Hydrogen Peroxide Induces Up-Regulation of Fas in Human Endothelial Cells
Hydrogen peroxide (H2O2), an oxidant generated by inflammatory cells, is an important mediator of injury of endothelial cells (ECs). Here we show that H2O2 induces up-regulation of the expression of Fas, a death signal, in human ECs in culture. Flow cytometric analysis with a mAb against human Fas showed that incubation for 24 h with H2O2 induced a dose-dependent increase in the level of Fas in ECs. Coincubation with catalase, which rapidly degrades H2O2, inhibited H2O2-induced up-regulation of Fas. H2O2 also induced a dose-dependent increase in Fas mRNA level. A significant increase in Fas mRNA levels was observed from 6 h after stimulation with H2O2. Vanadate, a protein phosphatase inhibitor, significantly enhanced Fas mRNA and protein levels in H2O2-treated ECs. On the other hand, genistein, a tyrosine kinase inhibitor, inhibited H2O2-induced Fas mRNA expression. Furthermore, a flow cytometric method with propidium iodide staining and electron microscopic analysis showed that incubation with an agonistic Ab against Fas (anti-Fas IgM) induced apoptosis in H2O2-treated cells. These findings suggest that H2O2 induces up-regulation of Fas in ECs and that activation of protein tyrosine kinase may be involved in the mechanism of H2O2-induced Fas expression. Therefore, Fas-mediated apoptosis may have a pathologic role in H2O2-induced EC injury and thereby provide a new therapeutic target.
Fas, also called APO-1 or CD95, is a type I membrane protein belonging to the TNF and nerve growth factor receptor family, which mediates a death signal (1). Triggering this pathway requires the cross-linking of Fas with either agonistic Abs to Fas or cells expressing Fas ligand (Fas-L).2 Various cells express Fas, whereas Fas-L is expressed predominantly in activated T cells. Malfunction of the Fas system causes lymphoproliferative disorders and accelerates autoimmune diseases, whereas its overactivity may cause tissue destruction. Therapeutic uses of the Fas system might include blocking the exacerbated Fas-based pathologic manifestations with the soluble form of Fas, neutralizing Abs to Fas or Fas-L, or with inhibitors of Fas-L induction or Fas-mediated apoptosis.
Injury of endothelial cells (ECs) is a critical event in the acute inflammatory process and the development of atherosclerosis (2, 3). In the genesis of inflammatory lesions, ECs can interact with macrophages, platelets, and vascular smooth muscle cells (VSMC) as well as T cells. One form of injury to ECs results from excessive levels of oxygen radicals released from leukocytes, macrophages, and ECs themselves (4, 5, 6, 7, 8, 9, 10). Increasing evidence suggests that oxidative stress is a mediator of apoptosis (11). This hypothesis is based on the findings that many agents that induce apoptosis are oxidants or stimulators of cellular oxidative metabolism, and that, conversely, many inhibitors of apoptosis have antioxidative activities or enhance cellular antioxidant defenses (12, 13, 14, 15, 16, 17). Furthermore, there is recent evidence that diffusable reactive oxygen intermediates such as nitric oxide (NO) and hydrogen peroxide (H2O2) can modulate cellular functions through altering signal transduction in many cell types, including ECs, VSMC, and T cells (18, 19, 20, 21, 22, 23, 24, 25). Recently, we demonstrated that NO induces up-regulation of Fas and apoptosis in VSMC (26). In the present study, we therefore examined whether H2O2 can affect the level of Fas in cultured ECs. We found that H2O2 up-regulates Fas expression through the activation of protein tyrosine kinase in ECs.
Hydrogen Peroxide Induces Up-Regulation of Fas in Human Endothelial Cells | The Journal of Immunology http://www.jimmunol.org/content/160/8/4042.full
Toshimitsu Suhara, Keisuke Fukuo, Tomosada Sugimoto, Shigeto Morimoto, Takeshi Nakahashi, Shigeki Hata, Masumi Shimizu and Toshio Ogihara
J Immunol April 15, 1998, 160 (8) 4042-4047;
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