Potential Interactions of Herbal Extracts of St. John´s Wort with Metabolites of Diazepam in an Organotypical Sandwich Culture of Primary Porcine Hepatocytes.

Extracts of St. John’s Wort (Hypericum perforatum) are known to cause interactions with certain conventional drugs. Herein, we focus on two clinically relevant concepts. First, St. John´s Wort has been used by people of all ages as an herbal treatment for depression without medical prescription. Second, diazepam-like substances called endogenous benzodiazepines are found in cases of acute liver failure without previous exposure to diazepam. Currently diazepam has over 500 brands name and well marketed throughout the world and became one most frequently prescribed medication in different form (oral, injectable, inhalation and rectal forms) for four decades. Based on this concept, we investigated diazepam biotransformation by incubation of a widely accepted in vitro organotypical culture model with Hypericum methanolic extract, powdered drug, infusion, oil, and with the pure Hypericum compounds hyperforin and hypericin. The amounts of the preparations and compounds were chosen according to the recommended daily medication doses. We measured the activities of ethoxyresorufin-O-deethylase (EROD), ethoxy coumarin O-deethylase (ECOD) and the potential induction of diazepam metabolites (desmethyldiazepam, temazepam and oxazepam) during biotransformation. None of the preparations or substances induced EROD or ECOD. However, different preparations did induce the formation of desmethyldiazepam and temazepam. The strongest activity was caused by the extract, followed by the powdered drug and hyperforin. All preparations and compounds increased the formation of the diazepam metabolite oxazepam, but only the extract, the drug powder and the pure compounds had marked effects. Therefore, we report here the potential interference of St. John´s Wort with all three metabolites of diazepam in an organotypical sandwich model that can be utilized to study potential interaction of metabolites of many drugs with herbal ingredients in preclinical stage of drug discovery process.

ESTABLISHMENT OF METHOD OF LARGE-SCALE CRYOPRESERVATION OF PORCINE HEPATOCYTES / 解放军医学杂志

Objective To establish a method of large-scale cryopreservation of porcine hepatocytes to meet the need of the biological artificial liver. Methods Porcine hepatocytes were isolated from Chinese miniature swines by collagenase disgestion. The cells were suspended in self-made nutrient solution supplemented with 10% DMSO. The hepatocytes (1?10 10~2?10 10) were stored in liquid nitrogen (-196℃) after being treated with two kinds of freezing methods. The cryopreserved hepatocytes were thawed in 39℃ water after one month and cultured. The morphology, viability and the function of thawed hepatocytes were observed. Results The cell viability of cryobreserved hepatocytes after both freezing methods were high. However, synthesis of urea nitrogen and glucose was higher in stepwise temperature lowering group compared with programmed temperature lowering group. Conclusion The results indicate that the cryopreservation method is feasible and simple, large amount porcine hepatocytes can be cryopreserved.

Experimental study on the treatment of BioLiver Ⅰ bioartificial liver support system to two types of acute liver failure canine models / 解放军医学杂志

Objective To assess the efficacy and safety of BioLiver Ⅰ bioartificial liver support system (BALSS) on the treatment of two types of acute liver failure (ALF) canine models. Methods The drug induced ALF canine model was established by multiple subcutaneous injections of acetaminophen. Another model was operated to resect 80 percent of the liver tissue. The primary hepatocytes were separated from the swine and cultured within the BALSS. The ALF models were treated by BioLiver ⅠBALSS for 6 hours. The changes of physiological, biochemical and histological parameters were observed before and after the treatment. Results The canines developed ALF 48 hours after injections of acetaminophen, the established rate of the model was 63 3%. While the other ALF canine model developed 24 hours after 80 percent liver resection and the established rate was 84 2%. Using our modified enzymatic digestion method, the yield of hepatocytes was (1 0～3 0)?10 10 per swine with high viability. BioLiver ⅠBALSS treatment resulted in beneficial effects on blood biochemical parameters. The pathological lesions of the liver were repaired. BALSS treatment was harmless to other organs. The ALF canines in drug group survived longer than in operation group. Conclusion This type of BALSS can provide safe and efficacious liver function support in the two types of ALF canine models, and it may be as an hepaful and important therapy to ALF