Biomedical Engineering and Bioengineering Commons^™

Image-Based Quantification Of Benzoporphyrin Derivative Uptake, Localization, And Photobleaching In 3d Tumor Models, For Optimization Of Pdt Parameters, Michael D. Glidden, Jonathan P. Celli, Iqbal Massodi, Imran Rizvi, Brian Pogue, Tayyaba Hasan

Dartmouth Scholarship

Photodynamic therapy (PDT) is a light-based treatment modality in which wavelength specific activation of a photosensitizer (PS) generates cytotoxic response in the irradiated region. PDT response is critically dependent on several parameters including light dose, PS dose, uptake time, fluence rate, and the mode of light delivery. While the systematic optimization of these treatment parameters can be complex, it also provides multiple avenues for en- hancement of PDT efficacy under diverse treatment conditions, provided that a rational framework is established to quantify the impact of parameter selection upon treatment re- sponse. Here we present a theranostic technique, combining the inherent …

A Digital X-Ray Tomosynthesis Coupled Near Infrared Spectral Tomography System For Dual-Modality Breast Imaging, Venkataramanan Krishnaswamy, Kelly E. Michaelsen, Brian W. Pogue, Steven P. Poplack, Ian Shaw, Ken Defrietas, Ken Brooks, Keith D. Paulsen

Dartmouth Scholarship

A Near Infrared Spectral Tomography (NIRST) system has been developed and integrated into a commercial Digital Breast Tomosynthesis (DBT) scanner to allow structural and functional imaging of breast in vivo. The NIRST instrument uses an 8-wavelength continuous wave (CW) laser-based scanning source assembly and a 75-element silicon photodiode solid-state detector panel to produce dense spectral and spatial projection data from which spectrally constrained 3D tomographic images of tissue chromophores are produced. Integration of the optical imaging system into the DBT scanner allows direct co-registration of the optical and DBT images, while also facilitating the synergistic use of x-ray contrast as …

Measurement Of The Oxidation State Of Mitochondrial Cytochrome C From The Neocortex Of The Mammalian Brain, Y. Sakata, M. Abajian, M. O. Ripple, R. Springett

Dartmouth Scholarship

Diffuse optical remission spectra from the mammalian neocortex at visible wavelengths contain spectral features originating from the mitochondria. A new algorithm is presented, based on analytically relating the first differential of the attenuation spectrum to the first differential of the chromophore spectra, that can separate and calculate the oxidation state of cytochrome c as well as the absolute concentration and saturation of hemoglobin. The algorithm is validated in phantoms and then tested on the neocortex of the rat during an anoxic challenge. Implementation of the algorithm will provide detailed information of mitochondrial oxygenation and mitochondrial function in physiological studies of …

Integrated Analysis Of Hydrothermal Flow Through Pretreatment, Veronique Archambault-Leger, Xiongjun Shao, Lee R. Lynd

Dartmouth Scholarship

Background:

The impact of hydrothermal flowthrough (FT) pretreatment severity on pretreatment and solubilization performance metrics was evaluated for three milled feedstocks (corn stover, bagasse, and poplar) and two conversion systems (simultaneous saccharification and fermentation using yeast and fungal cellulase, and fermentation by Clostridium thermocellum).

Results:

Compared to batch pretreatment, FT pretreatment consistently resulted in higher XMG recovery, higher removal of non-carbohydrate carbon and higher glucan solubilization by simultaneous saccharification and fermentation (SSF). XMG recovery was above 90% for FT pretreatment below 4.1 severity but decreased at higher severities, particularly for bagasse. Removal of non-carbohydrate carbon during FT pretreatment increased …

Ethanol And Anaerobic Conditions Reversibly Inhibit Commercial Cellulase Activity In Thermophilic Simultaneous Saccharification And Fermentation (Tssf), Kara K. Podkaminer, William R. Kenealy, Christopher D. Herring, David A. Hogsett, Lee R. Lynd

Dartmouth Scholarship

A previously developed mathematical model of low solids thermophilic simultaneous saccharification and fermentation (tSSF) with Avicel was unable to predict performance at high solids using a commercial cellulase preparation (Spezyme CP) and the high ethanol yield Thermoanaerobacterium saccharolyticum strain ALK2. The observed hydrolysis proceeded more slowly than predicted at solids concentrations greater than 50 g/L Avicel. Factors responsible for this inaccuracy were investigated in this study.

Computational Design And Characterization Of A Temperature-Sensitive Plasmid Replicon For Gram Positive Thermophiles, Daniel G. Olson, Lee R. Lynd

Dartmouth Scholarship

Temperature-sensitive (Ts) plasmids are useful tools for genetic engineering, but there are currently none compatible with the gram positive, thermophilic, obligate anaerobe, Clostridium thermocellum. Traditional mutagenesis techniques yield Ts mutants at a low frequency, and therefore requires the development of high-throughput screening protocols, which are also not available for this organism. Recently there has been progress in the development of computer algorithms which can predict Ts mutations. Most plasmids currently used for genetic modification of C. thermocellum are based on the replicon of plasmid pNW33N, which replicates using the RepB replication protein. To address this problem, we set out …

Developing Unique Engineering Solutions To Improve Patient Safety, Bradley Watts, Douglas Van Citters, Brian Shiner, Peter Mills

Dartmouth Scholarship

Many efforts to improve healthcare safety have focused on redesigning processes of care or retraining clinicians. Far less attention has been focused on the use of new technologies to improve safety. We present the results of a unique collaboration between the VA National Center for Patient Safety (NCPS) and the Thayer School of Engineering at Dartmouth College. Each year, the NCPS identifies safety problems across the VA that could be addressed with newly-engineered devices. Teams of Thayer students and faculty participating in a senior design course evaluate and engineer a solution for one of the problems. Exemplar projects have targeted …

Quantitative Cherenkov Emission Spectroscopy For Tissue Oxygenation Assessment, Johan Axelsson, Adam K. Glaser, David J. Gladstone, Brian W. Pogue

Dartmouth Scholarship

Measurements of Cherenkov emission in tissue during radiation therapy are shown to enable estimation of hemoglobin oxygen saturation non-invasively, through spectral fitting of the spontaneous emissions from the treated tissue. Tissue oxygenation plays a critical role in the efficacy of radiation therapy to kill tumor tissue. Yet in-vivo measurement of this has remained elusive in routine use because of the complexity of oxygen measurement techniques. There is a spectrally broad emission of Cherenkov light that is induced during the time of irradiation, and as this travels through tissue from the point of the radiation deposition, the tissue absorption and scatter …

Complete Genome Sequence Of Clostridium Clariflavum Dsm 19732, Javier A. Izquierdo, Lynne Goodwin, Karen W. Davenport, Hazuki Teshima

Dartmouth Scholarship

Clostridium clariflavum is a Cluster III Clostridium within the family Clostridiaceae isolated from thermophilic anaerobic sludge (Shiratori et al, 2009). This species is of interest because of its similarity to the model cellulolytic organism Clostridium thermocellum and for the ability of environmental isolates to break down cellulose and hemicellulose. Here we describe features of the 4,897,678 bp long genome and its annotation, consisting of 4,131 protein-coding and 98 RNA genes, for the type strain DSM 19732.

Enhanced Microbial Utilization Of Recalcitrant Cellulose By An Ex Vivo Cellulosome-Microbe Complex, Chun You, Xiao-Zhou Zhang, Noppadon Sathitsuksanoh, Lee R. Lynd

Dartmouth Scholarship

A cellulosome-microbe complex was assembled ex vivo on the surface of Bacillus subtilis displaying a miniscaffoldin that can bind with three dockerin-containing cellulase components: the endoglucanase Cel5, the processive endoglucanase Cel9, and the cellobiohydrolase Cel48. The hydrolysis performances of the synthetic cellulosome bound to living cells, the synthetic cellulosome, a noncomplexed cellulase mixture with the same catalytic components, and a commercial fungal enzyme mixture were investigated on low-accessibility recalcitrant Avicel and high accessibility regenerated amorphous cellulose (RAC). The cellbound cellulosome exhibited 4.5- and 2.3-fold-higher hydrolysis ability than cell-free cellulosome on Avicel and RAC, respectively. The cellulosome-microbe synergy was not completely …

Mutant Alcohol Dehydrogenase Leads To Improved Ethanol Tolerance In Clostridium Thermocellum, Steven D. Brown, Adam M. Guss, Tatiana V. Karpinets, Jerry M. Parks

Dartmouth Scholarship

Clostridium thermocellum is a thermophilic, obligately anaerobic, Gram-positive bacterium that is a candidate microorganism for converting cellulosic biomass into ethanol through consolidated bioprocessing. Ethanol intolerance is an important metric in terms of process economics, and tolerance has often been described as a complex and likely multigenic trait for which complex gene interactions come into play. Here, we resequence the genome of an ethanol-tolerant mutant, show that the tolerant phenotype is primarily due to a mutated bifunctional acetaldehyde-CoA/alcohol dehydrogenase gene (adhE), hypothesize based on structural analysis that cofactor specificity may be affected, and confirm this hypothesis using enzyme assays. …

Dual-Channel Imaging System For Singlet Oxygen And Photosensitizer For Pdt, Seonkyung Lee, Martin E. Isabelle, Kristin L. Gabally-Kinney, Brian W. Pogue, Steven J. Davis

Dartmouth Scholarship

A two-channel optical system has been developed to provide spatially resolved simultaneous imaging of singlet molecular oxygen (¹O₂) phosphorescence and photosensitizer (PS) fluorescence produced by the photodynamic process. The current imaging system uses a spectral discrimination method to differentiate the weak ¹O₂ phosphorescence that peaks near 1.27 μ m from PS fluorescence that also occurs in this spectral region. The detection limit of ¹O₂ emission was determined at a concentration of 500 nM benzoporphyrin derivative monoacid (BPD) in tissue-like phantoms, and these signals observed were proportional to the PS fluorescence. Preliminary in …

Singular Value Decomposition Metrics Show Limitations Of Detector Design In Diffuse Fluorescence Tomography, Frederic Leblond, Kenneth M. Tichauer, Brian W. Pogue

Dartmouth Scholarship

The spatial resolution and recovered contrast of images reconstructed from diffuse fluorescence tomography data are limited by the high scattering properties of light propagation in biological tissue. As a result, the image reconstruction process can be exceedingly vulnerable to inaccurate prior knowledge of tissue optical properties and stochastic noise. In light of these limitations, the optimal source-detector geometry for a fluorescence tomography system is non-trivial, requiring analytical methods to guide design. Analysis of the singular value decomposition of the matrix to be inverted for image reconstruction is one potential approach, providing key quantitative metrics, such as singular image mode spatial …

Importance Of Phase Unwrapping For The Reconstruction Of Microwave Tomographic Images, Tomasz M. Grzegorczyk, Paul M. Meaney, Soon Ik Jeon, Shireen D. Geimer, Keith D. Paulsen

Dartmouth Scholarship

Microwave image reconstruction is typically based on a regularized least-square minimization of either the complex-valued field difference between recorded and modeled data or the logarithmic transformation of these field differences. Prior work has shown anecdotally that the latter outperforms the former in limited surveys of simulated and experimental phantom results. In this paper, we provide a theoretical explanation of these empirical findings by developing closed form solutions for the fie

Deletion Of The Cel48s Cellulase From Clostridium Thermocellum, Daniel G. Olson, Shital A. Tripathi, Richard J. Giannone, Jonathan Lo, Nicky C. Caiazza, David A. Hogsett, Robert L. Hettich, Adam M. Guss, Genia Dubrovsky, Lee R. Lynd

Dartmouth Scholarship

Clostridium thermocellum is a thermophilic anaerobic bacterium that rapidly solubilizes cellulose with the aid of a multienzyme cellulosome complex. Creation of knockout mutants for Cel48S (also known as CelS, S(S), and S8), the most abundant cellulosome subunit, was undertaken to gain insight into its role in enzymatic and microbial cellulose solubilization. Cultures of the Cel48S deletion mutant (S mutant) were able to completely solubilize 10 g/L crystalline cellulose. The cellulose hydrolysis rate of the S mutant strain was 60% lower than the parent strain, with the S mutant strain also exhibiting a 40% reduction in cell yield. The cellulosome produced …

Development Of Pyrf-Based Genetic System For Targeted Gene Deletion In Clostridium Thermocellum And Creation Of A Pta Mutant, Shital A. Tripathi, Daniel G. Olson, D. Aaron Argyros, Bethany B. Miller, Trisha F. Barrett, Daniel M. Murphy, Jesse D. Mccool, Anne K. Warner, Vineet B. Rajgarhia, Lee R. Lynd, David A. Hogsett, Nicky C. Caiazza

Dartmouth Scholarship

We report development of a genetic system for making targeted gene knockouts in Clostridium thermocellum, a thermophilic anaerobic bacterium that rapidly solubilizes cellulose. A toxic uracil analog, 5-fluoroorotic acid (5-FOA), was used to select for deletion of the pyrF gene. The ΔpyrF strain is a uracil auxotroph that could be restored to a prototroph via ectopic expression of pyrF from a plasmid, providing a positive genetic selection. Furthermore, 5-FOA was used to select against plasmid-expressed pyrF, creating a negative selection for plasmid loss. This technology was used to delete a gene involved in organic acid production, namely pta, which encodes …

Characterizing Accuracy Of Total Hemoglobin Recovery Using Contrast-Detail Analysis In 3d Image-Guided Near Infrared Spectroscopy With The Boundary Element Method, Hamid R. Ghadyani, Subhadra Srinivasan, Brian W. Pogue, Keith D. Paulsen

Dartmouth Scholarship

The quantification of total hemoglobin concentration (HbT) obtained from multi-modality image-guided near infrared spectroscopy (IG-NIRS) was characterized using the boundary element method (BEM) for 3D image reconstruction. Multi-modality IG-NIRS systems use a priori information to guide the reconstruction process. While this has been shown to improve resolution, the effect on quantitative accuracy is unclear. Here, through systematic contrast-detail analysis, the fidelity of IG-NIRS in quantifying HbT was examined using 3D simulations. These simulations show that HbT could be recovered for medium sized (20mm in 100mm total diameter) spherical inclusions with an average error of 15%, for the physiologically …

Characterization Of An Implicitly Resistively-Loaded Monopole Antenna In Lossy Liquid Media, Colleen J. Fox, Paul M. Meaney, Fridon Shubitidze, Lincoln Potwin, Keith D. Paulsen

Dartmouth Scholarship

Abstract

Microwave tomographic imaging of the breast for cancer detection is a topic of considerable interest because of the potential to exploit the apparent high-dielectric property contrast between normal and malignant tissue. An important component in the realization of an imaging system is the antenna array to be used for signal transmission/detection. The monopole antenna has proven to be useful in our implementation because it can be easily and accurately modeled and can be positioned in close proximity to the imaging target with high-element density when configured in an imaging array. Its frequency response is broadened considerably when radiating in …

Automatic Exposure Control And Estimation Of Effective System Noise In Diffuse Fluorescence Tomography, Dax L. Kepshire, Hamid Dehghani, Frederic Leblond, Brian W. Pogue

Dartmouth Scholarship

A diffuse fluorescence tomography system, based upon time-correlated single photon counting, is presented with an automated algorithm to allow dynamic range variation through exposure control. This automated exposure control allows the upper and lower detection levels of fluorophore to be extended by an order of magnitude beyond the previously published performance and benefits in a slight decrease in system effective noise. The effective noise level is used as a metric to characterize the system performance, integrating both model-mismatch and calibration bias errors into a single parameter. This effective error is near 7% of the reconstructed fluorescent yield value, when imaging …

Statistical Hypothesis Testing For Postreconstructed And Postregistered Medical Images, Eugene Demidenko

Dartmouth Scholarship

Postreconstructed and postregistered medical images are typically treated as the raw data, implicitly assuming that those operations are error free. We question this assumption and explore how the precision of reconstruction and affine registration can be assessed by the image covariance matrix and confidence interval, called the confidence eigenimage, using a statistical model-based approach. Various hypotheses may be tested after image reconstruction and registration using classical statistical hypothesis testing vehicles: Is there a statistically significant difference between images? Does the intensity at a specific location or area of interest belong to the “normal” range? Is there a tumor? Does the …

Video-Rate Near Infrared Tomography To Image Pulsatile Absorption Properties In Thick Tissue, Zhiqiu Li, Venkataramanan Krishnaswamy, Scott C. Davis, Subhadra Srinivasan, Keith D. Paulsen, Brian W. Pogue

Dartmouth Scholarship

A high frame-rate near-infrared (NIR) tomography system was created to allow transmission imaging of thick tissues with spectral encoding for parallel source implementation. The design was created to maximize tissue penetration through up to 10 cm of tissue, allowing eventual use in human imaging. Eight temperature-controlled laser diodes (LD) are used in parallel with 1.5 nm shifts in their lasing wavelengths. Simultaneous detection is achieved with eight high-resolution, CCD-based spectrometers that were synchronized to detect the intensities and decode their source locations from the spectrum. Static and dynamic imaging is demonstrated through a 64 mm tissue-equivalent phantom, with acquisition rates …

In Vitro Ovarian Tumor Growth And Treatment Response Dynamics Visualized With Time-Lapse Oct Imaging, Conor L. Evans, Imran Rizvi, Tayyaba Hasan, Johannes F. De Boer

Dartmouth Scholarship

In vitro three-dimensional models for metastatic ovarian cancer have been useful for recapitulating the human disease. These spheroidal tumor cultures, however, can grow in excess of 1 mm in diameter, which are difficult to visualize without suitable imaging technology.Optical coherence tomography (OCT) is an ideal live imaging method for non-perturbatively visualizing these complex systems. OCT enabled detailed observations of the model at both nodular and cellular levels, revealing growth dynamics not previously observed. The development of a time-lapse OCT system, capable of automated, multidimensional acquisition, further provided insights into the growth and chemotherapeutic response of ovarian cancer.

Methodology Development For Three-Dimensional Mr-Guided Near Infrared Spectroscopy Of Breast Tumors, Colin M. Carpenter, Subhadra Srinivasan, Brian W. Pogue, Keith D. Paulsen

Dartmouth Scholarship

Combined Magnetic Resonance (MR) and Near Infrared Spectroscopy (NIRS) has been proposed as a unique method to quantify hemodynamics, water content, and cellular size and packing density of breast tumors, as these tissue constituents can be quantified with increased resolution and overlaid on the structural features identified by the MR. However, the choices in how to reconstruct and visualize this information can have a dramatic impact on the feasibility of implementing this modality in the clinic. This is especially true in 3 dimensions, as there is often limited optical sampling of the breast tissue, and methods need to accurately reflect …

Metabolic Engineering Of A Thermophilic Bacterium To Produce Ethanol At High Yield, A. Joe Shaw, Kara K. Podkaminer, Sunil G. Desai, John S. Bardsley, Stephen R. Rogers, Philip G. Thorne, David A. Hogsett, Lee R. Lynd

Dartmouth Scholarship

We report engineering Thermoanaerobacterium saccharolyticum, a thermophilic anaerobic bacterium that ferments xylan and biomass-derived sugars, to produce ethanol at high yield. Knockout of genes involved in organic acid formation (acetate kinase, phosphate acetyltransferase, and L-lactate dehydrogenase) resulted in a strain able to produce ethanol as the only detectable organic product and substantial changes in electron flow relative to the wild type. Ethanol formation in the engineered strain (ALK2) utilizes pyruvate:ferredoxin oxidoreductase with electrons transferred from ferredoxin to NAD(P), a pathway different from that in previously described microbes with a homoethanol fermentation. The homoethanologenic phenotype was stable for >150 generations …

Image-Guided Raman Spectroscopic Recovery Of Canine Cortical Bone Contrast In Situ, Subhadra Srinivasan, Matthew Schulmerich, Jacqueline H. Cole, Kathryn Dooley, Jaclynn M. Kreider, Brian W. Pogue

Dartmouth Scholarship

Raman scattering provides valuable biochemical and molecular markers for studying bone tissue composition with use in predicting fracture risk in osteoporosis. Raman tomography can image through a few centimeters of tissue but is limited by low spatial resolution. X-ray computed tomography (CT) imaging can provide high-resolution image-guidance of the Raman spectroscopic characterization, which enhances the quantitative recovery of the Raman signals, and this technique provides additional information to standard imaging methods. This hypothesis was tested in data measured from Teflon tissue phantoms and from a canine limb. Image-guided Raman spectroscopy (IG-RS) of the canine limb using CT images of the …

Fluorescence Tomography Characterization For Sub-Surface Imaging With Protoporphyrin Ix, Dax Kepshire, Scott C. Davis, Hamid Dehghani, Keith D. Paulsen, Brian W. Pogue

Dartmouth Scholarship

Optical imaging of fluorescent objects embedded in a tissue simulating medium was characterized using non-contact based approaches to fluorescence remittance imaging (FRI) and sub-surface fluorescence diffuse optical tomography (FDOT). Using Protoporphyrin IX as a fluorescent agent, experiments were performed on tissue phantoms comprised of typical in-vivo tumor to normal tissue contrast ratios, ranging from 3.5:1 up to 10:1. It was found that tomographic imaging was able to recover interior inclusions with high contrast relative to the background; however, simple planar fluorescence imaging provided a superior contrast to noise ratio. Overall, FRI performed optimally when the object was located on or …

N-Glycan Modification In Aspergillus Species, Elke Kainz, Andreas Gallmetzer, Christian Hatzl, Juergen H. Nett, Huijuan Li, Thorsten Schinko, Robert Pachlinger, Harald Berger, Yazmid Reyes-Dominguez, Andreas Bernreiter, Tillmann Gerngross, Stefan Wildt, Joseph Strauss

Dartmouth Scholarship

The production by filamentous fungi of therapeutic glycoproteins intended for use in mammals is held back by the inherent difference in protein N-glycosylation and by the inability of the fungal cell to modify proteins with mammalian glycosylation structures. Here, we report protein N-glycan engineering in two Aspergillus species. We functionally expressed in the fungal hosts heterologous chimeric fusion proteins containing different localization peptides and catalytic domains. . This strategy allowed the isolation of a strain with a functional -1,2-mannosidase producing increased amounts of N-glycans of the Man 5 GlcNAc 2 type. This strain was further engineered by the introduction of …

Image-Guided Diffuse Optical Fluorescence Tomography Implemented With Laplacian-Type Regularization, Scott C. Davis, Hamid Dehghani, Jia Wang, Shudong Jiang, Brian W. Pogue, Keith D. Paulsen

Dartmouth Scholarship

A promising method to incorporate tissue structural information into the reconstruction of diffusion-based fluorescence imaging is introduced. The method regularizes the inversion problem with a Laplacian-type matrix, which inherently smoothes pre-defined tissue, but allows discontinuities between adjacent regions. The technique is most appropriately used when fluorescence tomography is combined with structural imaging systems. Phantom and simulation studies were used to illustrate significant improvements in quantitative imaging and linearity of response with the new algorithm. Images of an inclusion containing the fluorophore Lutetium Texaphyrin (Lutex) embedded in a cylindrical phantom are more accurate than in situations where no structural information is …

Critical Computational Aspects Of Near Infrared Circular Tomographic Imaging: Analysis Of Measurement Number, Mesh Resolution And Reconstruction Basis, Phaneendra K. Yalavarthy, Hamid Dehghani, Brian W. Pogue, Keit D. Paulsen

Dartmouth Scholarship

The image resolution and contrast in Near-Infrared (NIR) tomographic image reconstruction are affected by parameters such as the number of boundary measurements, the mesh resolution in the forward calculation and the reconstruction basis. Increasing the number of measurements tends to make the sensitivity of the domain more uniform reducing the hypersensitivity at the boundary. Using singular-value decomposition (SVD) and reconstructed images, it is shown that the numbers of 16 or 24 fibers are sufficient for imaging the 2D circular domain for the case of 1% noise in the data. The number of useful singular values increases as the logarithm of …

Imaging Breast Adipose And Fibroglandular Tissue Molecular Signatures By Using Hybrid Mri-Guided Near-Infrared Spectral Tomography, Ben Brooksby, Brian W. Pogue, Shudong Jiang, Hamid Dehghani, Subhadra Srinivasan, Christine Kogel, Tor D. Tosteson, John Weaver, Steven P. Poplack, Keith D. Paulsen

Dartmouth Scholarship

Magnetic resonance (MR)-guided near-infrared spectral tomography was developed and used to image adipose and fibroglandular breast tissue of 11 normal female subjects, recruited under an institutional review board-approved protocol. Images of hemoglobin, oxygen saturation, water fraction, and subcellular scattering were reconstructed and show that fibroglandular fractions of both blood and water are higher than in adipose tissue. Variation in adipose and fibroglandular tissue composition between individuals was not significantly different across the scattered and dense breast categories. Combined MR and near-infrared tomography provides fundamental molecular information about these tissue types with resolution governed by MR T1 images.