Baicalein-induced apoptosis via endoplasmic reticulum stress through elevations of reactive oxygen species and mitochondria dependent pathway in mouse-rat hybrid retina ganglion cells (N18).
Studies were designed to investigate the effects of baicalein on mouse-rat hybrid retina ganglion cells (N18) to better understand its effect on apoptosis and apoptosis-related genes in vitro. Cell viability, reactive oxygen species (ROS), cytoplasmic Ca2+, mitochondrial membrane potential (MMP), apoptosis induction, and caspases-3 activity were examined by flow cytometric assay. Apoptosis-associated proteins such as p53, Bax, Bcl-2, cytochrome c, and caspase-3 were examined by Western blot. We demonstrated the increase in the levels of p53, Bax, and cytochrome c and decrease in the level of Bcl-2, which are associated with the induction of apoptotic cell death after 24 h treatment with baicalein in N18 cells. Baicalein induced an increase in the cytoplasmic levels of ROS and Ca2+ in 1 h and reached their peak at 3 h, and thereafter a loss of MMP by flow cytometry. We also demonstrated a release of the cytochrome c from mitochondria into cytosol and an activation of caspase-3, which led to the occurrence of apoptosis in N18 cells treated with baicalein by Western blot. Pretreatment was conducted with BAPTA (intracellular calcium chelator) in baicalein-treated cells, the decline of MMP was recovered, and the increase in the level of cytoplasmic Ca2+ was suppressed, and the proportion of apoptosis was also markedly diminished. In conclusion, our data suggests that oxidative stress and cellular Ca2+ modulates the baicalein-induced cell death via a Ca2+-dependent mitochondrial death pathway in N18 cells.
Li
Yu-Ching
YC
Department of Medical Laboratory Science and Biotechnology, Central Taiwan University of Science and Technology, Taichung, Taiwan, ROC.
Lin
Hui-Ju
HJ
Yang
Jen-Hung
JH
Yang
Jai-Sing
JS
Ho
Heng-Chien
HC
Chang
Shu-Jen
SJ
Hsai
Te-Chun
TC
Lu
Hsu-Feng
HF
Huang
An-Cheng
AC
Chung
Jing-Gung
JG
D000818:Animals; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000494:pharmacology*; D017209:Apoptosis; D002460:Cell Line; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D044950:Flavanones / Q000494:pharmacology*; D006822:Hybrid Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D051379:Mice; D008928:Mitochondria / Q000502:physiology*; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism*; D012165:Retinal Ganglion Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D015398:Signal Transduction
D000894:Anti-Inflammatory Agents, Non-Steroidal; D044950:Flavanones; D017382:Reactive Oxygen Species; C006680:baicalein