Matching Items (15)
Filtering by
- All Subjects: Bioengineering
- Creators: Wang, Xiao
- Creators: Buneo, Christopher
Description
The ultimate goal of human movement control research is to understand how natural movements performed in daily reaching activities, are controlled. Natural movements require coordination of multiple degrees of freedom (DOF) of the arm. Patterns of arm joint control were studied during daily functional tasks, which were performed through the rotation of seven DOF in the arm. Analyzed movements which imitated the following 3 activities of daily living: moving an empty soda can from a table and placing it on a further position; placing the empty soda can from initial position at table to a position at shoulder level on a shelf; and placing the empty soda can from initial position at table to a position at eye level on a shelf. Kinematic and kinetic analyses were conducted for these three movements. The studied kinematic characteristics were: hand trajectory in the sagittal plane, displacements of the 7 DOF, and contribution of each DOF to hand velocity. The kinetic analysis involved computation of 3-dimensional vectors of muscle torque (MT), interaction torque (IT), gravity torque (GT), and net torque (NT) at the shoulder, elbow, and wrist. Using the relationship NT = MT + GT + IT, the role of active control and passive factors (gravitation and inter-segmental dynamics) in rotation of each joint by computing MT contribution (MTC) to NT was assessed. MTC was computed using the ratio of the signed MT projection on NT to NT magnitude. Despite a variety of joint movements available across the different tasks, 3 patterns of shoulder and elbow coordination prevailed in each movement: 1) active rotation of the shoulder and predominantly passive rotation of the elbow; 2) active rotation of the elbow and predominantly passive rotation of the shoulder; and 3) passive rotation of both joints. Analysis of wrist control suggested that MT mainly compensates for passive torque and provides adjustment of wrist motion according to requirements of each task. In conclusion, it was observed that the 3 shoulder-elbow coordination patterns (during which at least one joint moved) passively represented joint control primitives, underlying the performance of well-learned arm movements, although these patterns may be less prevalent during non-habitual movements.
ContributorsSansgiri, Dattaraj (Author) / Dounskaia, Natalia (Thesis advisor) / Schaefer, Sydney (Thesis advisor) / Buneo, Christopher (Committee member) / Arizona State University (Publisher)
Created2018
Description
Recombinases are powerful tools for genome engineering and synthetic biology, however recombinases are limited by a lack of user-programmability and often require complex directed-evolution experiments to retarget specificity. Conversely, CRISPR systems have extreme versatility yet can induce off-target mutations and karyotypic destabilization. To address these constraints we developed an RNA-guided recombinase protein by fusing a hyperactive mutant resolvase from transposon TN3 to catalytically inactive Cas9. We validated recombinase-Cas9 (rCas9) function in model eukaryote Saccharomyces cerevisiae using a chromosomally integrated fluorescent reporter. Moreover, we demonstrated cooperative targeting by CRISPR RNAs at spacings of 22 or 40bps is necessary for directing recombination. Using PCR and Sanger sequencing, we confirmed rCas9 targets DNA recombination. With further development we envision rCas9 becoming useful in the development of RNA-programmed genetic circuitry as well as high-specificity genome engineering.
ContributorsStandage-Beier, Kylie S (Author) / Wang, Xiao (Thesis advisor) / Brafman, David A (Committee member) / Tian, Xiao-jun (Committee member) / Arizona State University (Publisher)
Created2018
Description
Growing understanding of the neural code and how to speak it has allowed for notable advancements in neural prosthetics. With commercially-available implantable systems with bi- directional neural communication on the horizon, there is an increasing imperative to develop high resolution interfaces that can survive the environment and be well tolerated by the nervous system under chronic use. The sensory encoding aspect optimally interfaces at a scale sufficient to evoke perception but focal in nature to maximize resolution and evoke more complex and nuanced sensations. Microelectrode arrays can maintain high spatial density, operating on the scale of cortical columns, and can be either penetrating or non-penetrating. The non-penetrating subset sits on the tissue surface without puncturing the parenchyma and is known to engender minimal tissue response and less damage than the penetrating counterpart, improving long term viability in vivo. Provided non-penetrating microelectrodes can consistently evoke perception and maintain a localized region of activation, non-penetrating micro-electrodes may provide an ideal platform for a high performing neural prosthesis; this dissertation explores their functional capacity.
The scale at which non-penetrating electrode arrays can interface with cortex is evaluated in the context of extracting useful information. Articulate movements were decoded from surface microelectrode electrodes, and additional spatial analysis revealed unique signal content despite dense electrode spacing. With a basis for data extraction established, the focus shifts towards the information encoding half of neural interfaces. Finite element modeling was used to compare tissue recruitment under surface stimulation across electrode scales. Results indicated charge density-based metrics provide a reasonable approximation for current levels required to evoke a visual sensation and showed tissue recruitment increases exponentially with electrode diameter. Micro-scale electrodes (0.1 – 0.3 mm diameter) could sufficiently activate layers II/III in a model tuned to striate cortex while maintaining focal radii of activated tissue.
In vivo testing proceeded in a nonhuman primate model. Stimulation consistently evoked visual percepts at safe current thresholds. Tracking perception thresholds across one year reflected stable values within minimal fluctuation. Modulating waveform parameters was found useful in reducing charge requirements to evoke perception. Pulse frequency and phase asymmetry were each used to reduce thresholds, improve charge efficiency, lower charge per phase – charge density metrics associated with tissue damage. No impairments to photic perception were observed during the course of the study, suggesting limited tissue damage from array implantation or electrically induced neurotoxicity. The subject consistently identified stimulation on closely spaced electrodes (2 mm center-to-center) as separate percepts, indicating sub-visual degree discrete resolution may be feasible with this platform. Although continued testing is necessary, preliminary results supports epicortical microelectrode arrays as a stable platform for interfacing with neural tissue and a viable option for bi-directional BCI applications.
The scale at which non-penetrating electrode arrays can interface with cortex is evaluated in the context of extracting useful information. Articulate movements were decoded from surface microelectrode electrodes, and additional spatial analysis revealed unique signal content despite dense electrode spacing. With a basis for data extraction established, the focus shifts towards the information encoding half of neural interfaces. Finite element modeling was used to compare tissue recruitment under surface stimulation across electrode scales. Results indicated charge density-based metrics provide a reasonable approximation for current levels required to evoke a visual sensation and showed tissue recruitment increases exponentially with electrode diameter. Micro-scale electrodes (0.1 – 0.3 mm diameter) could sufficiently activate layers II/III in a model tuned to striate cortex while maintaining focal radii of activated tissue.
In vivo testing proceeded in a nonhuman primate model. Stimulation consistently evoked visual percepts at safe current thresholds. Tracking perception thresholds across one year reflected stable values within minimal fluctuation. Modulating waveform parameters was found useful in reducing charge requirements to evoke perception. Pulse frequency and phase asymmetry were each used to reduce thresholds, improve charge efficiency, lower charge per phase – charge density metrics associated with tissue damage. No impairments to photic perception were observed during the course of the study, suggesting limited tissue damage from array implantation or electrically induced neurotoxicity. The subject consistently identified stimulation on closely spaced electrodes (2 mm center-to-center) as separate percepts, indicating sub-visual degree discrete resolution may be feasible with this platform. Although continued testing is necessary, preliminary results supports epicortical microelectrode arrays as a stable platform for interfacing with neural tissue and a viable option for bi-directional BCI applications.
ContributorsOswalt, Denise (Author) / Greger, Bradley (Thesis advisor) / Buneo, Christopher (Committee member) / Helms-Tillery, Stephen (Committee member) / Mirzadeh, Zaman (Committee member) / Papandreou-Suppappola, Antonia (Committee member) / Arizona State University (Publisher)
Created2018
Description
Neural interfacing applications have advanced in complexity, with needs for increasingly high degrees of freedom in prosthetic device control, sharper discrimination in sensory percepts in bidirectional interfaces, and more precise localization of functional connectivity in the brain. As such, there is a growing need for reliable neurophysiological recordings at a fine spatial scale matching that of cortical columnar processing. Penetrating microelectrodes provide localization sufficient to isolate action potential (AP) waveforms, but often suffer from recorded signal deterioration linked to foreign body response. Micro-Electrocorticography (μECoG) surface electrodes elicit lower foreign body response and show greater chronic stability of recorded signals, though they typically lack the signal localization necessary to isolate individual APs. This dissertation validates the recording capacity of a novel, flexible, large area μECoG array with bilayer routing in a feline implant, and explores the ability of conventional μECoG arrays to detect features of neuronal activity in a very high frequency band associated with AP waveforms.
Recordings from both layers of the flexible μECoG array showed frequency features typical of cortical local field potentials (LFP) and were shown to be stable in amplitude over time. Recordings from both layers also showed consistent, frequency-dependent modulation after induction of general anesthesia, with large increases in beta and gamma band and decreases in theta band observed over three experiments. Recordings from conventional μECoG arrays over human cortex showed robust modulation in a high frequency (250-2000 Hz) band upon production of spoken words. Modulation in this band was used to predict spoken words with over 90% accuracy. Basal Ganglia neuronal AP firing was also shown to significantly correlate with various cortical μECoG recordings in this frequency band. Results indicate that μECoG surface electrodes may detect high frequency neuronal activity potentially associated with AP firing, a source of information previously unutilized by these devices.
Recordings from both layers of the flexible μECoG array showed frequency features typical of cortical local field potentials (LFP) and were shown to be stable in amplitude over time. Recordings from both layers also showed consistent, frequency-dependent modulation after induction of general anesthesia, with large increases in beta and gamma band and decreases in theta band observed over three experiments. Recordings from conventional μECoG arrays over human cortex showed robust modulation in a high frequency (250-2000 Hz) band upon production of spoken words. Modulation in this band was used to predict spoken words with over 90% accuracy. Basal Ganglia neuronal AP firing was also shown to significantly correlate with various cortical μECoG recordings in this frequency band. Results indicate that μECoG surface electrodes may detect high frequency neuronal activity potentially associated with AP firing, a source of information previously unutilized by these devices.
ContributorsBarton, Cody David (Author) / Greger, Bradley (Thesis advisor, Committee member) / Santello, Marco (Committee member) / Buneo, Christopher (Committee member) / Graudejus, Oliver (Committee member) / Artemiadis, Panagiotis (Committee member) / Arizona State University (Publisher)
Created2018
Description
Intracellular voltage recordings from single neurons in vitro and in vivo have been fundamental to our understanding of neuronal function. Conventional electrodes and associated positioning systems for intracellular recording in vivo are large and bulky, which has largely restricted their use to single-channel recording from anesthetized animals. Further, intracellular recordings are very cumbersome, requiring a high degree of skill not readily achieved in a typical laboratory. This dissertation presents a robotic, head-mountable, MEMS (Micro-Electro-Mechanical Systems) based intracellular recording system to overcome the above limitations associated with form-factor, scalability and highly skilled and tedious manual operations required for intracellular recordings. This system combines three distinct technologies: 1) novel microscale, polycrystalline silicon-based electrode for intracellular recording, 2) electrothermal microactuators for precise microscale navigation of the electrode and 3) closed-loop control algorithm for autonomous movement and positioning of electrode inside single neurons. First, two distinct designs of polysilicon-based microscale electrodes were fabricated and tested for intracellular recordings. In the first approach, tips of polysilicon microelectrodes were milled to nanoscale dimensions (<300 nm) using focused ion beam (FIB) to develop polysilicon nanoelectrodes. Polysilicon nanoelectrodes recorded >1.5 mV amplitude, positive-going action potentials and synaptic potentials from neurons in the abdominal ganglion of Aplysia Californica. In the second approach, polysilicon microelectrodes were integrated with miniaturized glass micropipettes filled with electrolyte to fabricate glass-polysilicon microelectrodes. These electrodes consistently recorded high fidelity intracellular potentials from neurons in the abdominal ganglion of Aplysia Californica (Resting Potentials < -35 mV, Action Potentials > 60 mV) as well as the rat motor cortex (Resting Potentials < -50 mV). Next, glass-polysilicon microelectrodes were coupled with microscale electrothermal actuators and controller for autonomous intracellular recordings from single neurons in the abdominal ganglion. Consistent resting potentials (< -35 mV) and action potentials (> 60 mV) were recorded after each successful penetration attempt with the controller and microactuated glass-polysilicon microelectrodes. The success rate of penetration and quality of recordings achieved using electrothermal microactuators were comparable to that of conventional positioning systems. Finally, the feasibility of this miniaturized system to obtain intracellular recordings from single neurons in the motor cortex of rats in vivo is also demonstrated. The MEMS-based system offers significant advantages: 1) reduction in overall size for potential use in behaving animals, 2) scalable approach to potentially realize multi-channel recordings and 3) a viable method to fully automate measurement of intracellular recordings.
ContributorsSampath Kumar, Swathy (Author) / Muthuswamy, Jit (Thesis advisor) / Abbas, James (Committee member) / Hamm, Thomas (Committee member) / Christen, Jennifer Blain (Committee member) / Buneo, Christopher (Committee member) / Arizona State University (Publisher)
Created2018
Description
Motor behavior is prone to variable conditions and deviates further in disorders affecting the nervous system. A combination of environmental and neural factors impacts the amount of uncertainty. Although the influence of these factors on estimating endpoint positions have been examined, the role of limb configuration on endpoint variability has been mostly ignored. Characterizing the influence of arm configuration (i.e. intrinsic factors) would allow greater comprehension of sensorimotor integration and assist in interpreting exaggerated movement variability in patients. In this study, subjects were placed in a 3-D virtual reality environment and were asked to move from a starting position to one of three targets in the frontal plane with and without visual feedback of the moving limb. The alternating of visual feedback during trials increased uncertainty between the planning and execution phases. The starting limb configurations, adducted and abducted, were varied in separate blocks. Arm configurations were setup by rotating along the shoulder-hand axis to maintain endpoint position. The investigation hypothesized: 1) patterns of endpoint variability of movements would be dependent upon the starting arm configuration and 2) any differences observed would be more apparent in conditions that withheld visual feedback. The results indicated that there were differences in endpoint variability between arm configurations in both visual conditions, but differences in variability increased when visual feedback was withheld. Overall this suggests that in the presence of visual feedback, planning of movements in 3D space mostly uses coordinates that are arm configuration independent. On the other hand, without visual feedback, planning of movements in 3D space relies substantially on intrinsic coordinates.
ContributorsRahman, Qasim (Author) / Buneo, Christopher (Thesis director) / Helms Tillery, Stephen (Committee member) / Barrett, The Honors College (Contributor) / Harrington Bioengineering Program (Contributor)
Created2014-05
Description
Motor behavior is prone to variable conditions and deviates further in disorders affecting the nervous system. A combination of environmental and neural factors impacts the amount of uncertainty. Although the influence of these factors on estimating endpoint positions have been examined, the role of limb configuration on endpoint variability has been mostly ignored. Characterizing the influence of arm configuration (i.e. intrinsic factors) would allow greater comprehension of sensorimotor integration and assist in interpreting exaggerated movement variability in patients. In this study, subjects were placed in a 3-D virtual reality environment and were asked to move from a starting position to one of three targets in the frontal plane with and without visual feedback of the moving limb. The alternating of visual feedback during trials increased uncertainty between the planning and execution phases. The starting limb configurations, adducted and abducted, were varied in separate blocks. Arm configurations were setup by rotating along the shoulder-hand axis to maintain endpoint position. The investigation hypothesized: 1) patterns of endpoint variability of movements would be dependent upon the starting arm configuration and 2) any differences observed would be more apparent in conditions that withheld visual feedback. The results indicated that there were differences in endpoint variability between arm configurations in both visual conditions, but differences in variability increased when visual feedback was withheld. Overall this suggests that in the presence of visual feedback, planning of movements in 3D space mostly uses coordinates that are arm configuration independent. On the other hand, without visual feedback, planning of movements in 3D space relies substantially on intrinsic coordinates.
ContributorsRahman, Qasim (Author) / Buneo, Christopher (Thesis director) / Helms Tillery, Stephen (Committee member) / Barrett, The Honors College (Contributor) / Harrington Bioengineering Program (Contributor)
Created2014-05
Description
The advent of CRISPR/Cas9 revolutionized the field of genetic engineering and gave rise to the development of new gene editing tools including prime editing. Prime editing is a versatile gene editing method that mediates precise insertions and deletions and can perform all 12 types of point mutations. In turn, prime editing represents great promise in the design of new gene therapies and disease models where editing was previously not possible using current gene editing techniques. Despite advancements in genome modification technologies, parallel enrichment strategies of edited cells remain lagging behind in development. To this end, this project aimed to enhance prime editing using transient reporter for editing enrichment (TREE) technology to develop a method for the rapid generation of clonal isogenic cell lines for disease modeling. TREE uses an engineered BFP variant that upon a C-to-T conversion will convert to GFP after target modification. Using flow cytometry, this BFP-to-GFP conversion assay enables the isolation of edited cell populations via a fluorescent reporter of editing. Prime induced nucleotide engineering using a transient reporter for editing enrichment (PINE-TREE), pairs prime editing with TREE technology to efficiently enrich for prime edited cells. This investigation revealed PINE-TREE as an efficient editing and enrichment method compared to a conventional reporter of transfection (RoT) enrichment strategy. Here, PINE-TREE exhibited a significant increase in editing efficiencies of single nucleotide conversions, small insertions, and small deletions in multiple human cell types. Additionally, PINE-TREE demonstrated improved clonal cell editing efficiency in human induced pluripotent stem cells (hiPSCs). Most notably, PINE-TREE efficiently generated clonal isogenic hiPSCs harboring a mutation in the APOE gene for in vitro modeling of Alzheimer’s Disease. Collectively, results gathered from this study exhibited PINE-TREE as a valuable new tool in genetic engineering to accelerate the generation of clonal isogenic cell lines for applications in developmental biology, disease modeling, and drug screening.
ContributorsKostes, William Warner (Author) / Brafman, David (Thesis advisor) / Jacobs, Bertram (Committee member) / Lapinaite, Audrone (Committee member) / Tian, Xiaojun (Committee member) / Wang, Xiao (Committee member) / Arizona State University (Publisher)
Created2022
Description
Neurodegenerative diseases such as Alzheimer’s disease, Parkinson’s disease, or amyotrophic lateral sclerosis are defined by the loss of several types of neurons and glial cells within the central nervous system (CNS). Combatting these diseases requires a robust population of relevant cell types that can be employed in cell therapies, drug screening, or patient specific disease modeling. Human induced pluripotent stem cells (hiPSC)-derived neural progenitor cells (hNPCs) have the ability to self-renew indefinitely and differentiate into the various neuronal and glial cell types of the CNS. In order to realize the potential of hNPCs, it is necessary to develop a xeno-free scalable platform for effective expansion and differentiation. Previous work in the Brafman lab led to the engineering of a chemically defined substrate—vitronectin derived peptide (VDP), which allows for the long-term expansion and differentiation of hNPCs. In this work, we use this substrate as the basis for a microcarrier (MC)-based suspension culture system. Several independently derived hNPC lines were cultured on MCs for multiple passages as well as efficiently differentiated to neurons. Finally, this MC-based system was used in conjunction with a low shear rotating wall vessel (RWV) bioreactor for the integrated, large-scale expansion and neuronal differentiation of hNPCs. Finally, VDP was shown to support the differentiation of hNPCs into functional astrocytes. Overall, this fully defined and scalable biomanufacturing system will facilitate the generation of hNPCs and their derivatives in quantities necessary for basic and translational applications.
ContributorsMorgan, Daylin (Author) / Brafman, David (Thesis advisor) / Stabenfeldt, Sarah (Committee member) / Wang, Xiao (Committee member) / Arizona State University (Publisher)
Created2018
Description
Two distinct aspects of synthetic biology were investigated: the development of viral structures for new methods of studying self-assembly and nanomanufacturing, and the designs of genetic controls systems based on controlling the secondary structure of nucleic acids. Viral structures have been demonstrated as building blocks for molecular self-assembly of diverse structures, but the ease with which viral genomes can be modified to create specific structures depends on the mechanisms by which the viral coat proteins self-assemble. The experiments conducted demonstrate how the mechanisms that guide bacteriophage lambda’s self-assembly make it a useful and flexible platform for further research into biologically enabled self-assembly. While the viral platform investigations focus on the creation of new structures, the genetic control systems research focuses on new methods for signal interpretation in biological systems. Regulators of genetic activity that operate based on the secondary structure formation of ribonucleic acid (RNA), also known as riboswitches, are genetically compact devices for controlling protein translation. The toehold switch ribodevice can be modified to enable multiplexed logical operations with RNA inputs, requiring no additional protein transcription factors to regulate activity, but they cannot receive chemical inputs. RNA sequences generated to bind to specific chemicals, known as aptamers, can be used in riboswitches to confer genetic activity upon binding their target chemical. But attempts to use aptamers for logical operations and genetic circuits are difficult to generalize due to differences in sequence and binding strength. The experiments conducted demonstrate a ribodevice structure in which aptamers can be used semi-interchangeably to translate chemical inputs into the toehold switch paradigm, marrying the programmability and orthogonality of toehold switches with the broad sensing potential of aptamer-based ribodevices.
ContributorsMcCutcheon, Griffin Cooper (Author) / Green, Alexander (Thesis advisor) / Hariadi, Rizal (Committee member) / Stephanopoulos, Nicholas (Committee member) / Wang, Xiao (Committee member) / Arizona State University (Publisher)
Created2022