Bax-associated mechanisms underlying the response of embryonic cells to methotrexate.
Bax was shown previously to regulate apoptotic cell death in various experimental systems, however, its involvement in teratogen-induced apoptosis is not clear yet. Therefore, we explored the involvement of Bax in the response of mouse embryonic fibroblasts (MEFs) to the anti cancer drug methotrexate (MTX), using Bax wild type (WT) and knockout (Bax(-/-)) MEFs. Our results demonstrated a significant teratogen-induced dose- and time-dependant decrease in the survival and culture density of both cell lines, which were found to be somewhat more prominent in WT cells. Exposure to MTX resulted also in decreased cell proliferation of WT but not Bax(-/-) cells and accordingly, we observed an accumulation of cells in the S phase and an increased percentage of cells in the Sub-G(1) phase of the cell cycle and the appearance of condensed nuclei, which were found to be somewhat more prominent in WT MEFs. In parallel, WT MEFs demonstrated a MTX-induced increase in the percentage of Bax-positive cells and a significant decrease in the percentage of bcl-2-, p65- or IkappaBalpha-positive cells, which were not detected in Bax(-/-) MEFs. Altogether, the differential sensitivity of WT or Bax(-/-) MEFs to MTX suggests a possible involvement of this molecule in the response of embryonic cells to teratogens.
Savion
S
S
Department of Cell and Developmental Biology, Sackler School of Medicine, Tel Aviv University, Ramat Aviv, Tel Aviv 69978, Israel. shoshans@post.tau.ac.il
Shtelman
E
E
Orenstein
H
H
Torchinsky
A
A
Fein
A
A
Toder
V
V
D000818:Animals; D000964:Antimetabolites, Antineoplastic / Q000008:administration & dosage / Q000633:toxicity*; D002453:Cell Cycle / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects; D020672:I-kappa B Proteins / Q000378:metabolism; D008727:Methotrexate / Q000008:administration & dosage / Q000633:toxicity*; D051379:Mice; D018345:Mice, Knockout; D000072000:NF-KappaB Inhibitor alpha; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D013723:Teratogens / Q000633:toxicity*; D013997:Time Factors; D051996:Transcription Factor RelA / Q000378:metabolism; D051028:bcl-2-Associated X Protein / Q000378:metabolism*
D000964:Antimetabolites, Antineoplastic; C494827:Bax protein, mouse; D020672:I-kappa B Proteins; C000606205:Nfkbia protein, mouse; D019253:Proto-Oncogene Proteins c-bcl-2; D013723:Teratogens; D051996:Transcription Factor RelA; D051028:bcl-2-Associated X Protein; D000072000:NF-KappaB Inhibitor alpha; D008727:Methotrexate