Bcl-2 regulatory pathway is functional in chronic lymphocytic leukemia.
BACKGROUND
Chronic lymphocytic leukemia (CLL) is characterized by accumulation of clonal, malignant CD5(+), CD23(+) B cells. In vivo, these cells have an antiapoptotic phenotype (high levels of Bcl-2 and low levels of proapoptotic Bcl-2 family proteins, such as Bax). Abnormal B cells accumulate due to altered apoptosis regulation rather than to increased proliferation. However, it is unclear whether there are inherent Bcl-2 apoptotic pathway defects. With in vitro culture, these B cells rapidly apoptosis.


METHODS
To investigate apoptosis regulation, Bcl-2, Bax, mitochondrial membrane potential, annexin V, and caspase activation were simultaneous monitored in individual cells during in vitro apoptosis.


RESULTS
With in vitro culture, 30% to 50% of B cells were apoptotic at 24 h compared with fewer than 10% of T cells. Apoptotic B cells showed dramatic Bax upregulation and slight Bcl-2 decreases accompanied by decreased mitochondrial membrane potential and increased activated caspase-3 protein levels. Caspase-3 and caspase-9 activities were increased 18- to 51-fold and 6- to 11-fold, respectively, after 24 h of culture. Caspase-8 showed limited or no activation (less than fourfold).


CONCLUSIONS
These data show that in vitro apoptosis of CLL B cells occurs through a well-characterized Bcl-2 regulatory pathway consistent with that pathway being functional. Further, these cells' antiapoptotic phenotype is dependent on the in vivo environment, potentially involving paracrine/autocrine interactions.
Goolsby
Charles
C
Robert H. Lurie Comprehensive Cancer Center and Department of Pathology, Northwestern University Medical School, Chicago, Illinois, USA. c-goolsby@northwestern.edu
Paniagua
Mary
M
Tallman
Martin
M
Gartenhaus
Ronald B
RB
D000368:Aged; D017304:Annexin A5; D017209:Apoptosis / Q000502:physiology; D001402:B-Lymphocytes / Q000378:metabolism* / Q000473:pathology; D020169:Caspases / Q000096:biosynthesis; D002452:Cell Count; D002478:Cells, Cultured; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D016130:Immunophenotyping; D015451:Leukemia, Lymphocytic, Chronic, B-Cell / Q000378:metabolism* / Q000473:pathology; D008297:Male; D008564:Membrane Potentials; D008875:Middle Aged; D008928:Mitochondria; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism*; D051028:bcl-2-Associated X Protein
D017304:Annexin A5; C494826:BAX protein, human; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D020169:Caspases