It has been repeatedly suspected that telomere shortening might be one possible trigger of the p53-dependent cell cycle arrest, although the mechanism of this arrest remained unclear. Telomeres in human cells under mild oxidative stress accumulate single-strand damage faster than interstitial repetitive sequences. In MRC-5 ®bro-blasts and U87 glioblastoma cells, which both express wild-type p53, oxidative stress-mediated production of single-strand damage in telomeres is concomitant to the accumulation of p53 and p21 and to cell cycle arrest. This response can be modeled by treatment of cells with short single stranded telomeric G-rich DNA fragments. The arrest is transient in U87 cells. Recovery from it is accompanied by up-regulation of telomerase activity and elongation of telomeres. Overexpression of mutated p53 is su cient to reverse the phenotype of inhibition as well as the delayed activation of telomerase. These data suggest that the production of G-rich single stranded fragments during the course of telomere shortening is su cient to trigger a p53 dependent cell cycle arrest.

The induction of an adaptive immune response to a previously unencountered pathogen is a time-consuming process and initially the infection must be held in check by the innate immune system. In the case of an i.p. infection with Salmonella typhimurium, survival requires both CD14 and LPS-binding protein (LBP) which, together with Toll-like receptor 4 and myeloid differentiation protein 2, provide a sensitive means to detect bacterial LPS. In this study, we show that in the first hours after i.p. infection with Salmonella a local inflammatory response is evident and that concomitantly neutrophils flood into the peritoneum. This rapid neutrophil influx is dependent on TNF since it is 1) abolished in TNF KO mice and 2) can be induced by i.p. injection of TNF in uninfected animals. Neutrophil influx is not strictly dependent on the presence of either LBP or CD14. However, in their absence, no local inflammatory response is evident, neutrophil migration is delayed, and the mice succumb to the infection. Using confocal microscopy, we show that the neutrophils which accumulate in CD14 and LBP null mice, albeit with delayed kinetics, are nevertheless fully capable of ingesting the bacteria. We suggest that the short delay in neutrophil influx gives the pathogen a decisive advantage in this infection model.

The effect of grapefruit juice on the urinary excretion of 7-hydroxycoumarin after oral administration of 10 mg coumarin, as an index of cytochrome P450 dependent coumarin metabolism, has been investigated in an open, randomised cross over study in 13 healthy volunteers (7 female, 6 male). The percentage of 7-hydroxycoumarin found in urine was significantly decreased up to 8 h after simultaneous intake of 300 ml grapefruit juice. If the same volume of juice was swallowed 30 min prior to the administration of coumarin, 7-hydroxycoumarin excretion was delayed by up to 6 h. MRTexcr. of coumarin was 70% extended by coadministration of grapefruit juice. It appears that grapefruit flavonoids inhibit cytochrome P450 2A dependent metabolic pathways. The mechanism of cytochrome P450 inhibition by these flavonoids is still poorly understood.

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT• The analgesic activity of tilidine is mediated by its active metabolite, nortilidine, which easily penetrates the blood-brain barrier and binds to the m-opioid receptor as a potent agonist.• Tilidine undergoes an extensive first-pass metabolism, which has been suggested to be mediated by CYP3A4 and CYP2C19; furthermore, strong inhibition of CYP3A4 and CYP2C19 by voriconazole increased exposure of nortilidine, probably by inhibition of further metabolism.• The novel CYP2C19 gene variant CYP2C19*17 causes ultrarapid drug metabolism, in contrast to the *2 and *3 variants, which result in impaired drug metabolism. WHAT THIS STUDY ADDS• Using a panel study with CYP2C19 ultrarapid and poor metabolizers, a major contribution of polymorphic CYP2C19 on tilidine metabolic elimination can be excluded.• The potent CYP3A4 inhibitor ritonavir alters the sequential metabolism of tilidine, substantially reducing the partial metabolic clearances of tilidine to nortilidine and nortilidine to bisnortilidine, which increases the nortilidine exposure twofold.• The lowest clearance in overall tilidine elimination is the N-demethylation of nortilidine to bisnortilidine. Inhibition of this step leads to accumulation of the active nortilidine. AIMSTo investigate in vivo the effect of the CYP2C19 genotype on the pharmacokinetics of tilidine and the contribution of CYP3A4 and CYP2C19 to the formation of nortilidine using potent CYP3A4 inhibition by ritonavir. METHODSFourteen healthy volunteers (seven CYP2C19 poor and seven ultrarapid metabolizers) received ritonavir orally (300 mg twice daily) for 3 days or placebo, together with a single oral dose of tilidine and naloxone (100 mg and 4 mg, respectively). Blood samples and urine were collected for 72 h. Noncompartmental analysis was performed to determine pharmacokinetic parameters of tilidine, nortilidine, bisnortilidine and ritonavir. RESULTSTilidine exposure increased sevenfold and terminal elimination half-life fivefold during ritonavir treatment, but no significant differences were observed between the CYP2C19 genotypes. During ritonavir treatment, nortilidine area under the concentration-time curve was on average doubled, with no differences between CYP2C19 poor metabolizers [2242 h ng ml -1 (95% confidence interval 1811-2674) vs. 996 h ng ml CONCLUSIONSThe sequential metabolism of tilidine is inhibited by the potent CYP3A4 inhibitor, ritonavir, independent of the CYP2C19 genotype, with a twofold increase in the exposure of the active nortilidine.