Cellular basis of PVL in autopsied brains

该项目的一个关键概念是，脑室周围白质软化(PVL)是由于缺血/再灌注对围产期白质中脆弱的发育中的少突胶质细胞触发自由基损伤：结果是发育中的少突胶质细胞死亡，部分是通过凋亡，从而导致成熟的少突胶质细胞减少和髓鞘形成受损。这项研究的总体假设有两个方面：1)在PVL的高危期(24-32周)，大脑白质中发育的少突胶质细胞对氧化应激具有特定的脆弱性，这至少部分与抗氧化酶的表达发展和促氧化途径的发展之间的不匹配有关，例如获得少突胶质细胞分化所需的铁；以及2)氧化应激在PVL的发病机制中发挥重要作用。在基线研究中，我们将使用免疫细胞化学方法和阶段特异性少突胶质细胞的标志物来确定人脑白质中少突胶质细胞的时空成熟。我们还将使用组织化学、免疫细胞化学和免疫印迹来确定人类脑白质中选定的抗氧化酶和促氧化途径的标记物(即铁、一氧化氮合酶定位和12-脂氧合酶表达)的发育概况。在这里，我们将检验一种假设，即在PVL最危险的时期，抗氧化酶在未成熟的白质中低水平表达，并且促氧化途径的标志物的出现先于抗氧化酶的表达，从而表明潜在的自由基来源与清除它们的系统之间的发育不匹配。在PVL本身，我们将确定发育中的少突胶质细胞的密度，这表明这些细胞的靶向性死亡和丢失。我们还将确定氧化应激标志物是否在PVL中表达。最后，我们将确定选定的细胞因子在PVL中的参与。这项拟议的研究将直接为人类大脑内PVL中发育的少突胶质细胞的作用和与氧化应激相关的因素提供重要的见解。

A key concept of the program project is that periventricular leukomalacia (PVL) results from ischemia/reperfusion that triggers free radical injury to vulnerable developing oligodendrocytes in perinatal white matter: as a result, developing oligodendrocytes die, in part by apoptosis, thus resulting in decreased mature oligodendrocytes and impaired myelination. The overall hypothesis of the proposed study is two-fold: 1) there is a specific vulnerability of developing oligodendrocytes to oxidative stress in the cerebral white matter in the period of greatest risk for PVL (24-32 weeks) which relates, at least in part, to a mismatch between the development of the expression of antioxidant enzymes and the development of pro-oxidant pathways, such as the acquisition of iron necessary for oligodendrocyte differentiation; and 2) oxidative stress plays a major role in the pathogenesis of PVL. In baseline studies, we will determine the temporospatial maturation of oligodendrocytes in human cerebral white matter using immunocytochemical methods and markers to stage-specific oligodendrocytes. We will also determine the developmental profile of selected antioxidant enzymes and markers of pro-oxidant pathways (i.e., iron, nitric oxide synthase localization, and 12-lipoxygenase expression) in human cerebral white matter using histochemistry, immunocytochemistry, and immunoblotting. Here we will test the hypothesis that antioxidant enzymes are expressed at low levels in immature white matter during the period of greatest risk for PVL, and that the appearance of markers of pro-oxidant pathways precede the expression of antioxidant enzymes, thus denoting a developmental mismatch between potential sources of free radicals and the systems to clear them. In PVL itself, we will determine the densities of developing oligodendrocytes, indicative of a targeted death and loss of these cells. We will also determine if markers of oxidative stress are expressed in PVL. Finally, we will determine the involvement of selected cytokines in PVL. The proposed studies should provide important insight into the roles of developing oligodendrocytes and factors related to oxidative stress in PVL directly within the human brain.