Investigation of Brain Iron in Niemann-Pick Type C: A 7T Quantitative Susceptibility Mapping Study.

BACKGROUND AND PURPOSE: While brain iron dysregulation has been observed in several neurodegenerative disorders, its association with the progressive neurodegeneration in Niemann-Pick type C is unknown. Systemic iron abnormalities have been reported in patients with Niemann-Pick type C and in animal models of Niemann-Pick type C. In this study, we examined brain iron using quantitative susceptibility mapping MR imaging in individuals with Niemann-Pick type C compared with healthy controls. MATERIALS AND METHODS: A cohort of 10 patients with adolescent- and adult-onset Niemann-Pick type C and 14 age- and sex-matched healthy controls underwent 7T brain MR imaging with T1 and quantitative susceptibility mapping acquisitions. A probing whole-brain voxelwise comparison of quantitative susceptibility mapping between groups was conducted. Mean quantitative susceptibility mapping in the ROIs (thalamus, hippocampus, putamen, caudate nucleus, and globus pallidus) was further compared. The correlations between regional volume, quantitative susceptibility mapping values, and clinical features, which included disease severity on the Iturriaga scale, cognitive function, and the Social and Occupational Functioning Assessment Scale, were explored as secondary analyses. RESULTS: We observed lower volume in the thalamus and voxel clusters of higher quantitative susceptibility mapping in the pulvinar nuclei bilaterally in patients with Niemann-Pick type C compared with the control group. In patients with Niemann-Pick type C, higher quantitative susceptibility mapping in the pulvinar nucleus clusters correlated with lower volume of the thalamus on both sides. Moreover, higher quantitative susceptibility mapping in the right pulvinar cluster was associated with greater disease severity. CONCLUSIONS: Our findings suggest iron deposition in the pulvinar nucleus in Niemann-Pick type C disease, which is associated with thalamic atrophy and disease severity. This preliminary evidence supports the link between iron and neurodegeneration in Niemann-Pick type C, in line with existing literature on other neurodegenerative disorders.

Application of 7T MRS to High-Grade Gliomas.

MRS, including single-voxel spectroscopy and MR spectroscopic imaging, captures metabolites in high-grade gliomas. Emerging evidence indicates that 7T MRS may be more sensitive to aberrant metabolic activity than lower-field strength MRS. However, the literature on the use of 7T MRS to visualize high-grade gliomas has not been summarized. We aimed to identify metabolic information provided by 7T MRS, optimal spectroscopic sequences, and areas for improvement in and new applications for 7T MRS. Literature was found on PubMed using "high-grade glioma," "malignant glioma," "glioblastoma," "anaplastic astrocytoma," "7T," "MR spectroscopy," and "MR spectroscopic imaging." 7T MRS offers higher SNR, modestly improved spatial resolution, and better resolution of overlapping resonances. 7T MRS also yields reduced Cramér-Rao lower bound values. These features help to quantify D-2-hydroxyglutarate in isocitrate dehydrogenase 1 and 2 gliomas and to isolate variable glutamate, increased glutamine, and increased glycine with higher sensitivity and specificity. 7T MRS may better characterize tumor infiltration and treatment effect in high-grade gliomas, though further study is necessary. 7T MRS will benefit from increased sample size; reductions in field inhomogeneity, specific absorption rate, and acquisition time; and advanced editing techniques. These findings suggest that 7T MRS may advance understanding of high-grade glioma metabolism, with reduced Cramér-Rao lower bound values and better measurement of smaller metabolite signals. Nevertheless, 7T is not widely used clinically, and technical improvements are necessary. 7T MRS isolates metabolites that may be valuable therapeutic targets in high-grade gliomas, potentially resulting in wider ranging neuro-oncologic applications.

Introduction of a mobile device based tertiary survey application reduces missed injuries: A multi-center prospective study.

BACKGROUND: Missed injuries during the initial assessment are a major cause of morbidity after trauma. The tertiary survey is a head-to-toe exam designed to identify any injuries missed after initial resuscitation. We designed a novel mobile device application (Physician Assist Trauma Software [PATS]) to standardize performance and documentation of the tertiary survey. This study was undertaken to assess the feasibility of introducing PATS into routine clinical practice, as well as its capacity to reduce missed injuries. METHODS: Prior to implementation of PATS, the missed injury rates at a higher-volume and a medium-volume level I trauma center were assessed. The PATS program was implemented simultaneously at both centers. Missed injuries were tracked during the study period. Compliance and tertiary survey completion rates were evaluated as a marker of feasibility. RESULTS: At the higher-volume trauma center, the missed injury rated decreased from 1% to 0% with the introduction of the PATS program (p = 0.04). At the medium-volume trauma center, the missed injury rate decreased from 9% to 1% (p < 0.001). Compliance and documentation increased from 68% to 100%, and from no formal documentation to 60% compliance at the higher- and medium-volume centers respectively. CONCLUSIONS: The implementation of a mobile tertiary survey application significantly reduced missed injuries at both a higher- and medium-volume trauma center. The use of this application resulted in a significant improvement in compliance with documentation of the tertiary survey.

Glutamate quantification in patients with supratentorial gliomas using chemical shift imaging.

This study aimed to evaluate and validate chemical shift imaging (CSI) for in vivo glutamate (Glu) quantification in patients with supratentorial gliomas. If validated, CSI could become an extremely useful tool to investigate metabolic dysfunction of Glu in excitotoxic neuropathologies. Quantitative CSI estimates of Glu concentrations were compared with known concentrations of Glu in aqueous phantom solutions. Forty-one patients with known or likely supratentorial gliomas underwent preoperative CSI. The spectra obtained were analyzed for Glu concentrations and Glu to creatine (Cr) ratios. These in vivo measurements were correlated against ex vivo Glu content quantified by high performance liquid chromatography (HPLC) measured in 65 resected brain tumor and peritumoral brain specimens. For the phantom solutions the CSI estimates of Glu concentration and the Glu/Cr ratios were highly correlated with known Glu concentration (r² = 0.95, p = 0.002, and r² = 0.97, p < 0.0001, respectively). There was a modest, but statistically significant, correlation between the ex vivo measured Glu and in vivo spectroscopic Glu concentration (r² = 0.22, p = 0.04) and ratios of Glu to Cr (r² = 0.30, p = 0.002). Quantitative measurement of Glu content is feasible in patients with supratentorial gliomas using CSI. The in vitro and in vivo results suggest that this has the potential to be a reliable quantitative imaging assay for brain tumor patients. This may have wide clinical research applications in a number of neurological disorders where Glu excitotoxicity and metabolic dysfunction are known to play a role in pathogenesis, including tumor associated epilepsy, epilepsy, stroke and neurotrauma.

Fusion of the HSV-1 tegument protein vp22 to cytosine deaminase confers enhanced bystander effect and increased therapeutic benefit.

A major limitation in cancer gene therapy, specifically gene-dependent enzyme prodrug therapy (GDEPT), is inefficient gene delivery and expression. The suicide gene cytosine deaminase (CD) and its substrate, 5-fluorocytosine (5-FC), have been extensively explored due to the inherent 'bystander' effect achieved through diffusion of the toxic metabolite 5-fluorouracil (5-FU). In this study, we aimed to enhance this 'bystander' effect by fusing the Saccharomyces cerevisiae CD to the HSV-1 tegument protein vp22, a novel translocating protein. Two constructs were created: one with vp22 fused to CD (vp22CD) and a second wherein a truncated vp22, lacking the necessary residues for trafficking, fused to CD (delvp22CD). The generated 9L stable lines exhibited similar growth rates, enzyme expression, CD activity, and sensitivity to 5-FC and 5-FU. However, mixed population colony formation assays demonstrated greater bystander effect with the vp22CD fusion as compared to delvp22CD. This enhancement was maintained in vivo where 9L tumors expressing 20 or 50% vp22CD exhibited increased growth delay compared to the respective delvp22CD tumors. Moreover, adenoviral transduction of established wild-type 9L tumors showed increased growth delay with vp22CD (Ad-EF_vp22CD) as compared to equivalent CD (Ad-EF_CD) transduced tumors. Finally, confirming the increased efficacy, (19)F magnetic resonance spectroscopy (MRS) of vp22CD-expressing tumors demonstrated increased 5-FU levels as compared to tumors expressing the nontranslocating CD. These results together demonstrated that fusion of vp22 to CD resulted in CD translocation, which in turn amplified conversion of 5-FC to 5-FU in vivo and enhanced the therapeutic benefit of this GDEPT strategy.

Enhanced surveillance for childhood hepatitis B virus infection in Canada, 1999-2003.

Since hepatitis B virus (HBV) infection can have serious sequelae, especially if infection occurs during childhood, there is a continuing need to examine its epidemiology so as to inform control measures. We analyzed trends in disease incidence and patterns of hepatitis B virus (HBV) transmission in both Canadian-born and non-Canadian-born children from 1999 to 2003, through the Enhanced Hepatitis Strain Surveillance System. Amongst Canadian-born children, the incidence of newly identified HBV infection per 100,000 declined significantly during the study period from 1.4 in 1999, to 0.5 in 2003 (RR, 0.75 per year; 95% CI, 0.60-0.95). Amongst non-Canadian-born children, the incidence of HBV infection per 100,000 ranged from 9.4 to 16.3, during the study period (linear trend test, p=0.69). Poisson regression analysis revealed that non-Canadian-born children were more likely to have HBV infection (RR, 12.3; 95% CI, 7.6 to 19.8), than Canadian-born children. HBV infection was found to be more common among children emigrating from high endemic area, than among Canadian-born children. Current Canadian immunization policy should take into consideration the protection of all children against HBV infection, including those coming from countries where mass hepatitis B vaccination programs have still not been launched.

Diffusion imaging for evaluation of tumor therapies in preclinical animal models.

The increasing development of novel targeted therapies for treating solid tumors has necessitated the development of technology to determine their efficacy in preclinical animal models. One such technology that can non-invasively quantify early changes in tumor cellularity as a result of an efficacious therapy is diffusion MRI. In this overview we present some theories as to the origin of diffusion changes as a result of tumor therapy, a robust methodology for acquisition of apparent diffusion coefficient maps and some applications of determining therapeutic efficacy in a variety therapeutic regimens and animal models.

Anisotropic water transport in the human eye lens studied by diffusion tensor NMR micro-imaging.

We report in vitro measurements of effective diffusion tensors characterising the anisotropic transport of water in human eye lenses ranging in age from 13 to 86 years. The measurements were obtained by means of a pulsed field gradient spin echo (PFGSE) magnetic resonance imaging (MRI) technique at a spatial resolution of 218 x 218 x 1000 microm(3). The results show that water diffusion is both spatially inhomogeneous and highly anisotropic on the timescale of the measurements (approximately 15 msec). Diffusion parallel to the long axes of the lens fibre cells is relatively unrestricted, whereas that between cells is substantially inhibited by the cell membranes, particularly in the inner cortex region of the lens. The data confirm the presence of a diffusion barrier surrounding the lens nucleus, which inhibits transport of water and other small molecules into and out of the nucleus. The results shed light on factors that may influence the onset of presbyopia and senile cataract. They also have implications for delivery of drugs to the lens nucleus.

Age-related changes in refractive index distribution and power of the human lens as measured by magnetic resonance micro-imaging in vitro.

We report a new technique for non-invasively mapping the refractive index distribution through the eye lens using magnetic resonance micro-imaging. The technique is applied to map the refractive index distribution throughout the sagittal plane of 18 human eye lenses ranging in age from 14 to 82 years in vitro. The results are compared with standard models for the human eye lens. They confirm that the refractive index distribution, when plotted as a function of normalised lens radius, is a function of lens age and differs both between the equatorial and axial directions and between the anterior and posterior halves of the optical axis. The refractive index of the lens nucleus exhibits a significant reduction with age amounting to 3.4+/-0.6 x 10(-4) years(-1). The contribution of the gradient index (GRIN) to the lens power decreases by 0.286+/-0.067 D/year, accounting almost entirely for the estimated overall change in lens power with age for these lenses, which were probably in their most accommodated state. The results provide experimental verification of hypothesised changes in the GRIN that have previously been invoked as contributing to presbyopia and support the hypothesis that changes in the GRIN are sufficient to offset effects of increasing curvature of human lenses with age in their unaccommodated state.

The interpretation of multi-exponential water proton transverse relaxation in the human and porcine eye lens.

We report results of 1H NMR transverse relaxation experiments on human and porcine eye lenses. Several authors have reported that transverse relaxation is not mono-exponential when observed by the Carr-Purcell-Meiboom-Gill (CPMG) sequence and have interpreted the results by postulating the presence of "pools" of water molecules in different binding environments that do not exchange rapidly on the NMR timescale. We have compared CPMG data for intact lenses with results for lens homogenates and have combined a CPMG spectroscopic pulse train with NMR micro-imaging to study the nature of the transverse relaxation process in human and porcine lenses. Fast exchange of water protons with the lens proteins (crystallins) leads to an enhanced transverse relaxation rate that varies linearly with protein concentration. At the resolution of NMR micro-imaging the transverse relaxation process is mono-exponential. The results show that the multi-exponential CPMG data observed spectroscopically for whole lenses reflect spatial variations in crystallin content through the lens rather than the presence of distinct "bound" and "free" water pools.

Vascular endothelial cell growth factor-driven endothelial tube formation is mediated by vascular endothelial cell growth factor receptor-2, a kinase insert domain-containing receptor.

Vascular endothelial cell growth factor (VEGF) binds to 2 related receptor tyrosine kinases, known as kinase insert domain-containing receptor (KDR) and fms-like tyrosine kinase (Flt-1). The KDR has been shown to mediate VEGF-stimulated endothelial cell mitogenesis, migration, and permeability. The Flt-1 receptor has been suggested to mediate VEGF-stimulated endothelial branching morphogenesis, a process whereby endothelial cells, in the presence of a 3D milieu composed of extracellular matrix components and a mixture of growth factors, undergo a morphological transition into a tubular network with many lumina. In the present study, we have used 2 independent endothelial cell tube formation models and highly selective VEGF mutants for the KDR and Flt-1 receptors. We demonstrate that KDR, not Flt-1, stimulation is responsible for the induction of endothelial tubulogenesis. In addition, we demonstrate a modulatory role for Flt-1 in VEGF-mediated tube formation. We also report that VEGF-driven endothelial tube formation is inhibited by selective inhibitors of mitogen-activated protein kinase activation and p38 protein kinase.

Through the haze of cigarettes: teenage girls' stories about cigarette addiction.

Narrative inquiry was used to explore the meaning of nicotine addiction among teenage girls, age 14 to 17 years, who had recent experience with smoking. The following three narratives emerged: invincibility, giving in, and unanticipated addiction. Those who told a story of invincibility depicted how they were in control of their smoking and not addicted. Participants who gave accounts of giving in to smoking described yielding to external forces. In the narrative of unanticipated addiction, participants recounted their surprise at realizing that they were addicted. Two subnarratives, needing to quit and repeating history, were also uncovered. The study findings reveal the importance of semantics and identity issues as teenage girls talked about nicotine addiction. Listening to their stories is paramount in continued efforts in the reduction of tobacco consumption.

Analysis of biological effects and signaling properties of Flt-1 (VEGFR-1) and KDR (VEGFR-2). A reassessment using novel receptor-specific vascular endothelial growth factor mutants.

Endothelial cells express two related vascular endothelial growth factor (VEGF) receptor tyrosine kinases, KDR (kinase-insert domain containing receptor, or VEGFR-2) and Flt-1 (fms-like tyrosine kinase, or VEGFR-1). Although considerable experimental evidence links KDR activation to endothelial cell mitogenesis, there is still significant uncertainty concerning the role of individual VEGF receptors for other biological effects such as vascular permeability. VEGF mutants that bind to either KDR or Flt-1 with high selectivity were used to determine which of the two receptors serves to mediate different VEGF functions. In addition to mediating mitogenic signaling, selective KDR activation was sufficient for the activation of intracellular signaling pathways implicated in cell migration. KDR stimulation caused tyrosine phosphorylation of both phosphatidylinositol 3-kinase and phospholipase Cgamma in primary endothelial cells and stimulated cell migration. KDR-selective VEGF was also able to induce angiogenesis in the rat cornea to an extent indistinguishable from wild type VEGF. We also demonstrate that KDR, but not Flt-1, stimulation is responsible for the induction of vascular permeability by VEGF.

Characterization of novel neutralizing monoclonal antibodies specific to human neurturin.

Neurturin (NTN) a structural and functional relative of glial cell line-derived neurotrophic factor, was originally identified based on its ability to support the survival of sympathetic neurons in culture. Similar to glial cell line-derived neurotrophic factor (GDNF), Neurturin has been shown to bind to a high affinity glycosylphosphatidylinositol (GPI)-linked receptor (GFRalpha2) and induce phosphorylation of the tyrosine kinase receptor Ret, resulting in the activation of the mitogen activated protein kinase (MAPK) signalling pathway. A panel of six novel murine monoclonal antibodies (MAbs) specific to human Neurturin has been developed and characterized. Four of the MAbs tested inhibit, to varying degrees, binding of NTN to the GPI-linked GFRalpha2 receptor. Three MAbs cross-react with the murine homolog. These antibodies have been shown to be useful reagents for Western blotting, immunohistochemistry, and also for the development of a sensitive, quantitative enzyme-linked immunosorbent assay (ELISA) for human NTN. Novel, specific MAbs with varying epitope specificities and blocking activity will be valuable tools for both the in vitro and in vivo characterization of NTN and its relationship to the GFRalpha2 and Ret receptors.

Protection and regeneration of nigral dopaminergic neurons by neurturin or GDNF in a partial lesion model of Parkinson's disease after administration into the striatum or the lateral ventricle.

Both glial cell line-derived neurotrophic factor (GDNF) and its recently discovered congener, neurturin (NTN), have been shown to exert neuroprotective effects on lesioned nigral dopamine (DA) neurons when administered at the level of the substantia nigra. In the present study, we have explored the relative in vivo potency of these two neurotrophic factors using two alternative routes of administration, into the striatum or the lateral ventricle, which may be more relevant in a clinical setting. In rats subjected to an intrastriatal (IS) 6-hydroxydopamine (6-OHDA) lesion, GDNF and NTN were injected every third day for 3 weeks starting on the day after the 6-OHDA injection. GDNF provided almost complete (90-92%) protection of the lesioned nigral DA neurons after both IS and intracerebroventricular (ICV) administration. NTN, by contrast, was only partially effective after IS injection (72% sparing) and totally ineffective after ICV injection. Although the trophic factor injections protected the nigral neurons from lesion-induced cell death, the level of expression of the phenotypic marker, tyrosine hydroxylase (TH), was markedly reduced in the rescued cell bodies. The extent of 6-OHDA-induced DA denervation in the striatum was unaffected by both types of treatment; consistent with this observation, the high rate of amphetamine-induced turning seen in the lesioned control animals was unaltered by either GDNF or NTN treatment. In the GDNF-treated animals, and to a lesser extent also after IS NTN treatment, prominent axonal sprouting was observed within the globus pallidus, at the level where the lesioned nigrostriatal axons are known to end at the time of onset of the neurotrophic factor treatment. The results show that GDNF is highly effective as a neuroprotective and axon growth-stimulating agent in the IS 6-OHDA lesion model after both IS and ICV administration. The lower efficacy of NTN after IS, and particularly ICV, administration may be explained by the poor solubility and diffusion properties at neutral pH.

Age-related changes in the kinetics of water transport in normal human lenses.

Magnetic resonance microscopy (MRM) has been used to study the kinetics of water transport in human eye lenses. Fresh lenses obtained from the Queensland Eye Bank were incubated at 34.5 degrees C in artificial aqueous humour (AAH) containing nutrients and metabolites similar to those that are present in vivo. MR images were acquired over approximately a 20 hr period following replacement of H(2)O based AAH with deuterium oxide (D(2)O) based AAH. NMR signal intensity from the lenses decreased with time corresponding to a decrease in concentration of H(2)O within the lenses. A statistically significant correlation (P<0.001) was found between the rate of NMR signal loss from the lens nuclei and increasing age of the lenses. The results show that as lenses age, there is a reduction in the rate at which water and presumably also water soluble low molecular weight metabolites, can enter the cells of the lens nucleus via the epithelium and cortex. A decrease in the rate of transport of water, nutrients and anti-oxidants (e.g. glutathione) would be expected to lead to progressive oxidative damage to lenses with age, and may ultimately contribute to presbyopia and senile nuclear cataract.

Neurturin exerts potent actions on survival and function of midbrain dopaminergic neurons.

Glial cell line-derived neurotrophic factor (GDNF) exhibits potent effects on survival and function of midbrain dopaminergic (DA) neurons in a variety of models. Although other growth factors expressed in the vicinity of developing DA neurons have been reported to support survival of DA neurons in vitro, to date none of these factors duplicate the potent and selective actions of GDNF in vivo. We report here that neurturin (NTN), a homolog of GDNF, is expressed in the nigrostriatal system, and that NTN exerts potent effects on survival and function of midbrain DA neurons. Our findings indicate that NTN mRNA is sequentially expressed in the ventral midbrain and striatum during development and that NTN exhibits survival-promoting actions on both developing and mature DA neurons. In vitro, NTN supports survival of embryonic DA neurons, and in vivo, direct injection of NTN into the substantia nigra protects mature DA neurons from cell death induced by 6-OHDA. Furthermore, administration of NTN into the striatum of intact adult animals induces behavioral and biochemical changes associated with functional upregulation of nigral DA neurons. The similarity in potency and efficacy of NTN and GDNF on DA neurons in several paradigms stands in contrast to the differential distribution of the receptor components GDNF Family Receptor alpha1 (GFRalpha1) and GFRalpha2 within the ventral mesencephalon. These results suggest that NTN is an endogenous trophic factor for midbrain DA neurons and point to the possibility that GDNF and NTN may exert redundant trophic influences on nigral DA neurons acting via a receptor complex that includes GFRalpha1.

Persephin, a novel neurotrophic factor related to GDNF and neurturin.

A novel neurotrophic factor named Persephin that is approximately 40% identical to glial cell line-derived neurotrophic factor (GDNF) and neurturin (NTN) has been identified using degenerate PCR. Persephin, like GDNF and NTN, promotes the survival of ventral midbrain dopaminergic neurons in culture and prevents their degeneration after 6-hydroxydopamine treatment in vivo. Persephin also supports the survival of motor neurons in culture and in vivo after sciatic nerve axotomy and, like GDNF, promotes ureteric bud branching. However, in contrast to GDNF and NTN, persephin does not support any of the peripheral neurons that were examined. Fibroblasts transfected with Ret and one of the coreceptors GFRalpha-1 or GFRalpha-2 do not respond to persephin, suggesting that persephin utilizes additional, or different, receptor components than GDNF and NTN.

Solution structure of the fourth metal-binding domain from the Menkes copper-transporting ATPase.

Menkes disease is an X-linked disorder in copper transport that results in death during early childhood. The solution structures of both apo and Ag(I)-bound forms of the fourth metal-binding domain (mbd4) from the Menkes copper-transporting ATPase have been solved. The 72-residue mbd4 has a ferredoxin-like beta alpha beta beta alpha beta fold. Structural differences between the two forms are limited to the metal-binding loop, which is disordered in the apo structure but well ordered in the Ag(I)-bound structure. Ag(I) binds in a linear bicoordinate manner to the two Cys residues of the conserved GMTCxxC motif; Cu(I) likely coordinates in a similar manner. Menkes mbd4 is thus the first bicoordinate copper-binding protein to be characterized structurally. Sequence comparisons with other heavy-metal-binding domains reveal a conserved hydrophobic core and metal-binding motif.