Alterations in sulfated chondroitin glycosaminoglycans following controlled cortical impact injury in mice
ABSTRACT Chondroitin sulfate proteoglycans (CSPGs) play a pivotal role in many neuronal growth mechanisms including axon guidance and the modulation of repair processes following injury to the spinal cord or brain. Many actions of CSPGs in the central nervous system (CNS) are governed by the specific sulfation pattern on the glycosaminoglycan (GAG) chains attached to CSPG core proteins. To elucidate the role of CSPGs and sulfated GAG chains following traumatic brain injury (TBI), controlled cortical impact injury of mild to moderate severity was performed over the left sensory motor cortex in mice. Using immunoblotting and immunostaining, we found that TBI resulted in an increase in the CSPGs neurocan and NG2 expression in a tight band surrounding the injury core, which overlapped with the presence of 4-sulfated CS GAGs but not with 6-sulfated GAGs. This increase was observed as early as 7 days post injury (dpi), and persisted for up to 28 dpi. Labeling with markers against microglia/macrophages, NG2+ cells, fibroblasts, and astrocytes showed that these cells were all localized in the area, suggesting multiple origins of chondroitin-4-sulfate increase. TBI also caused a decrease in the expression of aggrecan and phosphacan in the pericontusional cortex with a concomitant reduction in the number of perineuronal nets. In summary, we describe a dual response in CSPGs whereby they may be actively involved in complex repair processes following TBI.
- SourceAvailable from: Thomas Dierks[Show abstract] [Hide abstract]
ABSTRACT: Bacterial chondroitinase ABC (ChaseABC) has been used to remove the inhibitory chondroitin sulfate chains from chondroitin sulfate proteoglycans to improve regeneration after rodent spinal cord injury. We hypothesized that the mammalian enzyme arylsulfatase B (ARSB) would also enhance recovery after mouse spinal cord injury. Application of the mammalian enzyme would be an attractive alternative to ChaseABC because of its more robust chemical stability and reduced immunogenicity. A one-time injection of human ARSB into injured mouse spinal cord eliminated immunoreactivity for chondroitin sulfates within five days, and up to 9 weeks after injury. After a moderate spinal cord injury, we observed improvements of locomotor recovery assessed by the Basso Mouse Scale (BMS) in ARSB treated mice, compared to the buffer-treated control group, at 6 weeks after injection. After a severe spinal cord injury, mice injected with equivalent units of ARSB or ChaseABC improved similarly and both groups achieved significantly more locomotor recovery than the buffer-treated control mice. Serotonin and tyrosine hydroxylase immunoreactive axons were more extensively present in mouse spinal cords treated with ARSB and ChaseABC, and the immunoreactive axons penetrated further beyond the injury site in ARSB or ChaseABC treated mice than in control mice. These results indicate that mammalian ARSB improves functional recovery after CNS injury. The structural/molecular mechanisms underlying the observed functional improvement remain to be elucidated.PLoS ONE 03/2013; 8(3):e57415. DOI:10.1371/journal.pone.0057415 · 3.53 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: The beneficial effect of interventions with chondroitinase-ABC enzyme to reduce axon growth-inhibitory chondroitin sulphate side-chains after CNS injuries has been mainly attributed to enhanced axonal sprouting. After traumatic brain injury it is unknown whether newly sprouting axons that occur as a result of interventional strategies are able to functionally contribute to existing circuitry, and it is uncertain if maladaptive sprouting occurs to increase the well-known risk for seizure activity after traumatic brain injury. Here we show that after a controlled cortical impact injury in rats, chondroitinase infusion into injured cortex at 30 min and 3 days reduced c-Fos+ cell staining resulting from the injury alone at one week post-injury, indicating that baseline, abnormal spontaneous activity is likely to be reduced, not increased with this type of intervention. c-Fos+ cell staining elicited by neural activity from stimulation of the affected forelimb one week after injury was significantly enhanced by chondroitinase, indicating a widespread effect on cortical map plasticity. Underlying this map plasticity was a larger contribution of neuronal rather than glial cells and an absence of c-Fos+ cells surrounded by perineuronal nets that were normally present in stimulated naïve rats. Following injury, CSPG digestion produced the expected increase in GAP43+ axons and perikarya, of which a significantly greater number were double-labelled for c-Fos after intervention with chondroitinase compared to vehicle. These data indicate that chondroitinase produces significant gains in cortical map plasticity after traumatic brain injury, and that either axonal sprouting and/or changes in perineuronal nets may underlie this effect. Chondroitinase dampens rather than increases non-specific c-Fos activity after brain injury, and the induction of axonal sprouting is not maladaptive, since greater numbers are functionally active and provide a significant contribution to the forelimb circuitry after brain injury.Journal of neurotrauma 03/2013; DOI:10.1089/neu.2012.2737 · 3.97 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: Cytoplasmic dynein plays important roles in mitosis and the intracellular transport of organelles, proteins, and mRNAs. Dynein function is particularly critical for survival of neurons, as mutations in dynein are linked to neurodegenerative diseases. Dynein function is also implicated in neuronal regeneration, driving the active transport of signaling molecules following injury of peripheral neurons. To enhance our understanding of dynein function and regulation in neurons, we established a novel knock-in mouse line in which the neuron-specific cytoplasmic dynein 1 intermediate chain 1 (IC-1)is tagged with both GFP and a 3xFLAG tag at its C-terminus.The fusion gene is under the control of IC-1's endogenous promoter and is integrated at the endogenous locus of the IC-1-encoding gene Dync1i1. The IC-1-GFP-3xFLAG fusion protein is incorporated into the endogenous dynein complex, and movements of GFP-labeled dynein expressed at endogenous levels can be observed in cultured neurons for the first time. The knock-in mouse line also allows isolation and analysisof dynein-bound proteins specifically from neurons. Using this mouse line we have found proteins, including 14-3-3 zeta, which physically interact with dynein upon injury of the brain cortex.Thus, we have created a useful tool for studying dynein function in the central nervous system under normal and pathologic conditions. © 2013 Wiley-Blackwell, Inc.Cytoskeleton 04/2013; 70(4). DOI:10.1002/cm.21102 · 3.01 Impact Factor