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Statistical flattening of MEG beamformer images.

We propose a method of correction for multiple comparisons in MEG beamformer based Statistical Parametric Maps (SPMs). We introduce a modification to the minimum-variance beamformer, in which beamformer weights and SPMs of source-power change are computed in distinct steps. This approach allows the calculation of image smoothness based on the computed weights alone. In the first instance we estimate image smoothness by looking at local spatial correlations in residual images generated using random data; we then go on to show how the smoothness of the SPM can be obtained analytically by measuring the correlations between the adjacent weight vectors. In simulations we show that the smoothness of the SPM is highly inhomogeneous and depends on the source strength. We show that, for the minimum variance beamformer, knowledge of image smoothness is sufficient to allow for correction of the multiple comparison problem. Per-voxel threshold estimates, based on the voxels extent (or cluster size) in flattened space, provide accurate corrected false positive error rates for these highly inhomogeneously smooth images.

Algorithms↗

Real-time fMRI using brain-state classification.

We have implemented a real-time functional magnetic resonance imaging system based on multivariate classification. This approach is distinctly different from spatially localized real-time implementations, since it does not require prior assumptions about functional localization and individual performance strategies, and has the ability to provide feedback based on intuitive translations of brain state rather than localized fluctuations. Thus this approach provides the capability for a new class of experimental designs in which real-time feedback control of the stimulus is possible-rather than using a fixed paradigm, experiments can adaptively evolve as subjects receive brain-state feedback. In this report, we describe our implementation and characterize its performance capabilities. We observed approximately 80% classification accuracy using whole brain, block-design, motor data. Within both left and right motor task conditions, important differences exist between the initial transient period produced by task switching (changing between rapid left or right index finger button presses) and the subsequent stable period during sustained activity. Further analysis revealed that very high accuracy is achievable during stable task periods, and that the responsiveness of the classifier to changes in task condition can be much faster than signal time-to-peak rates. Finally, we demonstrate the versatility of this implementation with respect to behavioral task, suggesting that our results are applicable across a spectrum of cognitive domains. Beyond basic research, this technology can complement electroencephalography-based brain computer interface research, and has potential applications in the areas of biofeedback rehabilitation, lie detection, learning studies, virtual reality-based training, and enhanced conscious awareness.

Adult↗

Comparison of functional brain PET images and intraoperative brain-mapping data using image-guided surgery.

OBJECTIVE: Knowledge about the spatial localization of eloquent brain areas is essential for resecting lesions in the vicinity of these areas. The classical approach is to perform surgery on the awake patient under local anesthesia using brain-mapping techniques. As an alternative, the location of eloquent areas can be visualized by preoperative functional brain-imaging techniques, for example, positron emission tomography (PET), functional magnetic resonance imaging (fMRI), or magnetoencephalography (MEG). Using functional activation PET, both methods were combined by integration into a frameless navigation system (BrainLAB) and used to map speech-eloquent areas. PATIENTS AND METHODS: Speech-eloquent areas were localized preoperatively in seven patients with a left-sided glioma using 2-[(18)F]-2-desoxy-D-glucose PET. Patients were scanned under silence conditions (i.e., with the patient remaining silent in a sound-proof cabin), and speech was activated using a verb-generation paradigm. The PET data were transferred to the neuronavigation workstation and matched with a preoperative 3D-MRI using an automatic image-fusion algorithm. Intraoperative speech localization was performed using brain-mapping techniques under local anesthesia with bipolar cortical stimulation. The stimulator position was mapped into the MRI/PET data set by neuronavigational tracking of the instrument. RESULTS: Functional PET images were integrated into the MRI-based neuronavigational system and could be transferred exactly to the operative field. By the additional integration of cortical stimulation, intraoperative electrophysiological findings can be directly compared with preoperative functional images. Seven patients with left-sided glioma were operated on using this protocol, confirming the technical feasibility. In three of seven patients, preoperative PET findings were not supported by intraoperative mapping. CONCLUSIONS: This matching and mapping technique is suitable for monitoring eloquent speech areas during surgical resection of extensive left-sided low-grade gliomas, allowing a direct comparison between intraoperative electrophysiological brain mapping and preoperative functional brain-imaging findings. The sensitivity and specificity of functional imaging techniques can now be evaluated by reconciling the data with the intraoperative stimulation results.

Adult↗

An autoregulatory enhancer element of the Drosophila homeotic gene Deformed.

The stable determination of different anterior-posterior regions of the Drosophila embryo is controlled by the persistent expression of homeotic selector genes. One mechanism that has been proposed to explain the persistent expression of the homeotic gene Deformed is an autoactivation circuit that would be used once Deformed expression had been established by earlier acting patterning genes. Here we show that a large cis-regulatory element mapping approximately 5 kb upstream of the Deformed transcription start has the properties predicted for a Deformed autoregulatory enhancer. This element provides late, spatially localized expression in the epidermal cells of the maxillary and mandibular segments which is wholly dependent upon endogenous Deformed function. In addition, the autoregulatory enhancer can be activated ectopically in embryos and in imaginal disc cells by ectopic expression of Deformed protein. Deletion analysis of the autoregulatory element indicates that it contains compartment specific sub-elements similar to those of other homeotic loci.

Animals↗

Engineering protein and cell adhesivity using PEO-terminated triblock polymers.

Previous studies on customizing cell culture environments have utilized a variety of microfabrication-based tools to control the spatial localization of adhesive proteins and subsequently mammalian cells. Others have used various methods to immobilize nonadhesive PEO-based polymers on surfaces to inhibit protein absorption and cell adhesion. In this study, we report the application of a well-characterized, commercially available, PEO-terminated triblock polymer (Pluronic F108) to create micropatterned nonadhesive domains on a variety of biomaterials that deter cell adhesion for up to 4 weeks in culture. The Pluronic can be applied using microfluidic tools or photolithographic techniques, and can be adsorbed to a variety of common surfaces including tissue culture polystyrene, methylated glass, silicone, and polylactic-co-glycolic acid. The effectiveness of the Pluronic in inhibiting cell adhesion in the presence of collagen I is also quantified. Finally, these patterning techniques are generalized to control tissue organization on a variety of common biomaterials. This simple method for micropatterning PEO and, therefore, proteins and cells should prove useful as a tool for biomolecular surface engineering.

3T3 Cells↗

Laser-layered microfabrication of spatially patterned functionalized tissue-engineering scaffolds.

Understanding cell behavior inside complex, three-dimensional (3D) microenvironments with controlled spatiotemporal patterning of physical and biochemical factors would provide significant insights into the basic biology of organ development and tissue functions. One of the fundamental limitations in studying such behavior has been the inability to create patterned microenvironments within 3D scaffold structures. Here a simple, layer-by-layer stereolithography (SL) method that can precisely pattern ligands, extracellular-matrix (ECM) components, and growth factors, as well as controlled release particles inside a single scaffold, has been developed. The process also allows fabrication of predesigned internal architectures and porosities. Photocrosslinkable poly(ethylene glycol) dimethacrylate (PEGDMA) was used as the basic structural component of these microfabricated scaffolds. PEG acrylates, covalently modified with the cell adhesive peptide arginine-glycine-aspartic acid (RGD) or the ECM component heparan sulfate, was incorporated within the scaffolds to facilitate cell attachment and to allow spatial sequestration of heparan-binding growth factors. Fluorescently labeled polymer microparticles and basic fibroblast growth factor (FGF-2) were chosen to illustrate the capability of SL to spatiotemporally pattern scaffolds. The results demonstrate that a precise, predesigned distribution of single or multiple factors within a single 3D structure can be created, and specific internal architectures can be fabricated. Functionalization of these scaffolds with RGD is demonstrated, and heparan sulfate allows efficient cell attachment and spatial localization of growth factors. Such patterned scaffolds might provide effective systems to study cell behavior in complex microenvironments and could eventually lead to engineering of complex, hybrid tissue structures through predesigned, multilineage differentiation of a single stem-cell population.

Animals↗

Interleukin-6, IL-6 receptor, and IL-6 nuclear factor gene expression in Paget's disease.

The cytokine interleukin-6 (IL-6) is considered an important regulator of bone cell function and may play a central role in bone disease states characterized by increased bone remodeling, such as Paget's disease. Indeed, recent in vitro data suggest that IL-6 may be an autocrine/paracrine factor for pagetic osteoclasts. However, its expression and role in vivo are not known. Using in situ hybridization we investigated the spatial localization of expression of IL-6, IL-6 receptor (IL-6R), and the transcription factor (NF-IL-6) in pagetic bone. Our results show that osteoblasts in the normal remodeling bone of osteoarthritis (controls) and in Paget's disease express IL-6, IL-6R, and NF-IL-6 genes with higher levels of IL-6 and IL-6R mRNA in pagetic bone. Osteoclasts in both osteoarthritic and pagetic bone express IL-6R mRNA and NF-IL-6, but only pagetic osteoclasts expressed IL-6, suggesting that in Paget's disease IL-6 can act as an autocrine factor on osteoclasts. These results provide evidence for a major role of the IL-6 regulatory pathway in the phenotype of the pagetic osteoclasts and lead us to suggest a model linking possible paramyxovirus infection and IL-6 regulation in the pagetic osteoclast.

Bone Remodeling↗

Some aspects of silica deposition in lithistid demosponge desmas.

Transmitted light microscope and SEM observations of various growth stages, including very young forms, of lithistid demosponge spicules called desmas provided a wealth of new observations on silica deposition in desmas of most lithistid demosponge groups. In typical (pachastrellid) demosponges a basic feature of silica deposition in young spicules is the formation of silica granules (100-160 nm in diameter) deposited in more or less regular concentric layers. Further growth stages in typical demosponges are similar, only silica granules are smaller and more densely packed. The shape of the spicule is controlled by an organic axial filament, while features of the outer spicule surface are also determined by silicalemma. In lithistid desmas the early stage of silica deposition is controlled by an organic axial filament or, in some cases, dispersed organic molecules only. The next step, after early arrest of axial filament growth and its total encasing by silica, is the deposition of various silica granules (40-300 nm in diameter), spheres, and/or cylinders (1,300-3,330 nm), which are either the result of precipitation or the effect of templating by organic molecules (proteins and polysaccharides), without direct control by the silicalemma. The later stages of desma growth are under direct control of the silicalemma, which molds secondary branches and/or elements of sculpture of the desmas. The tips of desmas, which articulate with older desmas, are also controlled by local spatial relationships. Differentiation of morphological forms of silica in desmas, which is at least genus-specific, clearly supports the polyphyletic nature of lithistid sponges.

Animals↗

A Xenopus maternal effect mutant gene affects oocyte meiotic reinitiation and fertilization.

No cleavage (nc) is a maternal effect mutant gene, recessive and sex limited. It affects the eggs laid by homozygous mutant females, independently of the male genotype. Contrary to normal oocytes, following germinal vesicle breakdown (GVBD) during maturation, the transient microtubular array (TMA) is not formed, nor are the meiotic spindles. Cytoplasmic asters with condensed chromosomes are present in the majority of oocytes, as well as microtubular bundles and sometimes cytoplasmic spindle-like asters. These mature oocytes exhibit a disturbance in yolk platelet arrangements. The white spot is rather irregular, and the maturation period is longer than normal. Transfers of cytoplasm from nc mature oocytes into normal stage VI oocytes resulted in abnormal maturation of the normal oocytes. Reciprocal transfers (cytoplasm from normal mature oocytes into nc stage VI oocytes) induce the formation of spindles, usually cytoplasmic; this indicates that the deficiency can be partly rescued. Following fertilization, the nc eggs show neither contraction nor rotation; polyspermy is present in the majority of cases. Even in the same egg, simultaneous spindles and nuclei can be observed, revealing a disturbance in the spatial localization of regulators of the cell cycle. Cytokinesis never occurs. Polyspermy results from the absence of cortical reaction following sperm entry. However, when mature nc oocytes are treated with PMA, they show cortical granule exocytosis and the formation of an altered vitelline envelope. The different factors possibly involved in these anomalies are discussed in relation to cytoarchitectural disorganization of the cell and abnormal cell cycle regulation.

Animals↗

Comparison of conventional single echo and multi-echo sequences with a fast spin-echo sequence for quantitative T2 mapping: application to the prostate.

The accuracy of water T2 maps generated from a fast spin-echo (FSE) sequence was compared with data obtained by conventional single and multi-echo spin-echo pulse sequences using a commercial gel phantom. Spatially localized stimulated echo acquisition mode (STEAM) proton spectroscopy was also used to confirm the reported water T2 values of the gels contained in the phantom. The FSE sequence was shown to be superior in accuracy to both the single and multi-echo spin echo sequences and comparable to STEAM, producing results that were within 10% of known values. The effectiveness of the FSE sequence was further demonstrated by generating T2 maps of the normal and diseased prostate in clinically acceptable imaging times, resulting in comparable T2 values to those obtained using STEAM. Accurate quantitative T2 maps can be produced with the FSE sequence.

Carcinoma↗

Direct analysis of drug candidates in tissue by matrix-assisted laser desorption/ionization mass spectrometry.

Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) has been used to directly analyze and image pharmaceutical compounds in intact tissue. The anti-tumor drug SCH 226374 was unambiguously determined in mouse tumor tissue using MALDI-QqTOFMS (QSTAR) by monitoring the dissociation of the protonated drug at m/z 695.4 to its predominant fragment at m/z 228.1. A second drug, compound A, was detected in slices of rat brain tissue following oral administration with doses ranging from 1-25 mg/kg. Quantitation of compound A from whole brain homogenates using routine high-performance liquid chromatography/tandem mass spectrometry (HPLC/MS/MS) procedures revealed that concentrations of the drug in whole brain varied from a low of 24 ng/g to a high of 1790 ng/g. The drug candidate was successfully detected by MALDI-QqTOF in samples from each dose, covering a range of approximately two orders of magnitude. In addition, good correlation was observed between the MALDI-QqTOFMS intensities at each dose with the HPLC/MS/MS results. Thus the MALDI-MS response is proportional to the amount of drug in tissue. Custom software was developed to facilitate the imaging of small molecules in tissue using the MALDI-QqTOF mass spectrometer. Images revealing the spatial localization of SCH 226374 in tumor tissue and compound A in brain tissue were acquired.

Animals↗

Fast proton spectroscopic imaging with high signal-to-noise ratio: spectroscopic RARE.

A new fast spectroscopic imaging (SI) method is presented which is based on spatial localization by the fast MRI method of rapid acquisition with relaxation enhancement (RARE) and encoding of the chemical shift information by shifting the position of a refocusing 180 pulse in a series of measurements. This method is termed spectroscopic RARE. In contrast to spectroscopic ultrafast low-angle RARE (U-FLARE), the formation of two echo families (odd and even) is suppressed by using a train of 180 RF pulses with an internal four-step phase cycle. By this means a high signal-to-noise ratio (SNR) per unit measurement time is obtained, because the separation of odd and even echoes, as well as dummy echoes to stabilize the echo amplitudes, is not needed anymore. The method is of particular interest for detecting signals of coupled spins, as effective homonuclear decoupling can be achieved by use of constant evolution time chemical shift encoding. The pulse sequence was implemented on a 4.7 T imaging system, tested on phantoms, and applied to the healthy rat brain in vivo. Spectroscopic RARE is particularly useful if T2* double less-than sign T2, which is typically fulfilled for in vivo proton SI measurements at high magnetic field strength.

Animals↗

Four-angle saturation transfer (FAST) method for measuring creatine kinase reaction rates in vivo.

A new fast method of measuring kinetic reaction rates for two-site chemical exchange is described. The method employs saturation transfer magnetic resonance spectroscopy (MRS) and acquisition of only four spectra under partially saturated, high signal-to-noise ratio (SNR) conditions. In two acquisitions one of the exchanging species is saturated; the other two employ a control saturation. Each pair of acquisitions is applied with two different flip angles, and the equilibrium magnetization, relaxation times, and reaction rates are calculated therefrom. This four-angle saturation transfer (FAST) method is validated theoretically using the Bloch equations modified for two-state chemical exchange. Potential errors in the rate measurements due to the effects of exchange are evaluated for creatine kinase (CK) metabolism modeled for skeletal and heart muscle, and are found to be < 5% for forward CK flux rates of 0.05 < or = k(f) < or = 1.0 s(-1), and up to a 90% depletion of phosphocreatine (PCr). The effect of too much or too little saturating irradiation on FAST appears to be comparable to that of the conventional saturation transfer method, although the relative performance deteriorates when spillover irradiation cuts the PCr signal by 50% or more. "FASTer" and " FASTest" protocols are introduced for dynamic CK studies wherein [PCr] and/or k(f) changes. These protocols permit the omission of one or two of the four acquisitions in repeat experiments, and the missing information is recreated from initial data via a new iterative algorithm. The FAST method is validated empirically in phosphorus ((31)P) MRS studies of human calf muscle at 1.5 T. FAST measurements of 10 normal volunteers yielded the same CK reaction rates measured by the conventional method (0.29 +/- 0.06 s(-1)) in the same subjects, but an average of seven times faster. Application of the FASTer algorithm to these data correctly restored missing information within seven iterations. Finally, the FAST method was combined with 1D spatially localized (31)P MRS in a study of six volunteers, yielding the same k(f) values independent of depth, in total acquisition times of 17-39 min. These timesaving FAST methods are enabling because they permit localized measurements of metabolic flux, which were previously impractical due to intolerably long scan times.

Algorithms↗

Functional magnetic resonance imaging of the human brain based on signal enhancement by extravascular protons (SEEP fMRI).

Functional magnetic resonance imaging (fMRI) studies of the human brain were carried out at 3 Tesla to investigate an fMRI contrast mechanism that does not arise from the blood oxygen-level dependent (BOLD) effect. This contrast mechanism, signal enhancement by extravascular protons (SEEP), involves only proton-density changes and was recently demonstrated to contribute to fMRI signal changes in the spinal cord. In the present study it is hypothesized that SEEP fMRI can be used to identify areas of neuronal activity in the brain with as much sensitivity and precision as can be achieved with BOLD fMRI. A detailed analysis of the areas of activity, signal intensity time courses, and the contrast-to-noise ratio (CNR), is also presented and compared with the BOLD fMRI results. Experiments were carried out with subjects performing a simple finger-touching task, or observing an alternating checkerboard pattern. Data were acquired using a conventional BOLD fMRI method (gradient-echo (GE) EPI, TE = 30 ms), a conventional method with reduced BOLD sensitivity (GE-EPI, TE = 12 ms), and SEEP fMRI (spin-echo (SE) EPI, TE = 22 ms). The results of this study demonstrate that SEEP fMRI may provide better spatial localization of areas of neuronal activity, and a higher CNR than conventional BOLD fMRI, and has the added benefit of lower sensitivity to field inhomogeneities.

Brain↗

19F NMR imaging of blood oxygenation in the brain.

Perfluorocarbon (PFC) emulsions have been developed as oxygen carrying blood substitutes. High fluorine concentrations allow them to be used in 19F NMR imaging of blood vessels. Furthermore, 19F relaxation times in PFCs are dependent on oxygen tension (pO2) so that NMR imaging may provide a noninvasive method of measuring spatially localized pO2 values in vivo. Using these principles, we have formed 19F images and calculated pO2 maps of the cat brain.

Animals↗

A general approach to selection of multiple cubic volume elements using the ISIS technique.

The ISIS method is used regularly for the selection of a single cubic volume of tissue for in vivo investigation by high-resolution NMR spectroscopy. This technique has been extended on a theoretical basis to include the simultaneous selection of a number of cubes, the signals from which can be either assessed individually or in certain circumstances coadded to produce improvement in signal-to-noise ratio. The modification requires additional selective RF pulses in the spatial encoding prepulse period, and spatially localized spectra are produced by addition and subtraction of NMR signals in a manner similar to the original ISIS technique.

Humans↗

In vivo proton spectroscopy in presence of eddy currents.

Spatially localized methods in spectroscopy often operate with magnetic field gradients for volume selection. The eddy currents induced by these gradients produce time-dependent shifts of the resonance frequency in the selected volume, which results in a distortion of the spectrum after Fourier transformation. In whole-body systems the complete compensation of eddy currents is a difficult procedure. To avoid this, a correction method is proposed for proton spectroscopy, which uses the signal of prominent water protons as a reference for the water-suppressed signal. The correction is performed in the time domain, dividing the water-suppressed signal by the phase factor of the water signal for each data point. The corrected spectra have a good resolution as shown by phantom measurements and brain and muscle spectra of volunteers.

Extracellular Space↗

1H spectroscopic imaging using a spectral-spatial excitation pulse.

Excellent water suppression is required to perform in vivo 1H spectroscopic experiments. However water suppression is difficult due to both B0 and RF inhomogeneities. These inhomogeneities are particularly troublesome in spectroscopic imaging experiments where water suppression is required throughout some large region of interest. In this paper, we propose the use of spectral-spatial excitation pulses for such experiments. These two-dimensional pulses are shown to provide water suppression that is insensitive to a range of B0 and RF variations while simultaneously providing spatial localization. Experimental results including images (with voxel volumes ranging from 3.4 to 1.5 cc) of various brain metabolites from both a normal volunteer and a patient with a metastatic lung carcinoma are presented.

Brain↗