Quantitative analysis of apoptotic cell death using proton nuclear magnetic resonance spectroscopy.
Quantification of apoptotic cell death in vivo has become an important area of investigation in patients with acute lymphoblastic leukemia (ALL). We have devised a noninvasive analytical method to estimate the percentage of apoptotic lymphoblasts in doxorubicin-treated Jurkat T-cell ALL cultures, using proton nuclear magnetic resonance spectroscopy (1H NMR). We have found that the ratio of the methylene (CH2) resonance (at 1.3 ppm) to the methyl (CH3) resonance (at 0.9 ppm) signal intensity, as observed by 1H NMR, is directly proportional to the percentage of apoptotic lymphoblasts in vitro. The correlation between the CH2/CH3 signal intensity ratio and the percentage of apoptotic lymphoblasts was optimal 24 to 28 hours after doxorubicin treatment (r2 = .947, N = 27 samples). There was also a direct temporal relationship between an increase in the CH2/CH3 signal intensity ratio and the onset of apoptosis as detected by nuclear morphologic analysis, fluorescein-annexin V flow cytometry, and DNA gel electrophoresis. Thin-layer chromatography confirmed that a dynamic and/or compositional change of the plasma membrane, rather than increases in lipase activity or fatty acid production, appears to account for the increase in the CH2/CH3 signal intensity ratio during apoptosis. 1H NMR may have clinical utility for the early noninvasive assessment of chemotherapeutic efficacy in patients with ALL.
Blankenberg
F G
FG
Department of Radiology, Stanford University School of Medicine, CA, USA.
Katsikis
P D
PD
Storrs
R W
RW
Beaulieu
C
C
Spielman
D
D
Chen
J Y
JY
Naumovski
L
L
Tait
J F
JF
D017304:Annexin A5 / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects; D015415:Biomarkers; D002462:Cell Membrane / Q000737:chemistry; D002467:Cell Nucleus / Q000648:ultrastructure; D002794:Choline / Q000032:analysis*; D002855:Chromatography, Thin Layer; D053938:DNA Fragmentation; D004273:DNA, Neoplasm / Q000032:analysis; D004317:Doxorubicin / Q000494:pharmacology; D005434:Flow Cytometry; D006801:Humans; D015459:Leukemia-Lymphoma, Adult T-Cell / Q000378:metabolism / Q000473:pathology; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008563:Membrane Lipids / Q000032:analysis*; D009363:Neoplasm Proteins / Q000032:analysis; D010718:Phosphatidylserines / Q000032:analysis*; D018000:Receptors, Peptide / Q000032:analysis; D011994:Recombinant Proteins / Q000378:metabolism; D014407:Tumor Cells, Cultured / Q000737:chemistry / Q000187:drug effects
D017304:Annexin A5; D015415:Biomarkers; D004273:DNA, Neoplasm; D008563:Membrane Lipids; D009363:Neoplasm Proteins; D010718:Phosphatidylserines; D018000:Receptors, Peptide; D011994:Recombinant Proteins; C080707:annexin V receptor; D004317:Doxorubicin; D002794:Choline