Bcl-2 and Bcl-XL serve an anti-inflammatory function in endothelial cells through inhibition of NF-κB
A.Z. Badrichani, … , F.H. Bach, C. Ferran
A.Z. Badrichani, … , F.H. Bach, C. Ferran
Published February 15, 1999
Citation Information: J Clin Invest. 1999;103(4):543-553. https://doi.org/10.1172/JCI2517.
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Article

Bcl-2 and Bcl-XL serve an anti-inflammatory function in endothelial cells through inhibition of NF-κB

Abstract

To maintain the integrity of the vascular barrier, endothelial cells (EC) are resistant to cell death. The molecular basis of this resistance may be explained by the function of antiapoptotic genes such as bcl family members. Overexpression of Bcl-2 or Bcl-XL protects EC from tumor necrosis factor (TNF)–mediated apoptosis. In addition, Bcl-2 or Bcl-XL inhibits activation of NF-κB and thus upregulation of proinflammatory genes. Bcl-2–mediated inhibition of NF-κB in EC occurs upstream of IκBα degradation without affecting p65-mediated transactivation. Overexpression of bcl genes in EC does not affect other transcription factors. Using deletion mutants of Bcl-2, the NF-κB inhibitory function of Bcl-2 was mapped to bcl homology domains BH2 and BH4, whereas all BH domains were required for the antiapoptotic function. These data suggest that Bcl-2 and Bcl-XL belong to a cytoprotective response that counteracts proapoptotic and proinflammatory insults and restores the physiological anti-inflammatory phenotype to the EC. By inhibiting NF-κB without sensitizing the cells (as with IκBα) to TNF-mediated apoptosis, Bcl-2 and Bcl-XL are prime candidates for genetic engineering of EC in pathological conditions where EC loss and unfettered activation are undesirable.

Authors

A.Z. Badrichani, D.M. Stroka, G. Bilbao, D.T. Curiel, F.H. Bach, C. Ferran

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(a) Structure/function relationships of Bcl-2 deletion mutants. Expressi...
(a) Structure/function relationships of Bcl-2 deletion mutants. Expression of Bcl-2 deletion mutants in BAEC. (b) The Bcl homology domains BH4 and BH2 are required for the inhibitory effect of Bcl-2 upon NF-κB activation after TNF and LPS stimulation, whereas (c) all BH domains and the NRD are required for the antiapoptotic function of Bcl-2. (a) Immunoblot detection of human wild-type and deletion mutants of Bcl-2 (lanes 2–6) in BAEC-transfected cells, using a polyclonal anti–Bcl-2 antibody. Mutant Δ1 is not recognized by this antibody and was detected using a monoclonal anti–Bcl-2 antibody (lane 9). (b) BAEC were cotransfected with 0.3 μg of β-gal, 0.6 μg of NF-κB reporter along with 0.7 μg of pAC (lanes A1–3), human Bcl-2 (lanes B1–3), Δ1 (lanes C1–3), Δ2 (lanes D1–3), Δ4 (lanes E1–3), Δ8 (lanes F1–3), and Δ12 (lanes G1–3). Cells were stimulated with 100 U/ml TNF (lanes 2) or 100 ng/ml LPS (lanes 3). Overexpression of Δ2, Δ4, or Δ8 inhibits the induction by TNF or LPS of a NF-κB reporter, in contrast to Δ1 or Δ12. Graph shown is representative of four experiments. Results are expressed in RLU. Error bars represent ± SE. (c) BAEC were cotransfected with a CMVβ-gal reporter (0.5 μg) and 1 μg of pAC (lanes 1 and 2), hBcl-2 (lanes 3 and 4), Δ1 (lanes 5 and 6), Δ2 (lanes 7 and 8), Δ4 (lanes 9 and 10), Δ8 (lanes 11 and 12), Δ12 (lanes 13 and 14). CHX was added to transfected cells (all lanes) that were subsequently stimulated (even lanes) or not (odd lanes) with TNF for 12 h. The percent cell survival was calculated as described in Methods. Results demonstrate that all the Bcl-2 deletion mutants lose their ability to protect EC from CHX-TNF mediated apoptosis. Error bars represent ± SE. Graph shown is representative of three experiments. NRD, negative regulatory domain.