Activation of the parieto-premotor network is associated with vivid motor imagery - a parametric FMRI study

The present study examined the neural basis of vivid motor imagery with parametrical functional magnetic resonance imaging. 22 participants performed motor imagery (MI) of six different right-hand movements that differed in terms of pointing accuracy needs and object involvement, i.e., either none, two big or two small squares had to be pointed at in alternation either with or without an object grasped with the fingers. After each imagery trial, they rated the perceived vividness of motor imagery on a 7-point scale. Results showed that increased perceived imagery vividness was parametrically associated with increasing neural activation within the left putamen, the left premotor cortex (PMC), the posterior parietal cortex of the left hemisphere, the left primary motor cortex, the left somatosensory cortex, and the left cerebellum. Within the right hemisphere, activation was found within the right cerebellum, the right putamen, and the right PMC. It is concluded that the perceived vividness of MI is parametrically associated with neural activity within sensorimotor areas. The results corroborate the hypothesis that MI is an outcome of neural computations based on movement representations located within motor areas.

Semantic network disconnection in formal thought disorder

Structural and functional findings in schizophrenic patients with formal thought disorder (FTD) show abnormalities within left-side semantic areas. The present study investigate the network function of the involved brain regions as a function of FTD severity.

Mitochondrial dysfunction and Purkinje cell loss in autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS)

Autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS) is a childhood-onset neurological disease resulting from mutations in the SACS gene encoding sacsin, a 4,579-aa protein of unknown function. Originally identified as a founder disease in Québec, ARSACS is now recognized worldwide. Prominent features include pyramidal spasticity and cerebellar ataxia, but the underlying pathology and pathophysiological mechanisms are unknown. We have generated an animal model for ARSACS, sacsin knockout mice, that display age-dependent neurodegeneration of cerebellar Purkinje cells. To explore the pathophysiological basis for this observation, we examined the cell biological properties of sacsin. We show that sacsin localizes to mitochondria in non-neuronal cells and primary neurons and that it interacts with dynamin-related protein 1, which participates in mitochondrial fission. Fibroblasts from ARSACS patients show a hyperfused mitochondrial network, consistent with defects in mitochondrial fission. Sacsin knockdown leads to an overly interconnected and functionally impaired mitochondrial network, and mitochondria accumulate in the soma and proximal dendrites of sacsin knockdown neurons. Disruption of mitochondrial transport into dendrites has been shown to lead to abnormal dendritic morphology, and we observe striking alterations in the organization of dendritic fields in the cerebellum of knockout mice that precedes Purkinje cell death. Our data identifies mitochondrial dysfunction/mislocalization as the likely cellular basis for ARSACS and indicates a role for sacsin in regulation of mitochondrial dynamics.

Subcortical electrostimulation to identify network subserving motor control

Objectives: Recent anatomical-functional studies have transformed our understanding of cerebral motor control away from a hierarchical structure and toward parallel and interconnected specialized circuits. Subcortical electrical stimulation during awake surgery provides a unique opportunity to identify white matter tracts involved in motor control. For the first time, this study reports the findings on motor modulatory responses evoked by subcortical stimulation and investigates the cortico-subcortical connectivity of cerebral motor control. Experimental design: Twenty-one selected patients were operated while awake for frontal, insular, and parietal diffuse low-grade gliomas. Subcortical electrostimulation mapping was used to search for interference with voluntary movements. The corresponding stimulation sites were localized on brain schemas using the anterior and posterior commissures method. Principal observations: Subcortical negative motor responses were evoked in 20/21 patients, whereas acceleration of voluntary movements and positive motor responses were observed in three and five patients, respectively. The majority of the stimulation sites were detected rostral of the corticospinal tract near the vertical anterior-commissural line, and additional sites were seen in the frontal and parietal white matter. Conclusions: The diverse interferences with motor function resulting in inhibition and acceleration imply a modulatory influence of the detected fiber network. The subcortical stimulation sites were distributed veil-like, anterior to the primary motor fibers, suggesting descending pathways originating from premotor areas known for negative motor response characteristics. Further stimulation sites in the parietal white matter as well as in the anterior arm of the internal capsule indicate a large-scale fronto-parietal motor control network. Hum Brain Mapp, 2012. © 2012 Wiley Periodicals, Inc.

Identification of a highly conserved valine-glycine-phenylalanine amino acid triplet required for HIV-1 Nef function

Background The Nef protein of HIV facilitates virus replication and disease progression in infected patients. This role as pathogenesis factor depends on several genetically separable Nef functions that are mediated by interactions of highly conserved protein-protein interaction motifs with different host cell proteins. By studying the functionality of a series of nef alleles from clinical isolates, we identified a dysfunctional HIV group O Nef in which a highly conserved valine-glycine-phenylalanine (VGF) region, which links a preceding acidic cluster with the following proline-rich motif into an amphipathic surface was deleted. In this study, we aimed to study the functional importance of this VGF region. Results The dysfunctional HIV group O8 nef allele was restored to the consensus sequence, and mutants of canonical (NL4.3, NA-7, SF2) and non-canonical (B2 and C1422) HIV-1 group M nef alleles were generated in which the amino acids of the VGF region were changed into alanines (VGF→AAA) and tested for their capacity to interfere with surface receptor trafficking, signal transduction and enhancement of viral replication and infectivity. We found the VGF motif, and each individual amino acid of this motif, to be critical for downregulation of MHC-I and CXCR4. Moreover, Nef’s association with the cellular p21-activated kinase 2 (PAK2), the resulting deregulation of cofilin and inhibition of host cell actin remodeling, and targeting of Lck kinase to the trans-golgi-network (TGN) were affected as well. Of particular interest, VGF integrity was essential for Nef-mediated enhancement of HIV virion infectivity and HIV replication in peripheral blood lymphocytes. For targeting of Lck kinase to the TGN and viral infectivity, especially the phenylalanine of the triplet was essential. At the molecular level, the VGF motif was required for the physical interaction of the adjacent proline-rich motif with Hck. Conclusion Based on these findings, we propose that this highly conserved three amino acid VGF motif together with the acidic cluster and the proline-rich motif form a previously unrecognized amphipathic surface on Nef. This surface appears to be essential for the majority of Nef functions and thus represents a prime target for the pharmacological inhibition of Nef.

Transient rhythmic network activity in the somatosensory cortex evoked by distributed input in vitro

The initiation and maintenance of physiological and pathophysiological oscillatory activity depends on the synaptic interactions within neuronal networks. We studied the mechanisms underlying evoked transient network oscillation in acute slices of the adolescent rat somatosensory cortex and modeled its underpinning mechanisms. Oscillations were evoked by brief spatially distributed noisy extracellular stimulation, delivered via bipolar electrodes. Evoked transient network oscillation was detected with multi-neuron patch-clamp recordings under different pharmacological conditions. The observed oscillations are in the frequency range of 2-5 Hz and consist of 4-12 mV large, 40-150 ms wide compound synaptic events with rare overlying action potentials. This evoked transient network oscillation is only weakly expressed in the somatosensory cortex and requires increased [K+]o of 6.25 mM and decreased [Ca2+]o of 1.5 mM and [Mg2+]o of 0.5 mM. A peak in the cross-correlation among membrane potential in layers II/III, IV and V neurons reflects the underlying network-driven basis of the evoked transient network oscillation. The initiation of the evoked transient network oscillation is accompanied by an increased [K+]o and can be prevented by the K+ channel blocker quinidine. In addition, a shift of the chloride reversal potential takes place during stimulation, resulting in a depolarizing type A GABA (GABAA) receptor response. Blockade of alpha-amino-3-hydroxy-5-methyl-4-isoxazole-proprionate (AMPA), N-methyl-D-aspartate (NMDA), or GABA(A) receptors as well as gap junctions prevents evoked transient network oscillation while a reduction of AMPA or GABA(A) receptor desensitization increases its duration and amplitude. The apparent reversal potential of -27 mV of the evoked transient network oscillation, its pharmacological profile, as well as the modeling results suggest a mixed contribution of glutamatergic, excitatory GABAergic, and gap junctional conductances in initiation and maintenance of this oscillatory activity. With these properties, evoked transient network oscillation resembles epileptic afterdischarges more than any other form of physiological or pathophysiological neocortical oscillatory activity.

Visual function in human ocular toxoplasmosis

AIM: To assess functional impairment in terms of visual acuity reduction and visual field defects in inactive ocular toxoplasmosis. METHODS: 61 patients with known ocular toxoplasmosis in a quiescent state were included in this prospective, cross-sectional study. A complete ophthalmic examination, retinal photodocumentation and standard automated perimetry (Octopus perimeter, program G2) were performed. Visual acuity was classified on the basis of the World Health Organization definition of visual impairment and blindness: normal (> or =20/25), mild (20/25 to 20/60), moderate (20/60 to 20/400) and severe (<20/400). Visual field damage was correspondingly graded as mild (mean defect <4 dB), moderate (mean defect 4-12 dB) or severe (mean defect >12 dB). RESULTS: 8 (13%) patients presented with bilateral ocular toxoplasmosis. Thus, a total of 69 eyes was evaluated. Visual field damage was encountered in 65 (94%) eyes, whereas only 28 (41%) eyes had reduced visual acuity, showing perimetric findings to be more sensitive in detecting chorioretinal damage (p<0.001). Correlation with the clinical localisation of chorioretinal scars was better for visual field (in 70% of the instances) than for visual acuity (33%). Moderate to severe functional impairment was registered in 65.2% for visual field, and in 27.5% for visual acuity. CONCLUSION: In its quiescent stage, ocular toxoplasmosis was associated with permanent visual field defects in >94% of the eyes studied. Hence, standard automated perimetry may better reflect the functional damage encountered by ocular toxoplasmosis than visual acuity.

European Working Group on Clinical Cell Analysis: Consensus protocol for the flow cytometric characterisation of platelet function

An increased or disturbed activation and aggregation of platelets plays a major role in the pathophysiology of thrombosis and haemostasis and is related to cardiovascular disease processes. In addition to qualitative disturbances of platelet function, changes in thrombopoiesis or an increased elimination of platelets, (e. g., in autoimmune thrombocytopenia), are also of major clinical relevance. Flow cytometry is increasingly used for the specific characterisation of phenotypic alterations of platelets which are related to cellular activation, haemostatic function and to maturation of precursor cells. These new techniques also allow the study of the in vitro response of platelets to stimuli and the modification thereof under platelet-targeted therapy as well as the characterisation of platelet-specific antibodies. In this protocol, specific flow cytometric techniques for platelet analysis are recommended based on a description of the current state of flow cytometric methodology. These recommendations are an attempt to promote the use of these new techniques which are at present broadly evaluated for diagnostic purposes. Furthermore, the definition of the still open questions primarily related to the technical details of the method should help to promote the multi-center evaluation of procedures with the goal to finally develop standardized operation procedures as the basis of interlaboratory reproducibility when applied to diagnostic testing.

Prosthetic treatment planning on the basis of scientific evidence

The objective of this report is to summarize the results on survival and complication rates of different designs of fixed dental prostheses (FDP) published in a series of systematic reviews. Moreover, the various parameters for survival and risk assessment are to be used in attempt to perform treatment planning on the basis of scientific evidence. Three electronic searches complemented by manual searching were conducted to identify prospective and retrospective cohort studies on FDP and implant-supported single crowns (SC) with a mean follow-up time of at least 5 years. Patients had to have been examined clinically at the follow-up visit. Failure and complication rates were analyzed using random-effects Poisson regression models to obtain summary estimates of 5- and 10-year survival proportions. Meta-analysis of the studies included indicated an estimated 5-year survival of conventional tooth-supported FDP of 93.8%, cantilever FDP of 91.4%, solely implant-supported FDP of 95.2%, combined tooth-implant-supported FDP of 95.5% and implant-supported SC of 94.5% as well as resin-bonded bridges 87.7%. Moreover, after 10 years of function the estimated survival decreased to 89.2% for conventional FDP, to 80.3% for cantilever FDP, to 86.7% for implant-supported FDP, to 77.8% for combined tooth-implant-supported FDP, to 89.4% for implant-supported SC and to 65% for resin-bonded bridges. When planning prosthetic rehabilitations, conventional end-abutment tooth-supported FDP, solely implant-supported FDP or implant-supported SC should be the first treatment option. Only as a second option, because of reasons such as financial aspects patient-centered preferences or anatomical structures cantilever tooth-supported FDP, combined tooth-implant-supported FDP or resin-bonded bridges should be chosen.

Oxidized low density lipoprotein impairs endothelial progenitor cell function by downregulation of E-selectin and integrin alpha(v)beta5

BACKGROUND: Oxidized low density lipoprotein (oxLDL) has been shown to induce apoptosis and senescence of endothelial progenitor cells (EPC). In the present study, we hypothesized that even sub-apoptotic concentrations of oxLDL impair the angiogenic potential of EPC and investigated if this effect is mediated by affecting adhesion and incorporation. METHODS: A co-culture system of human microvascular endothelial cells and EPC was used to study the effect of sub-apoptotic concentrations of native (nLDL) and oxLDL on cell-cell interaction. The expression and the functional role of angiogenic adhesion molecules and integrins was monitored by FACS and neutralizing assay, respectively. RESULTS: We observed an inhibition of tube formation and impairment of EPC integration into the vascular network of mature endothelial cells by oxLDL. In contrast, nLDL did not affect angiogenic properties of EPC. Incubation of EPC with sub-apoptotic oxLDL concentrations significantly decreased E-selectin and integrin alpha(v)beta(5) expression (37.6% positive events vs. 71.5% and 24.3% vs. 49.9% compared to control culture media without oxLDL). Interestingly, expression of alpha(v)beta(3), VE-cadherin and CD31 remained unchanged. Blocking of E-selectin and integrin alpha(v)beta(5) by neutralizing antibody effectively inhibited adhesion of EPC to differentiated endothelial cells (56.5% and 41.9% of control; p<0.001). CONCLUSION: In conclusion, oxidative alteration of LDL impairs angiogenic properties of EPC at sub-apoptotic levels by downregulation of E-selectin and integrin alpha(v)beta(5), both substantial mediators of EPC-endothelial cell interaction.

Differential GPS as a monitoring tool on Volcano Santa Ana (Illamatepec) and the Coatepeque Caldera, El Salvador

We used differential GPS measurements from a 13 station GPS network spanning the Santa Ana Volcano and Coatepeque Caldera to characterize the inter-eruptive activity and tectonic movements near these two active and potentially hazardous features. Caldera-forming events occurred from 70-40 ka and at Santa Ana/Izalco volcanoes eruptive activity occurred as recently as 2005. Twelve differential stations were surveyed for 1 to 2 hours on a monthly basis from February through September 2009 and tied to a centrally located continuous GPS station, which serves as the reference site for this volcanic network. Repeatabilities of the averages from 20-minute sessions taken over 20 hours or longer range from 2-11 mm in the horizontal (north and east) components of the inter-station baselines, suggesting a lower detection limit for the horizontal components of any short-term tectonic or volcanic deformation. Repeatabilities of the vertical baseline component range from 12-34 mm. Analysis of the precipitable water vapor in the troposphere suggests that tropospheric decorrelation as a function of baseline lengths and variable site elevations are the most likely sources of vertical error. Differential motions of the 12 sites relative to the continuous reference site reveal inflation from February through July at several sites surrounding the caldera with vertical displacements that range from 61 mm to 139 mm followed by a lower magnitude deflation event on 1.8-7.4 km-long baselines. Uplift rates for the inflationary period reach 300 mm/yr with 1σ uncertainties of +/- 26 – 119 mm. Only one other station outside the caldera exhibits a similar deformation trend, suggesting a localized source. The results suggest that the use of differential GPS measurements from short duration occupations over short baselines can be a useful monitoring tool at sub-tropical volcanoes and calderas.

IDENTIFICATION OF GENES CONTROLLING BIOLOGICAL PROCESSES AND PATHWAYS THROUGH STATISTICAL ANALYSIS AND NETWORK RECONSTRUCTION

The developmental processes and functions of an organism are controlled by the genes and the proteins that are derived from these genes. The identification of key genes and the reconstruction of gene networks can provide a model to help us understand the regulatory mechanisms for the initiation and progression of biological processes or functional abnormalities (e.g. diseases) in living organisms. In this dissertation, I have developed statistical methods to identify the genes and transcription factors (TFs) involved in biological processes, constructed their regulatory networks, and also evaluated some existing association methods to find robust methods for coexpression analyses. Two kinds of data sets were used for this work: genotype data and gene expression microarray data. On the basis of these data sets, this dissertation has two major parts, together forming six chapters. The first part deals with developing association methods for rare variants using genotype data (chapter 4 and 5). The second part deals with developing and/or evaluating statistical methods to identify genes and TFs involved in biological processes, and construction of their regulatory networks using gene expression data (chapter 2, 3, and 6). For the first part, I have developed two methods to find the groupwise association of rare variants with given diseases or traits. The first method is based on kernel machine learning and can be applied to both quantitative as well as qualitative traits. Simulation results showed that the proposed method has improved power over the existing weighted sum method (WS) in most settings. The second method uses multiple phenotypes to select a few top significant genes. It then finds the association of each gene with each phenotype while controlling the population stratification by adjusting the data for ancestry using principal components. This method was applied to GAW 17 data and was able to find several disease risk genes. For the second part, I have worked on three problems. First problem involved evaluation of eight gene association methods. A very comprehensive comparison of these methods with further analysis clearly demonstrates the distinct and common performance of these eight gene association methods. For the second problem, an algorithm named the bottom-up graphical Gaussian model was developed to identify the TFs that regulate pathway genes and reconstruct their hierarchical regulatory networks. This algorithm has produced very significant results and it is the first report to produce such hierarchical networks for these pathways. The third problem dealt with developing another algorithm called the top-down graphical Gaussian model that identifies the network governed by a specific TF. The network produced by the algorithm is proven to be of very high accuracy.

Distally based abductor hallucis muscle flap: anatomic basis and clinical application

Although rarely used, the abductor hallucis muscle has its indications in coverage of small defects at the medial aspect of the hindfoot as a proximally based muscle flap. The authors describe a 69-year-old female patient in whom the abductor hallucis muscle was used as a distally based flap to reconstruct a defect in the forefoot. An anatomic study was undertaken on two cadaveric feet to explore the practicality of the distally based abductor hallucis muscle flap before it was applied clinically. The distally based abductor muscle flap receives its blood supply from minor and major pedicles in a retrograde fashion from both the dorsal arterial network and the deep plantar system, through communicating branches with the medial plantar artery distally. Transposition of the distally based hallucis flap is only advisable in individuals who have no vascular compromise in the lower leg and foot. To the authors' knowledge, this modification has not yet been described in the available literature.

DENITRIFICATION IN SOILS: FROM GENES TO ENVIRONMENTAL OUTCOMES

Denitrification is an important process of global nitrogen cycle as it removes reactive nitrogen from the biosphere, and acts as the primary source of nitrous oxide (N2O). This thesis seeks to gain better understanding of the biogeochemistry of denitrification by investigating the process from four different aspects: genetic basis, enzymatic kinetics, environmental interactions, and environmental consequences. Laboratory and field experiments were combined with modeling efforts to unravel the complexity of denitrification process under microbiological and environmental controls. Dynamics of denitrification products observed in laboratory experiments revealed an important role of constitutive denitrification enzymes, whose presence were further confirmed with quantitative analysis of functional genes encoding nitrite reductase and nitrous oxide reductase. A metabolic model of denitrification developed with explicit denitrification enzyme kinetics and representation of constitutive enzymes successfully reproduced the dynamics of N2O and N2 accumulation observed in the incubation experiments, revealing important regulatory effect of denitrification enzyme kinetics on the accumulation of denitrification products. Field studies demonstrated complex interaction of belowground N2O production, consumption and transport, resulting in two pulse pattern in the surface flux. Coupled soil gas diffusion/denitrification model showed great potential in simulating the dynamics of N2O below ground, with explicit representation of the activity of constitutive denitrification enzymes. A complete survey of environmental variables showed distinct regulation regimes on the denitrification activity from constitutive enzymes and new synthesized enzymes. Uncertainties in N2O estimation with current biogeochemical models may be reduced as accurate simulation of the dynamics of N2O in soil and surface fluxes is possible with a coupled diffusion/denitrification model that includes explicit representation of denitrification enzyme kinetics. In conclusion, denitrification is a complex ecological function regulated at cellular level. To assess the environmental consequences of denitrification and develop useful tools to mitigate N2O emissions require a comprehensive understanding of the regulatory network of denitrification with respect to microbial physiology and environmental interactions.

Reduced beta-cell mass and altered glucose sensing impair insulin-secretory function in betaIRKO mice

Pancreatic beta-cell-restricted knockout of the insulin receptor results in hyperglycemia due to impaired insulin secretion, suggesting that this cell is an important target of insulin action. The present studies were undertaken in beta-cell insulin receptor knockout (betaIRKO) mice to define the mechanisms underlying the defect in insulin secretion. On the basis of responses to intraperitoneal glucose, approximately 7-mo-old betaIRKO mice were either diabetic (25%) or normally glucose tolerant (75%). Total insulin content was profoundly reduced in pancreata of mutant mice compared with controls. Both groups also exhibited reduced beta-cell mass and islet number. However, insulin mRNA and protein were similar in islets of diabetic and normoglycemic betaIRKO mice compared with controls. Insulin secretion in response to insulin secretagogues from the isolated perfused pancreas was markedly reduced in the diabetic betaIRKOs and to a lesser degree in the nondiabetic betaIRKO group. Pancreatic islets of nondiabetic betaIRKO animals also exhibited defects in glyceraldehyde- and KCl-stimulated insulin release that were milder than in the diabetic animals. Gene expression analysis of islets revealed a modest reduction of GLUT2 and glucokinase gene expression in both the nondiabetic and diabetic mutants. Taken together, these data indicate that loss of functional receptors for insulin in beta-cells leads primarily to profound defects in postnatal beta-cell growth. In addition, altered glucose sensing may also contribute to defective insulin secretion in mutant animals that develop diabetes.