[Cadmium induced apoptosis of HEK293 cells and its mitochondrial apoptosis pathway].
HEK293 cell was chose to study the kidney damage of cadmium and to explore the significance of caspase 3,Bcl-2 and AIF (apoptosis inducing factor) in the apoptosis of cells induced by cadmium. Inhibition of the cell proliferation was measured by MTT assay. The structure of apoptotic cells was observed by light microscopy and electron microscopy; moreover, apoptotic cells were detected by DNA electrophoresis, flow cytometry and confocal laser microscopy. Furthermore,the expressions of Pro-caspase-3, Bcl-2 and the location of AIF in cells (mitochondria,cytoplasm or nuclei) were tested by western blot and immunofluorescence assay. CdCl2 exhibited anti-proliferative activity in dosage and time-dependent manner. DNA ladders of HEK293 cells were showed on agarose gel electrophoresis and the fragments of DNA were integral of 180-200 bp. 6-9 hours after 30 micromol/L CdCl2 treatment,DNA ladders were distinct. However, mistiness DNA ladder or smear was found when HEK293 cells were treated with CdCl2 on higher concentration or treated longer. It suggests that necrosis may happen, and flow cytometry results confirmed it. Morphological examination showed cell shrinkage, chromosomal condensation, karyotheca margination, nucleus cracking, vacuoles formed in cytoplasm and the presence of apoptotic bodies. At the same time,mitochondrial membrane potential (MMP) decreased, and the expression of Pro-caspase-3, Bcl-2 were decreased in time-dependent manner. Furthermore, AIF was released from mitochondria,and then traveled to nuclei. It suggests that CdCl2 may induce the apoptosis of HEK293 cells involving mitochondrial disruption including AIF migration and Cyt c release through both caspase-independent and -dependent pathways, and Bcl-2 and Caspase-3 are important factors which participate in the processes.
Ye
Ji Lin
JL
College of Life Science, Nanjing Normal University, Nanjing 210097,China.
Mao
Wei Ping
WP
Wu
Ai Lian
AL
Zhao
Jie Ming
JM
Zhang
Chao
C
Zhang
Na Na
NN
Jiang
Ping
P
Tian
Ting
T
D017209:Apoptosis / Q000187:drug effects*; D002104:Cadmium / Q000494:pharmacology*; D053148:Caspase 3 / Q000378:metabolism; D002460:Cell Line; D005434:Flow Cytometry; D006801:Humans; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D018613:Microscopy, Confocal; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*
D019253:Proto-Oncogene Proteins c-bcl-2; D002104:Cadmium; D053148:Caspase 3