Analysis of signal transduction pathways involved in anti-CD30 mAb-induced human eosinophil apoptosis.
BACKGROUND
Activation of cell surface CD30 by immobilized anti-CD30 monoclonal antibodies (mAbs) induces extremely rapid and intense apoptosis in human eosinophils in vitro. This anti-CD30 mAb-induced eosinophil apoptosis was inhibited by the addition of inhibitors of p38 and ERK1/2 mitogen-activated protein kinases (MAPKs). However, the signal transduction pathways other than for MAPKs remain unclear. In the present study, we tried to clarify the molecules involved in the induction of apoptosis after cross-linking of CD30.


METHODS
Purified human eosinophils were suspended in Iscove's minimal essential medium supplemented with 10% FCS and 1 ng/ml IL-5 and then cultured for 4 h in plates precoated with anti-CD30 mAb (clone Ber-H8) in the presence or absence of various signal transduction inhibitors. Eosinophil apoptosis was assessed using annexin V and flow cytometry.


RESULTS
The addition of inhibitors of caspase-3, caspase-8, pan-caspases, nuclear factor kappaB and protein kinase C at reasonable concentrations did not alter anti-CD30 mAb-induced eosinophil apoptosis in vitro. However, apoptosis was inhibited by phosphatidylinositol 3-kinase (PI3K) inhibitors: wortmannin at very high concentrations (>1 microM) significantly and LY294002 at a reasonable concentration partially inhibited that apoptosis.


CONCLUSIONS
Anti-CD30 mAb-induced eosinophil apoptosis is likely to be mediated mainly through MAPKs and partially through PI3K, but independent of caspase activation. Downstream signaling molecules of MAPK activation in eosinophils have to be clarified in future studies.
Matsumoto
Kenji
K
Department of Allergy and Immunology, National Research Institute for Child Health and Development, Tokyo, Japan. kmatsumoto @ nch.go.jp
Terakawa
Maki
M
Fukuda
Shuhei
S
Saito
Hirohisa
H
D000911:Antibodies, Monoclonal / Q000276:immunology / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D061945:Caspase Inhibitors; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004804:Eosinophils / Q000166:cytology* / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D006801:Humans; D017730:Ki-1 Antigen / Q000276:immunology*; D016328:NF-kappa B / Q000037:antagonists & inhibitors; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects*
D000911:Antibodies, Monoclonal; D061945:Caspase Inhibitors; D004791:Enzyme Inhibitors; D017730:Ki-1 Antigen; D016328:NF-kappa B; D000081082:Phosphoinositide-3 Kinase Inhibitors; D047428:Protein Kinase Inhibitors; D011493:Protein Kinase C