The acetaminophen metabolite N-acetyl-p-benzoquinone imine (NAPQI) inhibits glutathione synthetase in vitro; a clue to the mechanism of 5-oxoprolinuric acidosis?

1. Metabolic acidosis due to accumulation of l-5-oxoproline is a rare, poorly understood, disorder associated with acetaminophen treatment in malnourished patients with chronic morbidity. l-5-Oxoprolinuria signals abnormal functioning of the γ-glutamyl cycle, which recycles and synthesises glutathione. Inhibition of glutathione synthetase (GS) by N-acetyl-p-benzoquinone imine (NAPQI) could contribute to 5-oxoprolinuric acidosis in such patients. We investigated the interaction of NAPQI with GS in vitro. 2. Peptide mapping of co-incubated NAPQI and GS using mass spectrometry demonstrated binding of NAPQI with cysteine-422 of GS, which is known to be essential for GS activity. Computational docking shows that NAPQI is properly positioned for covalent bonding with cysteine-422 via Michael addition and hence supports adduct formation. 3. Co-incubation of 0.77 µM of GS with NAPQI (25-400 µM) decreased enzyme activity by 16-89%. Inhibition correlated strongly with the concentration of NAPQI and was irreversible. 4. NAPQI binds covalently to GS causing irreversible enzyme inhibition in vitro. This is an important novel biochemical observation. It is the first indication that NAPQI may inhibit glutathione synthesis, which is pivotal in NAPQI detoxification. Further studies are required to investigate its biological significance and its role in 5-oxoprolinuric acidosis.

Macro- and micronutrient losses and nutritional status resulting from 44 days of total fasting in a non-obese man.

OBJECTIVE: We wanted to establish and understand how the fractional losses of fat, fat-free tissues, and selected nutrients compare with that of body mass during a 44-d voluntary starvation (water only) and measurements of nutrient status. METHODS: We used anthropometry, sequential measurements of urinary substances during the fast, and blood analytes at the end of the fast. RESULTS: At the start of the fast, body weight was 96.0 kg (20% fat) and body mass index was 28.36 kg/m(2). The changes in body mass and arm anthropometry and in the pattern of urinary excretion of creatinine, ammonia, sodium, and ketone bodies during the study were consistent with starvation. At the end of the fast, body mass had decreased by 25.5%, of which a quarter to a third was due to loss of fat and the remainder to fat-free mass, predominantly muscle. There was an estimated loss of 20% of total body protein, 20-25% of fat-free mass, and a greater fractional loss of fat. Total energy expenditure was estimated to be 1638-2155 kcal/d of which 13.0-17.1% was from protein oxidation. Differential losses of minerals in urine ranged from 1.2% of estimated initial body content for manganese to 17.3% for selenium and 40.5% for zinc. At the end of the study, plasma concentrations of zinc and vitamin B12 were increased, those of copper, selenium, and manganese were normal, and there was biochemical evidence of deficiency in thiamine, riboflavin, and vitamin K (prothrombin time). CONCLUSION: The data confirm and extend the available information on prolonged fasting in lean individuals and have relevance to the understanding of the physiologic responses to starvation and the associated homeostatic mechanisms.

Rapid and reliable self-screening for nutritional risk in hospital outpatients using an electronic system.

OBJECTIVE: This study examined an electronic nutritional self-screening procedure for feasibility and for reliability, rapidity, and ease of use by hospital outpatients. METHODS: One hundred sixty consecutive patients (ages 18-87 y) attending a gastroenterology clinic measured their weight and height using a modified digital weight and height machine, which transmitted results to a computer. Following input of reported weight loss in the previous 3 mo to 6 mo, malnutrition risk by the Malnutrition Universal Screening Tool (MUST) was instantaneously calculated. The duration and ease of undertaking screening were noted. Screening also was undertaken by a health care professional. RESULTS: Of the patients in the study, 21.3% were at risk for malnutrition (medium + high risk). There was perfect agreement (kappa = 1.00) between self-screening and health care professional screening, between test-retest self-screening, and between two methods of measuring height (facing toward and away from the stadiometer). A low within-patient coefficient of variation was found for measurement of weight (<0.2%), height (<0.35%) and body mass index (<0.4%), except for two measurements in which height was recorded before correct positioning of the sliding headpiece. The overall time to self-screen was 1.29 ± 0.57 min but it was 2.81 ± 0.92 min in those aged ≥ 75 y. Of the participants, 96.2% rated self-screening as very easy (71.9%) or easy (24.3%) and 3.8% (predominantly patients ages ≥ 75 y) difficult. CONCLUSION: The study provides evidence that electronic nutritional self-screening can be rapid, easy, reliable, and feasible in a clinical setting. Equipment specifically designed for self-screening and use in other types of patients and settings could facilitate appropriate and routine implementation of self-screening.

A change to pass/fail grading in the first two years at one medical school results in improved psychological well-being.

PURPOSE: To measure the impact of a change in grading system in the first two years of medical school, from graded (A, B, C, D, F) to pass/fail, on medical students' academic performance, attendance, residency match, satisfaction, and psychological well-being. METHOD: For both the graded and pass/fail classes, objective data were collected on academic performance in the first- and second-year courses, the clerkships, United States Medical Licensing Examination (USMLE) Steps 1 and 2 Clinical Knowledge (CK), and residency placement. Self-report data were collected using a Web survey (which included the Dupuy General Well-Being Schedule) administered each of the first four semesters of medical school. The study was conducted from 2002 to 2007 at the University of Virginia School of Medicine. RESULTS: The pass/fail class exhibited a significant increase in well-being during each of the first three semesters of medical school relative to the graded class, greater satisfaction with the quality of their medical education during the first four semesters of medical school, and greater satisfaction with their personal lives during the first three semesters of medical school. The graded and pass/fail classes showed no significant differences in performance in first- and second-year courses, grades in clerkships, scores on USMLE Step 1 and Step 2CK, success in residency placement, and attendance at academic activities. CONCLUSIONS: A change in grading from letter grades to pass/fail in the first two years of medical school conferred distinct advantages to medical students, in terms of improved psychological well-being and satisfaction, without any reduction in performance in courses or clerkships, USMLE test scores, success in residency placement, or level of attendance.

Modification of fetal plasma amino acid composition by placental amino acid exchangers in vitro.

Fetal growth is dependent on both the quantity and relative composition of amino acids delivered to the fetal circulation, and impaired placental amino acid supply is associated with restricted fetal growth. Amino acid exchangers can alter the composition, but not the quantity, of amino acids in the intra- and extracellular amino acid pools. In the placenta, exchangers may be important determinants of the amino acid composition in the fetal circulation. This study investigates the substrate specificity of exchange between the placenta and the feto-placental circulation. Maternal-fetal transfer of radiolabelled amino acids and creatinine were measured in the isolated perfused human placental cotyledon. Transfer of L-[14C]serine or L-[14C]leucine, and [3H]glycine, were measured in the absence of amino acids in the fetal circulation (transfer by non-exchange mechanisms) and following 10-20 micromol boluses of unlabelled amino acids into the fetal circulation to provide substrates for exchange (transfer by exchange and non-exchange mechanisms). The ability of fetal arterial boluses of L-alanine and L-leucine to stimulate release of amino acids from the placenta was also determined using HPLC in order to demonstrate the overall pattern of amino acid release. Experiments with radiolabelled amino acids demonstrated increased maternal-fetal transfer of L-serine and L-leucine, but not glycine, following boluses of specific amino acids into the fetal circulation. L-[14C]Leucine, but not L-[14C]serine or [3H]glycine, was transferred from the maternal to the fetal circulation by non-exchange mechanisms also (P<0.01). HPLC analysis demonstrated that fetal amino acid boluses stimulated increased transport of a range of different amino acids by 4-7 micromol l(-1) (P<0.05). Amino acid exchange provides a mechanism to supply the fetus with amino acids that it requires for fetal growth. This study demonstrates that these transporters have the capacity to exchange micromolar amounts of specific amino acids, and suggests that they play an important role in regulating fetal plasma amino acid composition.