Cytosolic acidification and lysosomal alkalinization during TNF-alpha induced apoptosis in U937 cells.
Apoptosis is often associated with acidification of the cytosol and since loss of lysosomal proton gradient and release of lysosomal content are early events during apoptosis, we investigated if the lysosomal compartment could contribute to cytosolic acidification. After exposure of U937 cells to tumor necrosis factor-alpha, three populations; healthy, pre-apoptotic, and apoptotic cells, were identified by flow cytometry. These populations were investigated regarding intra-cellular pH and apoptosis-associated events. There was a drop in cytosolic pH from 7.2 +/- 0.1 in healthy cells to 6.8 +/- 0.1 in pre-apoptotic, caspase-negative cells. In apoptotic, caspase-positive cells, the pH was further decreased to 5.7 +/- 0.04. The cytosolic acidification was not affected by addition of specific inhibitors towards caspases or the mitochondrial F(0)F(1)-ATPase. In parallel to the cytosolic acidification, a rise in lysosomal pH from 4.3 +/- 0.3, in the healthy population, to 4.8 +/- 0.3 and 5.5 +/- 0.3 in the pre-apoptotic- and apoptotic populations, respectively, was detected. In addition, lysosomal membrane permeability increased as detected as release of cathepsin D from lysosomes to the cytosol in pre-apoptotic and apoptotic cells. We, thus, suggest that lysosomal proton release is the cause of the cytosolic acidification of U937 cells exposed to TNF-alpha.
Nilsson
Cathrine
C
Division of Experimental Pathology, Faculty of Health Sciences, Linköping University, S-581 85 Linköping, Sweden. cathrine.nilsson@inr.liu.se
Johansson
Uno
U
Johansson
Ann-Charlotte
AC
Kågedal
Katarina
K
Ollinger
Karin
K
D000577:Amides / Q000494:pharmacology; D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3; D053181:Caspase 8; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D002402:Cathepsin D / Q000378:metabolism; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D048429:Cell Size / Q000187:drug effects; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism*; D003902:Detergents / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D007425:Intracellular Membranes / Q000187:drug effects / Q000502:physiology; D008247:Lysosomes / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D025261:Mitochondrial Proton-Translocating ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism; D009840:Oligomycins / Q000494:pharmacology; D010539:Permeability / Q000187:drug effects; D010718:Phosphatidylserines / Q000378:metabolism; D021381:Protein Transport / Q000187:drug effects; D011522:Protons; D012694:Serine / Q000031:analogs & derivatives / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D020298:U937 Cells
D000577:Amides; D000590:Amino Acid Chloromethyl Ketones; D061945:Caspase Inhibitors; D003902:Detergents; D004791:Enzyme Inhibitors; C407126:O-methyl-serine dodecylamide hydrochloride; D009840:Oligomycins; D010718:Phosphatidylserines; D011522:Protons; D014409:Tumor Necrosis Factor-alpha; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D012694:Serine; C505407:CASP3 protein, human; C505530:CASP8 protein, human; D053148:Caspase 3; D053181:Caspase 8; D020169:Caspases; D002402:Cathepsin D; D025261:Mitochondrial Proton-Translocating ATPases