3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) causes Akt phosphorylation and morphological changes in intracellular organellae in cultured rat astrocytes.
3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) is widely used for cell viability and cytotoxicity assays, but cell biological effects of MTT itself have not been investigated. In this paper we show that MTT induces a morphological change in an intracellular membranous compartment labeled with anti-Rab5 antibody, dissociation of early endosomal auto-antigen (EEA1) from the membrane fraction, and phosphorylation of Akt probably through a phosphatidylinositol-3-OH kinase [PI(3)K] pathway in cultured rat astrocytes. These findings suggest that MTT affects cellular functions and conditions to some extent, and such effects of MTT may cause some discrepancies of measurement of cell viability using MTT assay and other assays. That is, the effects of MTT on cells could influence the results of cell viability assay. Moreover, MTT or other tetrazolium salts could be used as interesting activators of Akt to investigate the mechanism by which Akt or PI(3)K is activated.
Isobe
I
I
Department of Dementia Research, National Institute for Longevity Sciences, Nagoya City University Medical School, Japan. iisobe@med.nagoya-cu.ac.jp
Yanagisawa
K
K
Michikawa
M
M
D000818:Animals; D001253:Astrocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D002451:Cell Compartmentation / Q000187:drug effects; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D007150:Immunohistochemistry; D007424:Intracellular Fluid / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D015388:Organelles / Q000187:drug effects* / Q000648:ultrastructure; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases; D021381:Protein Transport / Q000187:drug effects; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt; D051381:Rats; D017207:Rats, Sprague-Dawley; D013778:Tetrazolium Salts / Q000494:pharmacology*; D013844:Thiazoles / Q000494:pharmacology*; D033921:Vesicular Transport Proteins; D020696:rab5 GTP-Binding Proteins / Q000378:metabolism
D008565:Membrane Proteins; D019869:Phosphatidylinositol 3-Kinases; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D013778:Tetrazolium Salts; D013844:Thiazoles; D033921:Vesicular Transport Proteins; D020696:rab5 GTP-Binding Proteins; C094100:2,2'-di-4-nitrophenyl-5,5'-diphenyl-3,3'-(3,3'-dimethoxy-4,4'-diphenylene)ditetrazolium; C110656:2-(4-iodophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium; C093846:early endosome antigen 1; C494920:Akt1 protein, rat; C022616:thiazolyl blue