Premium
Small Interfering RNA (siRNA) Suppresses the Fas Response Amplification Loop in Sulfur Mustard (SM)‐Exposed Normal Human Bronchial/Tracheal Epithelial (NHBE) Cells
Author(s) -
Keyser Brian,
Andres Devon,
Benton Betty,
Ray Radharaman
Publication year - 2009
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.23.1_supplement.618.7
Subject(s) - small interfering rna , apoptosis , caspase , caspase 8 , microbiology and biotechnology , western blot , sulfur mustard , chemistry , cancer research , biology , programmed cell death , biochemistry , rna , gene , organic chemistry , toxicity
Our previous studies using NHBE cells and caspase type‐specific peptide inhibitors demonstrated that SM induces apoptosis via a dominant caspase‐8 (Fas response) mediated pathway (Ray et al., Drug & Chem. Toxicol. 37:137‐48, 2008). Here, we validate this observation by using a more specific siRNA technology to suppress the Fas receptor as well as caspases‐9 and ‐6. Caspase activation was measured by using a luminescent caspase substrate hydrolysis assay. The effectiveness of the siRNA constructs was determined at both protein (western blot) and functional (caspase activation) levels. We found that down regulation of caspase‐9 by siRNA was able to significantly (~60%) reduce caspase‐3 activation by CH11, monoclonal antibody that activates Fas (caspase‐8 pathway); similar results were seen in SM‐exposed NHBE cells. These results provide further support to an amplification loop between caspases‐8, ‐9, ‐6, and ‐3 in apoptosis due to SM (300 μM) exposure in NHBE cells. These findings may be useful in developing siRNA based therapeutics against SM injury. Disclaimer: The opinions, interpretations, conclusions, and recommendations are those of the authors and are not necessarily endorsed by the U.S. Army or the Department of Defense. This research was supported by the Defense Threat Reduction Agency ‐ Joint Science and Technology Office, Medical S&T Division.