[The effect of N-acetyl-L-cysteine on endoplasmic reticulum stress mediated apoptosis of HepG2 cells].
OBJECTIVE
To analyze the mechanisms of NAC on endoplasmic reticulum (ER) stress mediated cells apoptosis of HepG2 cells and to evaluate the potential role of NAC in the treatment of liver injury.


METHODS
HepG2 cells were treated with H2O2 to make a model of oxidative ER stress mediated apoptosis. To evaluate the apoptosis, various methods such as MTT, DNA ladder, Western blot and flow cytometry were used. Then the optimal dosage and incubation time of NAC intervention in apoptosis were ascertained, and the differences between induction and intervention of apoptosis, including the percentage of apoptosis, the expression of apoptotic protein (GRP78, Caspase-12, PARP) and the production of reactive oxygen species (ROS) were compared.


RESULTS
The activity of the cells decreased by H2O2 (0, 1, 3, 5 mmol/L) treatments in a dose-dependent manner. The ratio of apoptotic cells increased (0.7%+/-0.5%, 26.4%+/-1.8%, 29.7%+/-1.2% and 51.2%+/-9.4%, respectively) as did the production of ROS (14.0%+/-0.5%, 95.2%+/-0.1%, 97.5%+/-0.2% and 98.3%+/-0.2%, respectively). The HepG2 cells showed typical morphologic change of ER stress 6 hr after they were treated with 3 mmol/L H2O2. ER stress mediated-apoptosis was confirmed by Western blot. NAC (10 mmol/L and 20 mmol/L) protected cells from apoptosis. Typical features of ER stress apoptosis were seen accompanied by diminishing the ratio of apoptotic cells from 29.7%+/-1.2% to 23.3%+/-4.7% and 14.3%+/-1.2%. The production of ROS also decreased from 97.5%+/-0.2% to 52.2%+/-0.8% and 51.2%+/-2.9%. The effect was related to the concentration: 20 mmol/L NAC was more effective than 10 mmol/L.


CONCLUSIONS
As an oxidizing agent, H2O2 may induce ROS in cells and induce oxidative stress, causing ER stress and apoptosis. NAC can inhibit the procession of ROS directly and prevent injuries to the hepatocytes.
Liu
Yun-ye
YY
Department of Infectious Diseases, Ruijin Hospital, Shanghai Jiaotong University Medical School, Shanghai 200025, China.
Xie
Qing
Q
Wang
Hui
H
Lin
Lan-yi
LY
Jiang
Shan
S
Zhou
Xia-qiu
XQ
Yu
Hong
H
Guo
Qing
Q
D000111:Acetylcysteine / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D000091342:Endoplasmic Reticulum Chaperone BiP; D056945:Hep G2 Cells; D006801:Humans; D006861:Hydrogen Peroxide; D018384:Oxidative Stress; D017382:Reactive Oxygen Species / Q000009:adverse effects
D000091342:Endoplasmic Reticulum Chaperone BiP; C000717247:HSPA5 protein, human; D017382:Reactive Oxygen Species; D006861:Hydrogen Peroxide; D000111:Acetylcysteine