Endothelial nitric oxide synthase is essential for nitric oxide generation, L-type Ca2+ channel activation and survival in RBL-2H3 mast cells.
Recent pharmacological and molecular genetic approaches have revealed the existence of functional L-type Ca2+ channels (LTCCs) in a variety of hematopoietic cells. We previously reported that Ca(v)1.2 LTCCs are expressed on mast cell surfaces, activated by the high-affinity IgE receptor (FcvarepsilonRI) engagement and protect mast cells against activation-induced cell death (AICD). We also demonstrated that FcvarepsilonRI engagement evokes nitric oxide (NO) generation in a phosphatidylinositol-3-kinase- and NO synthase (NOS)-dependent manner, which is also required for mast cell survival. Here we demonstrate that this endogenous NO mediates Ca(v)1.2 LTCC activation. FcvarepsilonRI engagement but not thapsigargin, a potent Ca2+ release-activated Ca2+ (CRAC) channel agonist, induced Ca2+ influx via NOS-dependent NO generation. RT-PCR analyses revealed predominant expression of eNOS in mast cells. Subsequent experiments involving siRNA-mediated gene silencing of eNOS or Ca(v)1.2 LTCC revealed that eNOS was essential for NOS-dependent NO generation and Ca(v)1.2 LTCC activation but not CRAC channel activation. Similar to Ca(v)1.2 LTCCs, eNOS prevented the dissipation of the mitochondrial membrane potential and mitochondrial integrity collapse, thereby protecting mast cells against AICD. Taken together, the present findings demonstrate the key roles of the eNOS-NO-LTCC axis in mast cell survival after FcvarepsilonRI engagement.
Suzuki
Yoshihiro
Y
Division of Molecular Cell Immunology and Allergology, Nihon University Graduate School of Medical Science, 30-1 Oyaguchikami-cho Itabashi-Ku, Tokyo 173-8610, Japan. ysuzuki@med.nihon-u.ac.jp
Inoue
Toshio
T
Ra
Chisei
C
D000818:Animals; D020746:Calcium Channels, L-Type / Q000378:metabolism*; D020013:Calcium Signaling / Q000502:physiology*; D020169:Caspases / Q000378:metabolism; D002460:Cell Line; D002470:Cell Survival / Q000502:physiology*; D045304:Cytochromes c / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000378:metabolism; D008407:Mast Cells / Q000166:cytology / Q000502:physiology*; D053078:Membrane Potential, Mitochondrial; D009569:Nitric Oxide / Q000096:biosynthesis*; D052250:Nitric Oxide Synthase Type III / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017455:Receptors, IgE / Q000378:metabolism
D020746:Calcium Channels, L-Type; D004791:Enzyme Inhibitors; D034741:RNA, Small Interfering; D017455:Receptors, IgE; D009569:Nitric Oxide; D045304:Cytochromes c; D052250:Nitric Oxide Synthase Type III; D020169:Caspases