Creatine kinase B-driven energy transfer in the brain is important for habituation and spatial learning behaviour,mossy fibre field size and determination of seizure susceptibility

Creatine kinases are important in maintaining cellular-energy homeostasis, and neuroprotective effects have been attributed to the administration of creatine and creatine-like compounds. Herein we examine whether ablation of the cytosolic brain-type creatine kinase (B-CK) in mice has detrimental effects on brain development, physiological integrity or task performance. Mice deficient in B-CK (B-CK-/-) showed no gross abnormalities in brain anatomy or mitochondrial ultrastructure, but had a larger intra- and infrapyramidal mossy fibre area. Nuclear magnetic resonance spectroscopy revealed that adenosine triphosphate (ATP) and phosphocreatine (PCr) levels were unaffected, but demonstrated an apparent reduction of the PCr left arrow over right arrow ATP phosphorus exchange capacity in these mice. When assessing behavioural characteristics B-CK-/- animals showed diminished open-field habituation. In the water maze, adult B-CK-/- mice were slower to learn, but acquired the spatial task. This task performance deficit persisted in 24-month-old, aged B-CK-/- mice, on top of the age-related memory decline normally seen in old animals. Finally, a delayed development of pentylenetetrazole-induced seizures (creating a high-energy demand) was observed in B-CK-/- mice. It is suggested that the persistent expression of the mitochondrial isoform ubiquitous mitochondrial CK (UbCKmit) in the creatine/phospho-creatine shuttle provides compensation for the loss of B-CK in the brain. Our studies indicate a role for the creatine-phosphocreatine/CK circuit in the formation or maintenance of hippocampal mossy fibre connections, and processes that involve habituation, spatial learning and seizure susceptibility. However, for fuelling of basic physiological activities the role of B-CK can be compensated for by other systems in the versatile and robust metabolic-energy network of the brain.     

A clearly visible endoscopic instrument shaft on the monitor facilitates hand-eye coordination

Background: Passing an instrument through a small incision alters the kinematics of the instrument, thus hampering hand–eye coordination. Nevertheless, the incision provides a stable, nearly invariant, point of rotation for instrument movements. Therefore, we set out to evaluate the effects of the altered kinematics on hand–eye coordination. In addition, we assessed the hypothesis that the hand–eye coordination of laparoscopic surgeons incorporates the incision as a point of reference. Methods: Eight surgeons with experience in laparoscopy repeatedly performed a positioning task on a two-dimensional endoscopic manipulation simulator. Task time was measured. In the first experiment, normal endoscopic manipulation was compared to a condition in which the kinematic effects of the incision were compensated for. In the second experiment, the instrument shaft on the monitor was not visible during half of the trials, so that all visual information about the location of the incision was obscured. Results: Task performance improved significantly when the kinematic effects of the incision were compensated for (p = 0.001). Task performance improved when the instrument shaft was clearly visible on the monitor (p <0.05). Conclusions: Compensating for the kinematic effects introduced by the incision improves hand–eye coordination. The results of this study indicate that the incision provides a point of reference for hand–eye coordination during endoscopic manipulation.     

Estimation of the effect of the acute phase response on indicators of micronutrient status in Indonesian infants

Many indicators of micronutrient status change during infection because of the acute phase response. In this study, relationships between the acute phase response, assessed by measuring concentrations of C-reactive protein (CRP), alpha(1)-antichymotrypsin (ACT) and alpha(1)-acid glycoprotein (AGP), and indicators of micronutrient status were analyzed in 418 infants who completed a 6-mo randomized, double-blind, placebo-controlled, supplementation trial with iron, zinc and/or beta-carotene. The acute phase response, defined by raised CRP (plasma concentration >10 mg/L), raised AGP (>1.2 g/L), or both raised CRP and AGP, significantly affected indicators of iron, vitamin A and zinc status, independently of the effects of supplementation. Plasma ferritin concentrations were higher by 15.7 (raised AGP) to 21.2 (raised CRP and AGP) micro g/L in infants with elevated acute phase proteins compared with infants without acute phase response (P < 0.001). In contrast, plasma concentrations of retinol were lower by 0.07 (P < 0.05, raised AGP) to 0.12 (P < 0.01, raised CRP) micro mol/L, and of zinc lower by 1.49 (P < 0.01, raised AGP) to 1.89 (P < 0.05, raised CRP and AGP) micro mol/L. Hemoglobin concentrations and the modified relative dose response were not affected. Consequently, the prevalence of iron deficiency anemia was underestimated in infants with raised acute phase proteins by >15%, whereas the prevalence of vitamin A deficiency was overestimated by >16% compared with infants without acute phase response. Hence, using indicators of micronutrient status without considering the effects of the acute phase response results in a distorted estimate of micronutrient deficiencies, whose extent depends on the prevalence of infection in the population.     

An economic analysis of developmental detection methods

OBJECTIVE: To assess the costs and benefits of various approaches to early detection of developmental disabilities. DESIGN: Cost-benefit analyses based on data from previously published studies of developmental screening tests. SETTING: General pediatric practices and day care centers. PATIENTS AND OTHER PARTICIPANTS: A total of 247 parents and their 0- to 6-year-old children-103 from day care centers and 144 from pediatric practices. MAIN OUTCOME MEASURES: Licensed psychological examiners administered a screening test of parents' concerns about children's development and one or two direct screening tests: the Denver-II and/or the Battelle Developmental Inventory Screening Test. For the day care sample, examiners also administered to each child measures of intelligence, adaptive behavior, and language. In the pediatric sample, children were administered additional assessments. At the same time, diagnostic measures were administered to a randomly selected subsample to make determinations about developmental status. Each screening method was evaluated for its short-term costs (administration, interpretation, diagnosis, and treatment) and long-term benefits (impact of early intervention on adult functioning as inferred from longitudinal studies by other researchers). RESULTS: When the long-term costs and benefits were considered, none of the approaches emerged as markedly superior to another. When viewing the short-term costs, the various screening approaches differed markedly. The use of parents' concerns was by far the least costly for physicians to administer and interpret. CONCLUSION: Physicians can incur tremendous expenses when attempting to detect children with developmental problems. Although the benefits of early detection and intervention are substantial, physicians are not well-compensated for providing a critical service to society. Health policymakers and third-party payers must reconsider their minimal investment in early detection by health care providers. Nevertheless, our findings have encouraging implications for practice, because the use of parents' concerns as a screening technique offers substantial savings over and above other methods.     

Contribution of genetic and nutritional factors to DNA damage in heavy smokers

Prior epidemiological evidence suggests that genes controlling the metabolism of carcinogens and antioxidant/nutritional status are associated with lung cancer risk, possibly through their ability to modulate DNA damage by carcinogens. We performed a cross-sectional analysis of 159 heavy smokers from a cohort of subjects enrolled in a smoking cessation program. A total of 159 blood samples were analyzed to determine the relative contributions of genetic polymorphisms [CYP1A1 MspI and exon 7 and glutathione S-transferase M1 (GSTM1)] and plasma micronutrients to polycyclic aromatic hydrocarbon-DNA (PAH-DNA) adduct levels. DNA damage in smokers was affected by genetic polymorphisms and nutritional status. Smokers with the CYP1A1 exon 7 valine polymorphism had significantly higher (2-fold, P < or = 0.03) levels of DNA damage than those without. In parallel models, PAH-DNA adducts were inversely associated with plasma levels of retinol (beta = -0.93, P = 0.01), beta-carotene (beta = -0.18, P = 0.09), and alpha- tocopherol (beta = -0.28, P = 0.21) in 159 subjects. The association between smoking-adjusted plasma beta-carotene levels and DNA damage was only significant in those subjects lacking the GSTM1 detoxification gene (beta = -0.30, P = 0.05, n = 75). There was a statistical interaction between beta-carotene and alpha-tocopherol; when beta- carotene was low, alpha-tocopherol had a significant protective effect (beta = -0.78, P = 0.04) on adducts, but not when beta-carotene was high (beta = -0.16, P = 0.57). Plasma alpha-tocopherol was significantly correlated with beta-carotene (r = 0.36, P = 0.0005) and less strongly with retinol (r = 0.20, P = 0.0005). These results suggest that several micronutrients may act in concert to protect against DNA damage and highlight the importance of assessing overall antioxidant status. In conclusion, a subset of smokers may be at increased risk of DNA damage and possibly lung cancer due to the combined effect of low plasma micronutrients and genetic susceptibility factors. The use of biological markers to assess efficacy of interventions and to study mechanisms of micronutrients is timely given the current debate regarding the use of chemopreventive agents in high risk populations.      

Sensitive force transducer system for mechanical studies of single isolated vascular smooth muscle cells

To measure forces of the order of 0·1 μN, generated by single isolated vascular smooth muscle cells, a sensitive transducer system has been developed. A video displacement detection system and an inductive position detector were used to control the tip displacement of a glass microneedle. The bending of the microneedle was used as a measure of the force generated by a single smooth muscle cell attached to the tip. During isometric contractions the stiffness of the system was 0·7 μN μm⁻¹. A signal limiter in one of the feedback loops made it possible to reduce the stiffness of the system to 4 nN μm⁻¹ when an adjustable force limit was exceeded. These two conditions of the system made it possible to measure both isometric and isotonic contractions of a single muscle cell. The force limiter may also be used to protect the preparation against excessive forces developed during contraction. A mechanical model of the system and the vascular smooth muscle cell was simulated and verified. The parameters of the model for a single vascular smooth muscle cell, two viscoelastic springs in parallel, could be estimated from the measurements. The bandwidth of the system during isometric contraction was 17 Hz.     

Myotonic dystrophy kinase is a component of neuromuscular junctions

The clinical manifestation of myotonic dystrophy (DM) is correlatedto the extent of expansion of an unstable [CTG]_n DNA motif.Recent studies have demonstrated that this trinucleotide motifforms part of the last, 3' untranslated exon of a gene whichpotentially encodes multiple protein Isoforms of a serine/threonineprotein klnase (myotonic dystrophy protein kinase, DM-PK). Wereport here on the development of antisera against syntheticDM-PK peptide antigens and their use In blochemical and histochemlcalstudies. Immunoreactlve DM-kinase protein of 53 kD is presentat low levels In skeletal and cardiac muscle extracts of DMpatients and normal controls. Immunohlstochemical staining revealedthat DM-PK Is localised prominently at sites of neuromuscularand myotendinous junctions (NMJs and MTJs) of human and rodentskeletal muscles. Furthermore, very low levels of ImmunoreactiveDM-PK protein are present in the sarcoplasm of predominantlytype I fibres in various muscles. Strikingly, presence of theprotein can also be demonstrated for NMJs of muscular tissuesof adult and congenital cases of DM, with no gross changes Instructural organisation. Our findings provide a basis for furthercharacterisation of the role of the kinase in protein assemblyprocesses or signal mediation at synaptic sites and ultimatelyfor the understanding of the complex pathophysiology of DM.     

Cloning and characterization of DXS6673E, a candidate gene for X-linked mental retardation in Xq13.1

In several families with non-specific X-linked mental retardation (XLMR) linkage analyses have assigned the underlying gene defect to the pericentromeric region of the X chromosome, but none of these genes have been isolated so far. Here, we report on the cloning and characterization of a novel gene, DXS6673E, that maps to Xq13.1, is subject to X-inactivation and is disrupted in the 5' untranslated region by a balanced X;13 translocation in a mentally retarded female. The DXS6673E gene is highly conserved among vertebrates and its expression is most abundant in brain. It encodes a hydrophilic protein of 1358 amino acids (aa) that does not show sequence homology to other known proteins. A segment of this protein consisting of neutral and hydrophobic aa with a proline residue in every second position may represent a transmembrane domain. Almost complete sequence identity was found between the 3' end of the DXS6673E gene and two expressed sequence tags (ESTs) and between the 5' end of the DXS6673E gene and a third EST. Moreover, weaker sequence similarity was observed between coding regions and two other ESTs.      

Cloning and characterization of mCRIP2, a mouse LIM-only protein that interacts with PDZ domain IV of PTP-BL

BACKGROUND: In the mouse submembranous protein tyrosine phosphatase PTP-BL five PDZ domains are present in between the N-terminal FERM domain, which directs the protein to the cell cortex, and the C-terminal catalytic phosphatase domain. To understand more on the physical role of PTP-BL in this microenvironment, we started to search for PTP-BL PDZ domain-interacting proteins. RESULTS: Yeast two-hybrid screening for PTP-BL targets resulted in the identification of a novel mouse LIM-only protein termed CRIP2 that is highly homologous to rat ESP1 and human CRP2 sequences. Mouse CRIP2 has a predicted molecular weight of 23 kD and consists of two LIM domains spaced by 68 amino acids. The fourth PDZ domain of PTP-BL is responsible for the binding of CRIP2 protein. Both PTP-BL and CRIP2 mRNAs display a wide, overlapping tissue distribution. Western blot analysis revealed a more restricted expression pattern for CRIP2 with high expression in lung, heart and brain. CRIP2 protein is localized at cell cortical, actin-rich structures, which is concurrent with the subcellular localization of PTP-BL. CONCLUSIONS: The observed characteristics of the LIM domain-containing adaptor protein CRIP2 are consistent with a potential role of PTP-BL in the dynamics of the cortical actin cytoskeleton.