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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

Quantification of scar-reduction.Quantification of (A) the number of clusters, (B) the cluster area and (C) the cluster size after 7 days of TGF-β1 and potential scar-reducing treatments. DFO reduced cluster size, number and area. cAMP reduced the number of clusters but not their size. DFO + cAMP reduced the number of clusters significantly more than DFO or cAMP alone. (D) Effects of scar-reducing treatments on fibroblast proliferation. cAMP reduced the proliferation of the fibroblasts. Statistics: one way Anova with Bonferroni post-hoc test * p < 0.05, ** p < 0.01, *** p< 0.001.
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pone.0134371.g005: Quantification of scar-reduction.Quantification of (A) the number of clusters, (B) the cluster area and (C) the cluster size after 7 days of TGF-β1 and potential scar-reducing treatments. DFO reduced cluster size, number and area. cAMP reduced the number of clusters but not their size. DFO + cAMP reduced the number of clusters significantly more than DFO or cAMP alone. (D) Effects of scar-reducing treatments on fibroblast proliferation. cAMP reduced the proliferation of the fibroblasts. Statistics: one way Anova with Bonferroni post-hoc test * p < 0.05, ** p < 0.01, *** p< 0.001.

Mentions: Co- cultures treated with the H2O vehicle control contained on average 38 clusters of about 0.03 mm2 in size (Fig 5). Treatment of the co-cultures with DFO led to a reduction in the number of clusters by 50% (Fig 5A). The effects of the DFO treatment were quite variable, since in some cases there were around 10 regular-sized clusters and in other cases there were 40 very small clusters. Therefore, we also measured the total area the clusters occupied and the average cluster size using Image J software. Both the cluster size and area were significantly reduced for DFO (71% and 83% respectively, Fig 5B and 5C). BPY-DCA alone affected neither cluster number nor size. cAMP led to a 59% reduction in cluster area which was further reduced (79%) in co-cultures treated with DFO + cAMP. This combination was also significantly different from the single treatments, indicating that DFO and cAMP had additional effects. The combination of BPY-DCA + cAMP showed a significant reduction in cluster number and cluster area compared to the H2O control but had no additional effect compared to cAMP alone.


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)

Quantification of scar-reduction.Quantification of (A) the number of clusters, (B) the cluster area and (C) the cluster size after 7 days of TGF-β1 and potential scar-reducing treatments. DFO reduced cluster size, number and area. cAMP reduced the number of clusters but not their size. DFO + cAMP reduced the number of clusters significantly more than DFO or cAMP alone. (D) Effects of scar-reducing treatments on fibroblast proliferation. cAMP reduced the proliferation of the fibroblasts. Statistics: one way Anova with Bonferroni post-hoc test * p < 0.05, ** p < 0.01, *** p< 0.001.
© Copyright Policy
Related In: Results  -  Collection

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

pone.0134371.g005: Quantification of scar-reduction.Quantification of (A) the number of clusters, (B) the cluster area and (C) the cluster size after 7 days of TGF-β1 and potential scar-reducing treatments. DFO reduced cluster size, number and area. cAMP reduced the number of clusters but not their size. DFO + cAMP reduced the number of clusters significantly more than DFO or cAMP alone. (D) Effects of scar-reducing treatments on fibroblast proliferation. cAMP reduced the proliferation of the fibroblasts. Statistics: one way Anova with Bonferroni post-hoc test * p < 0.05, ** p < 0.01, *** p< 0.001.
Mentions: Co- cultures treated with the H2O vehicle control contained on average 38 clusters of about 0.03 mm2 in size (Fig 5). Treatment of the co-cultures with DFO led to a reduction in the number of clusters by 50% (Fig 5A). The effects of the DFO treatment were quite variable, since in some cases there were around 10 regular-sized clusters and in other cases there were 40 very small clusters. Therefore, we also measured the total area the clusters occupied and the average cluster size using Image J software. Both the cluster size and area were significantly reduced for DFO (71% and 83% respectively, Fig 5B and 5C). BPY-DCA alone affected neither cluster number nor size. cAMP led to a 59% reduction in cluster area which was further reduced (79%) in co-cultures treated with DFO + cAMP. This combination was also significantly different from the single treatments, indicating that DFO and cAMP had additional effects. The combination of BPY-DCA + cAMP showed a significant reduction in cluster number and cluster area compared to the H2O control but had no additional effect compared to cAMP alone.

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