Performance of laboratory tests used to measure blood phenylalanine for the monitoring of patients with phenylketonuria

Analysis of blood phenylalanine is central to the monitoring of patients with phenylketonuria (PKU) and age‐related phenylalanine target treatment‐ranges (0‐12 years; 120‐360 μmol/L, and >12 years; 120‐600 μmol/L) are recommended in order to prevent adverse neurological outcomes. These target treatment‐ranges are based upon plasma phenylalanine concentrations. However, patients are routinely monitored using dried bloodspot (DBS) specimens due to the convenience of collection. Significant differences exist between phenylalanine concentrations in plasma and DBS, with phenylalanine concentrations in DBS specimens analyzed by flow‐injection analysis tandem mass spectrometry reported to be 18% to 28% lower than paired plasma concentrations analyzed using ion‐exchange chromatography. DBS specimens with phenylalanine concentrations of 360 and 600 μmol/L, at the critical upper‐target treatment‐range thresholds would be plasma equivalents of 461 and 768 μmol/L, respectively, when a reported difference of 28% is taken into account. Furthermore, analytical test imprecision and bias in conjunction with pre‐analytical factors such as volume and quality of blood applied to filter paper collection devices to produce DBS specimens affect the final test results. Reporting of inaccurate patient results when comparing DBS results to target treatment‐ranges based on plasma concentrations, together with inter‐laboratory imprecision could have a significant impact on patient management resulting in inappropriate dietary change and potentially adverse patient outcomes. This review is intended to provide perspective on the issues related to the measurement of phenylalanine in blood specimens and to provide direction for the future needs of PKU patients to ensure reliable monitoring of metabolic control using the target treatment‐ranges

A putative relay circuit providing low-threshold mechanoreceptive input to lamina I projection neurons via vertical cells in lamina II of the rat dorsal horn

Background: Lamina I projection neurons respond to painful stimuli, and some are also activated by touch or hair movement. Neuropathic pain resulting from peripheral nerve damage is often associated with tactile allodynia (touch-evoked pain), and this may result from increased responsiveness of lamina I projection neurons to non-noxious mechanical stimuli. It is thought that polysynaptic pathways involving excitatory interneurons can transmit tactile inputs to lamina I projection neurons, but that these are normally suppressed by inhibitory interneurons. Vertical cells in lamina II provide a potential route through which tactile stimuli can activate lamina I projection neurons, since their dendrites extend into the region where tactile afferents terminate, while their axons can innervate the projection cells. The aim of this study was to determine whether vertical cell dendrites were contacted by the central terminals of low-threshold mechanoreceptive primary afferents. Results: We initially demonstrated contacts between dendritic spines of vertical cells that had been recorded in spinal cord slices and axonal boutons containing the vesicular glutamate transporter 1 (VGLUT1), which is expressed by myelinated low-threshold mechanoreceptive afferents. To confirm that the VGLUT1 boutons included primary afferents, we then examined vertical cells recorded in rats that had received injections of cholera toxin B subunit (CTb) into the sciatic nerve. We found that over half of the VGLUT1 boutons contacting the vertical cells were CTb-immunoreactive, indicating that they were of primary afferent origin. Conclusions: These results show that vertical cell dendritic spines are frequently contacted by the central terminals of myelinated low-threshold mechanoreceptive afferents. Since dendritic spines are associated with excitatory synapses, it is likely that most of these contacts were synaptic. Vertical cells in lamina II are therefore a potential route through which tactile afferents can activate lamina I projection neurons, and this pathway could play a role in tactile allodynia

Neuronal circuitry for pain processing in the dorsal horn

Neurons in the spinal dorsal horn process sensory information, which is then transmitted to several brain regions, including those responsible for pain perception. The dorsal horn provides numerous potential targets for the development of novel analgesics and is thought to undergo changes that contribute to the exaggerated pain felt after nerve injury and inflammation. Despite its obvious importance, we still know little about the neuronal circuits that process sensory information, mainly because of the heterogeneity of the various neuronal components that make up these circuits. Recent studies have begun to shed light on the neuronal organization and circuitry of this complex region

Barth syndrome without tetralinoleoyl cardiolipin deficiency:a possible ameliorated phenotype

Barth syndrome (BTHS) is an X-linked disorder characterised by cardiac and skeletal myopathy, growth delay, neutropenia and 3-methylglutaconic aciduria (3-MGCA). Patients have TAZ gene mutations which affect metabolism of cardiolipin, resulting in low tetralinoleoyl cardiolipin (CL(4)), an increase in its precursor, monolysocardiolipin (MLCL), and an increased MLCL/CL(4) ratio. During development of a diagnostic service for BTHS, leukocyte CL(4) was measured in 156 controls and 34 patients with genetically confirmed BTHS. A sub-group of seven subjects from three unrelated families was identified with leukocyte CL(4) concentrations within the control range. This had led to initial false negative disease detection in two of these patients. MLCL/CL(4) in this subgroup was lower than in other BTHS patients but higher than controls, with no overlap between the groups. TAZ gene mutations in these families are all predicted to be pathological. This report describes the clinical histories of these seven individuals with an atypical phenotype: some features were typical of BTHS (five have had cardiomyopathy, one family has a history of male infant deaths, three have growth delay and five have 3-MGCA) but none has persistent neutropenia, five have excellent exercise tolerance and two adults are asymptomatic. This report also emphasises the importance of measurement of MLCL/CL(4) ratio rather than CL(4) alone in the biochemical diagnosis of the BTHS

Hamstring contractures in children with spastic cerebral palsy result from a stiffer extracellular matrix and increased in vivo sarcomere length

Cerebral palsy (CP) results from an upper motoneuron (UMN)lesion in the developing brain. Secondary to the UMNl esion,which causes spasticity, is a pathological response by muscle - namely, contracture. However, the elements within muscle that increase passive mechanical stiffness, and therefore result in contracture, are unknown. Using hamstring muscle biopsies from pediatric patients with CP (n =33) and control (n =19) patients we investigated passive mechanical properties at the protein, cellular, tissue and architectural levels to identify the elements responsible for contracture. Titin isoform, the major load-bearing protein within muscle cells, was unaltered in CP. Correspondingly, the passive mechanics of individual muscle fibres were not altered. However, CP muscle bundles, which include fibres in their constituent ECM, were stiffer than control bundles. This corresponded to an increase in collagen content of CP muscles measured by hydroxyproline assay and observed using immunohistochemistry. In vivo sarcomere length of CP muscle measured during surgery was significantly longer than that predicted for control muscle. The combination of increased tissue stiffness and increased sarcomere length interact to increase stiffness greatly of the contracture tissue in vivo. These findings provide evidence that contracture formation is not the result of stiffening at the cellular level, but stiffening of the ECM with increased collagen and an increase of in vivo sarcomere length leading to higher passive stresses.ope