Overexpression of truncated IκBα potentiates TNF-α-induced apoptosis in mesangial cells

Junichi Hirahashi, Atsushi Takayanagi, Keiichi Hishikawa, Osamu Takase, Akihiro Chikaraishi, Matsuhiko Hayashi, Nobuyoshi Shimizu, Takao Saruta

研究成果: Article査読

34 被引用数 (Scopus)

抄録

Background. Dysregulation of apoptosis is one of the likely underlying mechanisms of mesangial proliferative glomerulonephritis (GN), a disease in which proinflammatory cytokines exhibit a wide range of biological activities. Among them, tumor necrosis factor-α (TNF-α) induces two conflicting pathways, one leading to activation of the nuclear factor-kappa B (NF-κB), and the other leading to caspase-mediated apoptosis. We investigated whether or not specific inhibition of NF-κB affects TNF-α- induced apoptosis in rat mesangial cells (MCs). Methods. To specifically inhibit NF-κB activation, we constructed a recombinant adenovirus vector expressing a truncated form of I kappa Bα (AdexIκBΔN) that lacks the phosphorylation sites essential for the activation of NF-κB. Electrophoretic mobility shift assay was performed to evaluate NF-κB activity. Nuclear morphology was observed by staining with Hoechst-33258. DNA fragmentation was detected using an ELISA kit with an antihistone antibody. To investigate the regulation of apoptosis, we measured caspase-3 and caspase-8 activity by ELISA, and examined the Bcl-2 and Bax protein level by Western blot. Results. TNF-α-induced NF-κB activation was blocked by overexpression of IκBΔN. Overexpression of IκBΔN potentiated TNF-α-induced apoptosis compared to mock transfection, and the potentiation was abolished by treatment with a caspase-3 inhibitor, Z-DEVD-FMK. Overexpression of IκBΔN augmented TNF-α- induced caspase-3 and caspase-8 activity, but did not affect Bcl-2 or Bax protein expression. Conclusion. Overexpression of IκBΔN potentiates TNF-α- induced apoptosis and augments caspase-8 and caspase-3 activity in rat MCs without changing Bcl-2 or Bax protein expression. These results suggest the potential usefulness of AdexIκBΔN to induce apoptosis in MCs under inflammatory conditions.

本文言語English
ページ(範囲)959-968
ページ数10
ジャーナルKidney international
57
3
DOI
出版ステータスPublished - 2000

ASJC Scopus subject areas

  • 腎臓病学

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