Rapid hydroxylation of methtryptoline (1-methyltetrahydro-β-carboline) in rat: Identification of metabolites by chiral gas chromatography-mass spectrometry

Summary Racemic methtryptoline (1-methyltetrahydro--carboline) and 5-hydroxymethtryptoline-9-carboxylic acid (6-hydroxy-1-methyltetrahydro--carboline-1-carboxylic acid) were administered intraperitoneally to rats and the components of their urine was subsequently investigated by chiral gas chromatography-mass spectrometry. Methtryptoline rapidly became hydroxylated in the 5- and 6-position and excreted in urine. There was about a ninefold predominance of the S(–) enantiomer over the other in the 5-hydroxylated species, while the 6-hydroxylation produced a small excess of the R(+) enantiomer. About 75% of the injected dose of methtryptoline was recovered in the urine as 5- and 6-hydroxylated compounds during the first 24 h period, demonstrating that hydroxylation represents the major metabolic pathway. Treatment with 6-hydroxymethtryptoline-9-carboxylic acid led to a fivefold increase in the urinary excretion of 5-hydroxymethtryptoline during the first 24 h period with a predominance of the S(–)-enantiomer, indicating a much smaller conversion rate than from methtryptoline. It was concluded that hydroxylation of methtryptoline is a likely pathway for the natural formation of 5-hydroxymethtryptoline.     

Work factors facilitating working beyond state pension age: Prospective cohort study with register follow-up

Objectives:The demographic changes in Europe underline the need for an extension of working lives. This study investigates the importance of physical work demands and psychosocial work factors for working beyond the state pension age (65 years).Methods:We combined data from three cohorts of the general working population in Denmark (DWECS 2005 and 2010, and DANES 2008), where actively employed workers aged 55–59 years replied to questionnaires about work environment and were followed until the age of 66 years in the Danish AMRun register of paid employment. Using logistic regression analyses, we calculated prevalence ratios (PR) and 95% confidence intervals (CI) for the association between physical and psychosocial work factors and working beyond state pension age, adjusted for age, sex, cohort, cohabiting, sector, income, vocational education, working hours, lifestyle, and previous sickness absence.Results:Of the 2884 workers aged 55–59 years, 1023 (35.5%) worked beyond the state pension age. Higher physical work demands was associated with a lower likelihood (PR 0.69, 95% CI 0.58–0.82) and a good psychosocial work environment was associated with higher likelihood (average of 7 items: PR 1.81, 95% CI 1.49–2.20) of working beyond state pension age. Stratified analyses did not change the overall pattern, ie, a good overall psychosocial work environment – as well as several specific psychosocial factors – increased the likelihood of working beyond state pension age, both for those with physically active and seated work.Conclusion:While high physical work demands was a barrier, a good psychosocial work environment seems to facilitate working beyond state pension age, also for those with physically active work.     

The use of natural interferon alpha conjugated to a monoclonal antibody anti mammary epithelial mucin (Mc5) for the treatment of human breast cancer xenografts

Summary An immunoconjugate composed of natural interferon (nIFN) bound in a noncleavable fashion to a monoclonal antibody (MoAb) recognizing a breast epithelial membrane mucin (Mc5) was used to treat xenografts of a human mammary carcinoma cell line (MCF-7) growing in nude mice. The immunoconjugate (nIFN/Mc5) was administered as 20 intralesional (i.l.) injections to 1 of 2 xenografts in each animal. It was found that nIFN/Mc5 produced a significant enhancement of the growth inhibitory actions of nIFN on the injected tumors. Further enhancement was obtained when nIFN or nIFN together with Mc5 (at a dose 10 times larger than that present in nIFN/Mc5) were added to the immunoconjugate. Biodistribution experiments showed that the uptake of¹²⁵I-nIFN/Mc5 by the tumors was greater and its elimination slower than for¹²⁵I-nIFN alone or conjugated to irrelevant mouse IgG₁. In addition, the immunoconjugate up-regulated the antigenic expression of a breast epithelial membrane mucin by the carcinoma cells, an up-regulation which was not significantly different from that produced by nIFN alone. The contralateral noninjected tumors exposed to systemic levels of the immunoconjugate showed an enhancement of antitumor effects, but to a lesser extent than the injected tumors. These findings suggest that the enhancement of the growth inhibitory action of the immunoconjugate was related to the specific binding of Mc5 which targeted the IFN to the carcinoma cells and impeded its elimination. It is likely that the targeting was favored by the IFN-mediated up-regulation of antigenic expression by the carcinoma cells, thereby producing a cascade of interrelated effects. The results of this study point out the feasibility and potential usefulness of IFN treatment by means of immunoconjugates as well as the worth of pursuing and improving this form of therapy.     

A comparative proteome analysis of hippocampal tissue from schizophrenic and Alzheimer's disease individuals

The proteins expressed by a genome have been termed the proteome. Comparative proteome analysis of brain tissue offers a novel means to identify biologically significant gene products that underlie psychopathology. In this study we collected post mortem hippocampal tissue from the brains of seven schizophrenic, seven Alzheimer's disease (AD) and seven control individuals. Hippocampal proteomes were visualised by two-dimensional gel electrophoresis of homogenised tissue. A mean of 549 (s.d. 35) proteins were successfully matched between each disease group and the control group. In comparison with the control hippocampal proteome, eight proteins in the schizophrenic hippocampal proteome were found to be decreased and eight increased in concentration, whereas, in the AD hippocampal proteome, 35 proteins were decreased and 73 were increased in concentration (P<0.05). One protein, which was decreased in concentration in both diseases, was characterised as diazepam binding inhibitor (DBI) by N-terminal sequence analysis. DBI can regulate the action of the GABA(A) receptor. Protein changes involved 6% of the assessed AD hippocampal proteome, whereas, in schizophrenia protein changes involved less than 1% of the assessed hippocampal proteome. We conclude that schizophrenia has a subtle neuropathological presentation and comparative proteome analysis is a viable means by which to investigate diseases of the brain at the molecular level.     

Cloning and functional expression of alternative spliced variants of the human metabotropic glutamate receptor 8

Two new spliced variants of the human metabotropic glutamate receptor 8 (HmGluR8), designated HmGluR8b and HmGluR8c, were identified in a human fetal brain cDNA library. The HmGluR8b and c differ from previously reported HmGluR8a by the out-of-frame insertions of 55-bp and 74-bp, respectively. The 55-bp insertion which contains a stop codon resulted in substitution of the last 16 amino acids in the C-terminus of HmGluR8a with 16 different amino acids in HmGluR8b. The 74-bp insertion introduces a frame shift in the predicted translation resulting in termination of the polypeptide before the putative seven transmembrane domains. Thus, the predicted HmGluR8c protein is 501 amino acids long and could represent a secreted isoform of the receptor. The pattern of mRNA expression of mGluR8 variants in human brain were analyzed by RT-PCR, Northern blot and in situ hybridization. Both HmGluR8a and b are expressed with similar abundance in fetal and adult brains. The in situ hybridization results indicate a predominantly glial cell expression of HmGluR8c in human brain. The three isoforms were transiently expressed in CHO cells from Semliki Forest Virus vectors. [3H]l-AP4 binding was performed on the cell membranes and the saturation curves showed the presence of a binding site with KD values of 249 and 182 nM and Bmax values of 13.6 and 10.5 pmoles/mg protein for HmGluR8a and b, respectively. For the six mGluR ligands studied, a similar rank order of potency was observed on both HmGluRa and b: l-AP4>l-SOP=l-CCG I>l-glutamate>DCG IV>LY 354740.     

Fat gram target to achieve high energy intake in cystic fibrosis

Higher fat and energy intakes confer a survival advantage in cystic fibrosis (CF). There is a need to develop effective nutrition programmes that ensure optimal energy intake in CF.

Methodology:

A cross-sectional measurement of clinical characteristics and energy and fat intakes in patients attending the CF outpatients clinic of the John Hunter Hospital, Newcastle was undertaken. Twenty-nine subjects, mean age 12 years (range 4.3–20.2), completed weighed food records to determine the contribution of fat to the percentage of the recommended energy intake obtained and to document use of pancreatic enzyme replacement therapy.

Results:

Diets with a high percentage of energy derived from fat did not guarantee that individuals with CF met their energy requirements. Subjects with total fat intakes of 100 g per day or greater, however, achieved in excess of 110% recommended daily intake (RDI) for energy. Up to 47% of subjects consumed more pancreatic enzyme replacement capsules than shown to give maximum effectiveness.

Conclusion:

Setting a 100 g daily fat target is a realistic way of ensuring high energy intakes in CF. Fat ready reckoners would identify the fat content of food and prescribe specific numbers of pancreatic enzyme replacement capsules to be consumed with each meal or food item.     

Intestinal permeability in kwashiorkor

Intestinal permeability can be assessed non-invasively using the lactulose-rhamnose (L-R) test, which is a reliable measure of small intestinal integrity. AIMS: To determine risk factors for abnormal intestinal permeability in kwashiorkor, and to measure changes in L-R ratios with inpatient rehabilitation. DESIGN: A case-control study of 149 kwashiorkor cases and 45 hospital controls. The L-R test was adapted to study kwashiorkor in Malawi, with testing at weekly intervals during nutritional rehabilitation. Urine sugars were measured by thin layer chromatography in London. RESULTS: The initial geometric mean L-R ratios (x100) (with 95% confidence interval) in kwashiorkor were 17.3 (15.0 to 19.8) compared with 7.0 (5.6 to 8.7) for controls. Normal ratios are < 5, so the high ratios in controls indicate tropical enteropathy syndrome. Abnormal permeability in kwashiorkor was associated with death, oliguria, sepsis, diarrhoea, wasting and young age. Diarrhoea and death were associated with both decreased L-rhamnose absorption (diminished absorptive surface area) and increased lactulose permeation (impaired barrier function) whereas nutritional wasting affected only L-rhamnose absorption. Despite, clinical recovery, mean L-R ratios improved little on treatment, with mean weekly ratios of 16.3 (14.0 to 19.0), 13.3 (11.1 to 15.9) and 14.4 (11.0 to 18.8). CONCLUSION: Abnormal intestinal permeability in kwashiorkor correlates with disease severity, and improves only slowly with nutritional rehabilitation.     

Respiratory syncytial virus genotypes and disease severity among children in hospital

OBJECTIVES: To determine the spectrum of N and G genotypes of respiratory syncytial virus (RSV) causing respiratory tract infection and whether particular genotypes are associated with severity of infection. PATIENTS AND METHODS: Nasopharyngeal aspirates (NPAs) were obtained from 114 infants with acute respiratory tract infection due to RSV over two seasons. Viral mRNA was extracted from NPAs or cultured virus, reverse transcribed, and the cDNA amplified by the polymerase chain reaction using primers directed to parts of the N and G gene respectively. Amplicons were separately digested with four different restriction endonucleases for each gene. The fragments were separated by agarose gel, electrophoresis, and the electrophoretic patterns used to assign the various genotypes. Disease severity was assessed as very mild (upper respiratory tract signs only), mild (coryza and signs of lower respiratory tract infection), moderate (requiring nasogastric or intravenous fluids), and severe (requiring oxygen or ventilation). RESULTS: Five of the six known N genotypes were detected, but NP4 and NP2 were found most frequently. There was no association between N genotype and disease severity. Six G (SHL) genotypes were detected. Significantly (p = 0.04) more of the infants infected with the SHL2 genotype had severe or moderate disease. CONCLUSIONS: During the seasonal peaks of RSV respiratory tract infection at least 10 different RSV genotypes cocirculated. While there is no association between N genotypes and disease severity, infection with the SHL2 G genotype appears to result in moderate to severe disease.