Chronic erythropoietin-mediated effects on the expression of astrocyte markers in a rat model of contusive spinal cord injury.
Using a standardized rat model of contusive spinal cord injury (SCI; [Gorio A, Gokmen N, Erbayraktar S, Yilmaz O, Madaschi L, Cichetti C, Di Giulio AM, Vardar E, Cerami A, Brines M (2002) Recombinant human erythropoietin counteracts secondary injury and markedly enhances neurological recovery from experimental spinal cord trauma. Proc Natl Acad Sci U S A 99:9450-9455]), we previously showed that the administration of recombinant human erythropoietin (rhEPO) improves both tissue sparing and locomotory outcome. In the present study, to better understand rhEPO-mediated effects on chronic astrocyte response to SCI in rat, we have used immunocytochemical methods combined with confocal and electron microscopy to investigate, 1 month after injury, the effects of a single rhEPO administration on the expression of a) aquaporin 4 (AQP4), the main astrocytic water channel implicated in edema development and resolution, and two molecules (dystrophin and syntrophin) involved in its membrane anchoring; b) glial fibrillary acidic protein (GFAP) and vimentin as markers of astrogliosis; c) chondroitin sulfate proteoglycans of the extracellular matrix which are upregulated after SCI and can inhibit axonal regeneration and influence neuronal and glial properties. Our results show that rhEPO administration after SCI modifies astrocytic response to injury by increasing AQP4 immunoreactivity in the spinal cord, but not in the brain, without apparent modifications of dystrophin and syntrophin distribution. Attenuation of astrogliosis, demonstrated by the semiquantitative analysis of GFAP labeling, was associated with a reduction of phosphacan/RPTP zeta/beta, whereas the levels of lecticans remained unchanged. Finally, the relative volume of a microvessel fraction was significantly increased, indicating a pro-angiogenetic or a vasodilatory effect of rhEPO. These changes were consistently associated with remarkable reduction of lesion size and with improvement in tissue preservation and locomotor recovery, confirming previous observations and underscoring the potentiality of rhEPO for the therapeutic management of SCI.
Dipartimento di Scienze Biomolecolari e Biotecnologie, Università degli Studi di Milano, Via Celoria, 26 I-20133 Milano, Italy. laura.vitellaro@unimi.it
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Vitellaro-Zuccarello
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Mazzetti
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Madaschi
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Bosisio
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Fontana
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Gorio
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De Biasi
D000818:Animals; D051401:Aquaporin 4 / Q000378:metabolism; D001253:Astrocytes / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D003288:Contusions / Q000378:metabolism* / Q000473:pathology; D016189:Dystrophin / Q000378:metabolism; D004921:Erythropoietin / Q000494:pharmacology*; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D005911:Gliosis / Q000188:drug therapy / Q000473:pathology; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D007202:Indicators and Reagents; D008297:Male; D018613:Microscopy, Confocal; D016253:Microscopy, Immunoelectron; D009043:Motor Activity / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins; D013119:Spinal Cord Injuries / Q000378:metabolism* / Q000473:pathology; D014746:Vimentin / Q000378:metabolism
D051401:Aquaporin 4; D016189:Dystrophin; D005904:Glial Fibrillary Acidic Protein; D007202:Indicators and Reagents; D011994:Recombinant Proteins; D014746:Vimentin; D004921:Erythropoietin