Degradation Of Chondroitin Sulfate Proteoglycans Research Articles

Chondroitin sulfate proteoglycans mRNA expression and degradation in the zebra finch following traumatic brain injury

Traumatic brain injury (TBI) is one of the leading causes of fatality and disability worldwide. From minutes to months following damage, injury can result in a complex pathophysiology that can lead to temporary or permanent deficits including an array of neurodegenerative symptoms. These changes can include behavioral dysregulation, memory dysfunctions, and mood changes including depression. The nature and severity of impairments resulting from TBIs vary widely given the range of injury type, location, and extent of brain tissue involved. In response to the injury, the brain induces structural and functional changes to promote repair and minimize injury size. Despite its high prevalence, effective treatment strategies for TBI are limited. PNNs are part of the neuronal extracellular matrix (ECM) that mediate synaptic stabilization in the adult brain and thus neuroplasticity. They are associated mostly with inhibitory GABAergic interneurons and are thought to be responsible for maintaining the excitatory/inhibitory balance of the brain. The major structural components of PNNs include multiple chondroitin sulfate proteoglycans (CSPGs) as well as other structural proteins. Here we examine the effects of injury on CSPG expression, specifically around the changes in the side change moieties. To investigate CSPG expression following injury, adult male and female zebra finches received either a bilateral penetrating, or no injury and qPCR analysis and immunohistochemistry for components of the CSPGs were examined at 1- or 7-days post-injury. Next, to determine if CSPGs and thus PNNs should be a target for therapeutic intervention, CSPG side chains were degraded at the time of injury with chondroitinase ABC (ChABC) CSPGs moieties were examined. Additionally, GABA receptor mRNA and aromatase mRNA expression was quantified following CSPG degradation as they have been implicated in neuronal survival and neurogenesis. Our data indicate the CSPG moieties change following injury, potentially allowing for a brief period of synaptic reorganization, and that treatments that target CSPG side chains are successful in further targeting this brief critical period by decreasing GABA mRNA receptor expression, but also decreasing aromatase expression.

Chondroitin sulfate proteoglycan is a potential target of memantine to improve cognitive function via the promotion of adult neurogenesis.

Chondroitin sulfate proteoglycan (CSPG) constitutes the neurogenic niche in the hippocampus. The reduction of hippocampal neurogenesis is involved in ageing-related cognitive decline and dementia. The purpose of this study is to find candidates that improve cognitive function by analysing the effects of memantine (MEM), a therapeutic agent for Alzheimer's disease, on CSPG and adult hippocampal neurogenesis. The effects of MEM on neurogenesis-related cells and CSPG content were assessed in the hippocampus of middle-aged mice. The MEM-induced alterations in gene expressions of neurotrophins and enzymes associated with biosynthesis and degradation of CSPG in the hippocampus also were measured. The effects of MEM on cognitive function were estimated using a behavioural test battery. The same set of behavioural tests was applied to evaluate the effects of pharmacological depletion of CSPG in the hippocampus. The densities of newborn granule cells and content of CSPG in the hippocampus were increased by MEM. The expression levels of the enzyme responsible for the biosynthesis CSPG were increased by MEM. The neurotrophin-related molecules were activated by MEM. Short- and long-term memory performance was improved by MEM. Pharmacological depletion of CSPG impairs the effects of MEM on cognitive improvement in middle-aged mice. MEM regulates the biosynthesis and degradation of CSPG, which may underlie the improvement of cognitive function via the promotion of adult hippocampal neurogenesis. These results imply that CSPG-related enzymes potentially may be attractive candidates for the treatment of ageing-related cognitive decline.

Alleviation of cognitive deficits via upregulation of chondroitin sulfate biosynthesis by lignan sesamin in a mouse model of neuroinflammation

Lignans are plant-derived compounds that act as partial estrogen agonists. Chondroitin sulfate proteoglycans (CSPGs) represent one of the major components of the extracellular matrix. Here we aimed to understand the role of sesamin (SES), a major lignan compound, in the biosynthesis and degradation of CSPGs in the mouse hippocampus because CSPGs play a key role in the regulation of cognitive functions through the promotion of adult neurogenesis. The expression of the pro-inflammatory cytokine interleukin-1β was decreased by SES administration in the hippocampus of lipopolysaccharide (LPS)-treated mice, a model of neuroinflammation-induced cognitive deficits. The expression of genes related to biosynthesis and degradation of CSPGs in the hippocampus of LPS-treated mice was both increased and decreased by SES administration. Further, the diffuse extracellular matrix labeling of CSPGs by Wisteria floribunda agglutinin (WFA) in the hippocampus of LPS-treated mice was increased by SES administration. The densities of neural stem cells, late transit-amplifying cells, and newborn-granule cells in the hippocampus of LPS-treated mice were also increased by SES administration. Moreover, SES-induced alterations in gene expression, WFA labeling, and adult neurogenesis in LPS-treated mice were more evident in the dorsal hippocampus (center of cognition) than in the ventral hippocampus (center of emotion). Neither LPS nor SES administration affected locomotor activity, anxiety-like behavior, and depression-related behavior. However, impairments in contextual memory and sensorimotor gating in LPS-treated mice were recovered by SES administration. Our results show that SES can promote adult hippocampal neurogenesis through the upregulation of CSPGs, which may alleviate cognitive deficits induced by neuroinflammation.

ChABC infusions into medial prefrontal cortex, but not posterior parietal cortex, improve the performance of rats tested on a novel, challenging delay in the touchscreen TUNL task.

Perineuronal nets (PNNs) are specialized extracellular matrix structures that surround subsets of neurons throughout the central nervous system (CNS). They are made up of chondroitin sulfate proteoglycans (CSPGs), hyaluronan, tenascin-R, and many other link proteins that together make up their rigid and lattice-like structure. Modulation of PNNs can alter synaptic plasticity and thereby affect learning, memory, and cognition. In the present study, we degraded PNNs in the medial prefrontal (mPFC) and posterior parietal (PPC) cortices of Long–Evans rats using the enzyme chondroitinase ABC (ChABC), which cleaves apart CSPGs. We then measured the consequences of PNN degradation on spatial working memory (WM) with a trial-unique, non-matching-to location (TUNL) automated touchscreen task. All rats were trained with a standard 6 sec delay and 20 sec inter-trial interval (ITI) and then tested under four different conditions: a 6 sec delay, a variable 2 or 6 sec delay, a 2 sec delay with a 1 sec ITI (interference condition), and a 20 sec delay. Rats that received mPFC ChABC treatment initially performed TUNL with higher accuracy, more selection trials completed, and fewer correction trials completed compared to controls in the 20 sec delay condition but did not perform differently from controls in any other condition. Rats that received PPC ChABC treatment did not perform significantly differently from controls in any condition. Posthumous immunohistochemistry confirmed an increase in CSPG degradation products (C4S stain) in the mPFC and PPC following ChABC infusions while WFA staining intensity and parvalbumin positive neuron number were decreased following mPFC, but not PPC, ChABC infusions. These findings suggest that PNNs in the mPFC play a subtle role in spatial WM, but PNNs in the PPC do not. Furthermore, it appears that PNNs in the mPFC are involved in adapting to a challenging novel delay, but that they do not play an essential role in spatial WM function.

Perturbing chondroitin sulfate proteoglycan signaling through LAR and PTP\u03c3 receptors promotes a beneficial inflammatory response following spinal cord injury

BackgroundTraumatic spinal cord injury (SCI) results in upregulation of chondroitin sulfate proteoglycans (CSPGs) by reactive glia that impedes repair and regeneration in the spinal cord. Degradation of CSPGs is known to be beneficial in promoting endogenous repair mechanisms including axonal sprouting/regeneration, oligodendrocyte replacement, and remyelination, and is associated with improvements in functional outcomes after SCI. Recent evidence suggests that CSPGs may regulate secondary injury mechanisms by modulating neuroinflammation after SCI. To date, the role of CSPGs in SCI neuroinflammation remains largely unexplored. The recent discovery of CSPG-specific receptors, leukocyte common antigen-related (LAR) and protein tyrosine phosphatase-sigma (PTPσ), allows unraveling the cellular and molecular mechanisms of CSPGs in SCI. In the present study, we have employed parallel in vivo and in vitro approaches to dissect the role of CSPGs and their receptors LAR and PTPσ in modulating the inflammatory processes in the acute and subacute phases of SCI.MethodsIn a clinically relevant model of compressive SCI in female Sprague Dawley rats, we targeted LAR and PTPσ by two intracellular functionally blocking peptides, termed ILP and ISP, respectively. We delivered ILP and ISP treatment intrathecally to the injured spinal cord in a sustainable manner by osmotic mini-pumps for various time-points post-SCI. We employed flow cytometry, Western blotting, and immunohistochemistry in rat SCI, as well as complementary in vitro studies in primary microglia cultures to address our questions.ResultsWe provide novel evidence that signifies a key immunomodulatory role for LAR and PTPσ receptors in SCI. We show that blocking LAR and PTPσ reduces the population of classically activated M1 microglia/macrophages, while promoting alternatively activated M2 microglia/macrophages and T regulatory cells. This shift was associated with a remarkable elevation in pro-regenerative immune mediators, interleukin-10 (IL-10), and Arginase-1. Our parallel in vitro studies in microglia identified that while CSPGs do not induce an M1 phenotype per se, they promote a pro-inflammatory phenotype. Interestingly, inhibiting LAR and PTPσ in M1 and M2 microglia positively modulates their inflammatory response in the presence of CSPGs, and harnesses their ability for phagocytosis and mobilization. Interestingly, our findings indicate that CSPGs regulate microglia, at least in part, through the activation of the Rho/ROCK pathway downstream of LAR and PTPσ.ConclusionsWe have unveiled a novel role for LAR and PTPσ in regulating neuroinflammation in traumatic SCI. Our findings provide new insights into the mechanisms by which manipulation of CSPG signaling can promote recovery from SCI. More importantly, this work introduces the potential of ILP/ISP as a viable strategy for modulating the immune response following SCI and other neuroinflammatory conditions of the central nervous system.

ADAMTS-4 in central nervous system pathologies.

ADAMTS-4 (a disintegrin and metalloproteinase with thrombospondin motifs type 4) is a metalloproteinase specialized in the degradation of chondroitin sulfate proteoglycans, contributing to cartilage breakdown during arthritis. In this review, we first focus on the modifications of ADAMTS-4 expression during CNS physiological and pathological conditions, including chronic diseases and injuries. Then, we discuss the contributions of ADAMTS-4 to mechanisms mediating neuroplasticity, neuroinflammation and neurodegeneration during spinal cord injury, ischemic stroke, amyotrophic lateral sclerosis and Alzheimer's disease. Here, we provide an overview of ADAMTS-4 functions and effects in the CNS, and we discuss directions for future studies and treatments. Overall, this review highlights that ADAMTS-4 is a unique multifaceted metalloproteinase which influences various CNS disease pathophysiologies. © 2017 Wiley Periodicals, Inc.

Combined chondroitinase and KLF7 expression reduce net retraction of sensory and CST axons from sites of spinal injury

Axon regeneration in the central nervous system is limited both by inhibitory extracellular cues and by an intrinsically low capacity for axon growth in some CNS populations. Chondroitin sulfate proteoglycans (CSPGs) are well-studied inhibitors of axon growth in the CNS, and degradation of CSPGs by chondroitinase has been shown to improve the extension of injured axons. Alternatively, axon growth can be improved by targeting the neuron-intrinsic growth capacity through forced expression of regeneration-associated transcription factors. For example, a transcriptionally active chimera of Krüppel-like Factor 7 (KLF7) and a VP16 domain improves axon growth when expressed in corticospinal tract neurons. Here we tested the hypothesis that combined expression of chondroitinase and VP16-KLF7 would lead to further improvements in axon growth after spinal injury. Chondroitinase was expressed by viral transduction of cells in the spinal cord, while VP16-KLF7 was virally expressed in sensory neurons of the dorsal root ganglia or corticospinal tract (CST) neurons. After transection of the dorsal columns, both chondroitinase and VP16-KLF7 increased the proximity of severed sensory axons to the injury site. Similarly, after complete crush injuries, VP16-KLF7 expression increased the approach of CST axons to the injury site. In neither paradigm however, did single or combined treatment with chondroitinase or VP16-KLF7 enable regenerative growth distal to the injury. These results substantiate a role for CSPG inhibition and low KLF7 activity in determining the net retraction of axons from sites of spinal injury, while suggesting that additional factors act to limit a full regenerative response.

Conditional Sox9 ablation improves locomotor recovery after spinal cord injury by increasing reactive sprouting

The absence of axonal regeneration after spinal cord injury (SCI) has been attributed to the up-regulation of axon-repelling molecules, such as chondroitin sulfate proteoglycans (CSPGs) present in the glial scar that forms post-SCI. We previously identified the transcription factor SOX9 as a key up-regulator of CSPG production and also demonstrated that conditional Sox9 ablation leads to decreased CSPG levels and improved recovery of hind limb function after SCI. We herein demonstrate increased neural input onto spinal neurons caudal to the lesion in spinal cord injured Sox9 conditional knock out mice as indicated by increased levels of the presynaptic markers synaptophysin and vesicular glutamate transporter 1 (VGLUT1) compared to controls. Axonal sparing, long-range axonal regeneration and reactive sprouting were investigated as possible explanations for the increase in neural inputs caudal to the lesion and for the improved locomotor outcomes in spinal cord-injured Sox9 conditional knock out mice. Whereas retrograde tract-tracing studies failed to reveal any evidence for increased axonal sparing or for long-range regeneration in the Sox9 conditional knock out mice, anterograde tract-tracing experiments demonstrated increased reactive sprouting caudal to the lesion after SCI. Finally we demonstrate that application of a broad spectrum MMP inhibitor to reduce CSPG degradation in Sox9 conditional knock out mice prevents the improvements in locomotor recovery observed in untreated Sox9 conditional knock out mice. These results suggest that improved recovery of locomotor function in Sox9 conditional knock out mice after SCI is due to increased reactive sprouting secondary to reduced CSPG levels distal to the lesion.

Editorial: Neuroplasticity and Extracellular Proteolysis.

Editorial: Neuroplasticity and Extracellular Proteolysis.

ADAMTS-4 promotes neurodegeneration in a mouse model of amyotrophic lateral sclerosis.

BackgroundA disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS) proteoglycanases are specialized in the degradation of chondroitin sulfate proteoglycans and participate in mechanisms mediating neuroplasticity. Despite the beneficial effect of ADAMTS-4 on neurorepair after spinal cord injury, the functions of ADAMTS proteoglycanases in other CNS disease states have not been studied. Therefore, we investigated the expression, effects and associated mechanisms of ADAMTS-4 during amyotrophic lateral sclerosis (ALS) in the SOD1G93A mouse model.ResultsADAMTS-4 expression and activity were reduced in the spinal cord of SOD1G93A mice at disease end-stage when compared to WT littermates. To counteract the loss of ADAMTS-4, SOD1G93A and WT mice were treated with saline or a recombinant ADAMTS-4 before symptom onset. Administration of ADAMTS-4 worsened the prognosis of SOD1G93A mice by accelerating clinical signs of neuromuscular dysfunctions. The worsened prognosis of ADAMTS-4-treated SOD1G93A mice was accompanied by increased degradation of perineuronal nets enwrapping motoneurons and increased motoneuron degeneration in the lumbar spinal cord. Motoneurons of ADAMTS-4-treated SOD1G93A mice were more vulnerable to degeneration most likely due to the loss of their extracellular matrix envelopes. The decrease of neurotrophic factor production induced by ADAMTS-4 in vitro and in vivo may also contribute to a hostile environment for motoneuron especially when devoid of a net.ConclusionsThis study suggests that the reduction of ADAMTS-4 activity during the progression of ALS pathology may be an adaptive change to mitigate its neurodegenerative impact in CNS tissues. Therapies compensating the compromized ADAMTS-4 activity are likely not promising approaches for treating ALS.

Effect of the combination of mesenchymal stromal cells and chondroitinase ABC on chronic spinal cord injury

Transplantation of mesenchymal stromal cells (MSCs) has been identified as a potential therapeutic modality for treating spinal cord injury (SCI). Degradation of chondroitin sulfate proteoglycans (CSPGs) using the enzyme chondroitinase ABC (chABC) can promote functional recovery after SCI. The effect of the simultaneous administration of MSCs and chABC on chronic SCI was investigated. Sixteen dogs were assigned to one of the following four groups: (i) canine adipose tissue-derived MSCs (cADMSCs), (ii) chABC, (iii) cADMSCs + chABC and (iv) control. Treatments were carried out 3 weeks after SCI; cADMSCs (1 × 107 cells suspended in 150 μL of PBS), chABC (5 U/mL, 150 μL), cADMSCs + chABC (1 × 107 cells suspended in 150 μL of chABC), or phosphate-buffered saline (150 μL) were injected into the spinal cord at three locations to a depth of 3 mm using a 30-gauge needle. The spinal cord was harvested 8 weeks after transplantation. In a behavioral assessment, dogs treated with cADMSCs + chABC and cADMSCs alone showed significantly better functional recovery 8 weeks after transplantation compared with the control and chABC groups (P < 0.05). In addition, the combination of cADMSCs and chABC increased the expression of digested CSPGs (2B6), β3 tubulin, and NF-M. However, the levels of COX2 (P < 0.05), and tumor necrosis factor-α was higher in the treatment groups than in the control. In conclusion, transplantation of cADMSCs + chABC was more effective in improving clinical signs and neural regeneration, but a strategy for anti-inflammation after the treatment for chronic SCI would be needed for further improvement.

Understanding the functions of endogenous and exogenous tissue-type plasminogen activator during stroke.

Alteplase (Actilyse or Activase) is the gold standard acute treatment of ischemic stroke1 and receives attention for hematoma resolution in hemorrhagic stroke. The active principle of Alteplase is the recombinant form of an endogenous protease, tissue-type plasminogen activator (tPA). Its intravascular thrombolytic activity is well known, but less are its multifaceted functions in the central nervous system (CNS). Endogenous tPA is not only released in the blood by endothelial cells but also expressed by many cells within the brain parenchyma (online-only Data Supplement), and it can act on all cell types of the brain virtually. Endogenous tPA has been involved in an ever-increasing number of brain functions, of which several are highly relevant during and after stroke. Importantly, knowledge on the mechanisms of action of endogenous tPA may hold true for recombinant tPA. In this review, we provide an up-to-date overview of the current knowledge on the enzymatic or cytokine-like effects of action of tPA in the CNS, its various molecular substrates or receptors, focusing on the processes occurring during and after ischemic or hemorrhagic stroke, including excitotoxicity, apoptosis, blood–brain barrier breakdown, inflammation, axonal damage, and demyelination. tPA is a mosaic protease of 527 amino acids consisting of 5 distinct modules: a Finger domain, an epidermal growth factor (EGF)–like domain, 2 kringle domains (K1 and K2), and a serine protease proteolytic domain. Through these domains, tPA can interact with a variety of binding proteins and receptors in the brain parenchyma, thus extending its functions above the conversion of plasminogen into plasmin (Figure 1). Figure 1. Main cellular and molecular targets of tissue-type plasminogen activator in the central nervous system. BBB indicates blood–brain barrier, EGFR, epidermal growth factor receptor; LRP, low-density lipoprotein receptor–related protein; NMDAR, N-methyl-d-aspartate receptors; and PDGFR, platelet-derived growth factor receptor. tPA is not fibrinolytic by itself. …

TPA promotes ADAMTS-4-induced CSPG degradation, thereby enhancing neuroplasticity following spinal cord injury

Although tissue plasminogen activator (tPA) is known to promote neuronal remodeling in the CNS, no mechanism of how this plastic function takes place has been reported so far. We provide here in vitro and in vivo demonstrations that this serine protease neutralizes inhibitory chondroitin sulfate proteoglycans (CSPGs) by promoting their degradation via the direct activation of endogenous type 4 disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS-4). Accordingly, in a model of compression-induced spinal cord injury (SCI) in rats, we found that administration of either tPA or its downstream effector ADAMTS-4 restores the tPA-dependent activity lost after the SCI and thereby, reduces content of CSPGs in the spinal cord, a cascade of events leading to an improved axonal regeneration/sprouting and eventually long term functional recovery. This is the first study to reveal a tPA–ADAMTS-4 axis and its function in the CNS. It also raises the prospect of exploiting such cooperation as a therapeutic tool for enhancing recovery after acute CNS injuries.

Combination of engineered Schwann cell grafts to secrete neurotrophin and chondroitinase promotes axonal regeneration and locomotion after spinal cord injury.

Transplantation of Schwann cells (SCs) is a promising therapeutic strategy for spinal cord repair. SCs introduced into lesions support axon regeneration, but because these axons do not exit the transplant, additional approaches with SCs are needed. Here, we transplanted SCs genetically modified to secrete a bifunctional neurotrophin (D15A) and chondroitinase ABC (ChABC) into a subacute contusion injury in rats. We examined the effects of these modifications on graft volume, SC number, degradation of chondroitin sulfate proteoglycans (CSPGs), astrogliosis, SC myelination of axons, propriospinal and supraspinal axon numbers, locomotor outcome (BBB scoring, CatWalk gait analysis), and mechanical and thermal sensitivity on the hind paws. D15A secreted from transplanted SCs increased graft volume and SC number and myelinated axon number. SCs secreting ChABC significantly decreased CSPGs, led to some egress of SCs from the graft, and increased propriospinal and 5-HT-positive axons in the graft. SCs secreting both D15A and ChABC yielded the best responses: (1) the largest number of SC myelinated axons, (2) more propriospinal axons in the graft and host tissue around and caudal to it, (3) more corticospinal axons closer to the graft and around and caudal to it, (4) more brainstem neurons projecting caudal to the transplant, (5) increased 5-HT-positive axons in the graft and caudal to it, (6) significant improvement in aspects of locomotion, and (7) improvement in mechanical and thermal allodynia. This is the first evidence that the combination of SC transplants engineered to secrete neurotrophin and chondroitinase further improves axonal regeneration and locomotor and sensory function.

Chondroitin sulfate proteoglycans inhibit oligodendrocyte myelination through PTPσ

CNS damage often results in demyelination of spared axons due to oligodendroglial cell death and dysfunction near the injury site. Although new oligodendroglia are generated following CNS injury and disease, the process of remyelination is typically incomplete resulting in long-term functional deficits. Chondroitin sulfate proteoglycans (CSPGs) are upregulated in CNS grey and white matter following injury and disease and are a major component of the inhibitory scar that suppresses axon regeneration. CSPG inhibition of axonal regeneration is mediated, at least in part, by the protein tyrosine phosphatase sigma (PTPσ) receptor. Recent evidence demonstrates that CSPGs inhibit OL process outgrowth, however, the means by which their effects are mediated remains unclear. Here we investigate the role of PTPσ in CSPG inhibition of OL function. We found that the CSPGs, aggrecan, neurocan and NG2 all imposed an inhibitory effect on OL process outgrowth and myelination. These inhibitory effects were reversed by degradation of CSPGs with Chondroitinase ABC prior to OL exposure. RNAi-mediated down-regulation of PTPσ reversed the inhibitory effect of CSPGs on OL process outgrowth and myelination. Likewise, CSPG inhibition of process outgrowth and myelination was significantly reduced in cultures containing PTPσ−/− OLs. Finally, inhibition of Rho-associated kinase (ROCK) increased OL process outgrowth and myelination during exposure to CSPGs. These results suggest that in addition to their inhibitory effects on axon regeneration, CSPGs have multiple inhibitory actions on OLs that result in incomplete remyelination following CNS injury. The identification of PTPσ as a receptor for CSPGs, and the participation of ROCK downstream of CSPG exposure, reveal potential therapeutic targets to enhance white matter repair in the damaged CNS.

Intraventricular hemorrhage induces deposition of proteoglycans in premature rabbits, but their in vivo degradation with chondroitinase does not restore myelination, ventricle size and neurological recovery

Intraventricular hemorrhage (IVH) results in white matter injury and hydrocephalus in premature infants. Chondroitin sulfate proteoglycans (CSPGs)–neuorcan, brevican, versican, aggrecan and phosphacan–are unregulated in the extracellular matrix after brain injury, and their degradation enhances plasticity of the brain. Therefore, we hypothesized that CSPG levels were elevated in the forebrain of premature infants with IVH and that in vivo degradation of CSPGs would enhance maturation of oligodendrocyte, augment myelination, promote neurological recovery, and minimize hydrocephalus. We found that levels of neurocan, brevican, aggrecan, phosphacan, and versican were elevated, whereas NG2 expression was reduced in premature rabbit pups and human infants with IVH compared to controls. Intracerebroventricular chondroitinase ABC (ChABC) reduced the expression of neuorcan, brevican, versican and aggrecan, but not NG2. However, ChABC treatment did not enhance maturation of oligodendrocytes, myelination, or neurological recovery in the pups with IVH. Moreover, ChABC did not reduce gliosis or ventriculomegaly. Our results demonstrate that IVH induces distinct changes in the components of CSPGs, and that reversing these changes by in vivo ChABC treatment neither promotes clinical recovery, myelination, nor reduces ventriculomegaly in preterm rabbit pups.

Extracellular matrix inhibits structural and functional plasticity of dendritic spines in the adult visual cortex

Brain cells are immersed in a complex structure forming the extracellular matrix. The composition of the matrix gradually matures during postnatal development, as the brain circuitry reaches its adult form. The fully developed extracellular environment stabilizes neuronal connectivity and decreases cortical plasticity as highlighted by the demonstration that treatments degrading the matrix are able to restore synaptic plasticity in the adult brain. The mechanisms through which the matrix inhibits cortical plasticity are not fully clarified. Here we show that a prominent component of the matrix, chondroitin sulfate proteoglycans (CSPGs), restrains morphological changes of dendritic spines in the visual cortex of adult mice. By means of in vivo and in vitro two-photon imaging and electrophysiology, we find that after enzymatic digestion of CSPGs, cortical spines become more motile and express a larger degree of structural and functional plasticity.

Effects of Digesting Chondroitin Sulfate Proteoglycans on Plasticity in Cat Primary Visual Cortex

Monocular deprivation (MD) during a critical period of postnatal development produces significant changes in the anatomy and physiology of the visual cortex, and the deprived eye becomes amblyopic. Extracellular matrix molecules have a major role in restricting plasticity such that the ability to recover from MD decreases with age. Chondroitin sulfate proteoglycans (CSPGs) act as barriers to cell migration and axon growth. Previous studies showing that degradation of CSPGs by the bacterial enzyme chondroitinase can restore plasticity in the adult rat visual cortex suggest a potential treatment for amblyopia. Here MD was imposed in cats from the start of the critical period until 3.5 months of age. The deprived eye was reopened, the functional architecture of the visual cortex was assessed by optical imaging of intrinsic signals, and chondroitinase was injected into one hemisphere. Imaging was repeated 1 and 2 weeks postinjection, and visually evoked potentials (VEPs) and single-cell activity were recorded. Immunohistochemistry showed that digestion of CSPGs had been successful. After 2 weeks of binocular exposure, some recovery of deprived-eye responses occurred when chondroitinase had been injected into the hemisphere contralateral to that eye; when injected into the ipsilateral hemisphere, no recovery was seen. Deprived-eye VEPs were no larger in the injected hemisphere than in the opposite hemisphere. The small number of neurons dominated by the deprived eye exhibited poor tuning characteristics. These results suggest that despite structural effects of chondroitinase in adult cat V1, plasticity was not sufficiently restored to enable significant functional recovery of the deprived eye.

Gene delivery to overcome astrocyte inhibition of axonal growth: An in vitro Model of the glial scar

After injury to the central nervous system, a glial scar develops that physically and biochemically inhibits axon growth. In the scar, activated astrocytes secrete inhibitory extracellular matrix, of which chondroitin sulfate proteoglycans (CSPGs) are considered the major inhibitory component. An inhibitory interface of CSPGs forms around the lesion and prevents axons from traversing the injury, and decreasing CSPGs can enhance axon growth. In this report, we established an in vitro interface model of activated astrocytes and subsequently investigated gene delivery as a means to reduce CSPG levels and enhance axon growth. In the model, a continuous interface of CSPG producing astrocytes was created with neurons seeded opposite the astrocytes, and neurite crossing, stopping, and turning were evaluated as they approached the interface. We investigated the efficacy of lentiviral delivery to degrade or prevent the synthesis of CSPGs, thereby removing CSPG inhibition of neurite growth. Lentiviral delivery of RNAi targeting two key CSPG synthesis enzymes, chondroitin polymerizing factor and chondroitin synthase-1, decreased CSPGs, and reduced inhibition by the interface. Degradation of CSPGs by lentiviral delivery of chondroitinase also resulted in less inhibition and more neurites crossing the interface. These results indicate that the interface model provides a tool to investigate interventions that reduce inhibition by CSPGs, and that gene delivery can be effective in promoting neurite growth across an interface of CSPG producing astrocytes.

Chondroitin sulfate proteoglycans in demyelinated lesions impair remyelination

Failure of remyelination is a critical impediment to recovery in multiple sclerosis (MS). Chondroitin sulfate proteoglycans (CSPGs) have been reported to accumulate in MS lesions, and we thus examined the functional roles of CSPGs on oligodendrocyte precursor cells (OPCs), oligodendrocytes, and remyelination. We evaluated the expression of CSPGs in lysolecithin-injected mouse spinal cord, an animal model of demyelination and spontaneous remyelination. The functional impact of CSPGs on OPCs and remyelination was investigated using cultured adult murine and human OPCs and by treating demyelinated mice with xyloside to reduce the CSPG deposition that occurred following injury. Early and robust upregulation of CSPGs following lysolecithin-induced demyelination was cleared during remyelination. In culture, CSPGs anchored onto the substratum reduced the adhesion of mouse and human OPCs and their subsequent morphological differentiation into process-bearing oligodendrocytes. Soluble CSPGs added to already adherent OPCs reduced the development of processes, whereas the acquisition of mature myelin proteins was unimpeded. Stripe assays of alternating CSPG and control substrata confirmed the nonpermissive nature of CSPGs for OPC adhesion and morphological differentiation. Enzymatic degradation of CSPGs with chondroitinase ABC was sufficient to overcome CSPG-dependent inhibition of human oligodendrocytes. Finally, in vivo xyloside treatment to reduce CSPG synthesis in lysolecithin-demyelinated mice increased numbers of OPCs and oligodendrocytes in lesions, and culminated in improved remyelination. These results identify CSPGs as a nonpermissive substrate for OPCs and oligodendrocytes, and as a prominent impediment to remyelination. The data suggest the requirement for the neutralization of CSPGs for repair after demyelination.