TY - JOUR
T1 - Evidence that photodynamic stress kills Zellweger fibroblasts by a nonapoptotic mechanism
AU - Spisni, Enzo
AU - Cavazzoni, Marika
AU - Griffoni, Cristiana
AU - Calzolari, Elisabetta
AU - Tomasi, Vittorio
PY - 1998/3/12
Y1 - 1998/3/12
N2 - Zellweger fibroblasts, which an devoid of peroxisomes and fail to synthesize plasmalogens, are very sensitive to the killing effect triggered by UV-activated 12-(1-pyrene) dodecanoic acid (P12). Although in some studies performed, it is assumed that reactive oxygen species (ROS) may damage plasma membrane causing necrosis, other studies suggest that ROS are involved in apoptotic cell death induced by a wide variety of stimuli. Analysing the P12 dose-response in Zellweger fibroblasts, we observed that at high doses (1-2 μM), more than 75% of the cells died after 24 h. This behaviour suggested that, at high doses, P12 kills the cells by unspecific lytic mechanisms or by necrosis, while at low doses (0.1-0.5 μM), an apoptotic mechanism could be involved. Cytofluorimetric analysis of Zellweger fibroblasts-treated with activated P12 (0.5 μM) did not show morphological modifications typical of apoptotic cell death. This was supported by comparative staining of fibroblast nuclei, DNA gel electrophoresis and identification of poly(ADP-ribose) polymerase (PARP) cleavage and Bcl-2 expression, assayed by Western blots. Thus, our results, while confirming the importance of plasmalogens in the protection against ROS, establish that apoptosis is not involved in photodynamic death induced by activated P12. Therefore, we can expect that in gene transfer experiments, the rescue of Zellweger cells will be dependent only on the correction of peroxisomal biogenesis.
AB - Zellweger fibroblasts, which an devoid of peroxisomes and fail to synthesize plasmalogens, are very sensitive to the killing effect triggered by UV-activated 12-(1-pyrene) dodecanoic acid (P12). Although in some studies performed, it is assumed that reactive oxygen species (ROS) may damage plasma membrane causing necrosis, other studies suggest that ROS are involved in apoptotic cell death induced by a wide variety of stimuli. Analysing the P12 dose-response in Zellweger fibroblasts, we observed that at high doses (1-2 μM), more than 75% of the cells died after 24 h. This behaviour suggested that, at high doses, P12 kills the cells by unspecific lytic mechanisms or by necrosis, while at low doses (0.1-0.5 μM), an apoptotic mechanism could be involved. Cytofluorimetric analysis of Zellweger fibroblasts-treated with activated P12 (0.5 μM) did not show morphological modifications typical of apoptotic cell death. This was supported by comparative staining of fibroblast nuclei, DNA gel electrophoresis and identification of poly(ADP-ribose) polymerase (PARP) cleavage and Bcl-2 expression, assayed by Western blots. Thus, our results, while confirming the importance of plasmalogens in the protection against ROS, establish that apoptosis is not involved in photodynamic death induced by activated P12. Therefore, we can expect that in gene transfer experiments, the rescue of Zellweger cells will be dependent only on the correction of peroxisomal biogenesis.
KW - Apoptosis
KW - Peroxisome
KW - Plasmalogen
KW - Pyrene dodecanoate
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U2 - 10.1016/S0167-4889(97)00148-1
DO - 10.1016/S0167-4889(97)00148-1
M3 - Article
C2 - 9551086
AN - SCOPUS:0032510295
VL - 1402
SP - 61
EP - 69
JO - Biochimica et Biophysica Acta - Molecular Cell Research
JF - Biochimica et Biophysica Acta - Molecular Cell Research
SN - 0167-4889
IS - 1
ER -