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Pharmacological Suppression of CNS Scarring by Deferoxamine Reduces Lesion Volume and Increases Regeneration in an In Vitro Model for Astroglial-Fibrotic Scarring and in Rat Spinal Cord Injury In Vivo.

Vogelaar CF, König B, Krafft S, Estrada V, Brazda N, Ziegler B, Faissner A, Müller HW - PLoS ONE (2015)

Bottom Line: DFO could be identified as a putative anti-scarring treatment for CNS trauma.We subsequently validated this by local application of DFO to a dorsal hemisection in the rat thoracic spinal cord.DFO treatment led to significant reduction of scarring, slightly increased regeneration of corticospinal tract as well as ascending CGRP-positive axons and moderately improved locomotion.

View Article: PubMed Central - PubMed

Affiliation: Molecular Neurobiology Laboratory, Department of Neurology, Heinrich-Heine-University of Duesseldorf, Duesseldorf, Germany; Institute of Microanatomy and Neurobiology, Johannes Gutenberg-University Mainz, Mainz, Germany.

ABSTRACT
Lesion-induced scarring is a major impediment for regeneration of injured axons in the central nervous system (CNS). The collagen-rich glial-fibrous scar contains numerous axon growth inhibitory factors forming a regeneration-barrier for axons. We demonstrated previously that the combination of the iron chelator 2,2'-bipyridine-5,5'-decarboxylic acid (BPY-DCA) and 8-Br-cyclic AMP (cAMP) inhibits scar formation and collagen deposition, leading to enhanced axon regeneration and partial functional recovery after spinal cord injury. While BPY-DCA is not a clinical drug, the clinically approved iron chelator deferoxamine mesylate (DFO) may be a suitable alternative for anti-scarring treatment (AST). In order to prove the scar-suppressing efficacy of DFO we modified a recently published in vitro model for CNS scarring. The model comprises a co-culture system of cerebral astrocytes and meningeal fibroblasts, which form scar-like clusters when stimulated with transforming growth factor-β (TGF-β). We studied the mechanisms of TGF-β-induced CNS scarring and compared the efficiency of different putative pharmacological scar-reducing treatments, including BPY-DCA, DFO and cAMP as well as combinations thereof. We observed modulation of TGF-β-induced scarring at the level of fibroblast proliferation and contraction as well as specific changes in the expression of extracellular matrix molecules and axon growth inhibitory proteins. The individual and combinatorial pharmacological treatments had distinct effects on the cellular and molecular aspects of in vitro scarring. DFO could be identified as a putative anti-scarring treatment for CNS trauma. We subsequently validated this by local application of DFO to a dorsal hemisection in the rat thoracic spinal cord. DFO treatment led to significant reduction of scarring, slightly increased regeneration of corticospinal tract as well as ascending CGRP-positive axons and moderately improved locomotion. We conclude that the in vitro model for CNS scarring is suitable for efficient pre-screening and identification of putative scar-suppressing agents prior to in vivo application and validation, thus saving costs, time and laboratory animals.

No MeSH data available.


Related in: MedlinePlus

Effects of treatments on neurite outgrowth.Neurite outgrowth from neonatal cortical neurons on the fibroblast and astrocyte cell layers as well as on the clusters in the co-cultures after treatment. cAMP and the combination treatments had a positive effect on outgrowth on the fibroblast layer, whereas only the combination treatments resulted in increased growth on the astrocytes (A, B). DFO was the only treatment resulting in increased growth of neurites on the clusters. Corrected for the cluster diameter, DFO showed an increase in the maximal length (C) and the sum length of the neurites (D). Representative pictures of clusters with neurites (E-H) that were traced using the Neuron J plugin of Image J (e-h, clusters marked by dotted black lines). Statistics: one way Anova with Dunnett’s post-hoc test * p < 0.05, ** p < 0.01, *** p< 0.001 Scale bar = 100 μm.
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pone.0134371.g007: Effects of treatments on neurite outgrowth.Neurite outgrowth from neonatal cortical neurons on the fibroblast and astrocyte cell layers as well as on the clusters in the co-cultures after treatment. cAMP and the combination treatments had a positive effect on outgrowth on the fibroblast layer, whereas only the combination treatments resulted in increased growth on the astrocytes (A, B). DFO was the only treatment resulting in increased growth of neurites on the clusters. Corrected for the cluster diameter, DFO showed an increase in the maximal length (C) and the sum length of the neurites (D). Representative pictures of clusters with neurites (E-H) that were traced using the Neuron J plugin of Image J (e-h, clusters marked by dotted black lines). Statistics: one way Anova with Dunnett’s post-hoc test * p < 0.05, ** p < 0.01, *** p< 0.001 Scale bar = 100 μm.

Mentions: In order to study putative axon growth-permitting properties of the scar-reducing treatments, we quantified neurite outgrowth of neonatal cortical neurons plated onto the co-cultures (Fig 7). On the astrocyte layer, only the combination of iron chelators with cAMP significantly increased the average neurite length per neuron. On the fibroblast layer, cAMP alone or in combination with iron chelators increased neurite length significantly. Only a non-significant trend towards longer neurites on fibroblasts was observed upon DFO-treatment.


Pharmacological Suppression of CNS Scarring by Deferoxamine Reduces Lesion Volume and Increases Regeneration in an In Vitro Model for Astroglial-Fibrotic Scarring and in Rat Spinal Cord Injury In Vivo.

Vogelaar CF, König B, Krafft S, Estrada V, Brazda N, Ziegler B, Faissner A, Müller HW - PLoS ONE (2015)

Effects of treatments on neurite outgrowth.Neurite outgrowth from neonatal cortical neurons on the fibroblast and astrocyte cell layers as well as on the clusters in the co-cultures after treatment. cAMP and the combination treatments had a positive effect on outgrowth on the fibroblast layer, whereas only the combination treatments resulted in increased growth on the astrocytes (A, B). DFO was the only treatment resulting in increased growth of neurites on the clusters. Corrected for the cluster diameter, DFO showed an increase in the maximal length (C) and the sum length of the neurites (D). Representative pictures of clusters with neurites (E-H) that were traced using the Neuron J plugin of Image J (e-h, clusters marked by dotted black lines). Statistics: one way Anova with Dunnett’s post-hoc test * p < 0.05, ** p < 0.01, *** p< 0.001 Scale bar = 100 μm.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4519270&req=5

pone.0134371.g007: Effects of treatments on neurite outgrowth.Neurite outgrowth from neonatal cortical neurons on the fibroblast and astrocyte cell layers as well as on the clusters in the co-cultures after treatment. cAMP and the combination treatments had a positive effect on outgrowth on the fibroblast layer, whereas only the combination treatments resulted in increased growth on the astrocytes (A, B). DFO was the only treatment resulting in increased growth of neurites on the clusters. Corrected for the cluster diameter, DFO showed an increase in the maximal length (C) and the sum length of the neurites (D). Representative pictures of clusters with neurites (E-H) that were traced using the Neuron J plugin of Image J (e-h, clusters marked by dotted black lines). Statistics: one way Anova with Dunnett’s post-hoc test * p < 0.05, ** p < 0.01, *** p< 0.001 Scale bar = 100 μm.
Mentions: In order to study putative axon growth-permitting properties of the scar-reducing treatments, we quantified neurite outgrowth of neonatal cortical neurons plated onto the co-cultures (Fig 7). On the astrocyte layer, only the combination of iron chelators with cAMP significantly increased the average neurite length per neuron. On the fibroblast layer, cAMP alone or in combination with iron chelators increased neurite length significantly. Only a non-significant trend towards longer neurites on fibroblasts was observed upon DFO-treatment.

Bottom Line: DFO could be identified as a putative anti-scarring treatment for CNS trauma.We subsequently validated this by local application of DFO to a dorsal hemisection in the rat thoracic spinal cord.DFO treatment led to significant reduction of scarring, slightly increased regeneration of corticospinal tract as well as ascending CGRP-positive axons and moderately improved locomotion.

View Article: PubMed Central - PubMed

Affiliation: Molecular Neurobiology Laboratory, Department of Neurology, Heinrich-Heine-University of Duesseldorf, Duesseldorf, Germany; Institute of Microanatomy and Neurobiology, Johannes Gutenberg-University Mainz, Mainz, Germany.

ABSTRACT
Lesion-induced scarring is a major impediment for regeneration of injured axons in the central nervous system (CNS). The collagen-rich glial-fibrous scar contains numerous axon growth inhibitory factors forming a regeneration-barrier for axons. We demonstrated previously that the combination of the iron chelator 2,2'-bipyridine-5,5'-decarboxylic acid (BPY-DCA) and 8-Br-cyclic AMP (cAMP) inhibits scar formation and collagen deposition, leading to enhanced axon regeneration and partial functional recovery after spinal cord injury. While BPY-DCA is not a clinical drug, the clinically approved iron chelator deferoxamine mesylate (DFO) may be a suitable alternative for anti-scarring treatment (AST). In order to prove the scar-suppressing efficacy of DFO we modified a recently published in vitro model for CNS scarring. The model comprises a co-culture system of cerebral astrocytes and meningeal fibroblasts, which form scar-like clusters when stimulated with transforming growth factor-β (TGF-β). We studied the mechanisms of TGF-β-induced CNS scarring and compared the efficiency of different putative pharmacological scar-reducing treatments, including BPY-DCA, DFO and cAMP as well as combinations thereof. We observed modulation of TGF-β-induced scarring at the level of fibroblast proliferation and contraction as well as specific changes in the expression of extracellular matrix molecules and axon growth inhibitory proteins. The individual and combinatorial pharmacological treatments had distinct effects on the cellular and molecular aspects of in vitro scarring. DFO could be identified as a putative anti-scarring treatment for CNS trauma. We subsequently validated this by local application of DFO to a dorsal hemisection in the rat thoracic spinal cord. DFO treatment led to significant reduction of scarring, slightly increased regeneration of corticospinal tract as well as ascending CGRP-positive axons and moderately improved locomotion. We conclude that the in vitro model for CNS scarring is suitable for efficient pre-screening and identification of putative scar-suppressing agents prior to in vivo application and validation, thus saving costs, time and laboratory animals.

No MeSH data available.


Related in: MedlinePlus