Robust CNS regeneration after complete spinal cord transection using aligned poly-L-lactic acid microfibers

International Center for Spinal Cord Injury, Hugo W. Moser Research Institute at Kennedy Krieger, Baltimore, MD 21205, USA.
Biomaterials (Impact Factor: 8.56). 05/2011; 32(26):6068-79. DOI: 10.1016/j.biomaterials.2011.05.006
Source: PubMed


Following spinal cord injury, axons fail to regenerate without exogenous intervention. In this study we report that aligned microfiber-based grafts foster robust regeneration of vascularized CNS tissue. Film, random, and aligned microfiber-based conduits were grafted into a 3 mm thoracic rat spinal cord gap created by complete transection. Over the course of 4 weeks, microtopography presented by aligned or random poly-L-lactic acid microfibers facilitated infiltration of host tissue, and the initial 3 mm gap was closed by endogenous cell populations. This bulk tissue response was composed of regenerating axons accompanied by morphologically aligned astrocytes. Aligned fibers promoted long distance (2055 ± 150 μm), rostrocaudal axonal regeneration, significantly greater than random fiber (1162 ± 87 μm) and film (413 ± 199 μm) controls. Retrograde tracing indicated that regenerating axons originated from propriospinal neurons of the rostral spinal cord, and supraspinal neurons of the reticular formation, red nucleus, raphe and vestibular nuclei. Our findings outline a form of regeneration within the central nervous system that holds important implications for regeneration biology.

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    • "In this work, the combination of laminin coated, aligned electrospun fibers with simultaneous exogenous electrical stimulation were explored to examine the impact of biophysical and topographical cues on neurite outgrowth. Previously, we demonstrated that neurite outgrowth is independently enhanced by both aligned electrospun PLLA fibers (Hurtado et al 2011) and by exogenous electrical stimulation (50 mV mm −1 ) (Koppes et al 2011). We hypothesize that the combination (topography with exogenous electrical stimulation) will synergistically enhance neurite outgrowth in a directional manner. "
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    ABSTRACT: Objective: Both electrical stimuli (endogenous and exogenous) and topographical cues are instructive to axonal extension. This report, for the first time, investigated the relative dominance of directional topographical guidance cues and directional electrical cues to enhance and/or direct primary neurite extension. We hypothesized the combination of electrical stimulation with electrospun fiber topography would induce longer neurite extension from dorsal root ganglia neurons than the presence of electrical stimulation or aligned topography alone. Approach: To test the hypothesis, neurite outgrowth was examined on laminin-coated poly-L-lactide films or electrospun fibers (2 µm in diameter) in the presence or absence of electrical stimulation. Immunostained neurons were semi-automatically traced using Neurolucida software and morphology was evaluated. Main results: Neurite extension increased 74% on the aligned fibers compared to film controls. Stimulation alone increased outgrowth by 32% on films or fibers relative to unstimulated film controls. The co-presentation of topographical (fibers) with biophysical (electrical stimulation) cues resulted in a synergistic 126% increase in outgrowth relative to unstimulated film controls. Field polarity had no influence on the directionality of neurites, indicating topographical cues are responsible for guiding neurite extension. Significance: Both cues (electrical stimulation and fiber geometry) are modular in nature and can be synergistically applied in conjunction with other common methods in regenerative medicine such as controlled release of growth factors to further influence axonal growth in vivo. The combined application of electrical and aligned fiber topographical guidance cues described herein, if translated in vivo, could provide a more supportive environment for directed and robust axonal regeneration following peripheral nerve injury.
    Journal of Neural Engineering 06/2014; 11(4):046002. DOI:10.1088/1741-2560/11/4/046002 · 3.30 Impact Factor
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    • "han , & Ramakrishna , 2008 ) . Furthermore , tuning the fiber dimension and pattern of PLLA fibers regulates the viability , proliferation , and neurite outgrowth of neonatal mouse cerebellum C17 . 2 cells ( He et al . , 2010 ) . Hur - tado et al . investigated the repair of a completely transected spinal cord with a 3 - mm defect in a rat model ( Hurtado et al . , 2011 ) by filling the spinal cord gap with cells enclosed in PLLA electrospun fiber conduits four weeks after surgery . Kim et al . carried out both in vitro and in vivo studies on the impacts of aligned electrospun poly ( acrylonitrile - co - methylacrylate ) fibers on neuron outgrowth and Schwann cell migration for the repair of a 17 - mm "
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    ABSTRACT: Based on accumulating evidence that the 3D topography and the chemical features of a growth surface influence neuronal differentiation, we combined these two features by evaluating the cytotoxicity, proliferation, and differentiation of the rat PC12 line and human neural stem cells (hNSCs) on chitosan (CS), cellulose acetate (CA), and polyethersulfone (PES)-derived electrospun nanofibers that had similar diameters, centered in the 200-500nm range. None of the nanofibrous materials were cytotoxic compared to 2D (e.g., flat surface) controls; however, proliferation generally was inhibited on the nanofibrous scaffolds although to a lesser extent on the polysaccharide-derived materials compared to PES. In an exception to the trend toward slower growth on the 3D substrates, hNSCs differentiated on the CS nanofibers proliferated faster than the 2D controls and both cell types showed enhanced indication of neuronal differentiation on the CS scaffolds. Together, these results demonstrate beneficial attributes of CS for neural tissue engineering when this polysaccharide is used in the context of the defined 3D topography found in electrospun nanofibers.
    01/2014; 99:483-90. DOI:10.1016/j.carbpol.2013.08.050
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    • "Thus, it had been suggested that pHPMA hydrogel had a strong stimulating effect on neurons and Schwann cells activity as well as supporting revascularization of the lesion site and having an apparent protective effect against Wallerian degeneration (Woerly et al., 1999, 2001a, 2001b). Finally, it had been suggested that because of its physico-chemical properties, the pHPMA hydrogel may have therapeutic interest in the repair of spinal cord lesion (Woerly et al., 1990, 1996a, 1996b) compared with various materials used to experimentally bridge the spinal cord lesion such as collagen matrix (Marchand and Woerly, 1990), guidance channels (tubes or cylinders) of poly(acrylonitrile-vinylchloride) (Xu et al., 1995), polycarbonate (Montgomery et al., 1996), poly(α-hydroxyacids) (Gautier et al., 1998; Oudega et al., 2001), silicone (Borgens, 1999), polyethylene glycol (Shi et al., 1999), poly(lactic acid) (Hurtado et al., 2011) and poly(lactic-co-glycolic acid) (Moore et al., 2006; Chen et al., 2009). "
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    ABSTRACT: There have been considerable interests in attempting to reverse the deficit due to a spinal cord injury by restoring neural pathways through the lesion and by rebuilding tissue network. In order to provide an appropriate micro-environment for regrowing axotomized neurons and proliferating and migrating cells, we have implanted small block of pHPMA hydrogel into the hemisected T10 rat spinal cord. Locomotor activity was evaluated once a week during 14 weeks with the BBB rating scale in an open field. At the 14th week after spinal cord injury, the reflexivity of the sub-lesional region was measured. We also monitored the ventilatory frequency during an electrically induced muscle fatigue known to elicit the muscle metaboreflex and increase the respiratory rate. Spinal cords were then collected, fixed and stained with anti-ED-1 and anti-NF-H antibodies, and FluoroMyelin. We show in this study that hydrogel implanted animals exhibit 1) an improved locomotor BBB score, 2) an improved breathing adjustments to electrically-evoked isometric contractions, 3) an H-reflex recovery close to Control animals. Qualitative histological results put in evidence higher accumulation of ED-1 positive cells (macrophages/monocytes) at the lesion border, a large number of NF-H positive axons penetrating the applied matrix, and myelin preservion rostrally and caudally to the lesion. Our data confirm that pHPMA hydrogel is a potent biomaterial that can be used for improving neuromuscular adaptive mechanisms and H-reflex responses after spinal cord injury.
    ASN Neuro 04/2013; 5(2). DOI:10.1042/AN20120082 · 4.02 Impact Factor
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