title: CD69 expression induced by thapsigargin, phorbol ester and ouabain on thymocytes is dependent on external Ca2+ entry.
abstract: In the present work murine thymocytes exposed to Thapsigargin (TG 10, 20 and 50 nM), Phorbol-12,13,20-triacetate (TPA16 nM) and Ouabain (OUA100 nM) exhibited an increased expression of CD69, a molecule related to cellular activation and associated to Ca(++) influx in other systems. The kinetics of CD69 appearance depended on the stimuli and dose used. TG 50 nM induced an increased expression by 6 h whereas with lower doses (10 and 20 nM) an increase was detected at 18 h. TPA maximal increase was evident at 6 h. OUA lead to an observable increase at 18 h. However, in the case of TPA or TG the presence of the stimuli was only necessary for the first 2 h of culture, whereas OUA needed to be present during the whole assay. It was also demonstrated that Ca(++) influx was an essential feature, as EGTA diminished or abolished CD69 increased expression. Nevertheless, EGTA was only capable of this effect when present at the time of the stimuli. No correlation of CD69 expression with thymocyte death was observed. Similarly, the agents under study did not promote the maturation from double-positive into single-positive thymocytes. TPA and Thapsigargin were capable of decreasing the level of CD4 molecules on the cell surface, probably due to the loss of these molecules. OUA, on the other hand, did not modify CD4/CD8 expression on these cells.
authors.affiliation: Laboratory of Tumoral Immunology, Department of Medical Biochemistry, Federal University of Rio de Janeiro, Rio de Janeiro, Brazil.
authors.forename: Sandra
authors.initials: S
authors.lastname: Rodrigues Mascarenhas
authors.forename: Juliana
authors.initials: J
authors.lastname: Echevarria-Lima
authors.forename: Neusa
authors.initials: N
authors.lastname: Fernandes dos Santos
authors.forename: Vivian M
authors.initials: VM
authors.lastname: Rumjanek
mesh_terms: D000818:Animals; D015703:Antigens, CD / Q000096:biosynthesis*; D000945:Antigens, Differentiation, T-Lymphocyte / Q000096:biosynthesis*; D015704:CD4 Antigens / Q000096:biosynthesis; D016827:CD8 Antigens / Q000096:biosynthesis; D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D004533:Egtazic Acid / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005434:Flow Cytometry; D007700:Kinetics; D037181:Lectins, C-Type; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D010042:Ouabain / Q000494:pharmacology*; D010703:Phorbol Esters / Q000494:pharmacology*; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors; D013601:T-Lymphocytes / Q000187:drug effects / Q000378:metabolism*; D019284:Thapsigargin / Q000494:pharmacology*; D000099192:CD69 Antigens
chemical_list: D015703:Antigens, CD; D000945:Antigens, Differentiation, T-Lymphocyte; D015704:CD4 Antigens; D016827:CD8 Antigens; D002118:Calcium; D000252:Calcium-Transporting ATPases; D002614:Chelating Agents; D004533:Egtazic Acid; D004791:Enzyme Inhibitors; D037181:Lectins, C-Type; D010042:Ouabain; D010703:Phorbol Esters; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000254:Sodium-Potassium-Exchanging ATPase; D019284:Thapsigargin; D000099192:CD69 Antigens