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[Quantitative method of assessing the effectiveness of radiotherapy of malignant neoplasms based on computerized tomography data].

Computed tomography assesses the linear coefficient of X-ray radiation decrease in the tissue and hence determines changes in its density thus allowing one to control the destruction of tumor cells and tissues during treatment. The paper proposes a procedure for determining the sizes and density of a tumor along the chosen linear direction crossing the image of a pathological focus. The whole procedure is performed by the special computer programme "Diaglmag". The equations that characterize the dynamics of using the parameters used before, during, and after treatment are presented. Baseline information on the optic image densities on a computer tomogram is obtained with a graphic scanner. The examples presented in the paper show it feasible to solve a difficult task to determine the effect of treatment. This enables a treatment regimen to be corrected in time or modified.

Evaluation Studies as Topic↗

Labelling of human adipose-derived stem cells for non-invasive in vivo cell tracking.

Human adipose-derived stem cells (ASC) can be expanded in an undifferentiated state or differentiated along the osteogenic, chondrogenic, adipogenic, myogenic, endothelial and neurogenic lineage. To test their in vivo and in situ regenerative potential, their fate needs to be traced after application in suitable defect models. Non-invasive imaging systems allow for real time tracking of labelled cells in the living animal. We have evaluated a bioluminescence cell tracking approach to visualise ASC labelled with luciferase in the living animal. Two procedures have been tested to efficiently label human stem cells with a reporter gene (luciferase, green fluorescent protein), namely lipofection with Lipofectamine 2000 and electroporation with a Nucleofector device. With both lipofection and nucleofection protocols, we have reached transfection efficiencies up to 60%. Reporter gene expression was detectable for 3 weeks in vitro and did not interfere with the phenotype and the stem cell properties of the cells. By means of a highly sensitive CCD camera, we were able to achieve real time imaging of cell fate for at least 20 days after application (intravenous, intramuscular, intraperitoneal, subcutaneous) in nude mice. Moreover, we were able to influence cell mobility by choosing different modes of application such as enclosure in fibrin matrix. The optical imaging system with transient transfection is an elegant cell-tracking concept to follow survival and fate of human stem cells in small animals.

Adipose Tissue↗

A single gold particle as a probe for apertureless scanning near-field optical microscopy.

We report on the fabrication, characterization and application of a probe consisting of a single gold nanoparticle for apertureless scanning near-field optical microscopy. Particles with diameters of 100 nm have been successfully and reproducibly mounted at the end of sharp glass fibre tips. We present the first optical images taken with such a probe. We have also recorded plasmon resonances of gold particles and discuss schemes for exploiting the wavelength dependence of their scattering cross-section for a novel form of apertureless scanning near-field optical microscopy.

Journal Article↗

Complex technical methodologies and their applications in the surgery of intracranial meningiomas.

As neurosurgery moves into the twenty-first century, improved visualization/localization techniques, neuromonitoring, and advanced instrumentation will become standards of care for all intracranial procedures. This article has focused on current available technologies that can be used to facilitate operations on intracranial meningiomas. Preoperative anatomic localization with MR imaging, CT, MR angiography, and angiography are standard techniques. Preoperative functional assessments with MR imaging, magnetic source imaging, PET, and functional MR imaging are crucial to recognize and preserve eloquent adjacent cortex. Pathologic correlations with preoperative imaging (i.e., MR imaging) may help to predict the histopathology. Perioperative rehearsal of the operation can be performed. Intraoperative anatomic localization is important to minimize the craniotomy, dural opening, and passage through normal neural structures. An impressive array of new technologies are currently available, including real-time ultrasonography, frame-based stereotaxy (CT, MR imaging PET), frameless stereotaxy (acoustic localization, neuronavigators, real-time visualization), robotics, neuroendoscopy, and intraoperative dye administration. Increased understanding of the function and individual variability of the human cortex underscores the importance of intraoperative functional localization by electrocorticography and optical imaging. Continuous intraoperative neuromonitoring of sensory (SSEPs, BAERs, visual evoked potentials) and motor evoked potentials is now standard during many intracranial procedures. Complex adjunctive instrumentation, such as the ultrasonic aspirator and the laser, are part of the contemporary armamentarium for meningioma surgery. As we have stated in a previous article, "Developmental trends imply realization of three major directions of technical neurosurgery: (1) precise preoperative simulation, (2) minimization or avoidance of transcranial operative corridors, [and] (3) increased refinement of technical adjuvants--both physical and molecular." The end point of these technical advances is to improve precision and safety and will enhance the outcome in each surgical procedure so that one day we will be able to operate on patients with intracranial lesions with minimal morbidity.

Brain↗

Imaging cortical correlates of illusion in early visual cortex.

Exploring visual illusions reveals fundamental principles of cortical processing. Illusory motion perception of non-moving stimuli was described almost a century ago by Gestalt psychologists. However, the underlying neuronal mechanisms remain unknown. To explore cortical mechanisms underlying the 'line-motion' illusion, we used real-time optical imaging, which is highly sensitive to subthreshold activity. We examined, in the visual cortex of the anaesthetized cat, responses to five stimuli: a stationary small square and a long bar; a moving square; a drawn-out bar; and the well-known line-motion illusion, a stationary square briefly preceding a long stationary bar presentation. Whereas flashing the bar alone evoked the expected localized, short latency and high amplitude activity patterns, presenting a square 60-100 ms before a bar induced the dynamic activity patterns resembling that of fast movement. The preceding square, even though physically non-moving, created gradually propagating subthreshold cortical activity that must contribute to illusory motion, because it was indistinguishable from cortical representations of real motion in this area. These findings demonstrate the effect of spatio-temporal patterns of subthreshold synaptic potentials on cortical processing and the shaping of perception.

Animals↗

A study and optimization of lumbar spine X-ray imaging systems.

A Monte Carlo program has been developed that incorporates a voxel phantom of an adult patient in a model of the complete X-ray imaging system, including the anti-scatter grid and screen-film receptor. This allows the realistic estimation of patient dose and the corresponding image (optical density map) for a wide range of equipment configurations. This paper focuses on the application of the program to lumbar spine anteroposterior and lateral screen-film examinations. The program has been applied to study the variation of physical image quality measures and effective dose for changing system parameters such as tube voltage, grid design and screen-film system speed. These variations form the basis for optimization of these system parameters. In our approach to optimization, the best systems are those that can match (or come close to) the calculated image quality measure of systems preferred in a recent European clinical trial, but with lower patient dose. The largest dose savings found were 21% for a 400 speed class system with a grid having a strip density of 40 cm(-1) and a grid ratio of 16. A further dose saving of 13% was possible when a 600 speed class system was employed. The best systems found from the optimization correspond to those recommended by the European Commission guidelines on image quality criteria for diagnostic radiographic images.

Adult↗

Application of charge-coupled device technology for measurement of laser light and fluorescence distribution in tumors for photodynamic therapy.

Laser and fluorescence light distributions with applications for photodynamic therapy were measured in mouse tumors using a non-invasive electronic optical imaging system. The system consists of a liquid-nitrogen-cooled, charge-coupled-device (CCD) array camera under computer control with 576 x 384 detection elements having dimensions of 23 microns x 23 microns. The available dynamic range of the array is approx. 10(3), and the effective wavelength range is 400-1000 nm. An interstitially placed cylindrical diffusing optical fiber was used to provide tumor illumination. The light distribution pattern from the fiber was determined by immersing the cylindrical diffusing tip in a fluorescing solution and recording the emission image. Fluorescence imaging facilitates an accurate measurement of light intensity distribution while avoiding problems associated with the directional nature of other detection methods used with diffusing fibers. Radiation-induced fibrosarcoma tumors on C3H mice were grown to about 1 cm diameter for in vivo recording of light distribution from the tumor volume and for determination of effective light penetration distance at 18 wavelengths in the range 458-995 nm. Endogenous tumor fluorescence and Photofrin II fluorescence intensity were measured over the wavelength range 585-725 nm to investigate the possible application of CCD imaging technology for drug distribution measurements. Model experiments were begun to evaluate the relative importance of potential distortions of light distribution measurements using this approach.

Animals↗

Postnatal changes in intrinsic optical responses to peripheral nerve stimulation in the in vivo rat spinal cord.

We previously applied an intrinsic optical imaging technique to the in vivo rat spinal cord, and showed its feasibility for analyzing neural functions. Three- to 7-week-old postnatal and adult (>10-week-old) rats were anesthetized and laminectomy was performed between C5 and Th1 to expose the dorsal surface of the cervical spinal cord. Optical reflectance changes in response to simultaneous stimulation of the median and ulnar nerves were recorded from the spinal cord at a depth corresponding to the dorsal horn (substantia gelatinosa), using a differential video acquisition system. The shape and extent of the area of the evoked optical signals varied during postnatal development. In 3- to 4-week-old rats, the response area was small and ellipsoidal, and constrictions between spinal segments were not clear. As postnatal ages proceeded, the area expanded in a rostrocaudal direction, and the constrictions became clear. In adult rats, the response area exhibited a segmental pattern. At all ages, the optical responses showed trial-to-trial variations. A semiquantitative analysis of the variations in one spinal segment showed that the variations in a mediolateral direction appeared to decrease slightly with development, although statistical differences were insignificant between ages. The dynamical changes in optical response patterns were discussed in relation to postnatal maturation of neural functions in the spinal cord.

Aging↗

GABA(B)-mediated presynaptic inhibition of excitatory transmission and synaptic vesicle dynamics in cultured hippocampal neurons.

Local recycling of synaptic vesicle membrane at nerve terminals is necessary to maintain a readily releasable pool of transmitter. To what extent are the dynamics of vesicle recycling subject to modulation? We examined the influence of presynaptic GABA(B) receptors on vesicle dynamics at single synapses using optical imaging of FM1-43 in cultured rat hippocampal neurons. The kinetics of FM1-43 destaining indicate that synapses from a single neuron have a unimodal distribution of release probabilities, and GABA(B)-mediated inhibition occurs uniformly at all sites. Electrical and optical recordings from single cells show that the inhibition of excitatory transmission is entirely accounted for by a rapidly reversible reduction of exocytosis. In contrast, GABA(B) receptors do not alter the rate or extent of endocytosis.

2-Amino-5-phosphonovalerate↗

Description of the third-order optical aberrations of near-circular pupil optical systems without symmetry.

Many authors, dating back to at least the 1950s, have presented mathematical expansions of the wave-front aberration function for optical systems without symmetry, typically based on limiting assumptions and simplifications, with some of the most recent work being done by Howard and Stone [Appl. Opt. 39, 3232 (2000)]. This paper reveals that in fact there are no new aberrations in imaging optical systems with near-circular aperture stops but otherwise without symmetry. What does occur is that the field dependence of an aberration often changes when symmetry is abandoned. Each aberration type develops a characteristic field behavior in a system without symmetry. Specifically, for example, astigmatism, develops a binodal field dependence; e.g., there are typically two points in the field with zero astigmatism, and typically neither point is on axis. This construct, nodal aberration theory, for understanding the aberrations in systems without symmetry becomes a direct extension of an optical designer's knowledge base. Through the use of real ray-based analysis methods, such as Zernike coefficients, it is possible to understand completely the aberrations of optical systems without symmetry in terms of rotationally symmetric aberration theory with the simple addition of the concept of field nodes.

Journal Article↗

Acoustic microscopy--principles and applications in the studies of biomaterial microstructure.

The acoustic microscopy is based on totally different physical concepts than both the optical and electron microscopes. Therefore, the information obtained with an acoustic microscope is significantly different from those obtainable by the other two. By employing elastic waves, detailed microscopic information regarding biomaterial properties like density, elasticity, viscosity and viscoelasticity are obtained with an acoustic microscope. Live tissue can be examined without fixing or chemical staining. The Scanning Laser Acoustic Microscope (SLAM) and the Scanning Acoustic Microscope (SAM) are the only acoustic microscopes commercially available today. The SAM operates at as high as 4.2 GHz frequency providing at least five times better resolution than the optical limit. The SLAM, however, operates generally at 100 or 500 MHz and can therefore examine thicker samples than the SAM, but at lower resolutions. For optically translucent samples, the SLAM can also provide an optical image simultaneously with the acoustical one. A laser beam is employed in the scanning mechanism in the SLAM, compared to mechanically translating the sample in the SAM. Consequently, the scanning rate is much faster in the SLAM at about 30 frames per second, compared to several seconds per frame for the SAM. Both qualitative as well as quantitative information is obtainable about the material under examination. As a qualitative tool, the information is used to classify and sort materials and detect and localize flaws and defects in optically opaque samples. A microscopic map of the specimen's mechanical properties can be produced using information from the acoustic image. This information may then lead to better understanding of biomaterial microstructure and also can be a valuable aid in characterization of certain subtle morphological differences.

Animals↗

Combined imaging and chemical sensing of L-glutamate release from the foregut plexus of the lepidopteran, Manduca sexta.

A new combined imaging and chemical detection sensor for the measurement of localized L-glutamate release at the insect neuromuscular junction (NMJ) is presented. The sensor is comprised of an L-glutamate-sensitive fluorescent gel, spin-coated onto the tip of an optical imaging fiber. The gel is composed of L-glutamate oxidase (GLOD); a pH-sensitive fluorescent dye, SNAFL; and poly(acrylamide-co-N-acryloxysuccinimide) (PAN). NH(3) is liberated from the interaction of L-glutamate with GLOD, which reversibly reduces the emitted fluorescence signal from SNAFL. This sensor has a spatial resolution of 3-4 micro m, and an L-glutamate detection limit of between 10 and 100 micro M. L-glutamate release and re-uptake from the foregut plexus of Manduca sexta was detected by the sensor in the presence of the L-glutamate re-uptake blocker dihydrokainate, and the post-synaptic L-glutamate receptor antagonist CNQX.

Animals↗

Single-molecule near-field optical energy transfer microscopy with dielectric tips.

The fluorescence lifetime and the fluorescence rate of single molecules are recorded as a function of the position of a Si3N4 atomic force microscopy tip with respect to the molecule. We observe a decrease of the excited state lifetime and the fluorescence rate when the tip apex is in close proximity to the molecule. These effects are attributed to the fact that the dielectric tip converts non-propagating near-fields to propagating fields within the dielectric tip effectively quenching the fluorescence. The spatial extension of the quenching area is of subwavelength dimensions. The results are discussed in terms of molecular fluorescence in a system of stratified media. The experiment provides surprising new insights into the interactions between a fluorescent molecule and a dielectric tip. The methodology holds promise for applications in ultra high-resolution near-field optical imaging at the level of single fluorophores.

Carbocyanines↗

Trapping cavitation bubbles with a self-focused laser beam.

We observed that laser-induced cavitation bubbles in water can be trapped in a self-focused laser beam. Both optical imaging and acoustic detection have been utilized to confirm bubble trapping. Transverse and longitudinal trapping forces were measured to be as large as 87 and 11 pN, respectively. This result is contrary to conventional wisdom, since the mechanism of trapping in conventional optical tweezers implies that a low-index particle (a bubble being the limiting case) should be antitrapped.

Equipment Design↗

Human cortical areas underlying the perception of optic flow: brain imaging studies.

In summary, we have reviewed electrophysiological and brain imaging studies of motion and optic-flow processing. Single-unit studies indicate that MST (V5a) is a site of optic-flow extraction and that this information can be used to guide pursuit eye movements and to estimate heading. The EEG and MEG studies point to a localized electrical dipole in occipitotemporal cortex evoked by visual motion. We have also discussed the evidence from functional imaging studies for response specificity of the rCBF and BOLD effects in posterior cortex to visual motion and optic flow. Focal attention modulates the amplitude of the BOLD signal evoked by visual motion stimulation. Retinotopic mapping techniques have been used to locate region borders within the visual cortex. Our results indicate that striate (V1) and extrastriate areas (V2, V3/V3a) respond robustly to optic flow. However, with exception of a more pronounced response in V3/V3a to random walk, we found little evidence for response selectivity with respect to flow type and disparity in these early visual areas. In a similar fashion, the human V5/V5a complex responds well to optic flow, but these responses do not vary significantly with the type of flow field and do not seem to depend on disparity. In contrast, the kinetic occipital area (KO/V3b) responds well to optic-flow information, and it is the only area that produces more pronounced activation to the disparity in the flow fields. These initial results are promising because they suggest that the fMRI method can be sensitive to changes in stimulus parameters that define flow fields. More work will be required to explore the extent to which these responses reflect the neuronal processing of optic flow. Eye position tracking is now possible during fMRI experiments. We have demonstrated that the eye movements affect the BOLD responses in motion-sensitive areas (Kimming et al., 1999). Further experiments in our laboratory are aimed at understanding the effects of eye movements on the neuronal coding of complex optic-flow fields (Schira et al., 1999).

Animals↗

Magnetic resonance imaging with an optical atomic magnetometer.

We report an approach for the detection of magnetic resonance imaging without superconducting magnets and cryogenics: optical atomic magnetometry. This technique possesses a high sensitivity independent of the strength of the static magnetic field, extending the applicability of magnetic resonance imaging to low magnetic fields and eliminating imaging artifacts associated with high fields. By coupling with a remote-detection scheme, thereby improving the filling factor of the sample, we obtained time-resolved flow images of water with a temporal resolution of 0.1 s and spatial resolutions of 1.6 mm perpendicular to the flow and 4.5 mm along the flow. Potentially inexpensive, compact, and mobile, our technique provides a viable alternative for MRI detection with substantially enhanced sensitivity and time resolution for various situations where traditional MRI is not optimal.

Magnetic Resonance Imaging↗

In vivo imaging and differential localization of lipid-modified GFP-variant fusions in embryonic stem cells and mice.

The visualization of live cell behaviors operating in situ combined with the power of mouse genetics represents a major step toward understanding the mechanisms regulating embryonic development, homeostasis, and disease progression in mammals. The availability of genetically encoded fluorescent protein reporters, combined with improved optical imaging modalities, have led to advances in our ability to examine cells in vivo. We developed a series of lipid-modified fluorescent protein fusions that are targeted to and label the secretory pathway and the plasma membrane, and that are amenable for use in mice. Here we report the generation of two strains of mice, each expressing a spectrally distinct lipid-modified GFP-variant fluorescent protein fusion. The CAG::GFP-GPI strain exhibited widespread expression of a glycosylphosphatidylinositol-tagged green fluorescent protein (GFP) fusion, while the CAG::myr-Venus strain exhibited widespread expression of a myristoyl-Venus yellow fluorescent protein fusion. Imaging of live transgenic embryonic stem (ES) cells, either live or fixed embryos and postnatal tissues demonstrated that glycosylphosphatidyl inositol- and myristoyl-tagged GFP-variant fusion proteins are targeted to and serve as markers of the plasma membrane. Moreover, our data suggest that these two lipid-modified protein fusions are dynamically targeted both to overlapping as well as distinct lipid-enriched compartments within cells. These transgenic strains not only represent high-contrast reporters of cell morphology and plasma membrane dynamics, but also may be used as in vivo sensors of lipid localization. Furthermore, combining these reporters with the study of mouse mutants will be a step forward in understanding the inter- and intracellular behaviors underlying morphogenesis in both normal and mutant contexts.

Animals↗

Dynamic imaging of collagen and its modulation in tumors in vivo using second-harmonic generation.

The content and structure of collagen is essential in governing the delivery of therapeutic molecules in tumors. Thus, simple histological staining of tumor tissue biopsies for collagen could be used to assess the accessibility of molecular therapeutics in tumors. Here we show that it is possible to optically image fibrillar collagen in tumors growing in mice using second-harmonic generation (SHG). Using this noninvasive technique, we estimated relative diffusive hindrance, quantified the dynamics of collagen modification after pharmacologic intervention and provided mechanistic insight into improved diffusive transport induced by the hormone relaxin. This technology could offer basic scientists and clinicians an enhanced ability to estimate the relative penetrabilities of molecular therapeutics.

Animals↗