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HB-GAM (pleiotrophin) reverses inhibition of neural regeneration by the CNS extracellular matrix

View Article: PubMed Central - PubMed

ABSTRACT

Chondroitin sulfate (CS) glycosaminoglycans inhibit regeneration in the adult central nervous system (CNS). We report here that HB-GAM (heparin-binding growth-associated molecule; also known as pleiotrophin), a CS-binding protein expressed at high levels in the developing CNS, reverses the role of the CS chains in neurite growth of CNS neurons in vitro from inhibition to activation. The CS-bound HB-GAM promotes neurite growth through binding to the cell surface proteoglycan glypican-2; furthermore, HB-GAM abrogates the CS ligand binding to the inhibitory receptor PTPσ (protein tyrosine phosphatase sigma). Our in vivo studies using two-photon imaging of CNS injuries support the in vitro studies and show that HB-GAM increases dendrite regeneration in the adult cerebral cortex and axonal regeneration in the adult spinal cord. Our findings may enable the development of novel therapies for CNS injuries.

No MeSH data available.


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HB-GAM overcomes the CSPG-dependent inhibition of neurite growth in PC12 cells.HB-GAM overcomes the inhibitory effect of neurocan and brain-derived CSPGs on neurite growth. (a) PC12 cells plated on unprecoated culture plastic, on the culture plastic precoated with aggrecan, with aggrecan + HB-GAM or with aggrecan + IgG. Immunoglobulin G (IgG) was used as an unspecific control in precoating. All samples were treated with NGF (100 ng/ml). (b) Neurite length measured in live cell images obtained 1 day after plating. (c–f) Cortical neurons cultured on different substrates in the absence or presence of HB-GAM. Neurons were cultured on unprecoated plastic, on neurocan (5 μg/ml) or on neurocan (5 μg/ml) + HB-GAM (25 μg/ml). (g–i) Hippocampal neurons from E17 rats were cultured on the substrate precoated with brain-derived CSPGs (50 μg/ml) or with brain-derived CSPGs (50 μg/ml) + HB-GAM (10 μg/ml) or on the substrate without precoating (uncoated in panel i). Live cells were imaged after 36 h in culture. The scale bar (20 μm) shown in a and c is valid for (a,c–e,g,h). One way ANOVA was used.
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f2: HB-GAM overcomes the CSPG-dependent inhibition of neurite growth in PC12 cells.HB-GAM overcomes the inhibitory effect of neurocan and brain-derived CSPGs on neurite growth. (a) PC12 cells plated on unprecoated culture plastic, on the culture plastic precoated with aggrecan, with aggrecan + HB-GAM or with aggrecan + IgG. Immunoglobulin G (IgG) was used as an unspecific control in precoating. All samples were treated with NGF (100 ng/ml). (b) Neurite length measured in live cell images obtained 1 day after plating. (c–f) Cortical neurons cultured on different substrates in the absence or presence of HB-GAM. Neurons were cultured on unprecoated plastic, on neurocan (5 μg/ml) or on neurocan (5 μg/ml) + HB-GAM (25 μg/ml). (g–i) Hippocampal neurons from E17 rats were cultured on the substrate precoated with brain-derived CSPGs (50 μg/ml) or with brain-derived CSPGs (50 μg/ml) + HB-GAM (10 μg/ml) or on the substrate without precoating (uncoated in panel i). Live cells were imaged after 36 h in culture. The scale bar (20 μm) shown in a and c is valid for (a,c–e,g,h). One way ANOVA was used.

Mentions: The effect of HB-GAM on the CSPG substrate is not restricted to primary CNS neurons. A similar effect was observed with PC12 cells cultured in the presence of nerve growth factor (NGF), which by itself was not able to promote neurite outgrowth on aggrecan (Fig. 2a,b). To examine whether the HB-GAM effect on the matrix side is restricted to the CSPG aggrecan, we tested neurocan (another major CSPG of the matrix)17 and a CSPG mixture prepared from brain in the same assays as aggrecan. HB-GAM was also found to overcome their inhibitory effects on neurite outgrowth (Fig. 2c–i). The HB-GAM effect on neurite outgrowth on the CSPG substrates may therefore depend on its interaction with the CS side chains of the CSPGs.


HB-GAM (pleiotrophin) reverses inhibition of neural regeneration by the CNS extracellular matrix
HB-GAM overcomes the CSPG-dependent inhibition of neurite growth in PC12 cells.HB-GAM overcomes the inhibitory effect of neurocan and brain-derived CSPGs on neurite growth. (a) PC12 cells plated on unprecoated culture plastic, on the culture plastic precoated with aggrecan, with aggrecan + HB-GAM or with aggrecan + IgG. Immunoglobulin G (IgG) was used as an unspecific control in precoating. All samples were treated with NGF (100 ng/ml). (b) Neurite length measured in live cell images obtained 1 day after plating. (c–f) Cortical neurons cultured on different substrates in the absence or presence of HB-GAM. Neurons were cultured on unprecoated plastic, on neurocan (5 μg/ml) or on neurocan (5 μg/ml) + HB-GAM (25 μg/ml). (g–i) Hippocampal neurons from E17 rats were cultured on the substrate precoated with brain-derived CSPGs (50 μg/ml) or with brain-derived CSPGs (50 μg/ml) + HB-GAM (10 μg/ml) or on the substrate without precoating (uncoated in panel i). Live cells were imaged after 36 h in culture. The scale bar (20 μm) shown in a and c is valid for (a,c–e,g,h). One way ANOVA was used.
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f2: HB-GAM overcomes the CSPG-dependent inhibition of neurite growth in PC12 cells.HB-GAM overcomes the inhibitory effect of neurocan and brain-derived CSPGs on neurite growth. (a) PC12 cells plated on unprecoated culture plastic, on the culture plastic precoated with aggrecan, with aggrecan + HB-GAM or with aggrecan + IgG. Immunoglobulin G (IgG) was used as an unspecific control in precoating. All samples were treated with NGF (100 ng/ml). (b) Neurite length measured in live cell images obtained 1 day after plating. (c–f) Cortical neurons cultured on different substrates in the absence or presence of HB-GAM. Neurons were cultured on unprecoated plastic, on neurocan (5 μg/ml) or on neurocan (5 μg/ml) + HB-GAM (25 μg/ml). (g–i) Hippocampal neurons from E17 rats were cultured on the substrate precoated with brain-derived CSPGs (50 μg/ml) or with brain-derived CSPGs (50 μg/ml) + HB-GAM (10 μg/ml) or on the substrate without precoating (uncoated in panel i). Live cells were imaged after 36 h in culture. The scale bar (20 μm) shown in a and c is valid for (a,c–e,g,h). One way ANOVA was used.
Mentions: The effect of HB-GAM on the CSPG substrate is not restricted to primary CNS neurons. A similar effect was observed with PC12 cells cultured in the presence of nerve growth factor (NGF), which by itself was not able to promote neurite outgrowth on aggrecan (Fig. 2a,b). To examine whether the HB-GAM effect on the matrix side is restricted to the CSPG aggrecan, we tested neurocan (another major CSPG of the matrix)17 and a CSPG mixture prepared from brain in the same assays as aggrecan. HB-GAM was also found to overcome their inhibitory effects on neurite outgrowth (Fig. 2c–i). The HB-GAM effect on neurite outgrowth on the CSPG substrates may therefore depend on its interaction with the CS side chains of the CSPGs.

View Article: PubMed Central - PubMed

ABSTRACT

Chondroitin sulfate (CS) glycosaminoglycans inhibit regeneration in the adult central nervous system (CNS). We report here that HB-GAM (heparin-binding growth-associated molecule; also known as pleiotrophin), a CS-binding protein expressed at high levels in the developing CNS, reverses the role of the CS chains in neurite growth of CNS neurons in vitro from inhibition to activation. The CS-bound HB-GAM promotes neurite growth through binding to the cell surface proteoglycan glypican-2; furthermore, HB-GAM abrogates the CS ligand binding to the inhibitory receptor PTPσ (protein tyrosine phosphatase sigma). Our in vivo studies using two-photon imaging of CNS injuries support the in vitro studies and show that HB-GAM increases dendrite regeneration in the adult cerebral cortex and axonal regeneration in the adult spinal cord. Our findings may enable the development of novel therapies for CNS injuries.

No MeSH data available.


Related in: MedlinePlus