Barton Samantha K, Melville Jacqueline M, Tolcos Mary, Polglase Graeme R, McDougall Annie R A, Azhan Aminath, Crossley Kelly J, Jenkin Graham, Moss Timothy J M
The Ritchie Centre, MIMR-PHI Institute of Medical Research, Clayton, Vic., Australia.
Dev Neurosci. 2015;37(4-5):338-48. doi: 10.1159/000371415. Epub 2015 Feb 25.
Preterm infants can be inadvertently exposed to high tidal volumes (VT) during resuscitation in the delivery room due to limitations of available equipment. High VT ventilation of preterm lambs produces cerebral white matter (WM) pathology similar to that observed in preterm infants who develop cerebral palsy. We hypothesized that human amnion epithelial cells (hAECs), which have anti-inflammatory and regenerative properties, would reduce ventilation-induced WM pathology in neonatal late preterm lamb brains.
Two groups of lambs (0.85 gestation) were used, as follows: (1) ventilated lambs (Vent; n = 8) were ventilated using a protocol that induces injury (VT targeting 15 ml/kg for 15 min, with no positive end-expiratory pressure) and were then maintained for another 105 min, and (2) ventilated + hAECs lambs (Vent+hAECs; n = 7) were similarly ventilated but received intravenous and intratracheal administration of 9 × 10(7) hAECs (18 × 10(7) hAECs total) at birth. Oxygenation and ventilation parameters were monitored in real time; cerebral oxygenation was measured using near-infrared spectroscopy. qPCR (quantitative real-time PCR) and immunohistochemistry were used to assess inflammation, vascular leakage and astrogliosis in both the periventricular and subcortical WM of the frontal and parietal lobes. An unventilated control group (UVC; n = 5) was also used for qPCR analysis of gene expression. Two-way repeated measures ANOVA was used to compare physiological data. Student's t test and one-way ANOVA were used for immunohistological and qPCR data comparisons, respectively.
Respiratory parameters were not different between groups. Interleukin (IL)-6 mRNA levels in subcortical WM were lower in the Vent+hAECs group than the Vent group (p = 0.028). IL-1β and IL-6 mRNA levels in periventricular WM were higher in the Vent+hAECs group than the Vent group (p = 0.007 and p = 0.001, respectively). The density of Iba-1-positive microglia was lower in the subcortical WM of the parietal lobes (p = 0.010) in the Vent+hAECs group but not in the periventricular WM. The number of vessels in the WM of the parietal lobe exhibiting protein extravasation was lower (p = 0.046) in the Vent+hAECs group. Claudin-1 mRNA levels were higher in the periventricular WM (p = 0.005). The density of GFAP-positive astrocytes was not different between groups.
Administration of hAECs at the time of birth alters the effects of injurious ventilation on the preterm neonatal brain. Further studies are required to understand the regional differences in the effects of hAECs on ventilation-induced WM pathology and their net effect on the developing brain.
由于现有设备的限制，早产儿在产房复苏过程中可能会无意中暴露于高潮气量（VT）。早产羔羊的高潮气量通气会产生与患脑瘫的早产儿中观察到的类似的脑白质（WM）病变。我们假设具有抗炎和再生特性的人羊膜上皮细胞（hAECs）会减少新生晚期早产羔羊大脑中通气诱导的WM病变。
使用两组羔羊（妊娠0.85），如下：（1）通气羔羊（Vent；n = 8）采用诱导损伤的方案进行通气（VT目标为15 ml/kg，持续15分钟，无呼气末正压），然后再维持105分钟，以及（2）通气+ hAECs羔羊（Vent+hAECs；n = 7）同样进行通气，但在出生时接受静脉和气管内注射9×10⁷ hAECs（共18×10⁷ hAECs）。实时监测氧合和通气参数；使用近红外光谱法测量脑氧合。qPCR（定量实时PCR）和免疫组织化学用于评估额叶和顶叶脑室周围和皮质下WM中的炎症、血管渗漏和星形胶质细胞增生。还使用未通气的对照组（UVC；n = 5）进行基因表达的qPCR分析。采用双向重复测量方差分析比较生理数据。分别使用学生t检验和单向方差分析进行免疫组织学和qPCR数据比较。
两组之间的呼吸参数无差异。Vent+hAECs组皮质下WM中的白细胞介素（IL）-6 mRNA水平低于Vent组（p = 0.028）。Vent+hAECs组脑室周围WM中的IL-1β和IL-6 mRNA水平高于Vent组（分别为p = 0.007和p = 0.001）。Vent+hAECs组顶叶皮质下WM中Iba-1阳性小胶质细胞的密度较低（p = 0.010），但在脑室周围WM中并非如此。Vent+hAECs组顶叶WM中出现蛋白外渗的血管数量较少（p = 0.046）。Claudin-1 mRNA水平在脑室周围WM中较高（p = 0.005）。两组之间GFAP阳性星形胶质细胞的密度无差异。
出生时给予hAECs会改变有害通气对早产新生儿大脑的影响。需要进一步研究以了解hAECs对通气诱导的WM病变影响的区域差异及其对发育中大脑的净效应。
