Stromal cells prevent apoptosis of AML cells by up-regulation of anti-apoptotic proteins.
The aim of this study was to study interactions between stromal bone marrow microenvironment and leukemic cells. We tested the hypothesis that stromal cells prevent apoptosis of AML cells by up-regulating anti-apoptotic proteins in leukemic blasts. In HL-60 and NB-4 cells, serum deprivation- and ara-C-induced apoptosis was diminished when cells were cocultured with murine MS-5 stromal cells (P < 0.02). This effect was reproduced with conditioned medium from MS-5 cells. Cocultivation with stromal cells induced Bcl-2 expression levels, both by PCR analysis and flow cytometry. In primary AML (n = 14), ara-C-induced apoptosis was significantly lower in cells cocultured with MS-5 cells than in controls (P < 0.001). This effect was partially preserved when leukemic cells were separated from stromal cells by a microporous insert (in 5/9 samples, P = 0.04). In addition, Bcl-2 levels were significantly higher in stroma-supported than in control CD34(+) AML cells (P < 0.01). Bcl-X(L) levels were higher in 5/7 samples grown on stromal layers. Of note, in AML patients resistant to induction chemotherapy (n = 6), Bcl-2 increased significantly after cultivation with stromal cells, but no such increase was noted in cells from chemotherapy-sensitive patients. In conclusion, MS-5 stromal cells prevented apoptosis in HL-60 cells and in primary AML blasts via modulation of Bcl-2 family proteins. The observed association of high Bcl-2 expression in stroma-supported AML blasts in vitro with resistance to chemotherapy in vivo suggests that the same mechanisms may be operational in vivo.
Section of Molecular Hematology and Therapy, Department of Blood and Marrow Transplantation, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA.
M
M
Konopleva
S
S
Konoplev
W
W
Hu
A Y
AY
Zaritskey
B V
BV
Afanasiev
M
M
Andreeff
D000208:Acute Disease; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000818:Animals; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D001854:Bone Marrow Cells / Q000187:drug effects / Q000473:pathology*; D002448:Cell Adhesion / Q000502:physiology; D002455:Cell Division / Q000187:drug effects; D016895:Culture Media, Serum-Free; D003561:Cytarabine / Q000494:pharmacology; D017931:DNA Primers / Q000737:chemistry; D005260:Female; D005456:Fluorescent Dyes; D018922:HL-60 Cells / Q000187:drug effects; D006801:Humans; D007951:Leukemia, Myeloid / Q000188:drug therapy / Q000378:metabolism* / Q000473:pathology; D008297:Male; D051379:Mice; D008875:Middle Aged; D009930:Organic Chemicals; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D012334:RNA, Neoplasm / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017154:Stromal Cells / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D014407:Tumor Cells, Cultured / Q000187:drug effects; D015854:Up-Regulation
D000964:Antimetabolites, Antineoplastic; D016895:Culture Media, Serum-Free; D017931:DNA Primers; D005456:Fluorescent Dyes; D009930:Organic Chemicals; C070080:PKH 26; D019253:Proto-Oncogene Proteins c-bcl-2; D012334:RNA, Neoplasm; D003561:Cytarabine