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KMID : 0357920020360010001
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Korean Journal of Pathology
2002 Volume.36 No. 1 p.1 ~ p.6
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A Study of Apoptosis Induced by Microinjection of Cytochrome c Protein into Mouse 3T3 Fibroblast
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Lee Min-Sup
Kang Gu
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Abstract
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Background: Microinjectors have been used for cell biology and development, and are useful for the study of cellular morphologic changes with response to the external milieu and intracellularly injected molecules.
Methods: This study was performed to confirm the apoptotic changes induced by intracytoplasmic microlnjection of cytochrome c （5 ㎎/㎖）to mouse 3T3 fibroblastswith and without pretreatment of Ac-DEVD-CHO(100μ㏖/mL), and BSA (bovine serum albumin, 5㎎/mL) as a control, and evaluate the usefulness of microinjection as a method to study apoptosis pathways.
Results: Mild focal cytoplasmic fragmentation was seen in the cells microinjected with cytochrome c as early as 10 min after the injection. Apoptotic morphology With apoptotic boby formation was observed at 60min after the injection, and then new apoptotic change of the injected cells was not seen. Cytochrome c-injected cells showed about 31% of apoptotic cells of the totoal injected cells 50-60 min after the injetctions. Caspase-3 inhibitor, Ac-DEVD-CHO-treated cells with cytochrome c microinjection exhibited lower apoptosis indices (average apoptosis index; 11±8.6%) than non-treated cells of the inhibitor (average apoptosis index; 11.5±8.6%).
Conclusions: It was observed that intracellular microinjection of cytochromic c induced apoptosis which was inhibited by Ac-DEVD-CHO, although apoptotic cells were so easily detached that further study could not be performed. However it is thought that microinjection should be a method to study apoptosis and signal transduction with the molecular biological techniques currently availble.
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KEYWORD
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Apoptosis, Caspases, Cytochrome c, Microinjections
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