2012 02 18 Ternary resin-bound dynamic combinatorial chemistry. The ability to carry out simultaneous orthogonal exchange chemistries has opened new opportunities for increasing the numerical and structural diversity accessible to Dynamic Combinatorial Chemistry. We present proof-of-concept experiments demonstrating this concept is transferrable to resin-bound DCC, facilitating the generation and analysis of libraries with greater structural diversity. 48 2131-3 AV Gromova JM Ciszewski BL Miller /people/?u=21977435 2012 02 07 Model-based elastography: a survey of approaches to the inverse elasticity problem. Elastography is emerging as an imaging modality that can distinguish normal versus diseased tissues via their biomechanical properties. This paper reviews current approaches to elastography in three areas-quasi-static, harmonic and transient-and describes inversion schemes for each elastographic imaging approach. Approaches include first-order approximation methods; direct and iterative inversion schemes for linear elastic; isotropic materials and advanced reconstruction methods for recovering parameters that characterize complex mechanical behavior. The paper's objective is to document efforts to develop elastography within the framework of solving an inverse problem, so that elastography may provide reliable estimates of shear modulus and other mechanical parameters. We discuss issues that must be addressed if model-based elastography is to become the prevailing approach to quasi-static, harmonic and transient elastography: (1) developing practical techniques to transform the ill-posed problem with a well-posed one; (2) devising better forward models to capture the complex mechanical behavior of soft tissues and (3) developing better test procedures to evaluate the performance of modulus elastograms. 57 R35-73 MM Doyley http://www.ece.rochester.edu/html/people/Doyley/Doyley.html 2012 02 02 Microfluidic nanoplasmonic-enabled device for multiplex DNA detection. We describe a rapid, quantitative, multiplex, self-labelled, and real-time DNA biosensor employing Ag nanoparticle-bound DNA hairpin probes immobilized in a microfluidic channel. Capture of complementary target DNAs by the microarrayed DNA hairpin probes results in a positive fluorescence signal via a conformational change of the probe molecules, signalling the presence of target DNAs. The device's capability for quantitative analyses was evaluated and a detection time as low as 6 min (with a target flow rate of 0.5 μl min(-1)) was sufficient to generate significant detection signals. This detection time translates to merely 3 μl of target solution consumption. An unoptimized sensitivity of 500 pM was demonstrated for this device. HI Peng CM Strohsahl BL Miller /people/?u=21977435 2012 02 01 Integration of crawling waves in an ultrasound imaging system. Part 2: signal processing and applications. This paper introduces methods to generate crawling wave interference patterns from the displacement fields generated from radiation force pushes on a GE Logiq 9 scanner. The same transducer and system provides both the pushing pulses to generate the shear waves and the tracking pulses to measure the displacements. Acoustic power and system limitations result in largely impulsive displacement fields. Measured displacements from pushes on either side of a region-of-interest (ROI) are used to calculate continuously varying interference patterns. This technique is explained along with a brief discussion of the conventional mechanical source-driven crawling waves for comparison. We demonstrate the method on three example cases: a gelatin-based phantom with a cylindrical inclusion, an oil-gelatin phantom and mouse livers. The oil-gelatin phantom and the mouse livers demonstrate not only shear speed estimation, but the frequency dependence of the shear wave speeds. 38 312-23 Z Hah C Hazard B Mills C Barry D Rubens /people/?u=21432965 K Parker 2012 02 01 Shear wave dispersion measures liver steatosis. Crawling waves, which are interfering shear wave patterns, can be generated in liver tissue over a range of frequencies. Some important biomechanical properties of the liver can be determined by imaging the crawling waves using Doppler techniques and analyzing the patterns. We report that the dispersion of shear wave velocity and attenuation, that is, the frequency dependence of these parameters, are strongly correlated with the degree of steatosis in a mouse liver model, ex vivo. The results demonstrate the possibility of assessing liver steatosis using noninvasive imaging methods that are compatible with color Doppler scanners and, furthermore, suggest that liver steatosis can be separated from fibrosis by assessing the dispersion or frequency dependence of shear wave propagations. 38 175-82 CT Barry B Mills Z Hah RA Mooney CK Ryan DJ Rubens /people/?u=21432965 KJ Parker http://www.ece.rochester.edu/html/people/Parker/Parker.html 2012 02 01 Unique angiogenic and vasculogenic properties of renal cell carcinoma in a xenograft model of bone metastasis are associated with high levels of vegf-a and decreased ang-1 expression. Management of various tumor metastases to bone has dramatically improved, but this is not so for renal cell carcinoma (RCC), which is a difficult surgical problem due to its great vascularity. Furthermore, the unique mechanisms that mediate RCC vasculogenesis in bone remain unknown. To understand this process we developed a xenograft model that recapitulates highly vascular RCC versus less vascular tumors that metastasize to bone. Human tumor cell lines of RCC (786-O), prostate cancer (PC3), lung cancer (A549), breast cancer (MDA-MB231), and melanoma (A375) were transduced with firefly luciferase (Luc), injected into the tibiae of nude mice, and differences in growth, osteolysis, and vascularity were assessed by longitudinal bioluminescent imaging, micro-CT for measurement of calcified tissues and vascularity and histology. The results showed that while RCC-Luc has reduced growth and osteolytic potential versus the other tumor lines, it displayed a significant increase in vascular volume (p < 0.05). This expansion was due to 3- and 5-fold increases in small and large vessel numbers respectively. In vitro gene expression profiling revealed that RCC-Luc expresses significantly (p < 0.05) more vegf-a (10-fold) and 20- to 30-fold less ang-1 versus the other lines. These data demonstrate the utility of this model to study the unique vasculogenic properties of RCC bone metastases. 30 325-33 C Xie EM Schwarz /people/?u=22215734 ER Sampson RS Dhillon D Li RJ O'Keefe W Tyler 2012 02 01 A fast majorize-minimize algorithm for the recovery of sparse and low-rank matrices. We introduce a novel algorithm to recover sparse and low-rank matrices from noisy and undersampled measurements. We pose the reconstruction as an optimization problem, where we minimize a linear combination of data consistency error, nonconvex spectral penalty, and nonconvex sparsity penalty. We majorize the nondifferentiable spectral and sparsity penalties in the criterion by quadratic expressions to realize an iterative three-step alternating minimization scheme. Since each of these steps can be evaluated either analytically or using fast schemes, we obtain a computationally efficient algorithm. We demonstrate the utility of the algorithm in the context of dynamic magnetic resonance imaging (MRI) reconstruction from sub-Nyquist sampled measurements. The results show a significant improvement in signal-to-noise ratio and image quality compared with classical dynamic imaging algorithms. We expect the proposed scheme to be useful in a range of applications including video restoration and multidimensional MRI. 21 742-53 Y Hu SG Lingala M Jacob /bme/people/faculty/bio/?id=243 2012 02 01 Hyperintensity on diffusion weighted image along ipsilateral cortical spinal tract after cerebral ischemic stroke: A diffusion tensor analysis. Hyperintensity along the ipsilateral cortical spinal tract (CST) on a diffusion weighted imaging (DWI) has been reported to may be associated with motor disability after brain infarction and can be misdiagnosed as a new infarction. However, the underlying patho-physiology related to this finding is not clear. The goal of our study was to analyze the diffusion tensor imaging (DTI) changes in patients with this hyperintensity. 81 292-7 X Liu W Tian L Li B Kolar X Qiu F Chen VS Dogra /people/?u=23018716 2012 01 19 Two-Photon and Second Harmonic Microscopy in Clinical and Translational Cancer Research. Application of two-photon microscopy (TPM) to translational and clinical cancer research has burgeoned over the last several years, as several avenues of pre-clinical research have come to fruition. In this review, we focus on two forms of TPM-two-photon excitation fluorescence microscopy, and second harmonic generation microscopy-as they have been used for investigating cancer pathology in ex vivo and in vivo human tissue. We begin with discussion of two-photon theory and instrumentation particularly as applicable to cancer research, followed by an overview of some of the relevant cancer research literature in areas that include two-photon imaging of human tissue biopsies, human skin in vivo, and the rapidly developing technology of two-photon microendoscopy. We believe these and other evolving two-photon methodologies will continue to help translate cancer research from the bench to the bedside, and ultimately bring minimally invasive methods for cancer diagnosis and treatment to therapeutic reality. SW Perry /bme/people/faculty/bio/?id=458 RM Burke EB Brown /bme/people/faculty/bio/?id=232 2012 01 17 Pro-apoptotic gene knockdown mediated by nanocomplexed siRNA reduces radiation damage in primary salivary gland cultures. A critical issue in the management of head and neck tumors is radioprotection of the salivary glands. We have investigated whether siRNA-mediated gene knock down of pro-apoptotic mediators can reduce radiation-induced cellular apoptosis in salivary gland cells in vitro. We used novel, pH-responsive nanoparticles to deliver functionally active siRNAs into cultures of salivary gland cells. The nanoparticle molecules are comprised of cationic micelles that electrostatically interact with the siRNA, protecting it from nuclease attack, and also include pH-responsive endosomolytic constituents that promote release of the siRNA into the target cell cytoplasm. Transfection controls with Cy3-tagged siRNA/nanoparticle complexes showed efficiently internalized siRNAs in more than 70% of the submandibular gland cells. We found that introduction of siRNAs specifically targeting the Pkcδ or Bax genes significantly blocked the induction of these pro-apoptotic proteins that normally occurs after radiation in cultured salivary gland cells. Furthermore, the level of cell death from subsequent radiation, as measured by caspase-3, TUNEL, and mitochondrial disruption assays, was significantly decreased. Thus, we have successfully demonstrated that the siRNA/ nanoparticle-mediated knock down of pro-apoptotic genes can prevent radiation-induced damage in submandibular gland primary cell cultures. J. Cell. Biochem. © 2012 Wiley Periodicals, Inc. S Arany Q Xu E Hernady DS Benoit /bme/people/faculty/bio/?id=385 S Dewhurst CE Ovitt 2012 01 06 Artifact versus reality-How astrocytes contribute to synaptic events? The neuronal doctrine, developed a century ago regards neuronal networks as the sole substrate of higher brain function. Recent advances in glial physiology have promoted an alternative hypothesis, which places information processing in the brain into integrated neuronal-glial networks utilizing both binary (neuronal action potentials) and analogue (diffusional propagation of second messengers/metabolites through gap junctions or transmitters through the interstitial space) signal encoding. It has been proposed that the feed-forward and feed-back communication between these two types of neural cells, which underlies information transfer and processing, is accomplished by the release of neurotransmitters from neuronal terminals as well as from astroglial processes. Understanding of this subject, however, remains incomplete and important questions and controversies require resolution. Here we propose that the primary function of perisynaptic glial processes is to create an "astroglial cradle" that shields the synapse from a multitude of extrasynaptic signaling events and provides for multifaceted support and long-term plasticity of synaptic contacts through variety of mechanisms, which may not necessarily involve the release of "glio" transmitters. © 2012 Wiley Periodicals, Inc. M Nedergaard /people/?u=23788299 A Verkhratsky 2012 01 04 Representation of 3D surface orientation by velocity and disparity gradient cues in area MT. Neural coding of the 3D orientation of planar surface patches may be an important intermediate step in constructing representations of complex 3D surface structure. Spatial gradients of binocular disparity, image velocity, and texture provide potent cues to the 3D orientation (tilt and slant) of planar surfaces. Previous studies have described neurons in both dorsal and ventral stream areas that are selective for surface tilt based on one or more of these gradient cues. However, relatively little is known about whether single neurons provide consistent information about surface orientation from multiple gradient cues. Moreover, it is unclear how neural responses to combinations of surface orientation cues are related to responses to the individual cues. We measured responses of middle temporal (MT) neurons to random-dot stimuli that simulated planar surfaces at a variety of tilts and slants. Four cue conditions were tested: disparity, velocity, and texture gradients alone, as well all three gradient cues combined. Many neurons showed robust tuning for surface tilt based on disparity and velocity gradients, with relatively little selectivity for texture gradients. Some neurons showed consistent tilt preferences for disparity and velocity cues, whereas others showed large discrepancies. Responses to the combined stimulus were generally well described as a weighted linear sum of responses to the individual cues, even when disparity and velocity preferences were discrepant. These findings suggest that area MT contains a rudimentary representation of 3D surface orientation based on multiple cues, with single neurons implementing a simple cue integration rule. TM Sanada JD Nguyenkim GC Deangelis http://www.bcs.rochester.edu/people/gdeangelis/ 2012 01 01 Epidermoid cyst of the zygomatic root. 33 e1-2 RE Roditi BT Crane /people/?u=27325469 2012 01 01 The susceptibility of the retina to photochemical damage from visible light. The photoreceptor/RPE complex must maintain a delicate balance between maximizing the absorption of photons for vision and retinal image quality while simultaneously minimizing the risk of photodamage when exposed to bright light. We review the recent discovery of two new effects of light exposure on the photoreceptor/RPE complex in the context of current thinking about the causes of retinal phototoxicity. These effects are autofluorescence photobleaching in which exposure to bright light reduces lipofuscin autofluorescence and, at higher light levels, RPE disruption in which the pattern of autofluorescence is permanently altered following light exposure. Both effects occur following exposure to visible light at irradiances that were previously thought to be safe. Photopigment, retinoids involved in the visual cycle, and bisretinoids in lipofuscin have been implicated as possible photosensitizers for photochemical damage. The mechanism of RPE disruption may follow either of these paths. On the other hand, autofluorescence photobleaching is likely an indicator of photooxidation of lipofuscin. The permanent changes inherent in RPE disruption might require modification of the light safety standards. AF photobleaching recovers after several hours although the mechanisms by which this occurs are not yet clear. Understanding the mechanisms of phototoxicity is all the more important given the potential for increased susceptibility in the presence of ocular diseases that affect either the visual cycle and/or lipofuscin accumulation. In addition, knowledge of photochemical mechanisms can improve our understanding of some disease processes that may be influenced by light exposure, such as some forms of Leber's congenital amaurosis, and aid in the development of new therapies. Such treatment prior to intentional light exposures, as in ophthalmic examinations or surgeries, could provide an effective preventative strategy. 31 28-42 JJ Hunter JI Morgan WH Merigan DH Sliney JR Sparrow DR Williams http://www.cvs.rochester.edu/williamslab/p_williams.html 2012 01 01 Neural correlates of reliability-based cue weighting during multisensory integration. Integration of multiple sensory cues is essential for precise and accurate perception and behavioral performance, yet the reliability of sensory signals can vary across modalities and viewing conditions. Human observers typically employ the optimal strategy of weighting each cue in proportion to its reliability, but the neural basis of this computation remains poorly understood. We trained monkeys to perform a heading discrimination task from visual and vestibular cues, varying cue reliability randomly. The monkeys appropriately placed greater weight on the more reliable cue, and population decoding of neural responses in the dorsal medial superior temporal area closely predicted behavioral cue weighting, including modest deviations from optimality. We found that the mathematical combination of visual and vestibular inputs by single neurons is generally consistent with recent theories of optimal probabilistic computation in neural circuits. These results provide direct evidence for a neural mechanism mediating a simple and widespread form of statistical inference. 15 146-54 CR Fetsch A Pouget GC DeAngelis http://www.bcs.rochester.edu/people/gdeangelis/ DE Angelaki 2012 01 01 Extracellular Ca2+ acts as a mediator of communication from neurons to glia. Defining the pathways through which neurons and astrocytes communicate may contribute to the elucidation of higher central nervous system functions. We investigated the possibility that decreases in extracellular calcium ion concentration ([Ca(2+)](e)) that occur during synaptic transmission might mediate signaling from neurons to glia. Using noninvasive photolysis of the photolabile Ca(2+) buffer diazo-2 {N-[2-[2-[2-[bis(carboxymethyl)amino]-5-(diazoacetyl)phenoxy]ethoxy]-4-methylphenyl]-N-(carboxymethyl)-, tetrapotassium salt} to reduce [Ca(2+)](e) or caged glutamate to simulate glutamatergic transmission, we found that a local decline in extracellular Ca(2+) triggered astrocytic adenosine triphosphate (ATP) release and astrocytic Ca(2+) signaling. In turn, activation of purinergic P2Y1 receptors on a subset of inhibitory interneurons initiated the generation of action potentials by these interneurons, thereby enhancing synaptic inhibition. Thus, astrocytic ATP release evoked by an activity-associated decrease in [Ca(2+)](e) may provide a negative feedback mechanism that potentiates inhibitory transmission in response to local hyperexcitability. 5 ra8 A Torres F Wang Q Xu T Fujita R Dobrowolski K Willecke T Takano M Nedergaard /people/?u=23788299 2012 01 01 Heterogeneity of astrocytic form and function. Astrocytes participate in all essential CNS functions, including blood flow regulation, energy metabolism, ion and water homeostasis, immune defence, neurotransmission, and adult neurogenesis. It is thus not surprising that astrocytic morphology and function differ between regions, and that different subclasses of astrocytes exist within the same brain region. Recent lines of work also show that the complexity of protoplasmic astrocytes increases during evolution. Human astrocytes are structurally more complex, larger, and propagate calcium signals significantly faster than rodent astrocytes. In this chapter, we review the diversity of astrocytic form and function, while considering the markedly expanded roles of astrocytes with phylogenetic evolution. We also define major challenges for the future, which include determining how astrocytic functions are locally specified, defining the molecular controls upon astrocytic fate and physiology and establishing how evolutionary changes in astrocytes contribute to higher cognitive functions. 814 23-45 NA Oberheim SA Goldman M Nedergaard /people/?u=23788299 2012 01 01 Cultured astrocytes do not release adenosine during hypoxic conditions. Recent reports based on a chemiluminescent enzymatic assay for detection of adenosine conclude that cultured astrocytes release adenosine during mildly hypoxic conditions. If so, astrocytes may suppress neural activity in early stages of hypoxia. The aim of this study was to reevaluate the observation using high-performance liquid chromatography (HPLC). The HPLC analysis showed that exposure to 20 or 120 minutes of mild hypoxia failed to increase release of adenosine triphosphate (ATP), adenosine diphosphate (ADP), adenosine monophosphate (AMP), and adenosine from cultured astrocytes. Similar results were obtained using a chemiluminescent enzymatic assay. Moreover, since the chemiluminescent enzymatic assay relies on hydrogen peroxide generation, release of free-radical scavengers from hypoxic cells can interfere with the assay. Accordingly, adenosine added to samples collected from hypoxic cultures could not be detected using the chemiluminescent enzymatic assay. Furthermore, addition of free-radical scavengers sharply reduced the sensitivity of adenosine detection. Conversely, use of a single-step assay inflated measured values due to the inability of the assay to distinguish adenosine and its metabolite inosine. These results show that cultured astrocytes do not release adenosine during mild hypoxia, an observation consistent with their high resistance to hypoxia. 32 T Fujita EK Williams TK Jensen NA Smith T Takano K Tieu M Nedergaard /people/?u=23788299 2012 01 01 Slow-light dispersion in periodically patterned silicon microring resonators. We demonstrate a novel periodically patterned ring resonator evanescently coupled with a coupler waveguide (CWG) on a silicon-on-insulator platform. In order to optimize the coupling, we phase match the ring resonator and the CWG by tuning the width of the CWG. In the transmission spectra, we measure a high extinction ratio of more than 20 dB and achieve a group index of ∼20 in the slow-light regime. Beating patterns in the calculated mode profiles suggest strong interference between degenerate modes. This device opens up the possibility of new applications in compact device integration in wavelength-division multiplexing system while reducing the in-band four-wave mixing cross talk. 37 58-60 JY Lee PM Fauchet http://www.ece.rochester.edu/html/people/Fauchet/Fauchet.html 2011 12 07 Binocular disparity tuning and visual-vestibular congruency of multisensory neurons in macaque parietal cortex. Many neurons in the dorsal medial superior temporal (MSTd) and ventral intraparietal (VIP) areas of the macaque brain are multisensory, responding to both optic flow and vestibular cues to self-motion. The heading tuning of visual and vestibular responses can be either congruent or opposite, but only congruent cells have been implicated in cue integration for heading perception. Because of the geometric properties of motion parallax, however, both congruent and opposite cells could be involved in coding self-motion when observers fixate a world-fixed target during translation, if congruent cells prefer near disparities and opposite cells prefer far disparities. We characterized the binocular disparity selectivity and heading tuning of MSTd and VIP cells using random-dot stimuli. Most (70%) MSTd neurons were disparity selective with monotonic tuning, and there was no consistent relationship between depth preference and congruency of visual and vestibular heading tuning. One-third of disparity-selective MSTd cells reversed their depth preference for opposite directions of motion [direction-dependent disparity tuning (DDD)], but most of these cells were unisensory with no tuning for vestibular stimuli. Inconsistent with previous reports, the direction preferences of most DDD neurons do not reverse with disparity. By comparison to MSTd, VIP contains fewer disparity-selective neurons (41%) and very few DDD cells. On average, VIP neurons also preferred higher speeds and nearer disparities than MSTd cells. Our findings are inconsistent with the hypothesis that visual/vestibular congruency is linked to depth preference, and also suggest that DDD cells are not involved in multisensory integration for heading perception. 31 17905-16 Y Yang S Liu SA Chowdhury GC DeAngelis http://www.bcs.rochester.edu/people/gdeangelis/ DE Angelaki 2011 12 01 Measuring intranodal pressure and lymph viscosity to elucidate mechanisms of arthritic flare and therapeutic outcomes. Rheumatoid arthritis (RA) is a chronic autoimmune disease with episodic flares in affected joints; the etiology of RA is largely unknown. Recent studies in mice demonstrated that alterations in lymphatics from affected joints precede flares. Thus, we aimed to develop novel methods for measuring lymph node pressure and lymph viscosity in limbs of mice. Pressure measurements were performed by inserting a glass micropipette connected to a pressure transducer into popliteal lymph nodes (PLN) or axillary lymph nodes (ALN) of mice; subsequently, we determined that the lymphatic pressures of water were 9 and 12 cm, respectively. We are also developing methods for measuring lymph viscosity in lymphatic vessels afferent to PLN, which can be measured by multiphoton fluorescence recovery after photobleaching (MP-FRAP) of fluorescein isothiocyanate-labeled bovine serum albumin (FITC-BSA) injected into the hind footpad. These results demonstrate the potential of lymph node pressure and lymph viscosity measurements, and future studies to test these outcomes as biomarkers of arthritic flare are warranted. 1240 47-52 EM Bouta RW Wood SW Perry /bme/people/faculty/bio/?id=458 EB Brown /bme/people/faculty/bio/?id=232 CT Ritchlin L Xing EM Schwarz /people/?u=22215734 2011 12 01 β-Adrenergic receptors (β-AR) regulate VEGF and IL-6 production by divergent pathways in high β-AR-expressing breast cancer cell lines. Activation of β-adrenergic receptors (β-AR) drives proangiogenic factor production in several types of cancers. To examine β-AR regulation of breast cancer pathogenesis, β-AR density, signaling capacity, and functional responses to β-AR stimulation were studied in four human breast adenocarcinoma cell lines. β-AR density ranged from very low in MCF7 and MB-361 to very high in MB-231 and in a brain-seeking variant of MB-231, MB-231BR. Consistent with β-AR density, β-AR activation elevated cAMP in MCF7 and MB-361 much less than in MB-231 and MB-231BR. Functionally, β-AR stimulation did not markedly alter vascular endothelial growth factor (VEGF) production by MCF7 or MB-361. In the two high β-AR-expressing cell lines MB-231 and MB-231BR, β-AR-induced cAMP and VEGF production differed considerably, despite similar β-AR density. The β(2)-AR-selective agonist terbutaline and the endogenous neurotransmitter norepinephrine decreased VEGF production by MB-231, but increased VEGF production by MB-231BR. Moreover, β(2)-AR activation increased IL-6 production by both MB-231 and MB-231BR. These functional alterations were driven by elevated cAMP, as direct activation of adenylate cyclase by forskolin elicited similar alterations in VEGF and IL-6 production. The protein kinase A antagonist KT5720 prevented β-AR-induced alterations in MB-231 and MB-231BR VEGF production, but not IL-6 production. Conclusions β-AR expression and signaling is heterogeneous in human breast cancer cell lines. In cells with high β-AR density, β-AR stimulation regulates VEGF production through the classical β-AR-cAMP-PKA pathway, but this pathway can elicit directionally opposite outcomes. Furthermore, in the same cells, β-AR activate a cAMP-dependent, PKA-independent pathway to increase IL-6 production. The complexity of breast cancer cell β-AR expression and functional responses must be taken into account when considering β-AR as a therapeutic target for breast cancer treatment. 130 747-58 KS Madden /bme/people/faculty/bio/?id=43 MJ Szpunar EB Brown /bme/people/faculty/bio/?id=232 2011 12 01 Dissociating motor cortex from the motor. During closed-loop control of a brain-computer interface, neurons in the primary motor cortex can be intensely active even though the subject may be making no detectable movement or muscle contraction. How can neural activity in the primary motor cortex become dissociated from the movements and muscles of the native limb that it normally controls? Here we examine circumstances in which motor cortex activity is known to dissociate from movement--including mental imagery, visuo-motor dissociation and instructed delay. Many such motor cortex neurons may be related to muscle activity only indirectly. Furthermore, the integration of thousands of synaptic inputs by individual α-motoneurons means that under certain circumstances even cortico-motoneuronal cells, which make monosynaptic connections to α-motoneurons, can become dissociated from muscle activity. The natural ability of motor cortex neurons under voluntarily control to become dissociated from bodily movement may underlie the utility of this cortical area for controlling brain-computer interfaces. 589 5613-24 MH Schieber /people/?u=22085467 2011 11 28 Direct measurement of tissue blood flow and metabolism with diffuse optics. Diffuse optics has proven useful for quantitative assessment of tissue oxy- and deoxyhaemoglobin concentrations and, more recently, for measurement of microvascular blood flow. In this paper, we focus on the flow monitoring technique: diffuse correlation spectroscopy (DCS). Representative clinical and pre-clinical studies from our laboratory illustrate the potential of DCS. Validation of DCS blood flow indices in human brain and muscle is presented. Comparison of DCS with arterial spin-labelled MRI, xenon-CT and Doppler ultrasound shows good agreement (0.50<r<0.95) over a wide range of tissue types and source detector distances, corroborating the potential of the method to measure perfusion non-invasively and in vivo at the microvasculature level. All-optical measurements of cerebral oxygen metabolism in both rat brain, following middle cerebral artery occlusion, and human brain, during functional activation, are also described. In both situations, the use of combined DCS and diffuse optical spectroscopy/near-infrared spectroscopy to monitor changes in oxygen consumption by the tissue is demonstrated. Finally, recent results spanning from gene expression-induced angiogenic response to stroke care and cancer treatment monitoring are discussed. Collectively, the research illustrates the capability of DCS to quantitatively monitor perfusion from bench to bedside, providing results that match up both with literature findings and with similar experiments performed with other techniques. 369 4390-406 RC Mesquita T Durduran G Yu EM Buckley MN Kim C Zhou R Choe /bme/people/faculty/bio/?id=416 U Sunar AG Yodh 2011 11 21 Investigating the impact of spatial priors on the performance of model-based IVUS elastography. This paper describes methods that provide pre-requisite information for computing circumferential stress in modulus elastograms recovered from vascular tissue-information that could help cardiologists detect life-threatening plaques and predict their propensity to rupture. The modulus recovery process is an ill-posed problem; therefore, additional information is needed to provide useful elastograms. In this work, prior geometrical information was used to impose hard or soft constraints on the reconstruction process. We conducted simulation and phantom studies to evaluate and compare modulus elastograms computed with soft and hard constraints versus those computed without any prior information. The results revealed that (1) the contrast-to-noise ratio of modulus elastograms achieved using the soft prior and hard prior reconstruction methods exceeded those computed without any prior information; (2) the soft prior and hard prior reconstruction methods could tolerate up to 8% measurement noise, and (3) the performance of soft and hard prior modulus elastograms degraded when incomplete spatial priors were employed. This work demonstrates that including spatial priors in the reconstruction process should improve the performance of model-based elastography, and the soft prior approach should enhance the robustness of the reconstruction process to errors in the geometrical information. 56 7223-46 MS Richards MM Doyley http://www.ece.rochester.edu/html/people/Doyley/Doyley.html 2011 11 01 Vascularization of three-dimensional collagen hydrogels using ultrasound standing wave fields. The successful fabrication of large, three-dimensional (3-D) tissues and organs in vitro requires the rapid development of a vascular network to maintain cell viability and tissue function. In this study, we utilized an application of ultrasound standing wave field (USWF) technology to vascularize 3-D, collagen-based hydrogels in vitro. Acoustic radiation forces associated with USWF were used to noninvasively organize human endothelial cells into distinct, multicellular planar bands within 3-D collagen gels. The formation and maturation of capillary-like endothelial cell sprouts were monitored over time and compared with sham-exposed collagen constructs, which were characterized by a homogeneous cell distribution. USWF-induced cell banding accelerated the formation and elongation of capillary-like sprouts, promoted collagen fiber alignment and resulted in the maturation of endothelial cell sprouts into lumen-containing, anastomosing networks found throughout the entire volume of the collagen gel. USWF-induced endothelial cell networks contained large, arteriole-sized lumen areas that branched into smaller, capillary-sized structures indicating the development of vascular tree-like networks. In contrast, sprout formation was delayed in sham-exposed collagen gels and endothelial cell networks were absent from sham gel centers and failed to develop into the vascular tree-like structures found in USWF-exposed constructs. Our results demonstrate that USWF technology leads to rapid and extensive vascularization of 3-D collagen-based engineered tissue and, therefore, provide a new strategy to vascularize engineered tissues in vitro. 37 1853-64 KA Garvin D Dalecki /bme/people/faculty/bio/?id=242 DC Hocking /people/?u=22430199 2011 11 01 Shear strain from irrotational tissue displacements near bubbles. Particle displacements can be much greater near bubbles than they would be in a homogeneous liquid or tissue when exposed to an acoustic wave. In a plane wave, shear and bulk strains are of the same order of magnitude. In contrast, for a bubble oscillating close to its resonance frequency, the shear strain in the medium near the bubble is roughly four orders of magnitude greater than the bulk strain. This can lead to shear strains of a few percent even with acoustic excitation pressures far below the pressure thresholds required to cause inertial cavitation. High shear strains near oscillating bubbles could potentially be the cause of bioeffects. After acoustic exposures at audio frequencies, hemorrhages in tissues as diverse as lung, liver, and kidney have been observed at shear strains on the order of 1%. 130 3467-71 E Carstensen SM Gracewski http://www.me.rochester.edu/Faculty/grace.html D Dalecki /bme/people/faculty/bio/?id=242 2011 11 01 Natural frequencies of two bubbles in a compliant tube: analytical, simulation, and experimental results. Motivated by various clinical applications of ultrasound contrast agents within blood vessels, the natural frequencies of two bubbles in a compliant tube are studied analytically, numerically, and experimentally. A lumped parameter model for a five degree of freedom system was developed, accounting for the compliance of the tube and coupled response of the two bubbles. The results were compared to those produced by two different simulation methods: (1) an axisymmetric coupled boundary element and finite element code previously used to investigate the response of a single bubble in a compliant tube and (2) finite element models developed in comsol Multiphysics. For the simplified case of two bubbles in a rigid tube, the lumped parameter model predicts two frequencies for in- and out-of-phase oscillations, in good agreement with both numerical simulation and experimental results. For two bubbles in a compliant tube, the lumped parameter model predicts four nonzero frequencies, each asymptotically converging to expected values in the rigid and compliant limits of the tube material. 130 3347-56 NW Jang A Zakrzewski C Rossi D Dalecki /bme/people/faculty/bio/?id=242 S Gracewski http://www.me.rochester.edu/Faculty/grace.html 2011 11 01 Estimating axial and lateral strain using a synthetic aperture elastographic imaging system. Model-based elastography is an emerging technique with clinical applications in imaging vascular tissues, guiding minimally invasive therapies and diagnosing breast and prostate cancers. Its usage is limited because ultrasound can measure only the axial component of displacement with high precision. The goal of this study was to assess the effect of lateral sampling frequency, lateral beam-width and the number of active transmission elements on the quality of axial and lateral strain elastograms. Elastographic imaging was performed on gelatin-based phantoms with a modified commercial ultrasound scanner. Three groups of radio-frequency (RF) echo frames were reconstructed from fully synthetic aperture data. In the first group, all 128 transmission elements (corresponding to a lateral beamwidth of 0.22 mm at the center of the field of view) were used to reconstruct RF echo frames with A-line densities that varied from 6.4 lines/mm to 51.2 lines/mm. In the second group, the size of the aperture was varied to produce RF echo frames with lateral beamwidths ranging from 0.22 mm to 0.43 mm and a fixed A-line density of 25.6 lines/mm. In the third group, sparse arrays with varying number of active transmission elements (from 2 to 128) were used to reconstruct RF echo frames, whose A-line density and lateral beamwidth were fixed to 25.6 lines/mm and 0.22 mm, respectively. Applying a two-dimensional (2-D) displacement estimator to the pre- and post-deformed RF echo frames produced displacement elastograms. Axial and lateral strain elastograms were computed from displacement elastograms with a least squares strain estimator. The quality of axial and lateral strain elastograms improved with increasing applied strain and A-line density but decreased with increasing lateral beamwidth and deteriorated as the number of active transmission elements in the sparse arrays were reduced. This work demonstrated that the variance incurred when estimating the lateral component of displacement was reduced considerably when elastography was performed with a synthetic aperture ultrasound imaging system. Satisfactory axial and lateral strain elastograms were produced using a sparse array with as few as 16 active transmission elements. 37 1893-908 S Korukonda MM Doyley http://www.ece.rochester.edu/html/people/Doyley/Doyley.html 2011 10 26 Spatiotemporal variation of multiple neurophysiological signals in the primary motor cortex during dexterous reach-to-grasp movements. To examine the spatiotemporal distribution of discriminable information about reach-to-grasp movements in the primary motor cortex upper extremity representation, we implanted four microelectrode arrays in the anterior bank and lip of the central sulcus in each of two monkeys. We used linear discriminant analysis to compare information, quantified as decoding accuracy, contained in various neurophysiological signals. For all signal types, decoding accuracy increased immediately after the movement cue, peaked around movement onset, and declined during the static hold. Spike recordings and local field potential (LFP) time domain amplitude provided more discriminable information than LFP frequency domain power. Discriminable information on movement type was distributed evenly across recording sites by LFP amplitude and 1-4 Hz power but unevenly by 100-170 Hz power and spike recordings. These latter two signal types provided higher decoding accuracies closer to the hemispheric surface than deep in the anterior bank and also provided accuracies that varied along the central sulcus. This variation in the distribution of movement-type information may be related to differences in the rostral versus caudal regions of the primary motor cortex and to its underlying somatotopic organization. The even distribution of information by LFP amplitude and 1-4 Hz power compared with the more localized distribution by 100-170 Hz power and spikes suggest that these different neurophysiological signals reflect different underlying processes that distribute information through the motor cortex during reach-to-grasp movements. 31 15531-43 M Mollazadeh V Aggarwal AG Davidson AJ Law NV Thakor MH Schieber /people/?u=22085467 2011 10 10 Regulation of human osteoclast development by dendritic cell-specific transmembrane protein (DC-STAMP). Osteoclasts (OC) are bone-resorbing, multinucleated cells that are generated via fusion of OC precursors (OCP). The frequency of OCP is elevated in patients with erosive inflammatory arthritis and metabolic bone diseases. Although many cytokines and cell surface receptors are known to participate in osteoclastogenesis, the molecular mechanisms underlying the regulation of this cellular transformation are poorly understood. Herein, we focused our studies on the dendritic cell-specific transmembrane protein (DC-STAMP), a seven-pass-transmembrane receptor-like protein known to be essential for cell-to-cell fusion during osteoclastogenesis. We identified an immunoreceptor tyrosine-based inhibitory motif (ITIM) in the cytoplasmic tail of DC-STAMP, and developed an anti-DC-STAMP monoclonal antibody 1A2 that detected DC-STAMP expression on human tumor giant cells, blocked OC formation in vitro, and distinguished four patterns of human PBMC with a positive correlation to OC potential. In freshly isolated monocytes, DC-STAMP(high) cells produced a higher number of OC in culture than DC-STAMPlow cells and the surface expression of DC-STAMP gradually declined during osteoclastogenesis. Importantly, we showed that DC-STAMP is phosphorylated on its tyrosine residues and physically interacts with SHP-1 and CD16, an SH2-domain-containing tyrosine phosphatase and an ITAM-associated protein, respectively. Taken together, these data show that DC-STAMP is a potential OCP biomarker in inflammatory arthritis. Moreover, in addition to its effect on cell fusion, DC-STAMP dynamically regulates cell signaling during osteoclastogenesis. © 2011 American Society for Bone and Mineral Research. YH Chiu KA Mensah EM Schwarz /people/?u=22215734 Y Ju M Takahata C Feng LA McMahon DG Hicks B Panepento PC Keng CT Ritchlin 2011 10 01 Temporal bone findings in a case of Susac's syndrome. To describe the histopathologic findings in the temporal bones of a patient with Susac's syndrome (SS). 32 1198-204 HW Francis C Makary C Halpin BT Crane /people/?u=27325469 SN Merchant 2011 10 01 Gaze shift duration, independent of amplitude, influences the number of spikes in the burst for medium-lead burst neurons in pontine reticular formation. Changes in the direction of the line of sight (gaze) allow successive sampling of the visual environment. Saccadic eye movements accomplish this goal when the head does not move. Medium-lead burst neurons (MLBs) in the paramedian pontine reticular formation (PPRF) discharge a high frequency burst of action potentials starting ~12 ms before the saccade begins. A subgroup of MLBs rostral of abducens nucleus monosynaptically excites oculomotor neurons. The number of spikes in the presaccadic burst is correlated with the amplitude of the horizontal component of the saccade, and the peak discharge rate is correlated with peak eye velocity. During head-unrestrained gaze shifts, a linear relationship between the number of action potentials in MLB bursts and gaze (but not eye) amplitude has been reported. The anatomical connection of MLBs to motor neurons and the similarity between the phasic motor neuron burst and MLB discharge have raised questions about the usefulness of counting spikes in MLBs to determine their role in eye-head coordination. We investigated this issue using a behavioral technique that permits a dissociation of eye movement amplitude and duration during constant vector gaze shifts. Surprisingly, during gaze shifts of constant amplitude and direction, we observe a nearly linear, positive correlation between saccade duration and spike number associated with a negative correlation between spike number and saccade amplitude. These data constrain models of the oculomotor controller and may further define the time-dependence of hypothesized neural integration in this system. 214 225-39 MM Walton EG Freedman /people/?u=23038240 2011 10 01 Tumor necrosis factor inhibits mesenchymal stem cell differentiation into osteoblasts via the ubiquitin E3 ligase Wwp1. Patients with chronic inflammatory disorders, such as rheumatoid arthritis, often have osteoporosis due to a combination of Tumor necrosis factor-induced increased bone resorption and reduced bone formation. To test if TNF inhibits bone formation by affecting the commitment and differentiation of mesenchymal stem cells (MSCs) into osteoblasts, we examined the osteogenic potential of MSCs from TNF transgenic (TNF-Tg) mice, a model of chronic inflammatory arthritis. MSC-enriched cells were isolated from bone marrow stromal cells using negative selection with anti-CD45 antibody coated magnetic beads. The expression profile of MSC surface markers the osteogenic, chondrogenic, and adipogenic properties of CD45(-) cells were confirmed by FACS and cell differentiation assays. MSC-enriched CD45(-) cells from TNF-Tg mice formed significantly decreased numbers of fibroblast and ALP(+) colonies and had a decreased expression of osteoblast marker genes. As TNF may upregulate ubiquitin ligases, which negatively regulate osteoblast differentiation, we examined the expression levels of several ubiquitin ligases and found that Wwp1 expression was significantly increased in MSC-enriched CD45(-) cells of TNF-Tg mice. Wwp1 knockdown rescued impaired osteoblast differentiation of TNF-Tg CD45(-) cells. Wwp1 promotes ubiquitination and degradation of JunB, an AP-1 transcription factor that positively regulates osteoblast differentiation. Injection of TNF into wild-type mice resulted in decreased osteoblast differentiation of MSCs and increased JunB ubiquitination, which was completely blocked in Wwp1(-/-) mice. Thus, Wwp1 targets JunB for ubiquitination and degradation in MSCs after chronic exposure to TNF, and inhibition of Wwp1 in MSCs could be a new mechanism to limit inflammation-mediated osteoporosis by promoting their differentiation into osteoblasts. 29 1601-10 L Zhao J Huang H Zhang Y Wang LE Matesic M Takahata H Awad /bme/people/faculty/bio/?id=240 D Chen L Xing 2011 09 28 Delayed short course treatment with teriparatide (PTH(1-34) ) improves femoral allograft healing by enhancing intramembranous bone formation at the graft-host junction. Clinical management of critical bone defects remains a major challenge. Despite pre-clinical work demonstrating teriparatide (PTH(1-34) ) effectiveness in small animals, inconclusive data from clinical trials have raised questions of dose and regimen. To address this, we completed a comprehensive study in the murine femoral allograft model, to assess the effects of dose (0.4, 4, and 40 µg/kg/day) and various treatment regimens on radiographic, histologic and biomechanical healing at 2, 4, and 9 weeks. Only the high dose (40 µg/kg) of PTH(1-34) demonstrated significant effects when given daily over 9 weeks. Remarkably, equivalent biomechanical results were obtained with delayed, short treatment from 2 to 6 weeks that did not induce a significant increase in endochondral bone formation and callus volume. In contrast, PTH(1-34) treatment from 1 to 5 weeks post-op demonstrated similar osteogenic effects as immediate-daily treatment for 9 weeks, but failed to achieve a significant increase in biomechanics at 9 weeks. Micro-CT and histologic analyses demonstrated that the 2-week delay in treatment allowed for timely completion of the endochondral phase, such that the prominent effects of PTH(1-34) were enhanced intramembranous bone formation and remodeling at the graft-host junction. These findings support the potential use of PTH(1-34) as an adjuvant therapy for massive allograft healing, and suggest that there may be an ideal treatment window in which a short course is administered following the endochondral phase to promote osteoblastic bone formation and remodeling to achieve superior union with modest callus formation. © 2011 American Society for Bone and Mineral Research. M Takahata EM Schwarz /people/?u=22215734 T Chen RJ O'Keefe HA Awad /bme/people/faculty/bio/?id=240 2011 09 21 Teriparatide as a chondroregenerative therapy for injury-induced osteoarthritis. There is no disease-modifying therapy for osteoarthritis, a degenerative joint disease that is projected to afflict more than 67 million individuals in the United States alone by 2030. Because disease pathogenesis is associated with inappropriate articular chondrocyte maturation resembling that seen during normal endochondral ossification, pathways that govern the maturation of articular chondrocytes are candidate therapeutic targets. It is well established that parathyroid hormone (PTH) acting via the type 1 PTH receptor induces matrix synthesis and suppresses maturation of chondrocytes. We report that the PTH receptor is up-regulated in articular chondrocytes after meniscal injury and in osteoarthritis in humans and in a mouse model of injury-induced knee osteoarthritis. To test whether recombinant human PTH(1-34) (teriparatide) would inhibit aberrant chondrocyte maturation and associated articular cartilage degeneration, we administered systemic teriparatide (Forteo), a Food and Drug Administration-approved treatment for osteoporosis, either immediately after or 8 weeks after meniscal/ligamentous injury in mice. Knee joints were harvested at 4, 8, or 12 weeks after injury to examine the effects of teriparatide on cartilage degeneration and articular chondrocyte maturation. Microcomputed tomography revealed increased bone volume within joints from teriparatide-treated mice compared to saline-treated control animals. Immediate systemic administration of teriparatide increased proteoglycan content and inhibited articular cartilage degeneration, whereas delayed treatment beginning 8 weeks after injury induced a regenerative effect. The chondroprotective and chondroregenerative effects of teriparatide correlated with decreased expression of type X collagen, RUNX2 (runt-related transcription factor 2), matrix metalloproteinase 13, and the carboxyl-terminal aggrecan cleavage product NITEGE. These preclinical findings provide proof of concept that Forteo may be useful for decelerating cartilage degeneration and inducing matrix regeneration in patients with osteoarthritis. 3 101ra93 ER Sampson MJ Hilton Y Tian D Chen EM Schwarz /people/?u=22215734 RA Mooney SV Bukata RJ O'Keefe H Awad /bme/people/faculty/bio/?id=240 JE Puzas RN Rosier MJ Zuscik 2011 09 16 Robust antigen-specific humoral immune responses to sublingually delivered adenoviral vectors encoding HIV-1 Env: association with mucoadhesion and efficient penetration of the sublingual barrier. The efficient induction of virus-specific mucosal antibodies is an important unmet objective in Human Immunodeficiency Virus Type-1 (HIV-1) vaccine research. One promising approach is sublingual (SL) immunization. We examined the effectiveness of SL delivery of two different viral vectors: (i) a recombinant adenovirus (rAd5), and (ii) a Herpes Simplex Virus Type-1 amplicon vector (HSV-1). Initial in vitro videomicroscopy experiments showed that rAd5 particles were trapped in saliva (i.e., that Ad5 was mucoadhesive) - unlike HSV-1 virions, which migrated freely in both saliva and water. In vivo imaging studies in mice revealed that only the rAd5 vector efficiently transduced the SL epithelium. Consistent with this, SL delivery of an rAd5 encoding HIV-1 envelope glycoprotein (Env) resulted in robust antigen-specific antibody responses in plasma and in vaginal washes, whereas SL delivery of a HSV-1 amplicon vector encoding HIV-1 Env failed to elicit Env-specific antibodies. In contrast, both vectors elicited equivalent humoral responses following intramuscular (IM) delivery. Finally, SL delivery of the rAd5:Env vector resulted in elevated levels of Env-specific serum IgA, and vaginal IgA and IgG, when compared to IM delivery of the same vector. These results findings shed light on vector properties (mucoadhesion, penetration of the sublingual barrier) which may be important for the induction of potent humoral immune responses following sublingual vector administration. Our data also show that SL delivery of an Env-encoding rAd5 vector can elicit a potent antigen-specific mucosal antibody response in the absence of adjuvant. Overall, these findings support the further exploration of the SL delivery route for HIV-1 vaccine delivery. 29 7080-9 W Domm L Brooks HL Chung C Feng WJ Bowers G Watson JL McGrath /bme/people/faculty/bio/?id=239 S Dewhurst 2011 09 02 Development of a statistical shape model of the patellofemoral joint for investigating relationships between shape and function. Patellofemoral (PF)-related pathologies, including joint laxity, patellar maltracking, cartilage degradation and anterior knee pain, affect nearly 25% of the population. Researchers have investigated the influence of articular geometry on kinematics and contact mechanics in order to gain insight into the etiology of these conditions. The purpose of the current study was to create a three-dimensional statistical shape model of the PF joint and to characterize relationships between PF shape and function (kinematics and contact mechanics). A statistical shape model of the patellar and femoral articular surfaces and their relative alignment was developed from magnetic resonance images. Using 15 shape parameters, the model characterized 97% of the variation in the training set. The first three shape modes primarily described variation in size, patella alta-baja and depth of the sulcus groove. A previously verified finite element model was used to predict kinematics and contact mechanics for each subject. Combining the shape and joint mechanics data, a statistical shape-function model was developed that established quantitative relations of how changes in the shape of the PF joint influence mechanics. The predictive capability of the shape-function model was evaluated by comparing statistical model and finite element predictions, resulting in kinematic root mean square errors of less than 3° and 2.5 mm. The key results of the study are dually in the implementation of a novel approach linking statistical shape and finite element models and the relationships elucidated between PF articular geometry and mechanics. 44 2446-52 CK Fitzpatrick MA Baldwin PJ Laz DP FitzPatrick AL Lerner /bme/people/faculty/bio/?id=241 PJ Rullkoetter 2011 09 01 TNF is required for the induction but not the maintenance of compression-induced BME signals in murine tail vertebrae: limitations of anti-TNF therapy for degenerative disc disease. While bone marrow edema (BME) is diagnostic of spondyloarthropathy, its nature remains poorly understood. In contrast, BME in ankylosing spondylitis is caused by tumor necrosis factor (TNF)-induced vascular and cellular changes. To investigate the relationship between chronic compression and TNF signaling in compression-induced BME we utilized a tail vertebrae compression model with WT, TNF-Tg, and TNFR1&2-/- mice to evaluate: (i) healing following release of chronic compression, (ii) induction of BME in the absence of TNFR, and (iii) efficacy of anti-TNF therapy. Compression-induced normalized marrow contrast enhancement (NMCE) in WT was significantly decreased threefold (p < 0.01) within 2 weeks of release, while the NMCE values in TNF-Tg vertebrae remained elevated, but had a significant decrease (p < 0.05) by 6 weeks after the release of compression. TNFR1&2-/- mice were resistant to compression-induced BME. Anti-TNF therapy did not affect NMCE versus placebo. Histological examination revealed that NMCE values significantly correlated with marrow vascularity and cellularity (p < 0.05), which account for 76% of the variability of NMCE. Collectively, these data demonstrate a critical role for TNF in the induction of chronic compression-induced BME, but not in its maintenance. Amelioration of BME is achieved through biomechanical stability, but is not affected by anti-TNF therapy. 29 1367-74 MO Papuga E Kwok Z You PT Rubery PE Dougherty G Pryhuber CA Beck MJ Hilton HA Awad /bme/people/faculty/bio/?id=240 EM Schwarz /people/?u=22215734 2011 09 01 Two-photon in vivo imaging of cells. In vivo imaging of cells gives a glimpse into the world of biology in a natural setting unparalleled by any other venue. Two-photon imaging of fluorescently labeled cells has become the standard to obtain high-resolution, dynamic images of living specimens with great specificity. This review focuses on providing the reader with a short history of, and impetus behind, two-photon imaging, its working mechanics, and emerging technologies related to biological multiphoton imaging. 26 1483-9 DJ Christensen M Nedergaard /people/?u=23788299 2011 09 01 Role of glucosyltransferase B in interactions of Candida albicans with Streptococcus mutans and with an experimental pellicle on hydroxyapatite surfaces. Candida albicans and mutans streptococci are frequently detected in dental plaque biofilms from toddlers afflicted with early childhood caries. Glucosyltransferases (Gtfs) secreted by Streptococcus mutans bind to saliva-coated apatite (sHA) and to bacterial surfaces, synthesizing exopolymers in situ, which promote cell clustering and adherence to tooth enamel. We investigated the potential role Gtfs may play in mediating the interactions between C. albicans SC5314 and S. mutans UA159, both with each other and with the sHA surface. GtfB adhered effectively to the C. albicans yeast cell surface in an enzymatically active form, as determined by scintillation spectroscopy and fluorescence imaging. The glucans formed on the yeast cell surface were more susceptible to dextranase than those synthesized in solution or on sHA and bacterial cell surfaces (P < 0.05), indicating an elevated α-1,6-linked glucose content. Fluorescence imaging revealed that larger numbers of S. mutans cells bound to C. albicans cells with glucans present on their surface than to yeast cells without surface glucans (uncoated). The glucans formed in situ also enhanced C. albicans interactions with sHA, as determined by a novel single-cell micromechanical method. Furthermore, the presence of glucan-coated yeast cells significantly increased the accumulation of S. mutans on the sHA surface (versus S. mutans incubated alone or mixed with uncoated C. albicans; P < 0.05). These data reveal a novel cross-kingdom interaction that is mediated by bacterial GtfB, which readily attaches to the yeast cell surface. Surface-bound GtfB promotes the formation of a glucan-rich matrix in situ and may enhance the accumulation of S. mutans on the tooth enamel surface, thereby modulating the development of virulent biofilms. 77 6357-67 S Gregoire J Xiao BB Silva I Gonzalez PS Agidi MI Klein KS Ambatipudi PL Rosalen R Bauserman RE Waugh /bme/people/faculty/bio/?id=166 H Koo 2011 08 25 Perceptual learning reduces interneuronal correlations in macaque visual cortex. Responses of neurons in early visual cortex change little with training and appear insufficient to account for perceptual learning. Behavioral performance, however, relies on population activity, and the accuracy of a population code is constrained by correlated noise among neurons. We tested whether training changes interneuronal correlations in the dorsal medial superior temporal area, which is involved in multisensory heading perception. Pairs of single units were recorded simultaneously in two groups of subjects: animals trained extensively in a heading discrimination task, and "naive" animals that performed a passive fixation task. Correlated noise was significantly weaker in trained versus naive animals, which might be expected to improve coding efficiency. However, we show that the observed uniform reduction in noise correlations leads to little change in population coding efficiency when all neurons are decoded. Thus, global changes in correlated noise among sensory neurons may be insufficient to account for perceptual learning. 71 750-61 Y Gu S Liu CR Fetsch Y Yang S Fok A Sunkara GC DeAngelis http://www.bcs.rochester.edu/people/gdeangelis/ DE Angelaki 2011 08 19 Insights into interstitial flow, shear stress, and mass transport effects on ECM heterogeneity in bioreactor-cultivated engineered cartilage hydrogels. Interstitial flow in articular cartilage is secondary to compressive and shear deformations during joint motion and has been linked with the well-characterized heterogeneity in structure and composition of its extracellular matrix. In this study, we investigated the effects of introducing gradients of interstitial flow on the evolution of compositional heterogeneity in engineered cartilage. Using a parallel-plate bioreactor, we observed that Poiseuille flow stimulation of chondrocyte-seeded agarose hydrogels led to an increase in glycosaminoglycan and type II collagen deposition in the surface region of the hydrogel exposed to flow. Experimental measurements of the interstitial flow fields based on the fluorescence recovery after photobleaching technique suggested that the observed heterogeneity in composition is associated with gradients in interstitial flow in a boundary layer at the hydrogel surface. Interestingly, the interstitial flow velocity profiles were nonlinearly influenced by flow rate, which upon closer examination led us to the original observation that the apparent hydrogel permeability decreased exponentially with increased interfacial shear stress. We also observed that interstitial flow enhances convective mass transport irrespective of molecular size within the boundary layer near the hydrogel surface and that the convective contribution to transport diminishes with depth in association with interstitial flow gradients. The implications of the nonlinearly inverse relationship between the interfacial shear stress and the interstitial flux and permeability and its consequences for convective transport are important for tissue engineering, since porous scaffolds comprise networks of Poiseuille channels (pores) through which interstitial flow must navigate under mechanical stimulation or direct perfusion. T Chen M Buckley I Cohen L Bonassar HA Awad /bme/people/faculty/bio/?id=240 2011 08 17 Representation of vestibular and visual cues to self-motion in ventral intraparietal cortex. Convergence of vestibular and visual motion information is important for self-motion perception. One cortical area that combines vestibular and optic flow signals is the ventral intraparietal area (VIP). We characterized unisensory and multisensory responses of macaque VIP neurons to translations and rotations in three dimensions. Approximately one-half of VIP cells show significant directional selectivity in response to optic flow, one-half show tuning to vestibular stimuli, and one-third show multisensory responses. Visual and vestibular direction preferences of multisensory VIP neurons could be congruent or opposite. When visual and vestibular stimuli were combined, VIP responses could be dominated by either input, unlike the medial superior temporal area (MSTd) where optic flow tuning typically dominates or the visual posterior sylvian area (VPS) where vestibular tuning dominates. Optic flow selectivity in VIP was weaker than in MSTd but stronger than in VPS. In contrast, vestibular tuning for translation was strongest in VPS, intermediate in VIP, and weakest in MSTd. To characterize response dynamics, direction-time data were fit with a spatiotemporal model in which temporal responses were modeled as weighted sums of velocity, acceleration, and position components. Vestibular responses in VIP reflected balanced contributions of velocity and acceleration, whereas visual responses were dominated by velocity. Timing of vestibular responses in VIP was significantly faster than in MSTd, whereas timing of optic flow responses did not differ significantly among areas. These findings suggest that VIP may be proximal to MSTd in terms of vestibular processing but hierarchically similar to MSTd in terms of optic flow processing. 31 12036-52 A Chen GC DeAngelis http://www.bcs.rochester.edu/people/gdeangelis/ DE Angelaki 2011 08 10 Convergence of vestibular and visual self-motion signals in an area of the posterior sylvian fissure. Convergence of visual motion information (optic flow) and vestibular signals is important for self-motion perception, and such convergence has been observed in the dorsal medial superior temporal (MSTd) and ventral intraparietal areas. In contrast, the parieto-insular vestibular cortex (PIVC), a cortical vestibular area in the sylvian fissure, is not responsive to optic flow. Here, we explore optic flow and vestibular convergence in the visual posterior sylvian area (VPS) of macaque monkeys. This area is located at the posterior end of the sylvian fissure, is strongly interconnected with PIVC, and receives projections from MSTd. We found robust optic flow and vestibular tuning in more than one-third of VPS cells, with all motion directions being represented uniformly. However, visual and vestibular direction preferences for translation were mostly opposite, unlike in area MSTd where roughly equal proportions of neurons have visual/vestibular heading preferences that are congruent or opposite. Overall, optic flow responses in VPS were weaker than those in MSTd, whereas vestibular responses were stronger in VPS than in MSTd. When visual and vestibular stimuli were presented together, VPS responses were dominated by vestibular signals, in contrast to MSTd, where optic flow tuning typically dominates. These findings suggest that VPS is proximal to MSTd in terms of vestibular processing, but distal to MSTd in terms of optic flow processing. Given the preponderance of neurons with opposite visual/vestibular heading preferences in VPS, this area may not play a major role in multisensory heading perception. 31 11617-27 A Chen GC DeAngelis http://www.bcs.rochester.edu/people/gdeangelis/ DE Angelaki 2011 08 05 Enantioselective, palladium-catalyzed α-arylation of N-Boc pyrrolidine: in situ react IR spectroscopic monitoring, scope, and synthetic applications. A comprehensive study of the enantioselective Pd-catalyzed α-arylation of N-Boc pyrrolidine has been carried out. The protocol involves deprotonation of N-Boc pyrrolidine using s-BuLi/(-)-sparteine in TBME or Et(2)O at -78 °C, transmetalation with ZnCl(2) and Negishi coupling using Pd(OAc)(2), t-Bu(3)P-HBF(4) and the aryl bromide. This paper reports several new features including in situ React IR spectroscopic monitoring of the process; use of (-)-sparteine and the (+)-sparteine surrogate to access products with opposite configuration; development of a catalytic asymmetric lithiation-Negishi coupling reaction; extension to a wide range of heteroaromatic bromides; total synthesis of (R)-crispine A, (S)-nicotine and (S)-SIB-1508Y via short synthetic routes; and examples of α-vinylation of N-Boc pyrrolidine using vinyl bromides exemplified by the total synthesis of naturally occurring (+)-maackiamine (thus establishing its configuration as (R)). In this way, the full scope and limitations of the methodology are delineated. 76 5936-53 G Barker JL McGrath /bme/people/faculty/bio/?id=239 A Klapars D Stead G Zhou KR Campos P O'Brien 2011 08 01 Establishment of an index with increased sensitivity for assessing murine arthritis. The goals of our study were to establish quantitative outcomes for assessing murine knee arthritis and develop an Arthritis Index that incorporates multiple outcomes into a single calculation that provides enhanced sensitivity. Using an accepted model of meniscal/ligamentous injury (MLI)-induced osteoarthritis (OA), we assessed mouse knee arthritis using several approaches. Histology-based methods were performed to visualize joint tissues including articular cartilage and subchondral bone. Accepted histologic scoring methods and histomorphometry were performed to grade cartilage degeneration and determine articular cartilage area, respectively. MicroCT was used to visualize and quantify the bony structures of the joint including osteophytes and joint bone volume. A statistical algorithm was then developed that combined histologic scores and cartilage areas into a single Arthritis Index. MLI induced progressive, OA-like articular cartilage degeneration characterized by increasing (worsening) histologic score and decreasing cartilage area. MicroCT revealed osteophytes and increased joint bone volume between the femoral and tibial physes following MLI. Lastly, an Arthritis Index calculation was established, which incorporated histologic scoring and cartilage area. The Arthritis Index provided enhanced quantitative sensitivity in assessing the level of joint degeneration compared to either histologic scoring or cartilage area determination alone; when using the Index, between 29% and 43% fewer samples are needed to establish statistical significance in studies of murine arthritis. Arthritis in the mouse knee can be quantitatively assessed by histologic scoring, measuring cartilage area, and determining joint bone volume. Enhanced sensitivity can be achieved by performing the Arthritis Index calculation, a novel method for quantitatively assessing mouse knee arthritis. 29 1145-51 ER Sampson CA Beck J Ketz KL Canary MJ Hilton H Awad /bme/people/faculty/bio/?id=240 EM Schwarz /people/?u=22215734 D Chen RJ O'Keefe RN Rosier MJ Zuscik 2011 08 01 Self-complementary AAV2.5-BMP2-coated femoral allografts mediated superior bone healing versus live autografts in mice with equivalent biomechanics to unfractured femur. Structural allografts used for critical bone defects have limited osteogenic properties for biointegration. Although ex vivo tissue-engineered constructs expressing bone morphogenetic protein-2 (BMP2) have demonstrated efficacy in critical defect models, similar success has not been achieved with off-the-shelf acellular approaches, including allografts coated with freeze-dried single-stranded adeno-associated virus (ssAAV-BMP2). To see whether the self-complementary AAV serotype 2.5 vector (scAAV2.5-BMP2) could overcome this, we performed side-by-side comparisons in vitro and in the murine femoral allograft model. Although ssAAV-BMP2 was unable to induce BMP2 expression and differentiation of C3H10T1/2 cells in culture, scAAV2.5-BMP2 transduction led to dose-dependent BMP2 expression and alkaline phosphatase activity, and displayed a 25-fold increased transduction efficiency in vivo. After 6 weeks, the ssAAV-BMP2 coating failed to demonstrate any significant effects. However, all allografts coated with 10(10) scAAV2.5-BMP2 formed a new cortical shell that was indistinguishable to that formed by live autografts. Additionally, coated allografts experienced reduced resorption resulting in a threefold increase in graft bone volume versus autograft. This led to biomechanical superiority versus both allografts and autografts, and equivalent torsional rigidity to unfractured femur. Collectively, these results demonstrate that scAAV2.5-BMP2 coating overcomes the major limitations of structural allografts, which can be used to heal critical defects of any size. 19 1416-25 C Yazici M Takahata DG Reynolds C Xie RJ Samulski J Samulski EJ Beecham AA Gertzman M Spilker X Zhang RJ O'Keefe HA Awad /bme/people/faculty/bio/?id=240 EM Schwarz /people/?u=22215734 2011 08 01 Raman spectroscopy detects deterioration in biomechanical properties of bone in a glucocorticoid-treated mouse model of rheumatoid arthritis. Although glucocorticoids are frequently prescribed for the symptomatic management of inflammatory disorders such as rheumatoid arthritis, extended glucocorticoid exposure is the leading cause of physician-induced osteoporosis and leaves patients at a high risk of fracture. To study the biochemical effects of glucocorticoid exposure and how they might affect biomechanical properties of the bone, Raman spectra were acquired from ex vivo tibiae of glucocorticoid- and placebo-treated wild-type mice and a transgenic mouse model of rheumatoid arthritis. Statistically significant spectral differences were observed due to both treatment regimen and mouse genotype. These differences are attributed to changes in the overall bone mineral composition, as well as the degree of phosphate mineralization in tibial cortical bone. In addition, partial least squares regression was used to generate a Raman-based prediction of each tibia's biomechanical strength as quantified by a torsion test. The Raman-based predictions were as accurate as those produced by microcomputed tomography derived parameters, and more accurate than the clinically-used parameter of bone mineral density. These results suggest that Raman spectroscopy could be a valuable tool for monitoring bone biochemistry in studies of bone diseases such as osteoporosis, including tests of drugs being developed to combat these diseases. 16 087012 JR Maher M Takahata HA Awad /bme/people/faculty/bio/?id=240 AJ Berger 2011 08 01 Identifying neuron communities during a reach and grasp task using an unsupervised clustering analysis. Recent advances in brain-machine interfaces (BMIs) have allowed for high density recordings using microelectrode arrays. However, these large datasets present a challenge in how to practically identify features of interest and discard non-task-related neurons. Thus, we apply a previously reported unsupervised clustering analysis to neural data acquired from a non-human primate as it performed a center-out reach-and-grasp task. Although neurons were recorded from multiple arrays across motor and premotor areas, neurons were found to cluster into only two groups which differ by their mean firing rate. No spatial distribution of neurons was evident in different groups, either across arrays or at different depths. Using a Kalman filter to decode arm, hand, and finger kinematics, we find that using neurons from only one of the groups resulted in higher decoding accuracy (r=0.73) than using randomly selected neurons (r=0.68). This suggests that the proposed method can be used to prune the input space and identify an optimal population of neurons for BMI tasks. 2011 6401-4 GI Newman V Aggarwal MH Schieber /people/?u=22085467 NV Thakor