Engineering Fiber-Based Nervous Tissue Constructs for Axon Regeneration

2021 ◽  
pp. 1-13
Author(s):  
Mevan L. Siriwardane ◽  
Kathleen Derosa ◽  
George Collins ◽  
Bryan J. Pfister
Keyword(s):  
Type I Collagen ◽  
Nerve Repair ◽  
Small Diameter ◽  
Axon Growth ◽  
Dorsal Root Ganglion Neurons ◽  
Type I ◽  
Adult Rats ◽  
Hydrogel Matrix ◽  
Ganglion Neurons

Biomaterial-based scaffolds used in nerve conduits including channels for confining regenerating axons and 3-dimensional (3D) gels as substrates for growth have made improvements in models of nerve repair. Many biomaterial strategies, however, continue to fall short of autologous nerve grafts, which remain the current gold standard in repairing severe nerve lesions (<20 mm). Intraluminal nerve conduit fibers have also shown considerable promise in directing regenerating axons in vitro and in vivo and have gained increasing interest for nerve repair. It is unknown, however, how growing axons respond to a fiber when encountered in a 3D environment. In this study, we considered a construct consisting of a compliant collagen hydrogel matrix and a fiber component to assess contact-guided axon growth. We investigated preferential axon outgrowth on synthetic and natural polymer fibers by utilizing small-diameter microfibers of poly-L-lactic acid and type I collagen representing 2 different fiber stiffnesses. We found that axons growing freely in a 3D hydrogel culture preferentially attach, turn and follow fibers with outgrowth rates and distances that far exceed outgrowth in a hydrogel alone. Wet-spun type I collagen from rat tail tendon performed the best, associated with highly aligned and accelerated outgrowth. This study also evaluated the response of dorsal root ganglion neurons from adult rats to provide data more relevant to axon regenerative potential in nerve repair. We found that ECM treatments on fibers enhanced the regeneration of adult axons indicating that both the physical and biochemical presentation of the fibers are essential for enhancing axon guidance and growth.

Collagen fibrillogenesis in vitro: interaction of types I and V collagen regulates fibril diameter

1990 ◽  
Vol 95 (4) ◽  
pp. 649-657 ◽  
Author(s):  
D.E. Birk ◽  
J.M. Fitch ◽  
J.P. Babiarz ◽  
K.J. Doane ◽  
T.F. Linsenmayer
Keyword(s):  
Type I Collagen ◽  
Small Diameter ◽  
Collagen Molecule ◽  
Type I ◽  
Collagen Types ◽  
Type V Collagen ◽  
Amino Terminal ◽  
Type V ◽  
Fibril Diameter

The small-diameter fibrils of the chick corneal stroma are heterotypic, composed of both collagen types I and V. This tissue has a high concentration of type V collagen relative to other type I-containing tissues with larger-diameter fibrils, suggesting that heterotypic interactions may have a regulatory role in the control of fibril diameter. The interactions of collagen types I and V were studied using an in vitro self-assembly system. Collagens were purified from lathyritic chick embryos in the presence of protease inhibitors. The type V collagen preparations contained higher molecular weight forms of the alpha 1(V) and alpha 2(V) chains constituting 60–70% of the total. Rotary-shadow electron micrographs showed a persistence of a small, pepsin-sensitive terminal region in an amount consistent with that seen by electrophoresis. In vitro, this purified type V collagen formed thin fibrils with no apparent periodicity, while type I collagen fibrils had a broad distribution of large diameters. However, when type I collagen was mixed with increasing amounts of type V collagen a progressive and significant decrease in both the mean fibril diameter and the variance was observed for D periodic fibrils. The amino-terminal domain of the type V collagen molecule was required for this regulatory effect and in its absence little diameter reducing activity was observed. Electron microscopy using collagen type-specific monoclonal antibodies demonstrated that the fibrils formed were heterotypic, containing both collagen types I and V. These data indicate that the interaction of type V with type I collagen is one mechanism modulating fibril diameter and is at least partially responsible for the regulation of collagen fibril formation.

scholarly journals Netrin Inhibits Regenerative Axon Growth of Adult Dorsal Root Ganglion Neurons in Vitro

2007 ◽  
Vol 22 (4) ◽  
pp. 641 ◽  
Author(s):  
Joo-In Park ◽  
In-Ae Seo ◽  
Hyun-Kyoung Lee ◽  
Hwan-Tae Park ◽  
Sung-Won Shin ◽  
...  
Keyword(s):  
Dorsal Root Ganglion ◽  
Dorsal Root ◽  
Axon Growth ◽  
Dorsal Root Ganglion Neurons ◽  
Ganglion Neurons

scholarly journals Chronic compression of mouse dorsal root ganglion alters voltage-gated sodium and potassium currents in medium-sized dorsal root ganglion neurons

2011 ◽  
Vol 106 (6) ◽  
pp. 3067-3072 ◽  
Author(s):  
Ni Fan ◽  
David F. Donnelly ◽  
Robert H. LaMotte
Keyword(s):  
Dorsal Root Ganglion ◽  
Dorsal Root ◽  
Small Diameter ◽  
Radicular Pain ◽  
Dorsal Root Ganglion Neurons ◽  
Delayed Rectifier ◽  
Drg Neurons ◽  
Voltage Dependent ◽  
Ganglion Neurons

Chronic compression (CCD) of the dorsal root ganglion (DRG) is a model of human radicular pain produced by intraforaminal stenosis and other disorders affecting the DRG, spinal nerve, or root. Previously, we examined electrophysiological changes in small-diameter lumbar level 3 (L3) and L4 DRG neurons treated with CCD; the present study extends these observations to medium-sized DRG neurons, which mediate additional sensory modalities, both nociceptive and non-nociceptive. Whole-cell patch-clamp recordings were obtained from medium-sized somata in the intact DRG in vitro. Compared with neurons from unoperated control animals, CCD neurons exhibited a decrease in the current threshold for action potential generation. In the CCD group, current densities of TTX-resistant and TTX-sensitive Na+ current were increased, whereas the density of delayed rectifier voltage-dependent K+ current was decreased. No change was observed in the transient or “A” current after CCD. We conclude that CCD in the mouse produces hyperexcitability in medium-sized DRG neurons, and the hyperexcitability is associated with an increased density of Na+ current and a decreased density of delayed rectifier voltage-dependent K+ current.

scholarly journals Overexpression of Reticulon 3 enhances CNS axon regeneration and functional recovery after injury

2021 ◽  
Author(s):  
Zubair Ahmed ◽  
Sharif Alhajlah ◽  
Adam Thompson
Keyword(s):  
Spinal Cord ◽  
Optic Nerve ◽  
Neurite Outgrowth ◽  
Functional Recovery ◽  
Axon Regeneration ◽  
Axon Growth ◽  
Dorsal Root Ganglion Neurons ◽  
Ganglion Neurons

CNS neurons are generally incapable of regenerating their axons after injury due to several intrinsic and extrinsic factors, including the presence of axon growth inhibitory molecules. One such potent inhibitor of CNS axon regeneration is Reticulon (RTN) 4 or Nogo-66. Here, we focused on RTN3 as its contribution in CNS axon regeneration is currently unknown. We found that RTN3 expression correlated with an axon regenerative phenotype in dorsal root ganglion neurons (DRGN) after injury to the dorsal columns, a model of spinal cord injury. Overexpression of RTN3 promoted disinhibited DRGN neurite outgrowth in vitro and dorsal column axon regeneration/sprouting and electrophysiological, sensory and locomotor functional recovery after injury in vivo. Knockdown of protrudin however, ablated RTN3-enhanced neurite outgrowth/axon regeneration in vitro and in vivo. Moreover, overexpression of RTN3 in a second model of CNS injury, the optic nerve crush injury model, enhanced retinal ganglion cell (RGC) survival, disinhibited neurite outgrowth in vitro and survival and axon regeneration in vivo, an effect that was also dependent on protrudin. These results demonstrate that RTN3 enhances neurite outgrowth/axon regeneration in a protrudin-dependent manner after both spinal cord and optic nerve injury.

scholarly journals Overexpression of Reticulon 3 Enhances CNS Axon Regeneration and Functional Recovery after Traumatic Injury

Cells ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 2015
Author(s):  
Sharif Alhajlah ◽  
Adam M Thompson ◽  
Zubair Ahmed
Keyword(s):  
Spinal Cord ◽  
Optic Nerve ◽  
Neurite Outgrowth ◽  
Functional Recovery ◽  
Axon Regeneration ◽  
Axon Growth ◽  
Dorsal Root Ganglion Neurons ◽  
Ganglion Neurons

CNS neurons are generally incapable of regenerating their axons after injury due to several intrinsic and extrinsic factors, including the presence of axon growth inhibitory molecules. One such potent inhibitor of CNS axon regeneration is Reticulon (RTN) 4 or Nogo-A. Here, we focused on RTN3 as its contribution to CNS axon regeneration is currently unknown. We found that RTN3 expression correlated with an axon regenerative phenotype in dorsal root ganglion neurons (DRGN) after injury to the dorsal columns, a well-characterised model of spinal cord injury. Overexpression of RTN3 promoted disinhibited DRGN neurite outgrowth in vitro and dorsal column axon regeneration/sprouting and electrophysiological, sensory and locomotor functional recovery after injury in vivo. Knockdown of protrudin, however, ablated RTN3-enhanced neurite outgrowth/axon regeneration in vitro and in vivo. Moreover, overexpression of RTN3 in a second model of CNS injury, the optic nerve crush injury model, enhanced retinal ganglion cell (RGC) survival, disinhibited neurite outgrowth in vitro and survival and axon regeneration in vivo, an effect that was also dependent on protrudin. These results demonstrate that RTN3 enhances neurite outgrowth/axon regeneration in a protrudin-dependent manner after both spinal cord and optic nerve injury.

Membrane properties and electrogenesis in the distal axons of small dorsal root ganglion neurons in vitro

2012 ◽  
Vol 108 (3) ◽  
pp. 729-740 ◽  
Author(s):  
Dmytro V. Vasylyev ◽  
Stephen G. Waxman
Keyword(s):  
Action Potential ◽  
Action Potentials ◽  
Dorsal Root ◽  
Small Diameter ◽  
Dorsal Root Ganglion Neurons ◽  
Membrane Properties ◽  
Resting Potential ◽  
Drg Neurons ◽  
Ganglion Neurons

Although it is generally thought that sensory transduction occurs at or close to peripheral nerve endings, with action potentials subsequently propagating along the axons of dorsal root ganglia (DRG) neurons toward the central nervous system, the small diameter of nociceptive axons and their endings have made it difficult to estimate their membrane properties and electrogenic characteristics. Even the resting potentials of nociceptive axons are unknown. In this study, we developed the capability to record directly with patch-clamp electrodes from the small-diameter distal axons of DRG neurons in vitro. We showed using current-clamp recordings that 1) these sensory axons have a resting potential of −60.2 ± 1 mV; 2) both tetrodotoxin (TTX)-sensitive (TTX-S) and TTX-resistant (TTX-R) Na+ channels are present and available for activation at resting potential, at densities that can support action potential electrogenesis in these axons; 3) TTX-sensitive channels contribute to the amplification of small depolarizations that are subthreshold with respect to the action potential in these axons; 4) TTX-R channels can support the production of action potentials in these axons; and 5) these TTX-R channels can produce repetitive firing, even at depolarized membrane potentials where TTX-S channels are inactivated. Finally, using voltage-clamp recordings with an action potential as the command, we confirmed the presence of both TTX-S and TTX-R channels, which are activated sequentially during action potential in these axons. These results provide direct evidence for the presence of TTX-S and TTX-R Na+ channels that are functionally available at resting potential and contribute to electrogenesis in small-diameter afferent axons.

scholarly journals Increased Na+ and K+ currents in small mouse dorsal root ganglion neurons after ganglion compression

2011 ◽  
Vol 106 (1) ◽  
pp. 211-218 ◽  
Author(s):  
Ni Fan ◽  
Parul Sikand ◽  
David F. Donnelly ◽  
Chao Ma ◽  
Robert H. LaMotte
Keyword(s):  
Dorsal Root Ganglion ◽  
Dorsal Root ◽  
Neuronal Response ◽  
Small Diameter ◽  
Neuronal Cell ◽  
Radicular Pain ◽  
Dorsal Root Ganglion Neurons ◽  
Delayed Rectifier ◽  
Ganglion Neurons

We investigated the effects of chronic compression (CCD) of the L3 and L4 dorsal root ganglion (DRG) on pain behavior in the mouse and on the electrophysiological properties of the small-diameter neuronal cell bodies in the intact ganglion. CCD is a model of human radicular pain produced by intraforaminal stenosis and other disorders affecting the DRG, spinal nerve, or root. On days 1, 3, 5, and 7 after the onset of compression, there was a significant decrease from preoperative values in the threshold mechanical force required to elicit a withdrawal of the foot ipsilateral to the CCD (tactile allodynia). Whole cell patch-clamp recordings were obtained, in vitro, from small-sized somata and, for the first time, in the intact DRG. Under current clamp, CCD neurons exhibited a significantly lower rheobase compared with controls. A few CCD but no control neurons exhibited spontaneous action potentials. CCD neurons showed an increase in the density of TTX-resistant and TTX-sensitive Na+ current. CCD neurons also exhibited an enhanced density of voltage-dependent K+ current, due to an increase in delayed rectifier K+ current, without a change in the transient or “A” current. We conclude that CCD in the mouse produces a model of radicular pain, as we have previously demonstrated in the rat. While the role of enhanced K+ current remains to be elucidated, we speculate that it represents a compensatory neuronal response to reduce ectopic or aberrant levels of neuronal activity produced by the injury.

Collagen fibrillogenesis in vitro: interaction of types I and V collagen

1986 ◽  
Vol 44 ◽  
pp. 210-211
Author(s):  
Arthur J. Wasserman ◽  
Kathy C. Kloos ◽  
David E. Birk
Keyword(s):  
Type I Collagen ◽  
Corneal Stroma ◽  
Minor Component ◽  
Homogeneous Distribution ◽  
Type I ◽  
Type V Collagen ◽  
Fibril Morphology ◽  
Type V ◽  
In Vitro Interaction

Type I collagen is the predominant collagen in the cornea with type V collagen being a quantitatively minor component. However, the content of type V collagen (10-20%) in the cornea is high when compared to other tissues containing predominantly type I collagen. The corneal stroma has a homogeneous distribution of these two collagens, however, immunochemical localization of type V collagen requires the disruption of type I collagen structure. This indicates that these collagens may be arranged as heterpolymeric fibrils. This arrangement may be responsible for the control of fibril diameter necessary for corneal transparency. The purpose of this work is to study the in vitro assembly of collagen type V and to determine whether the interactions of these collagens influence fibril morphology.

scholarly journals Restoration of Osteogenesis by CRISPR/Cas9 Genome Editing of the Mutated COL1A1 Gene in Osteogenesis Imperfecta

2021 ◽  
Vol 10 (14) ◽  
pp. 3141
Author(s):  
Hyerin Jung ◽  
Yeri Alice Rim ◽  
Narae Park ◽  
Yoojun Nam ◽  
Ji Hyeon Ju
Keyword(s):  
Osteogenesis Imperfecta ◽  
Type I Collagen ◽  
Disease Modeling ◽  
Bone Fragility ◽  
Osteogenic Potential ◽  
Type I ◽  
Gene Correction ◽  
Col1a1 Gene ◽  
Collagen Formation

Osteogenesis imperfecta (OI) is a genetic disease characterized by bone fragility and repeated fractures. The bone fragility associated with OI is caused by a defect in collagen formation due to mutation of COL1A1 or COL1A2. Current strategies for treating OI are not curative. In this study, we generated induced pluripotent stem cells (iPSCs) from OI patient-derived blood cells harboring a mutation in the COL1A1 gene. Osteoblast (OB) differentiated from OI-iPSCs showed abnormally decreased levels of type I collagen and osteogenic differentiation ability. Gene correction of the COL1A1 gene using CRISPR/Cas9 recovered the decreased type I collagen expression in OBs differentiated from OI-iPSCs. The osteogenic potential of OI-iPSCs was also recovered by the gene correction. This study suggests a new possibility of treatment and in vitro disease modeling using patient-derived iPSCs and gene editing with CRISPR/Cas9.

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