In vivo epithelial wound repair requires mobilization of endogenous intracellular and extracellular calcium.
We report that a localized intracellular and extracellular Ca(2+) mobilization occurs at the site of microscopic epithelial damage in vivo and is required to mediate tissue repair. Intravital confocal/two-photon microscopy continuously imaged the surgically exposed stomach mucosa of anesthetized mice while photodamage of gastric epithelial surface cells created a microscopic lesion that healed within 15 min. Transgenic mice with an intracellular Ca(2+)-sensitive protein (yellow cameleon 3.0) report that intracellular Ca(2+) selectively increases in restituting gastric epithelial cells adjacent to the damaged cells. Pretreatment with U-73122, indomethacin, 2-aminoethoxydiphenylborane, or verapamil inhibits repair of the damage and also inhibits the intracellular Ca(2+) increase. Confocal imaging of Fura-Red dye in luminal superfusate shows a localized extracellular Ca(2+) increase at the gastric surface adjacent to the damage that temporally follows intracellular Ca(2+) mobilization. Indomethacin and verapamil also inhibit the luminal Ca(2+) increase. Intracellular Ca(2+) chelation (1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid/acetoxymethyl ester, BAPTA/AM) fully inhibits intracellular and luminal Ca(2+) increases, whereas luminal calcium chelation (N-(2-hydroxyetheyl)-ethylendiamin-N,N,N'-triacetic acid trisodium, HEDTA) blocks the increase of luminal Ca(2+) and unevenly inhibits late-phase intracellular Ca(2+) mobilization. Both modes of Ca(2+) chelation slow gastric repair. In plasma membrane Ca-ATPase 1(+/-) mice, but not plasma membrane Ca-ATPase 4(-/-) mice, there is slowed epithelial repair and a diminished gastric surface Ca(2+) increase. We conclude that endogenous Ca(2+), mobilized by signaling pathways and transmembrane Ca(2+) transport, causes increased Ca(2+) levels at the epithelial damage site that are essential to gastric epithelial cell restitution in vivo.
['Aihara|Eitaro|E|', 'Hentz|Courtney L|CL|', 'Korman|Abraham M|AM|', 'Perry|Nicholas P J|NPJ|', 'Prasad|Vikram|V|', 'Shull|Gary E|GE|', 'Montrose|Marshall H|MH|']
D000818:Animals D000894:Anti-Inflammatory Agents, Non-Steroidal D001896:Boron Compounds D002118:Calcium D002121:Calcium Channel Blockers D020013:Calcium Signaling D000252:Calcium-Transporting ATPases D002614:Chelating Agents D004492:Edetic Acid D004533:Egtazic Acid D004963:Estrenes D005753:Gastric Mucosa D007213:Indomethacin D051379:Mice D018345:Mice, Knockout D010726:Phosphodiesterase Inhibitors D011760:Pyrrolidinones D014700:Verapamil D014945:Wound Healing
Calcium Two-photon Microscopy Photodamage Yellow Cameleon Calcium Imaging Calcium Signaling Cell Migration Fluorescence Resonance Energy Transfer (FRET) Fura Red