FGF-2 induced by interleukin-1β through the action of phosphatidylinositol 3-kinase mediates endothelial mesenchymal transformation in corneal endothelial cells
FGF-2 induced by interleukin-1β through the action of phosphatidylinositol 3-kinase mediates endothelial mesenchymal transformation in corneal endothelial cells
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DOI：
10.1074/jbc.m405208200
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发表时间：
2004-07-30
影响因子：
4.8
通讯作者：
Kay, EDP
中科院分区：
文献类型：
--
作者：
Lee, HT;Lee, JG;Kay, EDP
我们以前的工作表明，无论是多形核白细胞（PMN）和从PMN释放的蛋白质组分诱导从头合成的成纤维细胞生长因子2（FGF-2），这反过来又成为直接介质的内皮间充质转化中观察到的角膜内皮细胞（CEC）。为了鉴定蛋白质因子，我们使用蛋白质芯片技术。从中性粒细胞释放的条件培养基中获得的蛋白质组分通过固定化pH梯度条带的二维电泳进行解析。大部分的蛋白质点，分子量为17 kDa，依次进行凝胶内胰蛋白酶消化和质谱分析。17-kDa肽带被鉴定为白细胞介素-1 β（IL-1 β）。进一步测定IL-1 β的生物活性; IL-1 β以时间和剂量依赖性方式将CEC的形状从多边形改变为成纤维细胞形态，而中和IL-1 β抗体、FGF-2中和抗体和LY 294002阻断IL-1 β的作用。IL-1 β以时间和剂量依赖性方式显著增加FGF-2 mRNA水平; IL-1 β刺激FGF-2所有亚型的表达。IL-1 β最初诱导FGF-2在细胞核内积聚，并促进FGF-2向质膜和细胞外基质的转运。IL-1 β激活磷脂酰肌醇（PI）3-激酶，其酶活性在暴露于IL-1 β 5分钟后大大刺激。PI 3-激酶的这种早期和快速激活大大增强了CEC中FGF-2的产生;用LY 294002预处理阻碍了IL-1 β的诱导活性。这些观察结果表明，IL-1 β通过PI 3-激酶的作用，通过其对FGF-2的诱导潜能，参与CEC的内皮向间充质转化。
Our previous work demonstrated that both polymorphonuclear leukocytes (PMNs) and protein fractions released from PMNs induced de novo synthesis of fibroblast growth factor 2 (FGF-2), which in turn becomes the direct mediator of endothelial mesenchymal transformation observed in corneal endothelial cells (CECs). To identify the protein factor, we used ProteinChip Array technology. Protein fractions obtained from the conditioned medium released by PMNs were resolved by two-dimensional electrophoresis with immobilized pH gradient strips. Most of the protein spots, with molecular masses of 17 kDa, were sequentially subjected to in-gel trypsin digestion and mass spectrometry. The 17-kDa peptide band was identified as interleukin-1beta (IL-1beta). Biological activities of IL-1beta were further determined; IL-1beta altered the shape of CECs from polygonal to fibroblastic morphologies in a time- and dose-dependent manner, whereas neutralizing IL-1beta antibody, neutralizing antibody to FGF-2, and LY294002 blocked the action of IL-1beta. IL-1beta greatly increased the levels of FGF-2 mRNA in a time- and dose-dependent manner; IL-1beta stimulated expression of all isoforms of FGF-2. IL-1beta initially induced nuclear accumulation of FGF-2 and facilitated translocation of FGF-2 to plasma membrane and extracellular matrix. IL-1beta activated phosphatidylinositol (PI) 3-kinase, the enzyme activity of which was greatly stimulated after a 5-min exposure to IL-1beta. This early and rapid activation of PI 3-kinase greatly enhanced FGF-2 production in CECs; pretreatment with LY294002 hampered the induction activity of IL-1beta. These observations suggest that IL-1beta takes part in endothelial to mesenchymal transformation of CECs through its inductive potential on FGF-2 via the action of PI 3-kinase.