title: ATP-induced Ca2+ signals in bronchial epithelial cells.
abstract: Ca2+-dependent Cl- secretion in the respiratory tract occurs physiologically or under pathophysiological conditions when inflammatory mediators are released. The mechanism of intracellular Ca2+ release was investigated in the immortalized bronchial epithelial cell line 16HBE14o-. Experiments on both intact and permeabilized cells revealed that only inositol 1,4,5-trisphosphate (InsP3) receptors and not ryanodine receptors are involved in intracellular Ca2+ release. The expression pattern of the three InsP3 receptor isoforms was assessed both at the mRNA and at the protein level. The level of expression at the mRNA level was type 3 (92.5%) >> type 2 (5.4%) > type 1 (2.1%) and this rank order was also observed at the protein level. The ATP-induced Ca2+ signals in the intact cell, consisting of abortive Ca2+ spikes or fully developed [Ca2+] rises and intracellular Ca2+ waves, were indicative of positive feedback of Ca2+ on the InsP3 receptors. Low Ca2+ concentrations stimulated and high Ca2+ concentrations inhibited InsP3-induced Ca2+ release in permeabilized 16HBE14o- cells. We localized a cytosolic Ca2+-binding site between amino acid residues 2077 and 2101 in the type-2 InsP3 receptor and between amino acids 2030 and 2050 in the type-3 InsP3 receptor by expressing the respective parts of these receptors as glutathione S-transferase fusion proteins in bacteria. We conclude that the InsP3 receptor isoforms expressed in 16HBE14o- cells (mainly type-3 and type-2) are stimulated by Ca2+ and that this phenomenon contributes to the ATP-induced Ca2+ signals in intact 16HBE14o- cells.
authors.affiliation: Laboratorium voor Fysiologie, K.U. Leuven Campus Gasthuisberg, Herestraat 49, B-3000 Leuven, Belgium.
authors.forename: I
authors.initials: I
authors.lastname: Sienaert
authors.forename: S
authors.initials: S
authors.lastname: Huyghe
authors.forename: J B
authors.initials: JB
authors.lastname: Parys
authors.forename: M
authors.initials: M
authors.lastname: Malfait
authors.forename: K
authors.initials: K
authors.lastname: Kunzelmann
authors.forename: H
authors.initials: H
authors.lastname: De Smedt
authors.forename: G M
authors.initials: GM
authors.lastname: Verleden
authors.forename: L
authors.initials: L
authors.lastname: Missiaen
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology*; D001980:Bronchi / Q000187:drug effects / Q000378:metabolism*; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000235:genetics / Q000502:physiology; D002461:Cell Line, Transformed; D003600:Cytosol / Q000378:metabolism; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism; D005246:Feedback; D015870:Gene Expression; D005982:Glutathione Transferase / Q000235:genetics; D006801:Humans; D053496:Inositol 1,4,5-Trisphosphate Receptors; D012333:RNA, Messenger / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000502:physiology; D011993:Recombinant Fusion Proteins; D019837:Ryanodine Receptor Calcium Release Channel / Q000502:physiology; D015398:Signal Transduction / Q000187:drug effects*
chemical_list: D015220:Calcium Channels; C506926:ITPR1 protein, human; C506927:ITPR2 protein, human; D053496:Inositol 1,4,5-Trisphosphate Receptors; D012333:RNA, Messenger; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins; D019837:Ryanodine Receptor Calcium Release Channel; D002110:Caffeine; D000255:Adenosine Triphosphate; D005982:Glutathione Transferase; D002118:Calcium