Vitellaro-Zuccarello L, Mazzetti S, Madaschi L, Bosisio P, Fontana E, Gorio A, De Biasi S
Dipartimento di Scienze Biomolecolari e Biotecnologie, Università degli Studi di Milano, Via Celoria, 26 I-20133 Milano, Italy.
Neuroscience. 2008 Jan 24;151(2):452-66. doi: 10.1016/j.neuroscience.2007.11.004. Epub 2007 Nov 7.
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.
利用一种标准化的大鼠脊髓挫伤性损伤（SCI；[Gorio A, Gokmen N, Erbayraktar S, Yilmaz O, Madaschi L, Cichetti C, Di Giulio AM, Vardar E, Cerami A, Brines M (2002) 重组人促红细胞生成素可对抗继发性损伤并显著促进实验性脊髓创伤后的神经功能恢复。美国国家科学院院刊99:9450 - 9455]）模型，我们之前表明，给予重组人促红细胞生成素（rhEPO）可改善组织保留和运动功能结局。在本研究中，为了更好地理解rhEPO对大鼠脊髓损伤后慢性星形胶质细胞反应的介导作用，我们采用免疫细胞化学方法结合共聚焦显微镜和电子显微镜，在损伤后1个月研究单次给予rhEPO对以下方面表达的影响：a）水通道蛋白4（AQP4），这是参与水肿形成和消退的主要星形胶质细胞水通道，以及参与其膜锚定的两种分子（肌营养不良蛋白和肌营养不良蛋白相关蛋白）；b）胶质纤维酸性蛋白（GFAP）和波形蛋白作为星形胶质细胞增生的标志物；c）细胞外基质硫酸软骨素蛋白聚糖，其在脊髓损伤后上调，可抑制轴突再生并影响神经元和胶质细胞特性。我们的结果表明，脊髓损伤后给予rhEPO可通过增加脊髓而非大脑中的AQP4免疫反应性来改变星形胶质细胞对损伤的反应，而肌营养不良蛋白和肌营养不良蛋白相关蛋白的分布无明显改变。通过对GFAP标记的半定量分析证明的星形胶质细胞增生的减轻与磷酸软骨素/受体蛋白酪氨酸磷酸酶ζ/β的减少相关，而凝集素水平保持不变。最后，微血管部分的相对体积显著增加，表明rhEPO具有促血管生成或血管舒张作用。这些变化始终与损伤大小的显著减小以及组织保存和运动功能恢复的改善相关，证实了先前的观察结果，并强调了rhEPO在脊髓损伤治疗管理中的潜力。
