Sex Difference In Metabolism Research Articles

Evidence for an accumulation of mineral oil saturated hydrocarbons (MOSH) in human tissues: a re-evaluation of biopsy and autopsy data

AbstractConsumers are exposed to mineral oil hydrocarbons (MOH) e.g. through foodstuffs and cosmetics. Upon ingestion, MOH follow the absorption pathway of dietary lipids. Analytical chemistry has revealed the presence of the main fraction, designated as mineral oil saturated hydrocarbons (MOSH), in mesenteric lymph nodes (MLNs), liver, spleen, and adipose tissue. Recent results from animal studies raised concerns about a long-term, possibly irreversible accumulation of some MOSH in humans. To address this issue, we performed a statistical re-analysis of published biopsy and autopsy data regarding the age-dependence of MOSH levels in human tissue. MOSH concentrations in MLNs and adipose tissue showed a 1.2–1.4-fold increase per decade, pointing to very long-term accumulation in both tissues. There was no evidence for age-dependent MOSH concentrations in liver and spleen. There was no sex difference in the MOSH concentrations in MLNs, suggesting a similar oral exposure for men and women. On average, women had a 2.2–2.5-fold higher MOSH concentration in the liver, spleen and adipose tissue compared to men. This finding may point to a sex difference in metabolism, in line with animal data. The use of certain cosmetics was a relevant predictor in addition to age. Women that used cosmetics like lipstick, hand cream, and sun cream had an average 2.1-fold higher MOSH concentration in abdominal subcutaneous fat than non-users.

HPLC–NMR with severe column overloading: Fast-track metabolite identification in urine and bile samples from rat and dog treated with [ 14C]-ZD6126

The subject of this study was the determination of the major urinary and biliary metabolites of [ 14C]-ZD6126 following i.v. administration to female and male bile duct cannulated rats at 10 mg/kg and 20 mg/kg, respectively, and male bile duct cannulated dogs at 6 mg/kg by HPLC–NMR spectroscopy. ZD6126 is a phosphorylated pro-drug, which is rapidly hydrolysed to the active metabolite, ZD6126 phenol. The results presented here demonstrate that [ 14C]-ZD6126 phenol is subsequently metabolised extensively by male dogs and both, male and female rats. Recovery of the dose in bile and urine was determined utilising the radiolabel, revealing biliary excretion as the major route of excretion (93%) in dog, with the majority of the radioactivity recovered in both biofluids in the first 6 h. In the rat, greater than 92% recovery was obtained within the first 24 h. The major route of excretion was via the bile 51–93% within the first 12 h. The administered phosphorylated pro-drug was not observed in any of the excreta samples. Metabolite profiles of bile and urine samples were determined by high performance liquid chromatography with radiochemical detection (HPLC–RAD), which revealed a number of radiolabelled components in each of the biofluids. The individual metabolites were subsequently identified by HPLC–NMR spectroscopy and HPLC–MS. In the male dog, the major component in urine and bile was the [ 14C]-ZD6126 phenol glucuronide, which accounted for 3% and 77% of the dose, respectively. [ 14C]-ZD6126 phenol was observed in urine at 1% of dose, but was not observed in bile. A sulphate conjugate of demethylated [ 14C]-ZD6126 phenol was identified in bile by HPLC–NMR and confirmed by HPLC–MS. In the rat, the bile contained two major radiolabelled components. One was identified as the [ 14C]-ZD6126 phenol glucuronide, the other as a glucuronide conjugate of demethylated [ 14C]-ZD6126 phenol. However, a marked difference in the proportions of these two components was observed between male and female rats, either due to a sex difference in metabolism or a difference in dose level. The glucuronide conjugate of the demethylated [ 14C]-ZD6126 phenol was present at higher concentration in the bile of male rats (4–34%), while the phenol glucuronide was present at higher concentration in the bile of female rats (8–70%) over a 0–6 h collection period. A third component was only observed in the bile samples (0–6 h and 6–12 h) of male rats. This was identified as being the same sulphate conjugate of demethylated [ 14C]-ZD6126 phenol as the one observed in dog bile. The rat urines contained two main metabolites in greatly varying concentrations, namely the demethylated [ 14C]-ZD6126 phenol glucuronide and the glucuronide of [ 14C]-ZD6126 phenol. Again, the differences in relative amounts between male and female rats were observed, the major metabolite in the urines from male rats being the demethylated [ 14C]-ZD6126 phenol (0–17% in 0–24 h), whilst the phenol glucuronide, accounting for 0.5–50% of the dose over 0–24 h, was the major metabolite in females. Methanolic extracts of the pooled biofluid samples were submitted for HPLC–NMR for the quick identification of the major metabolites. Following a single injection of the equivalent of 6–28 ml of the biofluids directly onto the HPLC-column with minimal sample preparation, the metabolites could be largely successfully isolated. Despite severe column overloading, the major metabolites of [ 14C]-ZD6126 could be positively identified, and the results are presented in this paper.

Exercising women throw a wrench in the gears of the AMPK–lipid oxidation link

Several studies, including one in this issue of The Journal of Physiology (Roepstorff et al. 2006), have found that women oxidize more lipid than men during endurance exercise (Friedlander et al. 1999; Carter et al. 2001). The carefully controlled study reported in this issue (Roepstorff et al. 2006) evaluated AMPK signalling as a mechanism for a sex difference in metabolism by evaluating the potential role of AMPK in the regulation of skeletal muscle fat oxidation during exercise. It has been suggested that AMPK is a skeletal muscle energy sensor responding to elevated free AMP content during muscle contraction, which, in turn, inactivates acetyl CoA carboxylase, decreases malonyl CoA and removes its inhibitory effect on CPT1 activity (for review see Hardie & Sakamoto, 2006). Roepstorff et al. had moderately trained males and females complete 90 min of cycling at 60% VO2peak and found that skeletal muscle AMPK Thr172 and α2AMPK activity increased significantly with a concomitant increase in free AMP, the AMP/ATP ratio and free creatine in men, but not women (Roepstorff et al. 2006). These data suggested the higher lipid oxidation seen in women during submaximal endurance exercise was associated with lower ‘cellular energy stress’, but not with AMPK status (Roepstorff et al. 2006). Consequently, the results of the current study suggest that either the AMPK–fat oxidation relationship is more complicated than originally proposed, or that the regulation of fat oxidation is independent of AMPK in women. They also confirmed earlier reports of a higher proportion of type 1 muscle fibres in women (Carter et al. 2001), and added novel data showing a correspondingly higher capillarization (∼23%). These latter findings suggest that fibre type and capillarization can partially explain the sex difference in metabolism; however, there must be other factors at work given that acute 17-β-oestradiol administration can influence exercise substrate selection within days (Devries et al. 2005), which is far too rapid for capillarization or fibre type transformation to occur. In the future, it will be of great interest to determine the role of 17-β-oestradiol on AMPK activity and its ultimate effect on capillarization and fibre type. Initial insight into this could be gleaned from the AMPK mRNA and protein abundance and phosphorylation status in response to acute 17-β-oestradiol administration; however, fibre type and capillarization influences will likely require longer term animal studies. As is the case with every study done well, more questions and research ideas are generated, particularly with respect to the AMPK–fat oxidation relationship in human skeletal muscle. Researchers in every aspect of exercise physiology will be further reminded of the need to control for potential sex differences and metabolism in all aspects of research design.

Sex difference in the principal cytochrome P-450 for tributyltin metabolism in rats

Tributyltin is metabolized by cytochrome P-450 (CYP) system enzymes, and its metabolic fate may contribute to the toxicity of the chemical. In the present study, it is examined whether sex differences in the metabolism of tributyltin exist in rats. In addition, the in vivo and in vitro metabolism of tributyltin was investigated using rat hepatic CYP systems to confirm the principal CYP involved. A significant sex difference in metabolism occurred both in vivo and in vitro, suggesting that one of the CYPs responsible for tributyltin metabolism in rats is male specific or predominant at least. Eight cDNA-expressed rat CYPs, including typical phenobarbital (PB)-inducible forms and members of the CYP2C subfamily, were tested to determine their capability for tributyltin metabolism. Among the enzymes studied, a statistically significant dealkylation of tributyltin was mediated by CYP2C6 and 2C11. Furthermore, the sex difference in metabolism disappeared in vitro after anti-rat CYP2C11 antibody pretreatment because CYP2C11 is a major male-specific form in rats. These results indicate that CYP2C6 is the principal CYP for tributyltin metabolism in female rats, whereas CYP2C11 as well as 2C6 is involved in tributyltin metabolism in male rats, and it is suggested that CYP2C11 is responsible for the significant sex difference in the metabolism of tributyltin observed in rats.

Sex difference in metabolism of simvastatin by rat hepatic microsomes.

Metabolism of simvastatin (SV), a new cholesterol-lowering agent, by hepatic microsomes from male and female rats was investigated. After incubation of [14C]-SV with hepatic microsomes, radioactive metabolites were detected by HPLC. The main metabolite was 3' alpha-hydroxy-SV in male rats and the hydroxy open acid form of SV (SVA) in females. The 3"-hydroxy-SV and 3',3"-dihydroxy-SV which were observed in male rats were hardly detected in females. Specific activity for the metabolism of SV in male rats (3.97 nmol/mg protein/min) was about 9-times higher than that in females. Metabolic activity of hepatic microsomes in male rats was essentially unchanged with increase in age, whereas that in females decreased age-dependently and was very low or negligible after 7 weeks of age. Formation of 3"-hydroxy-SV and 3',3"-dihydroxy-SV in male rats was markedly increased with age, and that in females was negligible at all ages examined.

Glutathione conjugation and cytochrome P-450 metabolism of methyl chloride in vitro

Possible carcinogenic properties of methyl chloride (CH 3Cl) have been under discussion since an increase of renal tumours was observed in male B6C3F 1 mice after a 2-yr inhalation exposure to the substance. This was, however, only observed following exposure of male mice to the highest concentration level and not after exposure of females or F344 rats of both sexes. Accumulation of formaldehyde in the kidneys was thought to be responsible for tumour production. In the experiments presented here, cytosolic enzymes from the liver and kidneys of different mouse strains and F344 rats were incubated in head-space vials with methyl chloride or methyl bromide. Following equilibration, the decrease in the concentration of the gases was monitored by gas chromatography as a parameter for metabolic elimination. The metabolic turnover of the methyl halides was found to be significantly higher in female animals than in the males. In parallel experiments, the glutathione content of the liver and kidneys of mice exposed by inhalation to 1000 ppm methyl chloride was determined. In both organs, the glutathione content diminished rapidly after exposure to the methyl halide. The glutathione depletion was slightly greater in females than in males. Finally, the content of cytochromes P-450, P-420 and b5 was determined in liver and kidneys of different mouse strains by difference spectroscopy. Female mice were found to have a lower content of P-450 and b5 than males in the kidneys; there was no such sex difference in liver tissue. The results show that a sex difference in metabolism is unlikely to be responsible for the unique kidney tumour production in male B6C3F 1 mice. Other possible explanations are discussed.

Sex differences in cytochrome P-450 isozyme composition and activity in kidney microsomes of mature rainbow trout

Kidney microsomes from sexually mature male, as opposed to female, rainbow trout displayed an approximately 20-fold higher cytochrome P-450 specific content, NADPH-cytochrome c reductase activity, and rates of hydroxylation of lauric acid, testosterone, progesterone and aflatoxin B 1. Little or no sex difference in metabolism was observed with benzo[ a]pyrene or benzphetamine as substrates. A similar pattern was observed in hepatic microsomes from these fish, but the difference was much less striking (approximately 2-fold higher activity in males). Juvenile trout (both sexes) possessed activities intermediate between mature males and females. Sodium dodecyl sulfate polyacrylamide gel electrophoresis of kidney and liver microsomes of juvenile and sexually mature male and female trout suggested that the striking sex difference in kidney could be due to the high amount of trout P-450 isozyme LM 2 in sexually mature males. Immunoquantitation of LM 2, performed by Western Blotting and immunostaining with rabbit anti-trout LM 2-IgG, confirmed that mature male kidney contained much higher levels of P-450 LM 2 than juvenile or female kidney, or even of liver microsomes of all three groups. The amount of P-450 LM 2 in mature female kidney microsomes was barely detectable. The high amount of LM 2 in male trout kidney is consistent with the high activity of these microsomes towards lauric acid and aflatoxin B 1, which have been shown previously to be preferentially metabolized by trout P-450 LM 2. It is suggested that rainbow trout may serve as an alternative to the rat as an animal model for the study of sex-dependent differences in cytochromes P-450.

Metabolism of 2-hydroxy[4- 14C]estradiol by rat liver microsomes

The metabolism of 14C-labeled estradiol and 2-hydroxyestradiol by rat liver microsomes has been compared under a variety of experimental conditions. An active liver microsomal system as well as NADPH was required to obtain high percentage yields of water-soluble metabolites from both steroids. Spermine stimulated the formation of polar products in both cases but the sex-difference in metabolism observed with estradiol was less marked with 2-hydroxyestradiol as substrate.

Species, strain and sex differences in metabolism of hexobarbitone, amidopyrine, antipyrine and aniline

Species and strain differences in response to hexobarbitone and presumably to antipyrine, amidopyrine and other drugs can be expressed largely in terms of the activities of the drug-transforming enzymes in microsomes. However, variation in the inherent sensitivity of the central nervous system may also be a factor. The sex difference in the rat in response to hexobarbitone and presumably to other drugs is also a reflection of the activity of the enzyme system in liver microsomes. Sex hormones have a role in regulating sex difference in drug metabolism. The guinea pig and mouse show no sex difference in the metabolism of hexobarbitone. Implications of these findings to drug action and to the development of new drugs are discussed.