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RGD domains neuroprotect the immature brain by a glial-dependent mechanism

Authors

  • Hugo Peluffo PhD,

    Corresponding author
    1. Medical Histology, Department of Cell Biology, Physiology and Immunology, Institute of Neurosciences, Autonomous University of Barcelona
    • Medical Histology, Torre M5, Department of Cell Biology, Physiology and Anatomy and Neuroscience Institute, Autonomous University of Barcelona, Bellaterra 08193, Spain
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    • H.P. and P.G. contributed equally to this work.

  • Pau González MS,

    1. Medical Histology, Department of Cell Biology, Physiology and Immunology, Institute of Neurosciences, Autonomous University of Barcelona
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    • H.P. and P.G. contributed equally to this work.

  • Anna Arís PhD,

    1. Institute of Biotechnology and Biomedicine, Department of Genetics and Microbiology, CIBER Area of Biomaterials, Bioengineering and Biomedicine, Autonomous University of Barcelona, Bellaterra
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  • Laia Acarin PhD,

    Corresponding author
    1. Medical Histology, Department of Cell Biology, Physiology and Immunology, Institute of Neurosciences, Autonomous University of Barcelona
    • Medical Histology, Torre M5, Department of Cell Biology, Physiology and Anatomy and Neuroscience Institute, Autonomous University of Barcelona, Bellaterra 08193, Spain
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  • Josep Saura PhD,

    1. Department of Pharmacology and Toxicology Institut d'Investigaciones Biomèdiques de Barcelona (IIBB), Institut d'Investigaciones Biomèdiques August Pi i Sunyer (IDIBAPS), Consejo Superior de Investigaciones Cientificas (CSIC), Barcelona, Spain
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  • Antoni Villaverde PhD,

    1. Institute of Biotechnology and Biomedicine, Department of Genetics and Microbiology, CIBER Area of Biomaterials, Bioengineering and Biomedicine, Autonomous University of Barcelona, Bellaterra
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  • Bernardo Castellano PhD,

    1. Medical Histology, Department of Cell Biology, Physiology and Immunology, Institute of Neurosciences, Autonomous University of Barcelona
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  • Berta González PhD

    1. Medical Histology, Department of Cell Biology, Physiology and Immunology, Institute of Neurosciences, Autonomous University of Barcelona
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Abstract

Objective

Integrin binding to extracellular matrix ligands, including those presenting RGD motifs, modulate diverse cellular processes. In the brain, many endogenous RGD-containing molecules are induced after damage. Previously, the gene therapy vector termed NLSCt, which displays an RGD motif, was shown to neuroprotect after immature brain excitotoxicity. We analyze whether neuroprotection is mediated by the RGD motif.

Methods

RGD-containing synthetic peptide GPenGRGDSPCA (GPen) was injected 2 hours after N-methyl-D-aspartate–mediated excitotoxicity to the postnatal day 9 rat brain. Damage and glial/inflammatory response were evaluated 3 days later. In addition, the neuroprotective effect of GPen and NLSCt after N-methyl-D-aspartate–induced cell death was also analyzed in vitro using neuron-purified and mixed neuron-glia primary cultures. To further characterize whether the neuroprotective effect was mediated by glial-derived soluble factors, we also tested the protective ability of conditioned media from RGD-treated microglia, astrocyte, or mixed glia cultures.

Results

Animals treated with GPen peptide showed functional improvement, a significant reduction in lesion volume up to 28%, and a decrease in the number of degenerating neurons. In addition, N-methyl-D-aspartate–injected animals treated with both RGD-containing molecules at the neuroprotective doses showed a significant increase in microglial reactivity and microglia/macrophage cell number, but no differences in neutrophil infiltration and the astroglial response. Finally, in vitro studies showed that the neuroprotective effect was observed in mixed neuron-glia, but not in neuron-purified cultures. Conditioned media from RGD-treated microglial, astroglial, and mixed-glial cultures were not protective.

Interpretation

These results suggest that RGD-containing molecules neuroprotect by a glial-dependent mechanism. Ann Neurol 2007

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