Detection of VP-16-treated HL-60 cell apoptosis by TUNEL electron microscopy.
Apoptosis is essential to many physiological processes, including maturation of cells and the immune system. Deficient regulation of apoptosis may play an important role in many pathological conditions, such as autoimmunity, AIDS, and myelodysplastic syndrome. Several methods have been described to identify apoptotic cells. DNA strand breaks can be identified by labeling free 3'-OH termini with modified nucleotides by enzymatic reaction (TUNEL method). In this study, apoptosis was introduced in cultured HL-60 cells treated with VP-16, and the TUNEL method was adapted for electromicroscopy, called the EM TUNEL method. The results of the EM TUNEL method were compared with those of light microscopy and flow cytometry. Apoptotic cells following VP-16 (10 micrograms/mL) treatment were detected after 3 h by all methods (light microscopy, Erythrocin B, electron microscopy, flow cytometry, TUNEL, and EM TUNEL). EM TUNEL was the most sensitive method of detection, with a detection rate of 32%. Furthermore, EM TUNEL was more suitable for distinguishing cell lineage than light microscopy TUNEL. The results indicate that EM TUNEL can be used to detect apoptosis in bone marrow species in vivo.
Saitoh
T
T
Third Department of Internal Medicine, Gunma University School of Medicine, Japan.
Murakami
H
H
Hatsumi
N
N
Matsushima
T
T
Naruse
T
T
D017209:Apoptosis; D002452:Cell Count / Q000187:drug effects; D002843:Chromatin / Q000187:drug effects / Q000648:ultrastructure; D004923:Erythrosine / Q000032:analysis; D005047:Etoposide / Q000494:pharmacology*; D005069:Evaluation Studies as Topic; D005434:Flow Cytometry; D005456:Fluorescent Dyes / Q000032:analysis; D018922:HL-60 Cells / Q000187:drug effects / Q000378:metabolism / Q000473:pathology / Q000648:ultrastructure*; D006801:Humans; D020287:In Situ Nick-End Labeling; D008854:Microscopy, Electron / Q000379:methods*; D016252:Microscopy, Scanning Tunneling; D011237:Predictive Value of Tests; D012680:Sensitivity and Specificity
D002843:Chromatin; D005456:Fluorescent Dyes; D005047:Etoposide; D004923:Erythrosine