Freezing induces artificial cleavage of apoptosis-related proteins in human bone marrow cells.
The aim of this study was to investigate whether freeze-thawing of freshly isolated human mononuclear bone marrow cells (MNC) influences the integrity of apoptosis-related proteins as determined by immunoblot analyses. Our results show that bone marrow is more sensitive to this process than either myelomonocytoid leukemic P39 or Jurkat T-lymphocyte cell lines. Specifically, bone marrow cells displayed a high level of intrinsic proteolytic activity in response to a single freeze-thaw cycle, which led to the cleavage of various proteins involved in apoptosis cell signaling. This effect was completely blocked by the inclusion of broad-spectrum protease inhibitors in the freezing medium and subsequently thawing the cells on ice. Since differences in the freezing conditions (-80 degrees C vs. liquid nitrogen) did not alter the proteins of interest, we suggest that the thawing process is the critical point when proteolytic enzyme activity is elevated.
Schmidt-Mende
J
J
Division of Toxicology, Institute of Environmental Medicine, Karolinska Institutet, Box 210, S-171 77, Stockholm, Sweden.
Hellström-Lindberg
E
E
Joseph
B
B
Zhivotovsky
B
B
D017209:Apoptosis; D001854:Bone Marrow Cells / Q000166:cytology* / Q000378:metabolism*; D015925:Cryopreservation; D010450:Endopeptidases / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D066298:In Vitro Techniques; D019169:Jurkat Cells; D011506:Proteins / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D011506:Proteins; D010450:Endopeptidases