. The newly formed strand of nerve tissue was well vascularized and sheathed in connective tissue 6 weeks following implantation of the polymeric guide. Nerve tissue did not grow in the absence of the polymeric guide.
This communication presents an 11-center prospective randomized trial using the artificial dermis invented by Burke and Yannas. Patients with life-threatening burns who underwent primary excision and grafting within 7 days of injury had comparable sites randomized to receive either the artificial dermis (study site) or the investigator's usual skin grafting material (control site). Control materials were autograft, allograft, xenograft, or a synthetic dressing. Epidermal grafts were applied to the study site during a second operation, and surviving patients were followed for 1 year after grafting. One hundred thirty-nine sites on 106 patients were studied. Mean burn size was 46.5 +/- 15% mean total body surface (TBSA). Overall mortality was 13%, and mean hospital stay was 68 +/- 45 days. Median artificial dermis take was 80% compared with 95% for all comparative sites, but the take was equivalent to that of all nonautograft control materials. Results with the artificial dermis improved slightly as the investigators became more familiar with the material. Donor site thickness for the study site averaged .006" +/- .002" compared to .013" +/- .018" for control (p < .0001) and the epidermal donor site healed an average of 4 days sooner (10 +/- 6 vs. 14 +/- 8 days) (p < .0001). As the wounds matured during the first year, both patients and surgeons felt that both sites became more comparable in appearance and function. At the completion of the study, there was less hypertrophic scarring of the artificial dermis, and more patients preferred the artificial dermis to the control graft. Artificial dermis with an epidermal graft provides a permanent cover that is at least as satisfactory as currently available skin grafting techniques, and uses donor grafts that are thinner and donor sites that heal faster. (C) Lippincott-Raven Publishers.
The ulnar nerve of a 21-year old man was repaired at the wrist by a silicone chamber technique 10 days after a traumatic transection. A 3 mm gap was left between the nerve ends inside the chamber. At follow-up three years later, motor and sensory recovery was excellent. At exploration at that time a macro-scopically normal nerve was found in the tube.
Sciatic nerve regeneration through implanted silicone tubes was compared to allogenic nerve grafting in 30 adult female Sprague-Dawley rats. Regeneration was assessed at 10, 24, and 90 days post-transection. Axonal regrowth through the implanted neural prostheses was evaluated with electromyography and histologic examination.
Contractile fibroblasts were sought via electron microscopy in liver tissue from 12 patients with chronic alcoholic cirrhosis, and from 3 noncirrhotic patients. In 9 of the 12 cirrhotic livers, contractile fibroblasts (myofibroblasts) were seen containing the classic features of 60-80 A microfilament bundles with electron-dense bodies along with prominent microtubules. The remaining three cirrhotic liver specimens had fibroblasts containing microfilament bundles but without electron-dense bodies. Myofibroblasts were not found in any of the noncirrhotic livers. Just as in other types of scars, myofibroblasts are probably the active contractile force in the scarring and fibrosis which characterize chronic alcoholic cirrhosis.
Fibrous tissue capsules, from around silicone breast implants, were examined by light microscopy, and by transmission and scanning electron microscopy, the latter combined with energy dispersive X-ray analysis. Contractile fibroblasts (myofibroblasts) were found in most of these fibrous capsules, and especially in those recently formed or manipulated. The element silicon was positively identified by X-ray analysis in half the tissue specimens. This material was found only around gel-filled implants. In the specimens studied, the clinical hardness of the breast was unrelated to the presence of silicon in the capsule.
Sterile 7 mm. polyethylene implants were implanted in standard 6 mm. bilateral midfibular fracture gaps in thirty rats (sixty defects), and six rats were used as controls with identical bilateral midfibular fracture gaps. Clinical, radiologic, and histologic studies at a 12-week interval revealed osteogenesis at internal and external surfaces of implants resulting in bony union; however, gaps without implants resulted in fibrous union and nonunion.
A study was conducted to compare the regeneration of rat peroneal nerves across 0.5 cm gaps repaired with artificial nerve grafts (ANG) versus sutured autografts (SAG). The ANG model is composed of a synthetic biodegradable passive conduit made of glycolide trimethylene carbonate (GTMC) filled with a collagen matrix (predominantly Type I collagen, derived from calf skin, and with the telopeptide ends left intact). Axonal regeneration was studied in 11 long-term animals (two at 6 months and nine at 9 months). The nerves were studied by qualitative and quantitative histological, electrophysiological, and functional assays. Axonal regeneration with the ANG was equal to SAGs as measured by axonal diameters, physiological, and functional methods, although the SAG demonstrated statistically higher axonal counts.
Biodegradable polyurethane-based (PU) nerve guides, instilled with or without ACTH4-9 analog (a melanocortin) were used for bridging an 8 mm gap in the rat sciatic nerve and were evaluated for function and histological appearance after 16 weeks of implantation. Autologous nerve grafts functioned as controls. The guides successfully enabled the sciatic nerve to regenerate across the 8 mm gap, thus effectively reestablishing the contact between the proximal and distal nerve ends. The mean conduction velocity, motor latency, and muscle action potentials of all the nerve guides did not differ significantly from the autografts. The histological quality of the regeneration in the nerve guides was significantly better than in the autografts; in the nerve guides, a well-defined nerve cable of normal architecture had regenerated without extensive endoneural scarring as seen in the autografts. ACTH4-9 instilled in the nerve guides showed a slight, but significant, increase in the number of myelinated axons. It is concluded that biodegradable PU nerve guides result in similar functional recovery when compared with autografts, but their histological quality is significantly better. ACTH4-9 showed only slight, but significant, improved nerve growth promoting activity. Therefore biodegradable PU nerve guides with ACTH4-9 would appear to be promising alternatives to autografts for bridging nerve defects.
When a peripheral nerve is severed and left untreated, the most likely result is the formation of an endbulb neuroma; this tangled mass of disorganized nerve fibers blocks functional recovery following nerve injury. Although there are several different approaches for promoting nerve repair, which have been greatly refined over recent years, the clinical results of peripheral nerve repair remain very disappointing. In this paper we compare the results of a collagen nerve guide conduit to the more standard clinical procedure of nerve autografting to promote repair of transected peripheral nerves in rats and nonhuman primates.
In rats, we tested recovery from sciatic nerve transection and repair by (1) direct microsurgical suture, (2) 4 mm autograft, or (3) entubulation repair with collagen‐based nerve guide conduits. Evoked muscle action potentials (MAP), were recorded from the gastrocnemius muscle at 4 and 12 weeks following sciatic nerve transection. At 4 weeks the repair group of direct suture demonstrated a significantly greater MAP, compared to the other surgical repair groups. However, at 12 weeks all four surgical repair groups displayed similar levels of recovery of the motor response.
In six adult male Macaca fascicularis monkeys the median nerve was transected 2 cm above the wrist and repaired by either a 4 mm nerve autograft or a collagen‐based nerve guide conduit leaving a 4 mm gap between nerve ends. Serial studies of motor and sensory fibers were performed by recording the evoked MAP from the abductor pollicis brevis muscle (APB) and the sensory action potential (SAP) evoked by stimulation of digital nerves (digit II), respectively, up to 760 days following surgery. Evoked muscle responses returned to normal baseline levels in all cases. Statistical analysis of the motor responses, as judged by the slope of the recovery curves, indicated a significantly more rapid rate of recovery for the nerve guide repair group. The final level of recovery of the MAP amplitudes was not significantly different between the groups. In contrast, the SAP amplitude only recovered to the low normal range and there were no statistically significant differences between the two groups in terms of sensory recovery rates.
The rodent and primate studies suggest that in terms of recovery of physiological responses from target muscle and sensory nerves, entubulation repair of peripheral nerves with a collagen‐based nerve guide conduit over a short nerve gap (4 mm) is as effective as a standard nerve autograft. Furthermore, preliminary results show that entubulation repair with this material can support axon regeneration and maturation over a nerve gap distance of at least 15 mm.
Integra artificial skin is an effective means of treatment for full-thickness burns. In extensive burn injury the use of such skin substitutes may become the treatment of choice. The artificial skin consists of a dermal substitute of bovine collagen and chondroitin-6-sulfate and an epidermal layer of synthetic polysiloxane polymer (Silastic). Serial biopsy specimens were obtained from 131 patients during a period of 7 days to 2 years after application. In this histologic study, six sequential phases of repair were discerned. In addition, there were occasional unusual histologic features, eosinophilic infiltration, and/or macrophage-derived giant cell formation in the wound area; however, such findings did not clinically correlate with a negative response to Integra artificial skin. Good repair was obtained, with rare exceptions. An intact dermis was achieved as well as definitive closure of a complete epidermal layer with a minimum of scarring.
Prosthetic meniscal replacement offers the ability to stabilize the meniscectomized knee and provide prophylaxis against early degenerative arthritis. Since prosthetic meniscal replacement may be performed in the setting of normal articular cartilage, a prosthesis will be required to match the exact joint configuration, induce the same lubricity, produce the same coefficient of friction, and absorb and dampen the same joint forces (without incurring significant creep or abrasion) as does the normal meniscus. This feat is currently beyond the capabilities of artificial materials alone. Alternatively, collagen-based prostheses acting as resorbable regeneration templates offer the possibility of inducing regrowth of new menisci. This paper presents a summary of hypotheses, considerations, and laboratory evidence for the use of collagen-based, resorbable matrices as regeneration templates.
A large gap in peripheral nerve will not allow effective regeneration unless a grafting conduit is used to bridge the defect. Conventionally, nerve tissue has been used as such a conduit in nerve reconstruction; however, results from techniques using these grafts are often unsatisfactory. A number of recent investigations have indicated that nerve fibers will regenerate through a non-neural tube. The purpose of this review is: 1) to provide an overview of the various tubulation techniques previously reported for peripheral nerve gap repair; 2) to investigate new possibilities for enhancing the regenerative capacity of nerves following these tubulation techniques by drawing from technical innovations in microsurgery and recent progress in immunology and neurobiology. The interposed graft thus may perform a more positive role, not merely as a pathway for deregenerating axons, but as a source for neuronotrophic factors and neurite-promoting factors, which would nurture and guide the neurons and axons. Such modifications in graft materials may lead to clinical applications of tubulation of nerve defects that would result in an improvement in clinical results.
Three cases are reported to illustrate the potential danger of silicon-polymer intubulation of nerve for either nerve repair or following neurolysis. Since silicon-polymer intubulation of nerve is now a proven model for producing chronic nerve compression, its use clinically may be contraindicated where neural regeneration is the desired goal.
Sciatic nerve regeneration through implanted silicone tubes was compared to allogenic nerve grafting in 30 adult female Sprague-Dawley rats. Regeneration was assessed at 10, 24, and 90 days post-transection. Axonal regrowth through the implanted neural prostheses was evaluated with electromyography and histologic examination.
Regeneration of the dermis does not occur spontaneously in the adult mammal. The epidermis is regenerated spontaneously provided there is a dermal substrate over which it can migrate. Certain highly porous, crosslinked collagen-glycosaminoglycan copolymers have induced partial morphogenesis of skin when seeded with dermal and epidermal cells and then grafted on standard, full-thickness skin wounds in the adult guinea pig. A mature epidermis and a nearly physiological dermis, which lacked hair follicles but was demonstrably different from scar, were regenerated over areas as large as 16 cm2. These chemical analogs of extracellular matrices were morphogenetically active provided that the average pore diameter ranged between 20 and 125 microns, the resistance to degradation by collagenase exceeded a critical limit, and the density of autologous dermal and epidermal cells inoculated therein was greater than 5 x 10(4) cells per cm2 of wound area. Unseeded copolymers with physical structures that were within these limits delayed the onset of wound contraction by about 10 days but did not eventually prevent it. Seeded copolymers not only delayed contraction but eventually arrested and reversed it while new skin was being regenerated. The data identify a model extracellular matrix that acts as if it were an insoluble growth factor with narrowly specified physiochemical structure, functioning as a transient basal lamina during morphogenesis of skin.
Light and electron microscopy were used to investigate long-term regeneration in peripheral nerves regenerating across a 10 mm gap through silicone tubes. Schwann cells and axons co-migrated behind an advancing front of fibroblasts, bridging the 10 mm gap between 28 and 35 days following nerve transection. Myelination of regenerated fibres started between 14 and 21 days after transection and occurred in a manner similar to that reported during development. Although these early events were successful in producing morphologically normal-appearing regenerated fibres, complete maturation of many of these fibres was never achieved. Axonal distortion by neurofilaments, axonal degeneration and secondary demyelination were seen at 56 days following nerve transection. These changes progressed in severity with time as more axons advanced through the distal stump towards their peripheral target. Since regeneration occurs in the absence of endoneurial tubes, and because constrictive forces act on the nerve during regeneration, we suggest that these extrinsic factors limit the successful advancement of axons through the distal stump to their target organ.
These experiments present quantitative data concerning peripheral nerve regeneration in vivo. We used entubulation repair as a model to compare two different types of tubular prostheses, one nonbiodegradable and the other biodegradable. We modified the microenvironment of the regenerating axons within the tubular prostheses by adding a laminin-containing gel to the interior of the tube at the time of initial implantation. The data demonstrate that specific manipulations to the microenvironment of regenerating peripheral axons have quantitative effects on the rate and extent of nerve regeneration. Such effects were dependent on the composition of the tubular prosthesis and varied according to the survival time of the animals. For instance, the laminin gel within the biodegradable tubes enhanced nerve regeneration at 2 weeks but was inhibitory at 6 weeks. Furthermore, such manipulations may have different effects on the number of myelinated axons found within the regenerating nerve cable versus the number of primary motor and sensory neurons giving rise to such axons. We concluded that: the presence of a laminin-containing gel significantly increased the initial rate at which axons from primary sensory and motor neurons cross a transection site; an initial delay in axonal outgrowth at early time points did not necessarily predict diminished outgrowth at later times; and because of the potential for axonal branching the number of myelinated axons found in the midportion of a tubular prosthesis did not always correlate with the number of primary motor and sensory neurons which gave rise to those axons.
This study investigated nerve regeneration through a pseudosynovial sheath in a primate. After resecting a 3-cm segment of the ulnar nerve at the elbow, the two ends of the divided nerve were placed at either end of the pseudosynovial tube, positioning the nerve ends such that they were separated by a 3-cm gap. Histologic evaluation at 3, 5, 7, and 9 months demonstrated evidence of nerve regeneration across this gap. Nerve fiber diameter and density assessment demonstrated a maturation of the fiber pattern with time. The overall morphologic pattern of the regenerating nerve within the pseudosheath was that of multiple small fascicles, each within its own perineurial compartment. This pattern resembled neither the proximal nor distal nerve fascicular pattern.
In 6 cases of Dupuytren's disease and 1 of Ledderhose's disease, the nodules of the palmar and plantar aponeurosis were examined by light and electron microscopy. The cells composing these nodules, presumably fibroblasts, showed three significant ultrastructural features: (1) a fibrillar system similar to that of smooth muscle cells; (2) nuclear deformations such as are found in contracted cells, the severest being recognizable by light microscopy (cross-banded nuclei); (3) cell-to-cell and cell-to-stroma attachments. Based on these data and on recent information about the biology of the fibroblasts, it is suggested that these cells are fibroblasts that have modulated into contractile cells (myofibroblasts), and that their contraction plays a role in the pathogenesis of the contracture observed clinically.
Nontoxic, bioresorbable "nerve guide" tubes were used to bridge the transected optic nerves of adult rats. Nerve guides were fabricated as polymers of synthetic poly D,L-lactates with 2% triethyl citrate added as a plasticizer. The local environment was manipulated further by the addition of the proteins collagen, fibrinogen, and anti-Thy-1 antibody to the nerve guide lumens at the time of operation. Neovascular growth through the nerve guide lumens was quantified with the aid of a computer-controlled microscope. Neovascular growth was greater in the nerve guides to which proteins had been added, compared with initially empty nerve guides. These experiments demonstrated the effectiveness of these nerve guide tubes in supporting and directing neovascular growth in the mammalian central nervous system, and suggested that specific alterations of the local environment within the nerve guide lumen can affect the extent of neovascular growth.
An examination of the processes involved in wound healing, from reactions to injury through to scarring
Present techniques of nerve repair by suture are based on an anatomical approach. The severed layers of connective tissue are reapproximated. Another approach to nerve repair is to separate the specific cellular components of the peripheral nerve that contribute to fibrous healing and nerve regeneration. The perineurium normally separates the extrafascicular epineurium of mesodermal origin from the intrafascicular endoneurium of ectodermal origin. A cellular approach to nerve repair would use a tube around the fascicle as an artificial perineurium to separate fibrous healing from axonal regeneration until the perineurium reestablishes its continuity. Fascicular tubulization with polyglycolic acid tubes was studied in 25 rats. The polyglycolic acid tube is resorbed after the perineurium has reestablished its continuity. The repairs by fascicular tubulization demonstrated improved organization of the repair site compared to suture repairs. The diameter histograms of the regenerated myelinated axons were similar in repairs by tube and suture techniques. The total regenerated cross-sectional area of the myelinated axons was similar in the repairs by fascicular tubulization to repairs by fascicular suture.
An experimental model of a free, empty perineurial tube for use as a nerve graft is presented as an alternative to existing methods. The technique is described and the model compared with conventional nerve graft in the sciatic nerve of a rabbit. Results are evaluated with EMG studies, angiography, and histology. The experimental model compared favorably with the standard graft. Further avenues of investigation and clinical use are suggested.
The successful regeneration of a multifascicular, complete peripheral nerve through a tubular synthetic biodegradable nerve guide across a gap of 10 mm in the rat sciatic nerve is reported. The importance of the distal nerve as a source of target-derived neuronotrophic factors necessary for the successful regeneration of the proximal regenerating nerve is emphasized. A simplified research model for further investigation into and manipulation of the biological processes of nerve regeneration is described. The potential clinical utilization of this model in the management of peripheral nerve injuries and, ultimately, central nervous system lesions is mentioned.
The characteristics and long-term fate of the pseudosynovial sheath formed in response to a gliding tendon implant were examined. A primate model was chosen to reproduce the human clinical situation. Hunter passive gliding implants were implanted in 32 digits of eight Macque monkeys. Surgical syndactylism was created to the adjacent active digits to provide passive range of motion of the digits. No implanted digits demonstrated progressive flexion contractures. Radiographs revealed passive excursion of the implants of an average of 2.5 cm. Biopsies were taken at various time intervals and locations for histological examination. The pseudosynovial sheath has three descriptive layers: an intima, media, and adventitia. This sheath becomes mature and stable at 8 weeks. The intima cells contain a glycosaminoglycan substance and have a secretory capacity. The media cells have large amounts of collagen and provide structural and vascular support. The adventitia is a highly vascular structure composed of loose fibrous tissues that demonstrates clefts that may represent gliding planes. The pseudosynovial sheath was found to be a morphologically stable structure that showed no propensity for longitudinal contracture. The sheath appears to have the characteristics necessary to provide a good bed for a tendon graft.
Sciatic nerves of mice were cut and the early regenerative stages were studied after the stumps had been encased within plastic tubes and kept separate by a gap of 5 mm. Only isolated cells were seen inside the tube after 7 days; after 12 days active regeneration and myelination were seen proximally; more distally, cells with long processes formed large spaces filled with collagen and less numerous Schwann cells. Zonulae occludentes and segments of basal lamina became more evident at a later stage. One month after the operation an almost complete regeneration of the nerve had taken place and perineurial cells were lined by a continuous basal lamina. The regeneration of the perineurium seemed to take place from fibroblasts; their cytoplasm as well as that of Schwann cells contained fibrillary material at this stage, sometimes in relation to segments of basal lamina. The results of this study indicate that both types of cells take part in the formation of endoneurial structures and that the early arrangement of fibroblasts contributes to the orderly longitudinal alignment of collagen fibrils.
Severe nerve injuries may require microsurgical grafting to span a defect. Introduction of graft material into a highly vascular recipient bed is documented to aid in early regeneration of neuronal blood supply.
A silicone rod (SR)-induced fibrovascular sheath was employed to evaluate the regeneration of rat tibial nerve through 2-mmdiameter collagen tubes (CT) or contralateral nerve autografts (AUTO). At first operation, 5 mm of right tibial nerve was resected from 30 retired male breeder Sprague-Dawley rats. Resected nerve was replaced with either a 5 x 2 mm SR or the nerve ends were sutured to the intermuscular fascia.
Four weeks later, animals were repaired by replacing the SR with either a CT or a contralateral AUTO from the left tibial nerve. Three months later, EMG testing was performed, and histologic sections were prepared.
The EMG latency and the size of the compound action potential for sheathed or non-sheathed CT or AUTO were statistically superior to controls at the 95% confidence level. All other intergroup comparisons of latency and action potential size were statistically insignificant. The proportion of nerve fibers traversing the surgical sites was not influenced by the method of repair or by the presence or absence of sheathing. Tubulized repairs most closely resembled unoperated nerves, and autografted repairs had a large diameter, but much fibrosis, whereas controls displayed immaturity and disorganization.
Our observations suggest that there was no difference between repairs performed with or without a vascular pseudosheath. However, CT supported regeneration better than did AUTO repair.
The spatial-temporal progress of peripheral nerve regeneration across a 10-mm gap within a silicone chamber was examined with the light and electron microscope at 2-mm intervals. A coaxial, fibrin matrix was observed at 1 week with a proximal-distal narrowing that extended beyond the midpoint of the chamber. At 2 weeks, Schwann cells, fibroblasts, and endothelial cells had migrated into the matrix from both nerve stumps. There was a delay of 7-14 days after nerve transection and chamber implantation before regenerating axons appeared in the chamber. At 2 weeks, nonmyelinated axons were seen only in the proximal 1-5 mm of the chamber in association with Schwann cells. Axons reached the distal stump by 3 weeks and a proximal-distal gradient of myelination was observed. These observations define the parameters of a morphologic assay for regeneration in this chamber model which can be used to investigate cellular and molecular mechanisms underlying the success of peripheral nerve regeneration.
Defects in a sectioned tibial nerve were bridged by a new method using a polyglactin mesh-tube and compared with conventional nerve grafting in the rabbit. The capability of healing was evaluated by morphometrical observations and repeated EMG-recordings. Only minor differences between the two different techniques were observed and the possible advantage of the polyglactin method is discussed.
An experimental model is presented for studying nerve regeneration over gaps of various lengths between the both ends of a severed nerve. After transferring left and right sciatic nerves of rat to the back, the gap between the two nerve ends could be bridged by a preformed, tube-shaped mesothelial chamber of a desired length. When the gap length was 10 mm or less, a well developed nerve structure was generated in the chamber between the nerve ends, and axons from the left sciatic nerve reinnervated muscles in the right limb via the right sciatic nerve. When the gap length was extended to 15 mm or more no such regeneration occurred. When no distal nerve end was introduced into the chamber, there was a limited growth into this chamber over only 5-6 mm. This "limited growth phenomenon" is discussed with respect to a lack of "trophic" or cellular support from a distal nerve segment. It is also proposed that the termination of growth, seen under these circumstances, may suggest a new principle for avoiding the development of neuromas after nerve transections.
A bilayer artificial skin composed of a temporary Silastic epidermis and a porous collagen-chondroitn 6-sulfate fibrillar dermis, which is not removed, has been used to physiologically close up to 60% of the body surface following prompt excision of burn wounds in ten patients whose total burn size covered 50--95% body surface area (BSA). Following grafting, the dermal portion is populated with fibroblasts and vessels from the wound bed. The anatomic structure of the artificial dermis resembles normal dermis and serves as a template for the synthesis of new connective tissue and the formation of a "neodermis," while it is slowly biodegraded. This artificial skin has physiologically closed excised burn wounds for periods of time up to 46 days before the Silastic epidermis was removed. At the time of election when donor sites are ready for reharvesting, the Silastic epidermis is removed from the vascularized artificial dermis and replaced with 0.004 autoepidermal graft in sheet or meshed form. Clinical and histologic experience in a relatively short follow-up period (2--16 months) indicates that "neodermis" retains some of the anatomic characteristics and behavior of normal dermis, thus promising improvement in the functional and cosmetic results, as well as providing physiologic function as a skin substitute. The artificial skin is easily sterilized and stored at room temperature, capable of large scale production, and immediately available for grafting, indicating its potential for easy and relatively economic use in the burn patient.
The authors believe that the silicone chamber model discussed promises to be an outstanding new tool for neurobiological investigation of neural regeneration: One can contrast properties and behaviors in situations favoring or hindering the repair of motor and sensory (and sympathetic) nerves and, thus begin to recognize physical, biochemical, and cellular influences that bear on the outcome of a regenerative process. One can analyze the composition of the fluid environment; the matrix in which nerve fibers will or will not grow; the temporal changes occurring in the proximal and distal components; the reactions of the source neurons in the spinal cord and dorsal root ganglia, and of their partner glial cells in their various locations; and, the roles and responses of the end organs addressed by the nerve. One can introduce and investigate the effects of (among several other possible manipulations) different factors, antibody against them, hormones, nutrients, drugs; glial cells of PNS or CNS origin, as well as connective or other cells; and semisolid matrices or experimental terrains for neurite promotion and/or directional guidance. Obviously, a vast amount of work needs to be carried out before the model described here will fulfill even part of its promises. Eventually, however, one should be able to explore the applications of this model to selected problems of interest to both neurobiologists and neuropathologists, for example: Myelin degeneration and regeneration, peripheral neuropathies (including genetic defects expressed in mouse neurological mutants), compression damage on regenerated nerves, or the role of electrical stimulation in nerve repair. Lastly, one may be able to adapt this PNS model to CNS tissues such as optic nerve and spinal cord: if so, what one will have learned about repair processes and the humoral or cellular influences which control them in the PNA might become applicable at least in part to the much greater complexity of CNS regeneration.
Prompt and long-term closure of full-thickness skin wounds is guinea pigs and humans is achieved by applying a bilayer polymeric
membrane. The membrane comprises a top layer of a silicone elastomer and a bottom layer of a porous cross-linked network of
collagen and glycosaminoglycan. The bottom layer can be seeded with a small number of autologous basal cells before grafting.
No immunosuppression is used and infection, exudation, and rejection are absent. Host tissue utilizes the sterile membrane
as a culture medium to synthesize neoepidermal and neodermal tissue. A functional extension of skin over the entire wound
area is formed in about 4 weeks.
Polyglactin 910, a resorbable synthetic material, was used as a mesh-tube to bridge defects (7 to 9 mm in length) in a sectioned rabbit tibial nerve. After absorption of the mesh a new nerve sheath was formed which enclosed numerous minute fascicles of regenerating axons. The polyglactin tube influenced the direction taken by the regenerating axons and guided them into the distal segment. The tube also reduced the formation of neuromas and the growth of scar tissue from surrounding structures.
This study was performed to determine whether vein grafts might serve as a conduit for nerve regeneration. A 1 cm segment of sciatic nerve was removed bilaterally in 12 Sprague-Dawley rats. On one side the gap was not repaired, and on the other side a segment of femoral vein was used to bridge the nerve gap. Nerve conduction studies and necropsies were performed at intervals. Reconstitution of nerve trunk continuity and healing of plantar ulcers occurred only in the vein-grafted side. Histologic examination revealed orderly growth of nerve fibers within the lumen of the vein grafts as early as 1 month after repair. Most regenerating nerve fibers passed through the proximal junction in an orderly pattern and reached the distal stumps within 2 months after repair. Results of nerve conduction study at 4 months after operation demonstrated restoration of conduction through the vein-grafted sciatic nerves with muscle reinnervation. Nearly normal muscle fibers in the gastrocnemius on the repaired side were confirmed at necropsy. This study demonstrated that autogenous vein grafts can serve as a conduit for nerve regeneration in rodents.
We describe an experimental in vivo system for studying peripheral nerve regeneration, in which the proximal stump of a transected nerve regrows through a transparent silicone chamber toward the distal stump. Physical separation permits examination of the effects of the humoral and/or cellular influences from the distal stump on regenerating fibers before they invade the distal segment itself. A small segment of the rat sciatic nerve was resected, leaving a 6 mm gap which was then encased by a cylindrical silicone chamber. Within the first weeks, a nerve trunk regenerated along the central axis of the chamber bridged the gap between the proximal and distal stumps. When the distal nerve stump was omitted from the distal opening of the chamber, only a thin structure with a few small-caliber fibers extended across the gap. In each instance regenerating nerve appeared as a cord-like structure completely surrounded by clear fluid, a feature which permits easy collection of the extracellular fluid for analysis of its chemical properties and biological activity. This feature also allows in vivo manipulation of the humoral environment in which nerve regeneration occurs.
The range of growth-promoting influences from a distal nerve stump on a regenerating proximal stump was determined using an experimental system in which a gap between cross-anastomosed rat sciatic nerves was encased by a cylindrical silicone chamber. Two arrangements were examined after 1 month in situ: A proximal-distal (PD) system in which both proximal and distal stumps were introduced into the ends of the chamber, and a proximal-open (PO) system in which the distal stump was omitted. When the gap was 6 mm long, a regenerated nerve extended all the way through the chamber in both the PD and PO systems. When the gap was increased to 10 mm, a similar regrowth occurred in the PD chamber, whereas in the PO chamber proximal regrowth was partial or nonexistent. When the gap was increased to 15 mm, no regeneration occurred, even in the presence of the distal stump. These observations confirm that the distal stump influences proximal regeneration and indicate that this influence can act only over a limited distance or volume. Such an influence could consist of humoral agents which support nerve growth and/or outgrowth from the distal stump.