TNF-alpha mediates eosinophil cationic protein-induced apoptosis in BEAS-2B cells.
BACKGROUND
Eosinophilic granulocytes are important for the human immune system. Many cationic proteins with cytotoxic activities, such as eosinophil cationic protein (ECP) and eosinophil-derived neurotoxin (EDN), are released from activated eosinophils. ECP, with low RNase activity, is widely used as a biomarker for asthma. ECP inhibits cell viability and induces apoptosis to cells. However, the specific pathway underlying the mechanisms of ECP-induced cytotoxicity remains unclear. This study investigated ECP-induced apoptosis in bronchial epithelial BEAS-2B cells and elucidated the specific pathway during apoptosis.


RESULTS
To address the mechanisms involved in ECP-induced apoptosis in human BEAS-2B cells, investigation was carried out using chromatin condensation, cleavage of poly (ADP-ribose) polymerase (PARP), sub-G1 distribution in cell cycle, annexin V labeling, and general or specific caspase inhibitors. Caspase-8-dependent apoptosis was demonstrated by cleavage of caspase-8 after recombinant ECP treatment, accompanied with elevated level of tumor necrosis factor alpha (TNF-alpha). Moreover, ECP-induced apoptosis was effectively inhibited in the presence of neutralizing anti-TNF-alpha antibody.


CONCLUSIONS
In conclusion, our results have demonstrated that ECP increased TNF-alpha production in BEAS-2B cells and triggered apoptosis by caspase-8 activation through mitochondria-independent pathway.
Chang
Kun-Che
KC
Department of Life Science, Institute of Biotechnology, National Tsing Hua University, Hsinchu 30013, Taiwan.
Lo
Chih-Wei
CW
Fan
Tan-Chi
TC
Chang
Margaret Dah-Tsyr
MD
Shu
Chih-Wen
CW
Chang
Chuan-Hsin
CH
Chung
Cheng-Ta
CT
Fang
Shun-Lung
SL
Chao
Chih-Chung
CC
Tsai
Jaw-Ji
JJ
Lai
Yiu-Kay
YK
D017304:Annexin A5 / Q000378:metabolism; D000906:Antibodies / Q000378:metabolism; D017209:Apoptosis; D053181:Caspase 8 / Q000378:metabolism; D045744:Cell Line, Tumor; D047150:Eosinophil Cationic Protein / Q000235:genetics / Q000378:metabolism*; D047208:Eosinophil-Derived Neurotoxin / Q000378:metabolism; D004804:Eosinophils / Q000276:immunology / Q000378:metabolism*; D016193:G1 Phase; D006801:Humans; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000276:immunology / Q000378:metabolism*
D017304:Annexin A5; D000906:Antibodies; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha; D011065:Poly(ADP-ribose) Polymerases; D047208:Eosinophil-Derived Neurotoxin; D047150:Eosinophil Cationic Protein; D053181:Caspase 8