Beta2-adrenoceptor activation inhibits Shiga toxin2-induced apoptosis of renal tubular epithelial cells |
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Authors: | Nakamura Akio Imaizumi Akira Yanagawa Yukishige Niimi Ryo Kohsaka Takao Johns Edward J |
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Affiliation: | Department of Paediatrics, Teikyo University School of Medicine, 2-11-1 Kaga, Itabashi-ku, Tokyo 173, Japan. akio@med.teikyo-u.ac.jp |
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Abstract: | Apoptosis is regulated by several pathways, such as caspases, mitogen activated protein kinase (MAPK) and cAMP/cAMP-dependent protein kinase A (PKA) cascade. This study investigated the effect of beta(2)-adrenoceptor activation on Shiga toxin (Stx)2-induced apoptosis in renal tubular cells and the contribution of these signalling pathways. Cultured human adenocarcinoma-derived tubular cells were exposed to Stx2 (64 pg/mL) for 2-24hr following the addition of the beta(2)-adrenoceptor agonist (terbutaline) to the incubation medium. Stx2-induced apoptosis and its amelioration by beta(2)-adrenoceptor activation was confirmed using DNA degradation assays and by flow cytometry for annexin V, mitochondrial membrane potential and caspase(-3 and -7) activity. Exposure of cells to Stx2 for 24hr increased the DNA fragmentation to 11.6+/-0.9%, compared to 3.3+/-0.2% in control cells (P<0.05) but was decreased to approximately 5-7% (P<0.05) in the presence of terbutaline. Furthermore, Stx2-stimulated apoptosis, detected by TUNEL, annexin V and mitochondrial potential, was inhibited by terbutaline (P<0.05) which was prevented by cAMP-PKA inhibitors and a beta(2)-adrenoceptor antagonist. However, inhibition of Stx2-mediated caspase activity by terbutaline was partially blocked by cAMP-PKA inhibitors. On the other hand, p38MAPK inhibition by terbutaline prevented Stx2-induced apoptosis and caspase activity through a cAMP-independent pathway via beta(2)-adrenoceptor. These data indicate that beta(2)-adrenoceptor activation can inhibit Stx2-induced apoptosis of the cells, which may be caused by a reduction in caspase activity through cAMP-PKA activation and the p38MAPK pathway. |
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Keywords: | HUS, Haemolytic Uraemic Syndrome Stx, Shiga toxin MAPK, mitogen activated protein kinase JNK, c-Jun N-terminal protein kinase DMEM, Dulbecco’s modified Eagles medium PD, PD098059 SB, SB203580 ICI, ICI 118,551 KT, KT5720 FCS, foetal calf serum FITC, fluorescent isothiocyanate dbcAMP, dibutyryl cyclic AMP PARP, poly ADP-ribose polymerase Gb3, globotriaosylceramide |
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