CYP2E1-dependent hepatotoxicity and oxidative damage after ethanol administration in human primary hepatocytes

AIM： To observe the relationship between ethanol-induced oxidative damage in human primary cultured hepatocytes and cytochrome P450 2E1 （CYP2E1） activity, in order to address if inhibition of CYP2E1 could attenuate ethanol- induced cellular damage. METHODS： The dose-dependent （25-100 mmol/L） and time-dependent （0-24 h） exposures of primary human cultured hepatocytes to ethanol were carried out. CYP2E1 activity and protein expression were detected by spectrophotometer and Western blot analysis respectively. Hepatotoxicity was investigated by determination of lactate dehydrogenase （LDH） and aspartate transaminase （AST） level in hepatocyte culture supernatants, as well as the intracellular formation of malondialdehyde （MDA）. RESULTS： A dose-and time-dependent response between ethanol exposure and CYP2E1 activity in human hepatocytes was demonstrated. Moreover, there was a time-dependent increase of CYP2E1 protein after 100 mmol/L ethanol exposure. Meanwhile, ethanol exposure of hepatocytes caused a time-dependent increase of cellular MDA level, LDH, and AST activities in supernatants. Furthermore, the inhibitor of CYP2E1, diallyl sulfide （DAS） could partly attenuate the increases of MDA, LDH, and AST in human hepatocytes. CONCLUSION： A positive relationship between ethanolinduced oxidative damage in human primary cultured hepatocytes and CYP2E1 activity was exhibited, and the inhibition of CYP2E1 could partly attenuate ethanol-induced oxidative damage.

Acetyl-L-carnitine:An Effective Antioxidant against Cryo-damage on Human Spermatozoa with Asthenospermia

A variety of natural and artificial cryoprotectant extenders have been explored to enhance sperm recovery following cryopreservation-thawing process. The current investigation is aimed at evaluating the effect of acetyl-L-carnitine on human spermatozoa and reactive species oxygen（ROS） level after freezing-thawing process. The spermatozoa were collected from 35 male patients diagnosed as having asthenospermia. The cryopreservation of human spermatozoa treated with acetyl-L-carnitine at different concentrations（group B： 2.5 mmol/L, group C： 7.5 mmol/L, group D： 15 mmol/L） was compared with control（group A： no acetyl-L-carnitine given）. For the frozen-thawed spermatozoa, the viability, motility and DNA integrity were measured by comet assay, acrosome integrity by FITC-PNA staining and ROS level was determined in each group. The results showed that there were no significant differences in motility and viability between group A and group B, while the motility and viability of spermatozoa in group C and group D were significantly increased as compared with those in group A. As compared with group A, the values for DNA integrity parameters including comet rate（CR）, tail DNA percentage（TD）, tail length（TL） and Oliver tail moment（OTM） were significantly reduced in group C and group D. Group C and group D also displayed a higher proportion of intact acrosome than group A. No significant difference in ROS level was found between group A and group B, while with the increase in acetyl-L-carnitine concentration, the ROS level in groups C and D was significantly reduced as compared with that in group A. In conclusion, acetyl-L-carnitine at a concentration of 7.5 mmol/L is an effective antioxidant against cryo-damage on post-thawed human spermatozoa.

Hexabromocyclododecane-induced Genotoxicity in Cultured Human Breast Cells through DNA Damage

To investigate the genotoxicity and reveal the potential toxicological mechanisms of Hexabromocyclododecane （HBCD）, human breast cells HBL-100 were exposed to a sequence of HBCD concentrations （0, 5, 10, and 50 mg/L） for 24 h. With a series of zymology and molecular biology methods, we found that HBCD induced dose-dependent oxidative stress on HBL-100 DNA. As revealed in q RT-PCR, activated prognostic factor ATM down-regulated tumor suppressor gene BRCA1 and prompted DNA repair genes h OGG1 and h MTH1 expression in lower concentrations of HBCD （〈 10 mg/L）. However, DNA repair were inhibited as well as cell proliferation rate by higher concentrations of HBCD （50 mg/L）. The results inferred that the genotoxicity of HBCD was dose-dependent and related to DNA repair pathway.