Uk Namgung

Vimentin phosphorylation by Cdc2 in Schwann cell controls axon growth via β1-integrin activation

Although preconditioning injury on the peripheral nerve induces axonal regenerative capacity in neurons, it is not known whether similar lesion effects occur in glial cells. Here we demonstrate that Schwann cells are activated by peripheral nerve preinjury and primed to mediate axon regeneration. Cdc2, which was induced from Schwann cells after sciatic nerve injury, phosphorylated vimentin almost exclusively in the distal nerve area. Phospho‐vimentin‐positive Schwann cells showed increased migration activity and were in close contact with process outgrowth of co‐cultured neurons. Vimentin phosphorylation by Cdc2 was involved in β1‐integrin activation leading to FAK phoshorylation and associated with Erk1/2 activation in Schwann cells. Neurite outgrowth of dorsal root ganglion neurons was increased by co‐culture with activated Schwann cells, in which phospho‐vimentin signaling was transmitted into β1‐integrin activation. Then neurite outgrowth was suppressed by genetic depletion of phospho‐vimentin and β1 integrin as well as inhibition of vimentin phosphorylation by Cdc2 inhibitor purvalanol A. The sciatic nerve graft harboring activated Schwann cells into the spinal cord induced Schwann cell migration beyond the graft‐host barrier and facilitated regeneration of spinal axons, which was inhibited by purvalanol A pretreatment of the graft. This is the first report to our knowledge demonstrating that activation of phospho‐vimentin linked to β1‐integrin pathway may mediate transcellular signaling to promote axon growth.—Chang, I. A., Oh, M.‐J., Kim, M. H., Park, S.‐K., Kim, B. G., Namgung, U. Vimentin phosphorylation by Cdc2 in Schwann cell controls axon growth via β1‐integrin activation. FASEB J. 26, 2401‐2413 (2012). www.fasebj.org

Growth-promoting activity of Hominis Placenta extract on regenerating sciatic nerve

AbstractAim:Extract of Hominis Placenta (HP) has been used in oriental medicine as an agent for improving physiological function. The present study was conducted to investigate whether HP treatment in an experimental sciatic nerve injury animal model produces growth-promoting effects on regenerating peripheral nerve fibers after injury.Methods:After HP was injected into a sciatic nerve injury site, changes in protein levels were analyzed in the regenerating nerve area by Western blotting and immunofluorescence staining analyses. For quantitative assessment of axonal regeneration, a retrograde tracing technique was used to identify the neuronal cell bodies corresponding to regenerating axons, and the extent of neurite outgrowth in cultured dorsal root ganglia (DRG) sensory neurons prepared from animals that had experienced a sciatic nerve crush injury 7 d before neuron collection was analyzed.Results:Induction levels of axonal growth-associated protein (GAP-43) in the injured sciatic nerves were elevated by HP treatment. HP treatment also upregulated cell division cycle 2 (Cdc2) protein levels in the distal stump of the injured sciatic nerve. Induced Cdc2 protein was detected in Schwann cells, suggesting that Cdc2 kinase activity maybe involved in the growth-promoting activity of regenerating axons via Schwann cell proliferation. Cell body measurement by retrograde tracing indicated that HP treatment produced significant increases in regenerating motor axons. Finally, HP treatment of cultured DRG sensory neurons significantly increased neurite arborization and elongation.Conclusion:HP promotes the regeneration of injured sciatic axons by upregulating the synthesis of regeneration-related protein factors such as GAP-43 and Cdc2.

Endogenous Expression of Interleukin-4 Regulates Macrophage Activation and Confines Cavity Formation After Traumatic Spinal Cord Injury

Traumatic spinal cord injury (SCI) triggers inflammatory reactions in which various types of cells and cytokines are involved. Several proinflammatory cytokines are up‐regulated after SCI and play crucial roles in determining the extent of secondary tissue damage. However, relatively little is known about antiinflammatory cytokines and their roles in spinal cord trauma. Recent studies have shown that an antiinflammatory cytokine, interleukin‐4 (IL‐4), is expressed and exerts various modulatory effects in CNS inflammation. We found in the present study that IL‐4 was highly expressed at 24 hr after contusive SCI in rats and declined thereafter, with concurrent up‐regulation of IL‐4 receptor subunit IL‐4α. The majority of IL‐4‐producing cells were myeloperoxidase‐positive neutrophils. Injection of neutralizing antibody against IL‐4 into the contused spinal cord did not significantly affect the expression levels of proinflammatory cytokines such as IL‐1β, IL‐6, and tumor necrosis factor‐α or other antiinflammatory cytokines such as IL‐10 and transforming growth factor‐β. Instead, attenuation of IL‐4 activity led to a marked increase in the extent of ED1‐positive macrophage activation along the rostrocaudal extent at 7 days after injury. The enhanced macrophage activation was preceded by an increase in the level of monocyte chemoattractant protein‐1 (MCP‐1/CCL2). Finally, IL‐4 neutralization resulted in more extensive cavitation at 4 weeks after injury. These results suggest that endogenous expression of antiinflammatory cytokine IL‐4 regulates the extent of acute macrophage activation and confines the ensuing secondary cavity formation after spinal cord trauma.

Anti-Inflammatory Effects of Acupuncture Stimulation via the Vagus Nerve

Although acupuncture therapy is widely used in traditional Asian medicine for the treatment of diverse internal organ disorders, its underlying biological mechanisms are largely unknown. Here, we investigated the functional involvement of acupuncture stimulation (AS) in the regulation of inflammatory responses. TNF-α production in mouse serum, which was induced by lipopolysaccharide (LPS) administration, was decreased by manual acupuncture (MAC) at the zusanli acupoint (stomach36, ST36). In the spleen, TNF-α mRNA and protein levels were also downregulated by MAC and were recovered by using a splenic neurectomy and a vagotomy. c-Fos, which was induced in the nucleus tractus solitarius (NTS) and dorsal motor nucleus of the vagus nerve (DMV) by LPS and electroacupuncture (EAC), was further increased by focal administration of the AMPA receptor blocker CNQX and the purinergic receptor antagonist PPADS. TNF-α levels in the spleen were decreased by CNQX and PPADS treatments, implying the involvement of inhibitory neuronal activity in the DVC. In unanesthetized animals, both MAC and EAC generated c-Fos induction in the DVC neurons. However, MAC, but not EAC, was effective in decreasing splenic TNF-α production. These results suggest that the therapeutic effects of acupuncture may be mediated through vagal modulation of inflammatory responses in internal organs.

Role of Glial Cells in Axonal Regeneration

Axonal regeneration is critical for functional recovery following neural injury. In addition to intrinsic differences between regenerative responses of axons in peripheral versus central nervous systems, environmental factors such as glial cells and related molecules in the extracellular matrix (ECM) play an important role in axonal regeneration. Schwann cells in the peripheral nervous system (PNS) are recognized as favorable factors that promote axonal regeneration, while astrocytes and oligodendrocytes in the central nervous system (CNS) are not. In this review, we evaluate the roles of Schwann cells and astrocytes in axonal regeneration and examine recent evidence that suggests a dual function of astrocytes in regenerative responses. We also discuss the role of Cdc2 pathways in axonal regeneration, which is commonly activated in Schwann cells and astrocytes. Greater insight on the roles of glial cells in axonal regeneration is key to establishing baseline interventions for improving functional recovery following neural injury.

Ultrasound-stimulated peripheral nerve regeneration within asymmetrically porous PLGA/Pluronic F127 nerve guide conduit

Recently, we developed a novel method to fabricate a nerve guide conduit (NGC) with asymmetrical pore structure and hydrophilicity using poly(lactic-co-glycolic acid) (PLGA) and Pluronic F127 by a modified immersion precipitation method. From the animal study using a rat model (sciatic nerve defect of rat), we recognized that the unique PLGA/Pluronic F127 tube provided good environments for nerve regeneration. In this study, we applied low-intensity pulsed ultrasound as a simple and noninvasive stimulus at the PLGA/F127 NGC-implanted site transcutaneously in rats to investigate the feasibility of ultrasound for the enhanced nerve regeneration through the tube. The nerve regeneration behaviors within the ultrasound-stimulated PLGA/Pluronic F127 NGCs were compared with the NGCs without the ultrasound treatment as well as normal nerve by histological and immunohistochemical observations. It was observed that the PLGA/Pluronic F127 tube-implanted group applied with the ultrasound had more rapid nerve regeneration behavior (approximately 0.71 mm/day) than the tube-implanted group without the ultrasound treatment (approximately 0.48 mm/day). The ultrasound-treated tube group also showed greater neural tissue area as well as larger axon diameter and thicker myelin sheath than the tube group without the ultrasound treatment, indicating better nerve regeneration. The better nerve regeneration behavior in the our NGC/ultrasound system may be caused by the synergistic effect of the asymmetrically porous PLGA/Pluronic F127 tube with unique properties (selective permeability, hydrophilicity, and structural stability, which can provide good environment for nerve regeneration) and physical stimulus (stimulation of the Schwann cells and activation of the neurotrophic factors).

Acupuncture-Stimulated Activation of Sensory Neurons

Acupuncture is one of the key therapeutics in clinical oriental medicine, and recent studies using experimental animals have begun to provide the pathophysiological basis for the efficacy of acupuncture. Here, we investigated neuronal responses in rodent models given acupuncture stimulation. In both mice and rats, acupuncture stimulation at zusanli (ST36) generated an increased expression of axonal growth-associated protein (GAP-43) in the sensory neurons of the dorsal root ganglion (DRG). Electroacupuncture stimulation at ST36 in rats induced GAP-43 mRNA and protein expression in DRG neurons at the levels of lumbar 4 and 5. Stimulation on a non-acupuncture site as a sham control induced GAP-43 expression as well, but the induction level was lower than it was with acupuncture. We further found that acupuncture stimulation upregulated phospho-Erk1/2 signals in DRG neurons. Electroacupuncture stimulation induced c-Fos expression in the neurons of the dorsal motor nucleus of the vagus nerve (DMV), which was identified by retrograde tracing. These data suggest that acupuncture stimulation may generate physiological effects on the autonomic nervous system via the activation of a somatosensory pathway.

Effects of inhalable microparticle of flower of Lonicera japonica in a mouse model of COPD

ETHNOPHARMACOLOGICAL RELEVANCE Flower of Lonicera japonica (FLJ) is a traditional herbal medicine widely used in East Asia as an anti-inflammatory and anti-oxidative agent. The purpose of this study is to develop an inhalable powder formulation of FLJ and to evaluate its biological effects in a mouse model of chronic obstructive pulmonary disease (COPD). METHODS Inhalable dry powder containing FLJ was produced by spray-drying with leucine as an excipient. Its aerodynamic properties and anti-inflammatory activities were evaluated using the Anderson cascade impactor (ACI) and a mouse model of COPD, respectively. RESULTS FLJ microparticle (FLJmp) had a hollow spherical shape in electron microscopy and showed aerodynamic properties suitable for inhalation (fine particle fraction of 54.0 ± 4.68% and mass median aerodynamic diameter of 4.6 ± 0.34μm). FLJmp decreased TNF-α and IL-6 expression in RAW264.7 cells activated by lipopolysaccharide (LPS). In mice challenged with LPS and cigarette smoke solution (CSS) to develop COPD, FLJmp decreased the levels of TNF-α and IL-6 in bronchoalveolar fluidas well as the number of inflammatory cells including neutrophils in peripheral blood. In addition, FLJmp induced recovery of elastin and collagen distribution, reduction of caspase-3 expression in lung tissues of COPD mice. CONCLUSIONS Inhalational delivery of FLJ using a microparticle system is a promising strategy for the treatment of COPD.

Growth-promoting Activity of Sanyak (Dioscoreae rhizoma) Extract on Injured Sciatic Nerve in Rats

The present study evaluates the potential effects of Sanyak (Dioscoreae rhizome) on the regenerative capacity of the peripheral sciatic nerve after crush injury in rats. Focal application of Sanyak extract at the injury site increased GAP-43 and Cdc2 protein levels in the distal portion of the injured nerve. Immunohistochemical analysis showed that the signals of phospho-vimentin as Cdc2 substrate protein were almost colocalized with Cdc2. Retrograde DiI-tracing revealed enhancement in distal elongation of regenerating axons. Furthermore, the number of non-neuronal cells was higher in Sanyak-treated animals than saline controls. Thus, these data suggest that Sanyak extract is effective for promoting regenerative responses in injured peripheral neurons.

Beneficial Function of Cell Division Cycle 2 Activity in Astrocytes on Axonal Regeneration after Spinal Cord Injury

Migrating activity of reactive astrocytes induced after spinal cord injury (SCI) controls glial scar formation by limiting inflammatory responses around the injury area, and, therefore, can be beneficial for regenerative responses of spinal axons. Recently, we found that cell division cycle 2 (cdc2) activity in primary astrocytes facilitated neurite outgrowth of co-cultured neurons. Here, we investigated the effects of cdc2 activity on regenerative processes in vivo after SCI. Administration of the cdc2 inhibitor purvalanol A restricted compaction of the injury cavity and astrocyte infiltration into the cavity. After SCI, regenerative responses of anterogradely labeled corticospinal tract (CST) axons were attenuated by purvalanol A treatment. Using the polymeric tube that was implanted into the spinal cord as a nerve guide conduit, we found that purvalanol A treatments reduced astrocyte migration into the tube graft and, in parallel, retarded the extension of spinal axons into the tube. These results suggest that astrocytes with cdc2 activity may play a permissive role in mediating regrowth of spinal axons after lesion.