title: Enhanced Ca²⁺ entry and Na+/Ca²⁺ exchanger activity in dendritic cells from AMP-activated protein kinase-deficient mice.
abstract: In dendritic cells (DCs), chemotactic chemokines, such as CXCL12, rapidly increase cytosolic Ca(2+)concentrations ([Ca(2+)](i)) by triggering Ca(2+) release from intracellular stores followed by store-operated Ca(2+) (SOC) entry. Increase of [Ca(2+)](i) is blunted and terminated by Ca(2+) extrusion, accomplished by K(+)-independent Na(+)/Ca(2+) exchangers (NCXs) and K(+)-dependent Na(+)/Ca(2+) exchangers (NCKXs). Increased [Ca(2+)](i) activates energy-sensing AMP-activated protein kinase (AMPK), which suppresses proinflammatory responses of DCs and macrophages. The present study explored whether AMPK participates in the regulation of DC [Ca(2+)](i) and migration. DCs were isolated from AMPKα1-deficient (ampk(-/-)) mice and, as control, from their wild-type (ampk(+/+)) littermates. AMPKα1, Orai1-2, STIM1-2, and mitochondrial calcium uniporter protein expression was determined by Western blotting, [Ca(2+)](i) by Fura-2 fluorescence, SOC entry by inhibition of endosomal Ca(2+) ATPase with thapsigargin (1 μM), Na(+)/Ca(2+) exchanger activity from increase of [Ca(2+)](i), and respective whole-cell current in patch clamp following removal of extracellular Na(+). Migration was quantified utilizing transwell chambers. AMPKα1 protein is expressed in ampk(+/+) DCs but not in ampk(-/-) DCs. CXCL12 (300 ng/ml)-induced increase of [Ca(2+)](i), SOC entry, Orai 1 protein abundance, NCX, and NCKX were all significantly higher in ampk(-/-) DCs than in ampk(+/+) DCs. NCX and NCKX currents were similarly increased in ampk(-/-) DCs. Moreover, CXCL12 (50 ng/ml)-induced DC migration was enhanced in ampk(-/-) DCs. AMPK thus inhibits SOC entry, Na(+)/Ca(2+) exchangers, and migration of DCs.
authors.affiliation: Department of Physiology, University of Tübingen, Tübingen, Germany.
authors.forename: Meerim K
authors.initials: MK
authors.lastname: Nurbaeva
authors.forename: Evi
authors.initials: E
authors.lastname: Schmid
authors.forename: Kalina
authors.initials: K
authors.lastname: Szteyn
authors.forename: Wenting
authors.initials: W
authors.lastname: Yang
authors.forename: Benoit
authors.initials: B
authors.lastname: Viollet
authors.forename: Ekaterina
authors.initials: E
authors.lastname: Shumilina
authors.forename: Florian
authors.initials: F
authors.lastname: Lang
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000172:deficiency* / Q000235:genetics; D000818:Animals; D001483:Base Sequence; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000378:metabolism; D020013:Calcium Signaling; D002465:Cell Movement; D054377:Chemokine CXCL12 / Q000494:pharmacology; D017931:DNA Primers / Q000235:genetics; D003713:Dendritic Cells / Q000187:drug effects / Q000378:metabolism*; D066298:In Vitro Techniques; D051379:Mice; D018345:Mice, Knockout; D000071741:ORAI2 Protein; D019831:Sodium-Calcium Exchanger / Q000378:metabolism*
chemical_list: D015220:Calcium Channels; D054377:Chemokine CXCL12; C515256:Cxcl12 protein, mouse; D017931:DNA Primers; D000071741:ORAI2 Protein; C520431:Orai2 protein, mouse; D019831:Sodium-Calcium Exchanger; C548179:Prkaa1 protein, mouse; D055372:AMP-Activated Protein Kinases; D002118:Calcium