hsBAFF-upregulated intracellular free Ca(2+) homeostasis regulates ERK1/2 activity and cell proliferation in B cells in vitro.
We studied hsBAFF activity in in vitro mouse splenic B cells. hsBAFF effects on intracellular free Ca(2+) concentration ([Ca(2+)](i)) were assayed, using a laser scanning confocal microscope with fluorescent probe, Fluo-3/AM. We showed that treatment of B cells with 0.5-5 microg/ml hsBAFF resulted in significantly higher [Ca(2+)](i) levels in a dose-dependent fashion at 12 and 24 h, respectively (p<0.05 or p<0.01 vs. control). Furthermore, we noticed that 2.5 microg/ml hsBAFF-treated cells were significantly resistant to decrease of cellular viability induced by thapsigargin (Tg), an endoplasmic reticulum (ER) Ca(2+)-ATPase inhibitor (p<0.05 hsBAFF plus Tg group vs. Tg group). Thus hsBAFF may promote B cell survival by direct upregulation of [Ca(2+)](i) physiological homeostasis contributing to prevention of [Ca(2+)](i) dysfunction. Using immunocytochemistry and Western blot analysis, we found that the activation of ERK1/2 due to hsBAFF was triggered by a [Ca(2+)](i) -dependent pathway, leading to elevation of B cell proliferation. This is supported by the findings that intracellular Ca(2+) chelator BAPTA/AM attenuated phosphorylated ERK1/2 expression and cell proliferation in hsBAFF-stimulated B cells. hsBAFF-stimulated B cell proliferation was obviously reduced by mitogen extracellular kinase 1/2 (MEK1/2, upstream of ERK1/2) inhibitor U0126. Taken together, the main finding of this study is that hsBAFF elicits higher but homeostatic [Ca(2+)](i) levels, which regulates ERK1/2 activity and cell proliferation in in vitro B cells.
Liang
J Q
JQ
College of Life Sciences, Nanjing Normal University, Jiangsu Province Key Laboratory for Molecular and Medical Biotechnology, Nanjing 20046, P. R. China.
Zhang
W
W
Wen
L
L
Gao
W
W
Zhang
S Q
SQ
Chen
L
L
D000818:Animals; D053264:B-Cell Activating Factor / Q000378:metabolism*; D001402:B-Lymphocytes / Q000201:enzymology*; D002070:Butadienes / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D049109:Cell Proliferation; D002470:Cell Survival; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006706:Homeostasis; D006801:Humans; D051379:Mice; D008813:Mice, Inbred ICR; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D009570:Nitriles / Q000494:pharmacology; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism; D019284:Thapsigargin / Q000494:pharmacology; D013997:Time Factors
D053264:B-Cell Activating Factor; D002070:Butadienes; D002614:Chelating Agents; D004791:Enzyme Inhibitors; D009570:Nitriles; D047428:Protein Kinase Inhibitors; C505719:TNFSF13B protein, human; C113580:U 0126; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D004533:Egtazic Acid; D019284:Thapsigargin; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D002118:Calcium