[Stimulation of human hepatic stellate cells by cytochrome P4502E1-mediated oxidative stress].
OBJECTIVE
To explore the stimulation of human hepatic stellate cells by Cytochrome P4502E1-mediated oxidative stress.


METHODS
HepG2-line was transfected with human CYP2E1 plasmid (HepG2/CYP2E1) and empty plasmid (HepG2/PCI) respectively. The CYP2E1 expression was evaluated with RT-PCR and Western blot. MDA was measured in culture medium of HepG2 cell lines. LX2 was co-incubated with HepG2/CYP2E1, HepG2/PCI and HepG2 respectively. The level of hydroxyproline in culture medium was examined in 48 hours and the cells were lysated and total RNA and protein were extracted. COL-1 and MMP2 mRNA levels were detected by RT-PCR and analyzed semi-quantitatively. PICP proteins were measured by ELISA. Zymography was performed to investigate MMP2 enzymatic activities.


RESULTS
(1) MDA from the HepG2 which (HepG2/CYP2E1)express human CYP2E1 (6.51+/-0.25) was significantly higher than that from the HepG2 which do not (HepG2/PCI) express human CYP2E1 (3.07+/-0.29) and HepG2 alone (2.57+/-0.29). (F=22.66, all P<0.01). (2) After co-incubated for 48 hours,the level of hydroxyproline in culture medium (35.24+/-3.52) excreted from CYP2E1/LX2 could significantly increase (F=58.89, P is less than 0.01). PICP protein (540.01+/-11.38) excreted from CYP2E1/LX2 was significantly increased (F=124.97, P<0.01). Zymography showed MMP2 gene expression and enzymatic activities of MMP2 had no difference among the groups (F=0.29, P>0.05) (F=0.33, P>0.05).


CONCLUSIONS
CYP2E1 derived oxidative stress mediated stimulation of collagen I synthesis by hepatic stellate cells. Hydroxyproline excreted by LX2 was increased by CYP2E1. COL-1mRNA had no difference among the groups (F=0.73, P>0.05).
Li
Jing
J
Department of Internal Medicine, Beijing Tiantan Hospital, Capital Medical University, Beijing 100050, China.
Liu
Tian-hui
TH
You
Hong
H
Xu
You-qing
YQ
Wang
Chen
C
D024042:Collagen Type I / Q000096:biosynthesis; D019392:Cytochrome P-450 CYP2E1 / Q000378:metabolism*; D056945:Hep G2 Cells; D055166:Hepatic Stellate Cells / Q000378:metabolism*; D006801:Humans; D006909:Hydroxyproline / Q000378:metabolism; D008103:Liver Cirrhosis / Q000378:metabolism*; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D018384:Oxidative Stress
D024042:Collagen Type I; D019392:Cytochrome P-450 CYP2E1; C522361:MMP2 protein, human; D020778:Matrix Metalloproteinase 2; D006909:Hydroxyproline