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J Verhaagen

Publications and source records attributed to J Verhaagen.

At least 19 recordsLinked to original sources
PubMed2006

International spinal research trust research strategy. III: A discussion document.

STUDY DESIGN: Discussion document. OBJECTIVES/METHODS: To review the Research Strategy of the International Spinal Research Trust (ISRT), which identifies key areas of basic and clinical research that are likely to be beneficial in developing potential treatments for spinal cord injury for funding. This strategy is intended to both guide the programme of research towards areas of priority and stimulate discussion of the different avenues of research. This latest document has been developed to take into account the scientific progress in the 6 years since publication of the previous Research Strategy. RESULTS/DISCUSSION: The latest scientific developments in research designed to repair the spinal cord and restore function following injury and how they might impact on spinal cord injury research are highlighted.

Biomedical Research↗
PubMed2006

AAV-mediated expression of CNTF promotes long-term survival and regeneration of adult rat retinal ganglion cells.

We compared the effects of intravitreal injection of bi-cistronic adeno-associated viral (AAV-2) vectors encoding enhanced green fluorescent protein (GFP) and either ciliary neurotrophic factor (CNTF), brain-derived neurotrophic factor (BDNF) or growth-associated protein-43 (GAP43) on adult retinal ganglion cell (RGC) survival and regeneration following (i) optic nerve (ON) crush or (ii) after ON cut and attachment of a peripheral nerve (PN). At 7 weeks after ON crush, quantification of betaIII-tubulin immunostaining revealed that, compared to AAV-GFP controls, RGC survival was not enhanced by AAV-GAP43-GFP but was increased in AAV-CNTF-GFP (mean RGCs/retina: 17 450+/-358 s.e.m.) and AAV-BDNF-GFP injected eyes (10 200+/-4064 RGCs/retina). Consistent with increased RGC viability in AAV-CNTF-GFP and AAV-BDNF-GFP injected eyes, these animals possessed many betaIII-tubulin- and GFP-positive fibres proximal to the ON crush. However, only in the AAV-CNTF-GFP group were regenerating RGC axons seen in distal ON (1135+/-367 axons/nerve, 0.5 mm post-crush), some reaching the optic chiasm. RGCs were immunoreactive for CNTF and quantitative RT-PCR revealed a substantial increase in CNTF mRNA expression in retinas transduced with AAV-CNTF-GFP. The combination of AAV-CNTF-GFP transduction of RGCs with autologous PN-ON transplantation resulted in even greater RGC survival and regeneration. At 7 weeks after PN transplantation there were 27 954 (+/-2833) surviving RGCs/retina, about 25% of the adult RGC population. Of these, 13 352 (+/-1868) RGCs/retina were retrogradely labelled after fluorogold injections into PN grafts. In summary, AAV-mediated expression of CNTF promotes long-term survival and regeneration of injured adult RGCs, effects that are substantially enhanced by combining gene and cell-based therapies/interventions.

Animals↗
PubMed2003

Adeno-associated viral vector-mediated neurotrophin gene transfer in the injured adult rat spinal cord improves hind-limb function.

To foster axonal growth from a Schwann cell bridge into the caudal spinal cord, spinal cells caudal to the implant were transduced with adeno-associated viral (AAV) vectors encoding for brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (AAV-NT-3). Control rats received AAV vectors encoding for green fluorescent protein or saline. AAV-BDNF- and AAV-NT-3-transduced 293 human kidney cells produced and secreted BDNF or NT-3, respectively, in vitro. The secreted neurotrophins were biologically active; they both promoted outgrowth of sensory neurites in vitro. In vivo, transgene expression was observed predominantly in neurons for at least 16 weeks after injection. Compared with controls, a modest though significant improvement in hind-limb function was found in rats that received AAV-BDNF and AAV-NT-3. Retrograde tracing demonstrated that twice as many neurons with processes extending toward the Schwann cell graft were present in the second lumbar cord segment of AAV-BDNF- and AAV-NT-3-injected animals compared with controls. We found no evidence, however, for growth of regenerated axons from the Schwann cell implant into the caudal cord. Our results suggest that AAV vector-mediated overexpression of BDNF and NT-3 in the cord caudal to a Schwann cell bridge modified the local lumbar axonal circuitry, which was beneficial for locomotor function.

Adenoviridae↗
PubMed2002

Injury-induced class 3 semaphorin expression in the rat spinal cord.

In this study we evaluate the expression of all members of the class 3 semaphorins and their receptor components following complete transection and contusion lesions of the adult rat spinal cord. Following both types of lesions the expression of all class 3 semaphorins is induced in fibroblast in the neural scar. The distribution of semaphorin-positive fibroblasts differs markedly in scars formed after transection or contusion lesion. In contusion lesions semaphorin expression is restricted to fibroblasts of the meningeal sheet surrounding the lesion, while after transection semaphorin-positive fibroblast penetrate deep into the center of the lesion. Two major descending spinal cord motor pathways, the cortico- and rubrospinal tract, continue to express receptor components for class 3 semaphorins following injury, rendering them potentially sensitive to scar-derived semaphorins. In line with this we observed that most descending spinal cord fibers were not able to penetrate the semaphorin positive portion of the neural scar formed at the lesion site. These results suggest that the full range of secreted semaphorins contributes to the inhibitory nature of the neural scar and thereby may inhibit successful regeneration in the injured spinal cord. Future studies will focus on the neutralization of class 3 semaphorins, in order to reveal whether this creates a more permissive environment for regeneration of injured spinal cord axons.

Animals↗
PubMed2002

Intravitreal injection of adeno-associated viral vectors results in the transduction of different types of retinal neurons in neonatal and adult rats: a comparison with lentiviral vectors.

Replication-deficient viral vectors encoding the marker gene green fluorescent protein (GFP) were injected into the vitreous of newborn, juvenile (P14), and adult rats. We tested two different types of modified virus: adeno-associated viral-2-GFP (AAV-GFP) and lentiviral-GFP vectors (LV-GFP). The extent of retinal cell transduction in different-aged animals was compared 7, 21, and 70 days after eye injections. At all postinjection times, LV-GFP transduction was mostly limited to pigment epithelium and cells in sclera and choroid. In contrast, transduction of large numbers of neural retinal cells was seen 21 and 70 days after AAV-GFP injections. AAV-GFP predominantly transduced neurons, although GFP-positive Müller cells were seen. All neuronal classes were labeled, but the extent of transduction for a given class varied depending on injection age. After P0 injections about 50% of transduced cells were photoreceptors and 30-40% were amacrine or bipolar cells. After adult injections 60-70% of transduced cells were retinal ganglion cells. In adults many GFP-positive retinal axons were traced through the optic nerve/tract and terminal arbors were visualized in central targets.

Adenoviridae↗
PubMed2002

Viral vector-mediated gene expression in olfactory ensheathing glia implants in the lesioned rat spinal cord.

Implantation of olfactory ensheathing glia (OEG) is a promising strategy to augment long-distance regeneration in the injured spinal cord. In this study, implantation of OEG following unilateral hemisection of the dorsal cervical spinal cord was combined with ex vivo gene transfer techniques. We report, to our knowledge for the first time, that purified cultures of primary OEG are capable of expressing a foreign gene following adenoviral (AdV) and lentiviral (LV) vector-mediated gene transfer. OEG implants subjected to AdV vector-mediated gene transfer expressed high levels of transgenic protein in both intact and lesioned spinal cord at 7 days after implantation. However, the levels of transgene expression gradually declined between 7 and 30 days after implantation in lesioned spinal cord. Infection with LV vectors resulted in stable transduction of primary OEG cultures and transgene expression persisted for at least 4 months after implantation. Genetic engineering of OEG opens the possibility of expressing additional neurotrophic genes and create optimal 'bridging' substrates to support spinal axon regeneration. Furthermore, stable transduction of OEG allows us to reliably study the behaviour of implanted cells and to obtain better understanding of their regeneration supporting properties.

Adenoviridae↗
PubMed2002

Amelioration of chronic neuropathic pain after partial nerve injury by adeno-associated viral (AAV) vector-mediated over-expression of BDNF in the rat spinal cord.

Changing the levels of neurotrophins in the spinal cord micro-environment after nervous system injury has been proposed to recover normal function, such that behavioral response to peripheral stimuli does not lead to chronic pain. We have investigated the effects of recombinant adeno-associated viral (rAAV)-mediated over-expression of brain-derived neurotrophic factor (BDNF) in the spinal cord on chronic neuropathic pain after unilateral chronic constriction injury (CCI) of the sciatic nerve. The rAAV-BDNF vector was injected into the dorsal horn at the thirteenth thoracic spinal cord vertebra (L(1) level) 1 week after CCI. Allodynia and hyperalgesia induced by CCI in the hindpaws were permanently reversed, beginning 1 week after vector injection, compared with a similar injection of a control rAAV-GFP vector (green fluorescent protein) or saline. In situ hybridization for BDNF demonstrated that both dorsal and ventral lumbar spinal neurons contained an intense signal for BDNF mRNA, at 1 to 8 weeks after vector injection. There was no similar BDNF mRNA over-expression associated with either injections of saline or rAAV-GFP. These data suggest that chronic neuropathic pain is sensitive to early spinal BDNF levels after partial nerve injury and that rAAV-mediated gene transfer could potentially be used to reverse chronic pain after nervous system injuries in humans.

Animals↗
PubMed2001

Adenoviral vector-mediated expression of neurotrophin-3 increases neuronal survival in suprachiasmatic nucleus grafts.

To improve transplantation results of fetal suprachiasmatic nucleus (SCN) in SCN-lesioned (SCNX) rats, grafts were ex vivo transduced with an adenoviral vector encoding for neurotrophin-3 (AdNT-3) before implantation. Mock- and AdLacZ-transduced grafts were used as controls. First, transplants were evaluated microscopically and by image analysis for the presence of vasopressinergic (VPergic) and vasoactive intestinal polypeptidergic (VIPergic) SCN neurons at 10 weeks or later postgrafting. Ex vivo AdNT-3-transduced transplants displayed increased volume areas of VPergic and VIPergic SCN cells in comparison with those in mock- and AdLacZ-transduced transplants, but significantly improved graft-to-host VPergic and VIPergic SCN fiber growth was not reached (though AdNT-3-transduced transplants tended to grow more VPergic fibers into the brain of VP-deficient SCNX Brattleboro rat recipients, which were chosen as recipients to circumvent the presence of non-SCN VP fiber staining). Second, a small group of arrhythmic Wistar rats received AdNT-3- or control-treated SCN grafts while continuously on-line for the monitoring of overt circadian activities in the pre- and postgrafting periods. The results indicated that ex vivo transduced SCN grafts can still restore arrhythmia, but that the NT-3-mediated anatomical improvements of the grafting results were not sufficient to enhance efficacy of reinstatement of circadian rhythm in SCN-lesioned rats. However, in this group VIP staining volume area, not VP staining volume area, correlated significantly with reinstatement of circadian rhythm.

Activity Cycles↗
PubMed2001

Emerging roles for semaphorins in neural regeneration.

Progressive axon outgrowth during neural development contrasts with the failure of regenerative neurite growth in the mature mammalian central nervous system (CNS). During neuroembryogenesis, spatiotemporal patterns of repellent and attractant activities in the vicinity of the growth cone favor neurite outgrowth. In the mature CNS, however, a relative balance between forces supporting and restricting axon growth has been established, only allowing subtle morphological changes in existing neuritic arbors and synapses. Following CNS injury, this balance shifts towards enhanced expression of growth-inhibiting molecules and diminished availability of their growth-promoting counterparts. Evidence is now emerging that the proteins governing developmental axon guidance critically contribute to the failure of injured central neurons to regenerate. As a first step toward elucidation of the role of chemorepulsive axon guidance signals in axonal regeneration, the effects of lesions of the central and peripheral nervous system on the expression of Semaphorin3A, the prototype and founding member of the semaphorin family of axon guidance signals, and of the Semaphorin3A receptor proteins neuropilin-1 and plexin-A1 have recently been examined. Here we review the first evidence indicating that (i) lesion-induced changes in the expression of chemorepulsive semaphorins relate to the success or failure of injured neurons to regenerate and (ii) semaphorins may represent important molecular signals controlling multiple aspects of the cellular response that follows CNS injury. In the future, genetic manipulation of the injury-induced changes in the availability of semaphorins and/or of their receptors will provide further insight into the mechanisms by which semaphorins influence neural regeneration.

Animals↗
PubMed2001

Peripheral nerve injury fails to induce growth of lesioned ascending dorsal column axons into spinal cord scar tissue expressing the axon repellent Semaphorin3A.

We have investigated the hypothesis that the chemorepellent Semaphorin3A may be involved in the failure of axonal regeneration after injury to the ascending dorsal columns of adult rats. Following transection of the thoracic dorsal columns, fibroblasts in the dorsolateral parts of the lesion site showed robust expression of Semaphorin3A mRNA. In addition, dorsal root ganglion (DRG) neurons with projections through the dorsal columns to the injury site persistently expressed both Semaphorin3A receptor components, neuropilin-1 and plexin-A1. These ascending DRG collaterals failed to invade scar regions occupied by Semaphorin3A-positive fibroblasts, even in animals which had received conditioning lesions of the sciatic nerve to enhance regeneration. Other axon populations in the dorsal spinal cord were similarly unable to penetrate Semaphorin3A-positive scar tissue. These data suggest that Semaphorin3A may create an exclusion zone for regenerating dorsal column fibres and that enhancing the intrinsic regenerative response of DRG neurons has only limited effects on axonal regrowth. Tenascin-C and chondroitin sulphate proteoglycans were also detected at the injury site, which was largely devoid of central nervous system (CNS) myelin, showing that several classes of inhibitory factors, including semaphorins, with only partially overlapping spatial and temporal patterns of expression are in a position to participate in preventing regenerative axonal growth in the injured dorsal columns. Interestingly, conditioning nerve injuries enabled numerous ascending DRG axons to regrow across areas of strong tenascin-C and chondroitin sulphate proteoglycan expression, while areas containing Semaphorin3A and CNS myelin were selectively avoided by (pre)primed axonal sprouts.

Animals↗
PubMed2001

Mutant ubiquitin expressed in Alzheimer's disease causes neuronal death.

Ubiquitin-B+1 (UBB+1) is a mutant ubiquitin that accumulates in the neurones of patients with Alzheimer's disease (AD). Here we report on the biochemical and functional differences between ubiquitin and UBB+1 and the effect of the mutant protein on neuronal cells. UBB+1 lacks the capacity to ubiquitinate, and although it is ubiquitinated itself, UBB+1 is not degraded by the ubiquitin-proteasomal system and is quite stable in neuronal cells. Overexpression of UBB+1 in neuroblastoma cells significantly induces nuclear fragmentation and cell death. Our results demonstrate that accumulation of UBB+1 in neurones is detrimental and may contribute to neuronal dysfunction in AD patients.

Adenosine Triphosphate↗
PubMed2000

Tropism of AAV-2 vectors for neurons of the globus pallidus.

A recombinant AAV-2 vector encoding the green fluorescent protein (gfp) under the control of the cytomegalovirus (CMV) promoter was injected into the striatum at varying antero-posterior coordinates. When the virus was delivered to the anterior part of the striatum, transduction efficiency was low and limited to the vicinity of the needle tract. In contrast, after injection into the posterior part of the striatum, in addition to a localized transduced area in the striatum, efficient and widespread transduction was observed at distance from the injection site, in the globus pallidus. In the latter case, labelled cells were also detected in the internal capsule and in the stria terminalis. The number of transduced cells in the striatum increased up to I month and then decreased whereas in the globus pallidus, transduction was maximal as early as 2 weeks post-injection. In the striatum and in the globus pallidus, the labelled cells had a neuron-like morphology. In contrast, in the internal capsule, labelled cells had a glial-like morphology.

Animals↗
PubMed2000

Comparison of neurotrophin and repellent sensitivities of early embryonic geniculate and trigeminal axons.

Geniculate (gustatory) and trigeminal (somatosensory) afferents take different routes to the tongue during rat embryonic development. To learn more about the mechanisms controlling neurite outgrowth and axon guidance, we are studying the roles of diffusible factors. We previously profiled the in vitro sensitivity of trigeminal axons to neurotrophins and target-derived diffusible factors and now report on these properties for geniculate axons. GDNF, BDNF, and NT-4, but not NT-3 or NGF, stimulate geniculate axon outgrowth during the ages investigated, embryonic days 12-14. Sensitivity to effective neurotrophins is developmentally regulated and different from that of the trigeminal ganglion. In vitro coculture studies revealed that geniculate axons were repelled by branchial arch explants that were previously shown to be repellent to trigeminal axons (Rochlin and Farbman [1998] J Neurosci 18:6840-6852). In addition, some branchial arch explants and untransfected COS7 cells repelled geniculate but not trigeminal axons. Sema3A, a ligand for neuropilin-1, is effective in repelling geniculate and trigeminal axons, and antineuropilin-1, but not antineuropilin-2, completely blocks the repulsion by arch explants that repel axon outgrowth from both ganglia. Sema3A mRNA is concentrated in branchial arch epithelium at the appropriate time to mediate the repulsion. In Sema3A knockout mice, geniculate and trigeminal afferents explore medial regions of the immature tongue and surrounding territories not explored in heterozygotes, supporting our previous hypothesis that Sema3A-based repulsion mediates the early restriction of sensory afferents away from midline structures.

Animals↗
PubMed2000

Ectopic adenoviral vector-directed expression of Sema3A in organotypic spinal cord explants inhibits growth of primary sensory afferents.

Sema3A (Sema III, SemD, collapsin-1) can induce neuronal growth cone collapse and axon repulsion of distinct neuronal populations. To study Sema3A function in patterning afferent projections into the developing spinal cord, we employed the recombinant adenoviral vector technique in embryonic rat spinal cord slices. Virus solution was injected in the dorsal aspect of organotypic spinal cord cultures with segmentally attached dorsal root ganglia (sc-DRG). In cultures grown in the presence of nerve growth factor (NGF), injected either with the control virus AdCMVLacZ or with vehicle only, afferent innervation patterns were similar to those of control. However, unilateral injection of AdCMVSema3A/AdCMVLacZ in sc-DRG slices revealed a strong inhibitory effect on NGF-dependent sensory afferent growth. Ectopic Sema3A in the dorsal spinal cord, the target area of NGF-responsive DRG fibers in vivo, created an exclusion zone for these fibers and as a result they failed to reach and innervate their appropriate target zones. Taken together, gain of Sema3A function in the dorsal aspect of sc-DRG cultures revealed a dominant inhibitory effect on NGF-dependent, nociceptive sensory DRG afferents, an observation in line with the model proposed by E. K. Messersmith et al. (1995, Neuron 14, 949-959), suggesting that Sema3A secreted by spinal cord cells can act to repel central sensory fibers during the formation of lamina-specific connections in the spinal cord.

Adenoviridae↗
PubMed2000

P75 neurotrophin receptor in the nucleus basalis of meynert in relation to age, sex, and Alzheimer's disease.

In a previous study we showed that the staining of tyrosine kinase receptors (trks), which are high-affinity neurotrophin receptors (NTRs), is strongly diminished in the nucleus basalis of Meynert (NBM) of Alzheimer's disease (AD) patients, which may explain the lack of effect of NGF therapy in AD patients so far. Since the literature regarding the expression of low-affinity NTRs was rather controversial, the aim of the present study was to examine (i) possible changes in the staining of low-affinity NTRs, i.e., p75 in the human NBM, an area that is severely affected in AD; and (ii) alterations of these receptors in relation to risk factors for AD, e. g., age, sex, and menopause. Brain material of 31 controls and 30 AD patients was obtained at autopsy, embedded in paraffin, and stained immunocytochemically. Using an image analysis system, we quantified p75 immunoreactivity in both cell bodies and fibers at the level of the NBM. Our results showed a significant diminishment of p75 immunoreactivity in both cell bodies and fibers of NBM neurons in AD. We did not find any relationship between age or sex and the expression of p75 receptor in cell bodies. However, there was a clearly positive relationship between age and fiber staining in AD patients which suggests the occurrence of a p75 transport disorder as an early event in the process of AD. These observations and the earlier reported decreased staining of trk receptors show that degeneration of NBM neurons in AD is associated with a decreased neurotrophin responsiveness of NBM neurons in AD and that therapeutic strategies should be directed toward upregulation of receptors or facilitation of transport before an effect of neurotrophins in AD may be expected.

Age Factors↗
PubMed2000

Intercostal nerve implants transduced with an adenoviral vector encoding neurotrophin-3 promote regrowth of injured rat corticospinal tract fibers and improve hindlimb function.

Following injury to central nervous tissues, damaged neurons are unable to regenerate their axons spontaneously. Implantation of peripheral nerves into the CNS, however, does result in axonal regeneration into these transplants and is one of the most powerful strategies to promote CNS regeneration. In the present study implantation of peripheral nerve bridges following dorsal hemisection is combined with ex vivo gene transfer with adenoviral vectors encoding neurotrophin-3 (Ad-NT-3) to examine whether this would stimulate regeneration of one of the long descending tracts of the spinal cord, the corticospinal tract (CST), into and beyond the peripheral nerve implant. We chose to use an adenoviral vector encoding NT-3 because CST axons are sensitive to this neurotrophin and Schwann cells in peripheral nerve implants do not express this neurotrophin. At 16 weeks postimplantation of Ad-NT-3-transduced intercostal nerves, approximately three- to fourfold more of the anterogradely traced corticospinal tract fibers had regrown their axons through gray matter below the lesion site when compared to control animals. Regrowth of CST fibers occurred over more than 8 mm distal to the lesion site. No regenerating CST fibers were, however, observed into the transduced peripheral implant. Animals with a peripheral nerve transduced with Ad-NT-3 also exhibited improved function of the hindlimbs when compared to control animals treated with an adenoviral vector encoding LacZ. Thus, transient overexpression of NT-3 in peripheral nerve tissue bridges is apparently sufficient to stimulate regrowth of CST fibers and to promote recovery of hindlimb function, but does not result in regeneration of CST fibers into such transplants. Taken together, combining an established neurotransplantation approach with viral vector-gene transfer promotes the regrowth of injured CST fibers through gray matter and improves the recovery of hindlimb function.

Adenoviridae↗
PubMed1999

Transgenic expression of B-50/GAP-43 in mature olfactory neurons triggers downregulation of native B-50/GAP-43 expression in immature olfactory neurons.

The adult mammalian olfactory neuroepithelium is an unusual neural tissue, since it maintains its capacity to form new neurons throughout life. Newly formed neurons differentiate in the basal layers of the olfactory neuroepithelium and express B-50/GAP-43, a protein implicated in neurite outgrowth. During maturation these neurons migrate into the upper portion of the epithelium, upregulate expression of olfactory marker protein (OMP) and concomitantly downregulate the expression of B-50/GAP-43. Transgenic mice that exhibit OMP-promoter directed expression of B-50/GAP-43 in mature olfactory neurons display an unexpected decrease in the complement of B-50/GAP-43-positive cells in the lower region of the olfactory epithelium [A.J.G.D. Holtmaat, P.A. Dijkhuizen, A.B. Oestreicher, H. J. Romijn, N.M.T. Van der Lugt, A. Berns, F.L. Margolis, W.H. Gispen, J. Verhaagen, Directed expression of the growth-associated protein B-50/GAP-43 to olfactory neurons in transgenic mice results in changes in axon morphology and extraglomerular growth, J. Neurosci. 15 (1995) 7953-7965]. We have investigated whether the decrement in B-50/GAP-43-positive cells in this region was due to a dislocation of the immature neurons to other regions of the olfactory epithelium or to a downregulation of B-50/GAP-43 synthesis in these immature neurons. In eight of nine independent transgenic mouse lines that express the transgene in different numbers of olfactory neurons, a decline in the number of B-50/GAP-43-expressing neurons in the basal portion of the olfactory neuroepithelium was observed, both at the protein level and the mRNA level. An alternative marker for immature cells, a juvenile form of tubulin, was normally expressed in this location, indicating that the olfactory epithelium of OMP-B-50/GAP-43 transgenic mice contains a normal complement of immature olfactory neurons and that most of these neurons display a downregulation of B-50/GAP-43 expression.

Animals↗