Calcium and the heat-shock response in the human monocytic line U-937.
In the human monocytic line U-937, 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] increases cytosolic free calcium concentration ([Ca2+]i). 1,25-(OH)2D3 also enhances the synthesis of heat-shock proteins (HSPs) when U-937 cells are exposed to elevated temperatures. To establish whether these two effects of 1,25-(OH)2D3 are related, we examined the effects of calcium on the heat-shock (HS) response, as well as the influence of 1,25-(OH)2D3 on this system. The equilibrium dissociation constant (Kd) of the fluorescent probe used to measure [Ca2+]i, fura-2, at 37 and 45 degrees C was found to be 191 and 234 nM, respectively. Exposure of U-937 cells to 45 degrees C did not increase [Ca2+]i under conditions in which active efflux of the dye was prevented by the organic anion transport inhibitor probenecid (1 mM). In cells preincubated in calcium-free medium, with subsequent addition of 4 mM EGTA before HS, or exposed to the calmodulin antagonist N-(6-aminohexyl)-5-chloro-1-naphthalene sulfonamide (W-7), the increase in HSPs synthesis was not affected. Cell viability, assessed by [3H]thymidine uptake, was not different between cells exposed to HS in calcium-containing or calcium-free media. Moreover, the effects of 1,25-(OH)2D3 on the HS response were also observed in a calcium-depleted medium, indicating that the effects of 1,25-(OH)2D3 on HSP synthesis were not mediated by [Ca2+]i.
Respiratory Division and Allergy Unit, Geneva University Hospital, Switzerland.
S
S
Kantengwa
A M
AM
Capponi
J V
JV
Bonventre
B S
BS
Polla
D001572:Benzofurans; D002117:Calcitriol / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D005456:Fluorescent Dyes; D016257:Fura-2; D006360:Heat-Shock Proteins / Q000096:biosynthesis* / Q000302:isolation & purification; D006358:Hot Temperature; D006801:Humans; D007700:Kinetics; D016403:Lymphoma, Large B-Cell, Diffuse; D009000:Monocytes; D011339:Probenecid / Q000494:pharmacology; D013050:Spectrometry, Fluorescence; D013449:Sulfonamides / Q000494:pharmacology; D014407:Tumor Cells, Cultured / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*
D001572:Benzofurans; D005456:Fluorescent Dyes; D006360:Heat-Shock Proteins; D013449:Sulfonamides; C017967:W 7; D002117:Calcitriol; D011339:Probenecid; D002118:Calcium; D016257:Fura-2