Zhang Chenyang, Harder David R
Cardiovascular Research Center, Department of Physiology, Medical College of Wisconsin, Milwaukee, USA.
Stroke. 2002 Dec;33(12):2957-64. doi: 10.1161/01.str.0000037787.07479.9a.
Background and Purpose- Epoxyeicosatrienoic acids (EETs) are products of cytochrome P450 epoxygenation of arachidonic acid. We have previously demonstrated that astrocyte-conditioned medium induced mitogenesis in brain capillary endothelial cells. The goals of the present studies are to further define the mechanism through which this can occur and to confirm that EETs are derived from astrocytes, through which astrocytic activity can regulate cerebral angiogenesis in response to neuronal activation.
Astrocytes and cerebral capillary endothelial cells in primary cultures were cocultured to examine the interaction of the 2 cell types. We used multiple immunohistochemical techniques to characterize the multicellular nature of the capillaries, which is not simply an artifact related to the culture conditions. The mitogenic effect of EETs was determined by (3)H-thymidine incorporation and cell proliferation assay. Endothelial tube formation was examined in vitro and in vivo with the use of a reconstituted basement membrane (Matrigel) assay.
In cocultures of astrocytes and capillary endothelium, we observed morphological changes in both cell types such that each assumed certain physiological characteristics, ie, endothelial networks and astrocytes with "footlike" projections as well as intermittent gap junctions forming within the endothelial cells. EETs from astrocytes as well as synthetic EETs promoted mitogenesis of endothelial cells, a process sensitive to inhibition of tyrosine kinase with genistein. Treatments with exogenous EETs were sufficient for endothelial cells to differentiate into capillary-like structures in culture as well as in vivo in a Matrigel matrix.
The 2 major conclusions from these data are that astrocytes may play an important role in regulating angiogenesis in the brain and that cytochrome P450-derived EETs from astrocytes are mitogenic and angiogenic.
背景与目的——环氧二十碳三烯酸（EETs）是细胞色素P450对花生四烯酸进行环氧化作用的产物。我们之前已经证明，星形胶质细胞条件培养基可诱导脑毛细血管内皮细胞发生有丝分裂。本研究的目的是进一步明确其发生机制，并证实EETs来源于星形胶质细胞，通过这一途径，星形胶质细胞的活性可响应神经元激活来调节脑内血管生成。
将原代培养的星形胶质细胞和脑毛细血管内皮细胞共培养，以检测这两种细胞类型之间的相互作用。我们使用多种免疫组织化学技术来表征毛细血管的多细胞性质，这并非仅仅是与培养条件相关的假象。通过³H-胸腺嘧啶核苷掺入法和细胞增殖试验来确定EETs的促有丝分裂作用。利用重组基底膜（基质胶）试验在体外和体内检测内皮管形成情况。
在星形胶质细胞与毛细血管内皮细胞的共培养中，我们观察到两种细胞类型均出现形态变化，各自呈现出某些生理特征，即内皮网络和具有“足样”突起的星形胶质细胞，以及内皮细胞内形成的间歇性缝隙连接。来自星形胶质细胞以及合成的EETs均可促进内皮细胞的有丝分裂，这一过程对染料木黄酮抑制酪氨酸激酶敏感。用外源性EETs处理足以使内皮细胞在培养物中以及在基质胶基质的体内环境中分化为毛细血管样结构。
这些数据得出的两个主要结论是，星形胶质细胞可能在调节脑内血管生成中发挥重要作用，并且来自星形胶质细胞的细胞色素P450衍生的EETs具有促有丝分裂和血管生成作用。
