PMID- 12879458
OWN - NLM
STAT- MEDLINE
DCOM- 20040210
LR  - 20161118
IS  - 1552-4922 (Print)
IS  - 1552-4922 (Linking)
VI  - 54
IP  - 2
DP  - 2003 Aug
TI  - NF-kappaB inhibitor sesquiterpene parthenolide induces concurrently atypical 
      apoptosis and cell necrosis: difficulties in identification of dead cells in such 
      cultures.
PG  - 118-24
AB  - BACKGROUND: Apoptosis and necrosis ("accidental cell death") are distinct modes 
      of cell death. The feature that often distinguishes apoptotic from necrotic cells 
      is preservation of the plasma membrane integrity, reflected by ability of the 
      former cells to exclude cationic dyes such as propidium iodide (PI) for a certain 
      length of time. During necrosis, the plasma membrane is rapidly ruptured and 
      necrotic cells stain intensely with PI. While studying cytostatic effects of the 
      anti-inflammatory sesquiterpene parthenolide (PRT), we have noticed that, 
      concurrent with apoptosis, the cells were dying by necrosis in the same cultures. 
      Furthermore, because apoptosis was atypical, reflected by rapid loss of plasma 
      membrane integrity, it was difficult to distinguish apoptotic from necrotic cells 
      based on this feature. METHODS: Three methods were used to distinguish apoptosis 
      from necrosis: (a) HL-60 cells treated with PRT were subjected to analysis of 
      caspases activation using antibody that detects activated (cleaved) caspase-3, 
      (b) apoptotic cells were identified by binding of fluorochrome-labeled inhibitor 
      of caspases FAM-VAD-FMK combined with the PI exclusion assay, and (c) cellular 
      DNA and RNA were differentially stained with acridine orange (AO). RESULTS: 
      Apoptotic cells were characterized by (a) caspase-3 activation detected 
      immunocytochemically and (b) binding of FAM-VAD-FMK followed by or concurrent 
      with (c) loss of ability to exclude PI, (d) deficit in DNA content, and (e) 
      relatively little changed RNA content. Necrotic cells showed (a) no evidence of 
      caspase-3 activation, (b) no binding of FAM-VAD-FMK, (c) inability to exclude PI, 
      (d) rapid loss of RNA, and (e) unchanged DNA content. CONCLUSIONS: Identification 
      of apoptotic cells versus necrotic cells was possible either based on the 
      evidence of caspase-3 activation, by labeling with FAM-VAD-FMK combined with PI 
      or by differential staining of cellular DNA and RNA with AO. The data indicate 
      that plasma membrane appears to be one of the targets of PRT, because its 
      integrity is lost very early during cell death, which is reflected by atypical 
      apoptosis and by primary necrosis (lysis of the membrane).
CI  - Copyright 2003 Wiley-Liss, Inc.
FAU - Pozarowski, Piotr
AU  - Pozarowski P
AD  - Brander Cancer Research Institute, New York Medical College, Valhalla, New York, 
      USA.
FAU - Halicka, Dorota H
AU  - Halicka DH
FAU - Darzynkiewicz, Zbigniew
AU  - Darzynkiewicz Z
LA  - eng
GR  - R01 CA028704/CA/NCI NIH HHS/United States
GR  - R01 CA028704-27/CA/NCI NIH HHS/United States
GR  - CA 28704/CA/NCI NIH HHS/United States
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, P.H.S.
PL  - United States
TA  - Cytometry A
JT  - Cytometry. Part A : the journal of the International Society for Analytical 
      Cytology
JID - 101235694
RN  - 0 (Anti-Inflammatory Agents)
RN  - 0 (Anti-Inflammatory Agents, Non-Steroidal)
RN  - 0 (Coloring Agents)
RN  - 0 (NF-kappa B)
RN  - 0 (Sesquiterpenes)
RN  - 2RDB26I5ZB (parthenolide)
RN  - 36015-30-2 (Propidium)
RN  - 63231-63-0 (RNA)
RN  - 9007-49-2 (DNA)
RN  - EC 3.4.22.- (CASP3 protein, human)
RN  - EC 3.4.22.- (Caspase 3)
RN  - EC 3.4.22.- (Caspases)
RN  - F30N4O6XVV (Acridine Orange)
SB  - IM
MH  - Acridine Orange/pharmacology
MH  - Anti-Inflammatory Agents/pharmacology
MH  - Anti-Inflammatory Agents, Non-Steroidal/pharmacology
MH  - *Apoptosis
MH  - Caspase 3
MH  - Caspases/metabolism
MH  - Cell Membrane/metabolism
MH  - Coloring Agents/pharmacology
MH  - DNA/chemistry/metabolism
MH  - Flow Cytometry/*methods
MH  - HL-60 Cells
MH  - Humans
MH  - Immunohistochemistry
MH  - NF-kappa B/*antagonists & inhibitors
MH  - *Necrosis
MH  - Propidium/pharmacology
MH  - RNA/chemistry/metabolism
MH  - Sesquiterpenes/*pharmacology
MH  - Time Factors
EDAT- 2003/07/25 05:00
MHDA- 2004/02/11 05:00
CRDT- 2003/07/25 05:00
PHST- 2003/07/25 05:00 [pubmed]
PHST- 2004/02/11 05:00 [medline]
PHST- 2003/07/25 05:00 [entrez]
AID - 10.1002/cyto.a.10057 [doi]
PST - ppublish
SO  - Cytometry A. 2003 Aug;54(2):118-24. doi: 10.1002/cyto.a.10057.