Biochemical mechanisms in drug-induced liver injury:Certainties and doubts

Drug-induced liver injury is a significant and still unresolved clinical problem.Limitations to knowledge about the mechanisms of toxicity render incomplete the detection of hepatotoxic potential during preclinical development.Several xenobiotics are lipophilic substances and their transformation into hydrophilic compounds by the cytochrome P-450 system results in production of toxic metabolites.Aging,preexisting liver disease,enzyme induction or inhibition,genetic variances,local O2 supply and,above all,the intrinsic molecular properties of the drug may affect this process.Necrotic death follows antioxidant consumption and oxidation of intracellular proteins,which determine increased permeability of mitochondrial membranes,loss of potential,decreased ATP synthesis,inhibition of Ca2+-dependent ATPase,reduced capability to sequester Ca2+ within mitochondria,and membrane bleb formation.Conversely,activation of nucleases and energetic participation of mitochondria are the main intracellular mechanisms that lead to apoptosis.Non-parenchymal hepatic cells are inducers of hepatocellular injury and targets for damage.Activation of the immune system promotes idiosyncratic reactions that result in hepatic necrosis or cholestasis,in which different HLA genotypes might play a major role.This review focuses on current knowledge of the mechanisms of drug-induced liver injury and recent advances on newly discovered mechanisms of liver damage.Future perspectives including new frontiers for research are discussed.

Silybin counteracts lipid excess and oxidative stress in cultured steatotic hepatic cells

AIM: To investigate in vitro the therapeutic effect and mechanisms of silybin in a cellular model of hepatic steatosis.METHODS: Rat hepatoma Fa O cells were loaded with lipids by exposure to 0.75 mmol/L oleate/palmitate for 3 h to mimic liver steatosis. Then, the steatotic cells were incubated for 24 h with different concentrations(25 to 100 μmol/L) of silybin as phytosome complex with vitamin E. The effects of silybin on lipid accumulation and metabolism, and on indices of oxidative stress were evaluated by absorption and fluorescence microscopy, quantitative real-time PCR, Western blot, spectrophotometric and fluorimetric assays.RESULTS: Lipid-loading resulted in intracellular triglyceride(TG) accumulation inside lipid droplets, whose number and size increased. TG accumulation was mediated by increased levels of peroxisome proliferator-activated receptors(PPARs) and sterol regulatory element-binding protein-1c(SREBP-1c). The lipid imbalance was associated with higher production of reactive oxygen species(ROS) resulting in increased lipid peroxidation, stimulation of catalase activity and activation of nuclear factor kappa-B(NF-κB). Incubation of steatotic cells with silybin 50 μmol/L significantly reduced TG accumulation likely by promoting lipid catabolism and by inhibiting lipogenic pathways, as suggested by the changes in carnitine palmitoyltransferase 1(CPT-1), PPAR and SREBP-1c levels. The reduction in fat accumulation exerted by silybin in the steatotic cells was associated with the improvement of the oxidative imbalance caused by lipid excess as demonstrated by the reduction in ROS content, lipid peroxidation, catalase activity and NF-κB activation.CONCLUSION: We demonstrated the direct antisteatotic and anti-oxidant effects of silybin in steatotic cells, thus elucidating at a cellular level the encouraging results demonstrated in clinical and animal studies.

Therapy of gallstone disease:What it was,what it is,what it will be

Cholesterol gallstone disease is a common clinical condition influenced by genetic factors,increasing age,female gender,and metabolic factors.Although laparoscopic cholecystectomy is currently considered the gold standard in treating patients with symptomatic gallstones,new perspectives regarding medical therapy of cholelithiasis are currently under discussion,also taking into account the pathogenesis of gallstones,the natural history of the disease and the analysis of the overall costs of therapy.A careful selection of patients may lead to successful nonsurgical therapy in symptomatic subjects with a functioning gallbladder harboring small radiolucent stones.The classical oral litholysis by ursodeoxycholic acid has been recently paralleled by new experimental observations,suggesting that cholesterol-lowering agents which inhibit cholesterol synthesis (statins) or intestinal cholesterol absorption (ezetimibe),or drugs acting on specific nuclear receptors involved in cholesterol and bile acid homeostasis,might be proposed as additional approaches for treating cholesterol gallstones.In this review we discuss old,recent and future perspectives on medical treatment of cholesterol cholelithiasis.

A silybin-phospholipids complex counteracts rat fatty liver degeneration and mitochondrial oxidative changes

AIM:To investigate the effectiveness of antioxidant compounds in modulating mitochondrial oxidative alterations and lipids accumulation in fatty hepatocytes.METHODS:Silybin-phospholipid complex containing vitamin E(Realsil) was daily administered by gavage(one pouch diluted in 3 mL of water and containing 15 mg vitamin E and 47 mg silybin complexed with phospholipids) to rats fed a choline-deprived(CD) or a high fat diet [20% fat,containing 71% total calories as fat,11% as carbohydrate,and 18% as protein,high fat diet(HFD)] for 30 d and 60 d,respectively.The control group was fed a normal semi-purified diet containing adequate levels of choline(35% total calories as fat,47% as carbohydrate,and 18% as protein).Circulating and hepatic redox active and nitrogen regulating molecules(thioredoxin,glutathione,glutathione peroxidase),NO metabolites(nitrosothiols,nitrotyrosine),lipid peroxides [malondialdehyde-thiobarbituric(MDA-TBA)],and pro-inflammatory keratins(K-18) were measured on days 0,7,14,30,and 60.Mitochondrial respiratory chain proteins and the extent of hepatic fatty infiltration were evaluated.RESULTS:Both diet regimens produced liver steatosis(50% and 25% of liver slices with CD and HFD,respectively) with no signs of necro-inflammation:fat infiltration ranged from large droplets at day 14 to disseminated and confluent vacuoles resulting in microvesicular steatosis at day 30(CD) and day 60(HFD).In plasma,thioredoxin and nitrosothiols were not significantly changed,while MDA-TBA,nitrotyrosine(from 6 ± 1 nmol/L to 14 ± 3 nmol/L day 30 CD,P < 0.001,and 12 ± 2 nmol/L day 60 HFD,P < 0.001),and K-18(from 198 ± 20 to 289 ± 21 U/L day 30 CD,P < 0.001,and 242 ± 23 U/L day 60 HFD,P < 0.001) levels increased significantly with ongoing steatosis.In the liver,glutathione was decreased(from 34.0 ± 1.3 to 25.3 ± 1.2 nmol/mg prot day 30 CD,P < 0.001,and 22.4 ± 2.4 nmol/mg prot day 60 HFD,P < 0.001),while thioredoxin and glutathione peroxidase were initially increased and then decreased.Nitrosothiols were constantly increased.MDA-TBA levels were five-fold increased from 9.1 ± 1.2 nmol/g to 75.6 ± 5.4 nmol/g on day 30,P < 0.001(CD) and doubled with HFD on day 60.Realsil administration significantly lowered the extent of fat infiltration,maintained liver glutathione levels during the first half period,and halved its decrease during the second half.Also,Realsil modulated thioredoxin changes and the production of NO derivatives and significantly lowered MDA-TBA levels both in liver(from 73.6 ± 5.4 to 57.2 ± 6.3 nmol/g day 30 CD,P < 0.01 and from 27.3 ± 2.1 nmol/g to 20.5 ± 2.2 nmol/g day 60 HFD,P < 0.01) and in plasma.Changes in mitochondrial respiratory complexes were also attenuated by Realsil in HFD rats with a major protective effect on Complex Ⅱ subunit CII-30.CONCLUSION:Realsil administration effectively contrasts hepatocyte fat deposition,NO derivatives formation,and mitochondrial alterations,allowing the liver to maintain a better glutathione and thioredoxin antioxidant activity.