[Waiting List Management and Organ Allocation for Lung Transplantation 2018]. / Wartelistenführung und Organvermittlung zur Lungentransplantation 2018 ­ Aktuelles für den Pneumologen.

In 2017 an amendment to the German transplant law concerning organ allocation and waiting list management became effective. This implies important consequences on lung transplant centers. Crucial innovations concern the transplant conference, indications for lung transplantation and waiting list management. Certain medical conditions now imply a restriction of waiting list enrollment for patients and there are new options for size-matching of donor lung and recipient. Moreover, the new amendment describes in detail how the clinical parameters, on which the lung allocation score (LAS) is based, are defined and how the essential physical examinations have to be performed. Furthermore, the current article provides a summary of the process of organ allocation by the organ exchange organization.

Skin metabolism of aminophenols: human keratinocytes as a suitable in vitro model to qualitatively predict the dermal transformation of 4-amino-2-hydroxytoluene in vivo.

4-Amino-2-hydroxytolune (AHT) is an aromatic amine ingredient in oxidative hair colouring products. As skin contact occurs during hair dyeing, characterisation of dermal metabolism is important for the safety assessment of this chemical class. We have compared the metabolism of AHT in the human keratinocyte cell line HaCaT with that observed ex-vivo in human skin and in vivo (topical application versus oral (p.o.) and intravenous (i.v.) route). Three major metabolites of AHT were excreted, i.e. N-acetyl-AHT, AHT-sulfate and AHT-glucuronide. When 12.5 mg/kg AHT was applied topically, the relative amounts of each metabolite were altered such that N-acetyl-AHT product was the major metabolite (66% of the dose in comparison with 37% and 32% of the same applied dose after i.v. and p.o. administration, respectively). N-acetylated products were the only metabolites detected in HaCaT cells and ex-vivo whole human skin discs for AHT and p-aminophenol (PAP), an aromatic amine known to undergo N-acetylation in vivo. Since N-acetyltransferase 1 (NAT1) is the responsible enzyme, kinetics of AHT was further compared to the standard NAT1 substrate p-aminobenzoic acid (PABA) in the HaCaT model revealing similar values for K(m) and V(max). In conclusion NAT1 dependent dermal N-acetylation of AHT represents a 'first-pass' metabolism effect in the skin prior to entering the systemic circulation. Since the HaCaT cell model represents a suitable in vitro assay for addressing the qualitative contribution of the skin to the metabolism of topically-applied aromatic amines it may contribute to a reduction in animal testing.

Alcohol and a high-fat diet: a combination favoring overfeeding.

The effects of alcohol and dietary fat on spontaneous energy and macronutrient intakes were investigated in eight male subjects who participated in a protocol including four randomly assigned 2-d sessions during which they ate ad libitum. In each session they had free access to either high- or low-fat foods, with alcohol or a placebo. The high-fat diet was associated with a substantial increase in daily energy intake. Alcohol had no inhibitory effect on food intake and its energy content was thus associated with an additional increase in energy intake. The enhancing effects of alcohol and dietary fat on daily energy intake were additive so that overfeeding was maximal (2.8 MJ/d) under the high-fat diet+alcohol condition. To further examine the effects of alcohol on energy and macronutrient intakes, reported food intake was studied in 351 men and 360 women who had been tested in the Québec Family Study. The results showed that a high alcohol intake was associated with a high daily energy intake and had no inhibitory effect on lipid intake. In conclusion, a dietary regimen providing a high fraction of energy as alcohol and fat increases the risk for positive energy balance under free-living conditions.

With medium-chain triglycerides, higher and faster oxygen radical production by stimulated polymorphonuclear leukocytes occurs.

BACKGROUND: Parenteral lipid emulsions are suspected of suppressing the immune function. However, study results are contradictory and mainly concern the conventional long-chain triglyceride emulsions. METHODS: Polymorphonuclear leukocytes were preincubated with parenteral lipid emulsions. The influence of the lipid emulsions on the production of oxygen radicals by these stimulated leukocytes was studied by measuring chemiluminescence. Three different parenteral lipid emulsions were tested: long-chain triglycerides, a physical mixture of medium- and long-chain triglycerides, and structured triglycerides. Structured triglycerides consist of triglycerides where the medium- and long-chain fatty acids are attached to the same glycerol molecule. RESULTS: Stimulated polymorphonuclear leukocytes preincubated with the physical mixture of medium- and long-chain triglycerides showed higher levels of oxygen radicals (p < .005) and faster production of oxygen radicals (p < .005) compared with polymorphonuclear leukocytes preincubated with long-chain triglycerides or structured triglycerides. Additional studies indicated that differences in results of various lipid emulsions were not caused by differences in emulsifier. The overall production of oxygen radicals was significantly lower after preincubation with the three lipid emulsions compared with controls without lipid emulsion. CONCLUSIONS: A physical mixture of medium- and long-chain triglycerides induced faster production of oxygen radicals, resulting in higher levels of oxygen radicals, compared with long-chain triglycerides or structured triglycerides. This can be detrimental in cases where oxygen radicals play either a pathogenic role or a beneficial one, such as when rapid phagocytosis and killing of bacteria is needed. The observed lower production of oxygen radicals by polymorphonuclear leukocytes in the presence of parenteral lipid emulsions may result in immunosuppression by these lipids.

Metabolic capacity and interindividual variation in toxicokinetics of styrene in volunteers.

The aim of the present study was to assess the interindividual variation in styrene toxicokinetics and to correlate this variation with the individual metabolic capacity for cytochrome P450 (CYP), CYP2E1, CYP1A2 and CYP2D6. Twenty male volunteers were exposed on separate occasions to 104+/-3 and 360+/-20 mg/m3 of styrene for 1 h while performing 50 W physical exercise on a bicycle ergometer. Styrene concentrations in blood and mandelic (MA) and phenylglyoxylic acid (PGA) in urine were measured. The metabolic capacity was assessed by phenotyping with chlorzoxazone (CYP2E1), caffeine (CYP1A2), dextromethorphan (CYP2D6) and antipyrine (CYP450). In addition, for the main styrene-metabolising enzyme, CYP2E1, genotyping for the genetic polymorphisms of the gene was performed. The average pulmonary retention of styrene was 62 +/- 7% at both exposure concentrations, and the 24-h excretion of MA and PGA accounted for 58% of the dose at both concentrations. The interindividual variation in styrene kinetics ranged from 19% for the terminal half-life (t(1/2,beta)) of styrene to 41% for the cumulative excretion of MA and PGA. However, no correlation between the apparent blood clearance of styrene (CLapp), t(1/2,beta) of styrene or excretion of MA and PGA on one hand, and the individual metabolic capacity on the other hand was found. Although other explanations cannot be excluded, this lack of correlation might be due to the high apparent blood clearance (1.4 l/min) of styrene, indicating that styrene metabolism is liver-blood-flow-dependent.

Determination of mandelic acid enantiomers in urine by gas chromatography and electron-capture or flame ionisation detection.

A sensitive and stereospecific GC method was developed for the analysis of R- and S-enantiomers of mandelic acid (MA) in urine, using a chiral CP Chirasil-Dex-CB column. The enantiomers of MA were derivatised with isopropanol into their corresponding isopropyl esters and determined either directly with flame ionisation detection (FID) or after subsequent derivatisation of a hydroxy group with pentafluoropropionic anhydride with electron-capture detection (ECD). Both derivatisation steps proceeded with negligible inversion of enantiomers (<1%). The limit of detection of the FID determination was 8 and 5 mg/l for R-MA and S-MA, respectively and of the ECD determination 1 mg/l for both enantiomers. Repeatability (within-day precision) and reproducibility (day-to-day precision) was for both enantiomers below 7.5% for the FID and below 5.8% for the ECD analysis. The method was applied to urine of volunteers exposed to 105 and 420 mg styrene/m3 air. In the urine of the exposed volunteers, the S-enantiomer showed higher excretion compared to that of the R-enantiomer, with marked interindividual differences in excretion of both enantiomers.

Metabolism of styrene-7,8-oxide in human liver in vitro: interindividual variation and stereochemistry.

Styrene is an industrial solvent which is mainly oxidized by cytochrome P450 to an electrophilic, chiral epoxide metabolite: styrene-7,8-oxide (SO). SO has cytotoxic and genotoxic properties; the (R)-enantiomer is more mutagenic to Salmonella typhimurium TA 100 in the Ames test than the (S)-enantiomer. Detoxication proceeds via microsomal epoxide hydrolase (mEH). Interindividual differences in mEH activity as well as differences in mEH enantioselectivity are important factors for toxic effects of SO. To study the extent of the interindividual variation, microsomal preparations of 20 human livers were incubated with (R)- and (S)-SO separately (1-2000 microM) and Michaelis-Menten kinetics were determined. In addition, samples were genotyped for two genetic polymorphisms of the mEH gene. V(max), K(m) and V(max)/K(m) values of both enantiomers differed three- to fivefold between the livers. No association of the enzyme constants with the genetic polymorphisms of the epoxide hydrolase gene was found. Hydrolysis of the styrene oxide enantiomers proceeded in an enantioselective manner, with the (S)-enantiomer having an approximately six times higher K(m) and five times higher V(max) than the (R)-enantiomer. In vivo, both SO enantiomers are formed; therefore, time course incubations with racemic SO were carried out in vitro to investigate possible interactions between the enantiomers. When racemic SO was used as a substrate, the (R)-enantiomer acted as an inhibitor on the hydrolysis of the (S)-enantiomer. These results indicate that mEH-mediated hydrolysis of SO is subject to appreciable interindividual variation and that hydrolysis of the more toxic enantiomer is favored.

Stereochemical metabolism of styrene in volunteers.

OBJECTIVES: To study the stereochemistry of styrene metabolism in volunteers, and its interindividual variability. METHODS: Twenty healthy male volunteers (aged 18-37 years) were exposed to 360 mg/m3 styrene for 1 h while they performed 50 W physical exercise. Venous blood was drawn during and for up to 2 h after exposure. Urine was collected at time-intervals up to 24 h after exposure. The following parameters were determined: styrene, free and conjugated styrene glycol (SG) in blood, and conjugated SG, mandelic acid (MA) and phenylglyoxylic acid (PGA) in urine. RESULTS: Average pulmonary retention of styrene was 62%. Excretion of the acidic metabolites MA and PGA accounted for 58% of the pulmonary uptake. The average maximum concentration (Cmax) and area under the curve (AUC) of free (R)-SG in blood were 1.3 and 1.7 times higher than those of (S)-SG respectively; the half-life of (R)-SG was longer (82 vs 62 min, P < 0.005). Cmax and AUC of the conjugated SG enantiomers in blood did not differ, but again half-life for (R)-SG was longer (72 vs 64 min, P < 0.05). Cumulative excretion and renal clearance of conjugated (S)-SG in urine were three and four times higher, respectively, than that of (R)-SG. Cumulative excretion of (S)-MA was 1.6 times higher than (R)-MA. Interindividual differences in the kinetic parameters of the metabolites were two- to threefold. CONCLUSIONS: The enantiomeric excess found was different for each metabolite under study, implying different enantioselectivity and/or enantiospecificity of the enzymes and carrier-proteins involved in the biotransformation and excretion. The use of these metabolites as biological indicators for prediction of the enantiomeric excess of the toxic metabolite styrene-7,8-oxide (SO) is therefore not justified. Interindividual differences in the stereochemical metabolism of styrene are moderate.

Metabolism of styrene in the human liver in vitro: interindividual variation and enantioselectivity.

1. The interindividual variation and enantioselectivity of the in vitro styrene oxidation by cytochrome P450 have been investigated in 20 human microsomal liver samples. Liver samples were genotyped for the CYP2E1*6 and CYP2E1*5B alleles. 2. Kinetic analysis indicated the presence of at least two forms of styrene-metabolizing cytochrome P450. The enzyme constants for the high-affinity component were subject to appreciable interindividual variation, i.e. Vmax1 ranged from 0.39 to 3.20 nmol mg protein(-1) min(-1) (0.96+/-0.63) and Km1 ranged from 0.005 to 0.03 mM (0.011+/-0.006). Inhibition studies with chemical inhibitors of CYP2E1, CYP1A2, CYP2C8/9 and CYP3A4 demonstrated that CYP2E1 was the primary enzyme involved in the high-affinity component of styrene oxidation. No relationship between the interindividual variation in Vmax1 and Km1 and the genetic polymorphisms of the CYP2E1 gene was found. 3. Cytochrome P450-mediated oxidation of styrene demonstrated a moderate enantioselectivity, with an enantiomeric excess (ee) of (S)-styrene oxide of 15% (range 4-27%) at low styrene concentration and an ee of (R)-styrene oxide of 7% (range -11 to +22%) at high styrene concentration. This points towards the involvement of at least two cytochrome P450, with different enantioselectivities. 4. The data indicate that cytochrome P450-mediated styrene oxidation is subject to considerable interindividual variation, but only to a moderate product enantioselectivity.

Gas chromatography-electron capture determination of styrene-7,8-oxide enantiomers.

The enantiomers of styrene-7,8-oxide (phenyloxirane, SO) were determined using a method based on base catalysed hydrolysis with sodium methoxide. The oxirane ring opening resulted in formation, without racemisation, of the enantiomeric pairs of the two regional isomers, 2-methoxy-1-phenylethanol and 2-methoxy-2-phenylethanol. The structure of these regional isomers was confirmed by gas chromatography-mass spectrometry (GC-MS) and proton nuclear magnetic resonance (1H-NMR). To improve sensitivity of determination, the formed methoxy alcohols were subsequently derivatised with pentafluoropropionic anhydride enabling electron capture detection. This derivatization proceeded also without racemisation and the formed pentafluoropropionyl derivatives were separated on two serially coupled columns, a non-chiral AT 1705 and a chiral CP Chirasil-Dex-CB. As internal standard 2S,3S-(-)-2-methyl-3-phenyloxirane was used. The limit of quantitation of the method was 0.2 microM. The repeatability of the method was assessed at two concentration levels (2.5 and 25 microM) and ranged from 6 to 9% for both enantiomers. The method was applied to the determination of the rate and enantioselectivity of the cytochrome P-450 dependent oxidation of styrene to SO enantiomers in human liver microsomes.