McDougall Annie R A, Hale Nadia, Rees Sandra, Harding Richard, De Matteo Robert, Hooper Stuart B, Tolcos Mary
The Ritchie Centre, Hudson Institute of Medical ResearchClayton, VIC, Australia.
Department of Obstetrics and Gynaecology, Monash UniversityClayton, VIC, Australia.
Front Cell Neurosci. 2017 Jul 28;11:224. doi: 10.3389/fncel.2017.00224. eCollection 2017.
Erythropoietin (EPO) ameliorates inflammation-induced injury in cerebral white matter (WM). However, effects of inflammation on the cerebellum and neuroprotective effects of EPO are unknown. Our aims were to determine: (i) whether lipopolysaccharide (LPS)-induced intrauterine inflammation causes injury to, and/or impairs development of the cerebellum; and (ii) whether recombinant human EPO (rhEPO) mitigates these changes. At 107 ± 1 days gestational age (DGA; 0.7 of term), fetal sheep received LPS (0.9 μg/kg; i.v.) or an equivalent volume of saline, followed 1 h later with 5000 IU/kg rhEPO (i.v.) or an equivalent volume of saline (i.v.). This generated the following experimental groups: control (saline + saline; = 6), LPS (LPS + saline, = 8) and LPS + rhEPO ( = 8). At necropsy (116 ± 1 DGA; ~0.8 of term) the brain was perfusion-fixed and stained histologically (H&E) and immunostained to identify granule cells (Neuronal Nuclei, NeuN), granule cell proliferation (Ki67), Bergmann glia (glial fibrillary acidic protein, GFAP), astrogliosis (GFAP) and microgliosis (Iba-1). In comparison to controls, LPS fetuses had an increased density of Iba-1-positive microglia ( < 0.005) in the lobular WM; rhEPO prevented this increase ( < 0.05). The thickness of both the proliferative (Ki67-positive) and post-mitotic zones (Ki67-negative) of the EGL were increased in LPS-exposed fetuses compared to controls ( < 0.05), but were not different between controls and LPS + rhEPO fetuses. LPS also increased ( < 0.001) the density of granule cells (NeuN-positive) in the internal granule layer (IGL); rhEPO prevented the increase ( < 0.01). There was no difference between groups in the areas of the vermis (total cross-section), molecular layer (ML), IGL or WM, the density of NeuN-positive granule cells in the ML, the linear density of Bergmann glial fibers, the areal density or somal area of the Purkinje cells, the areal coverage of GFAP-positive astrocytes in the lobular and deep WM, the density of Iba-1-positive microglia in the deep WM or the density of apopotic cells in the cerebellum. LPS-induced intrauterine inflammation caused microgliosis and abnormal development of granule cells. rhEPO ameliorated these changes, suggesting that it is neuroprotective against LPS-induced inflammatory effects in the cerebellum.
促红细胞生成素（EPO）可改善脑白质（WM）炎症诱导的损伤。然而，炎症对小脑的影响以及EPO的神经保护作用尚不清楚。我们的目的是确定：（i）脂多糖（LPS）诱导的宫内炎症是否会导致小脑损伤和/或损害小脑发育；（ii）重组人促红细胞生成素（rhEPO）是否能减轻这些变化。在妊娠107±1天胎龄（DGA；约足月的0.7）时，给胎羊静脉注射LPS（约0.9μg/kg）或等体积生理盐水，1小时后再静脉注射5000IU/kg rhEPO或等体积生理盐水。这产生了以下实验组：对照组（生理盐水+生理盐水；n = 6）、LPS组（LPS+生理盐水，n = 8）和LPS+rhEPO组（n = 8）。尸检时（116±1 DGA；约足月的0.8），对大脑进行灌注固定，进行组织学（苏木精-伊红染色，H&E）染色和免疫染色，以识别颗粒细胞（神经元核，NeuN）、颗粒细胞增殖（Ki67）细胞、伯格曼胶质细胞（胶质纤维酸性蛋白，GFAP）、星形胶质细胞增生（GFAP）和小胶质细胞增生（离子钙结合衔接分子1，Iba-1）。与对照组相比，LPS组胎羊小叶白质中Iba-1阳性小胶质细胞密度增加（P<0.005）；rhEPO可防止这种增加（P<0.05）。与对照组相比，暴露于LPS的胎羊的外颗粒层（EGL）增殖区（Ki67阳性）和有丝分裂后区（Ki67阴性）的厚度均增加（P<0.05），但对照组和LPS+rhEPO组胎羊之间无差异。LPS还增加了（P<0.001）内颗粒层（IGL）中颗粒细胞（NeuN阳性）的密度；rhEPO可防止这种增加（P<0.01）。各组在蚓部（总横截面积）、分子层（ML）、IGL或WM的面积、ML中NeuN阳性颗粒细胞的密度、伯格曼胶质纤维的线性密度、浦肯野细胞的面积密度或体细胞面积、小叶和深部WM中GFAP阳性星形胶质细胞的面积覆盖率、深部WM中Iba-1阳性小胶质细胞的密度或小脑中凋亡细胞的密度方面均无差异。LPS诱导的宫内炎症导致小胶质细胞增生和颗粒细胞异常发育。rhEPO可改善这些变化，表明其对LPS诱导的小脑炎症具有神经保护作用。
