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Control Approaches for Exosuits in Clinical Populations during Diverse Activities
No full textIndividuals with musculoskeletal or neurological diseases often experience mobility challenges, including reduced walking efficiency and limited automaticity. For example, gait patterns of individuals post-stroke are hemiparesis, characterized with slowness and asymmetry; more than half of individuals with Parkinson's Disease (PD) experience Freezing of Gait (FoG), characterized with brief, episodic reduction or absence of forward progression. Despite existing solutions such as rehabilitation and passive assistive devices, these impairments elevate injury risk and diminish quality of life for clinical populations.
Wearable electromechanical devices have been developed to restore mobility, compensate for impairments, and enhance rehabilitation outcomes for clinical populations. Specifically, joint-targeting devices have achieved notable success in clinical populations to reduce physical effort, increase practice volume, and retrain regular biomechanics. Nevertheless, controllers of joint-targeting devices have been tuned for continuous, steady walking, relying on gait phase estimation or gait event detection algorithms only reliable under supervised, controlled conditions. Pathological gait poses additional challenges in controllers with atypical and varying joint kinematics, short and intermittent walking bouts, and impaired endurance.
This thesis seeks to address the challenge in assisting pathological gait in real world settings by designing and evaluating control systems that are robust in diverse activities. First, we deployed a unilateral hip flexion exosuit for inpatient gait retraining in clinics. We expanded the use of exosuit to non-ambulatory patients and non-walking repetition practices during actual rehabilitation sessions by implementing assistance triggered by pysical therapisyt (PT). The exosuit eliminated the needs for manual limb support from PTs and increased participants' gait endurance or speed. We also executed a community Robotic Exosuit Augmented Locomotion program to support community walking practices with a semi-active ankle exosuit. Four participants in the chronic stage of stroke independently used the ankle exosuit for 4 weeks. Two participants significantly increased their unassisted paretic propulsion and daily steps following the program.
Second, we developed a turning-specific controller to provide assistance during transitional activities and reduce FoG for individuals with PD This is an advancement from a case study of a single individual performing straight-line walking to a full study of nine participants performing turning and freeze-provoking activity. We developed a controller for a bilateral hip flexion exosuit to accommodate asymmetry and reduced range of motion during turning. The turning controller successfully assisted various activities, including individual-specific hotspots, and led to a significant reduction in freeze severity and improvements in turn quality.
Third, we further innovated the controller for individuals with PD and established a control strategy combining adaptive oscillators and machine learning models to accommodate complex freeze-provoking trajectories at home. We introduced a phase-coupling term in adaptive oscillators to greatly improve the reliability and adaptability of gait phase estimation in transitional activities. As gait further deteriorated due to FoG, we switched to a kinetic-based gait phase realized by a machine learning (ML)-estimated weight distribution, along with ML-estimated FoG probability to determine FoG in real time. This control strategy was validated in complex at-home hotspots, demonstrating reliable assistance across various gait representations and a successful reduction in freeze severity and improvement in functional outcome.
Together, this thesis advances control systems for wearable assistive devices that consistently assist pathological gait patterns, validated in real-world deployment in clinics and communities for individuals post-stroke, and versatilely adapt to diverse activities and complex trajectories at home, demonstrating a life changing reduction in FoG for individuals with PD. We removed several key barriers and established a practical path to implement exosuits from laboratory to clinics and home.Engineering and Applied Sciences - Engineering Science
Essays in Macroeconomics and Labor Markets
No full textLabor markets are a central channel through which households are exposed to macroeconomic shocks. This dissertation comprises three essays that examine and quantify the impact of macroeconomic fluctuations on labor markets in the United States. Chapter 1 studies how individual job destruction decisions spill over to other workers in the labor market. Using variation in the nationwide layoff decisions of large firms across local labor markets, we find that labor market congestion, caused by many firms simultaneously destroying jobs, significantly amplifies the earnings losses of laid-off workers during economic downturns. We further interpret these findings through the lens of a heterogeneous-agent quantitative model with labor market frictions. Chapter 2 investigates how a decline in the compensation investors demand for bearing firm default risk affects the allocation of labor. We show that loose credit market conditions lead more workers to take jobs at financially risky firms. Using variation from both labor market and credit market segmentation, we find that taking jobs at these firms increases workers’ labor income in the short run, but results in large and persistent earnings losses once credit conditions tighten. Chapter 3 studies how the provision of mortgage forbearance helped stabilize labor markets following the 2020 COVID-19 recession. Using variation in financial intermediation frictions across mortgage servicers, we find that mortgage payment deferrals under the federal forbearance program accelerated the recovery of local employment once economic lockdowns were lifted. Our estimates underscore the importance of household liquidity in boosting labor demand during economic downturns.Economic
From Cell Type to Structure: A Multiscale Framework for Discovering Psychiatric Genes, Pathways and Mechanisms
No full textHuman genetics is perhaps the most fundamental biomarker for neuropsychiatric disorders. While genetic studies have identified numerous risk loci for neuropsychiatric disorders, the biological mechanisms they perturb remain unclear. We generated single-nucleus RNA sequencing (snRNA-seq) data from across multiple human brain regions to weight neuronal co-expression patterns by polygenic heritability, enabling the identification of disease-relevant pathways from common variant architecture. This framework was robustly validated through convergence with rare variant signals from large-scale exome sequencing data across multiple neuropsychiatric disorders, revealing Ca2+ homeostasis as a central and recurrent axis of genetic vulnerability.
Within this pathway, we identified ATP2B2 – a P-type ATPase responsible for Ca2+ extrusion – as consistently downregulated in the prefrontal cortex in donors with schizophrenia compared to controls, both synaptic proteomes and snRNA-seq. This reduction is specific to excitatory neurons, pointing to a cell type-specific loss-of-function mechanism linking ATP2B2 to schizophrenia risk.
ATP2B2 displays a striking enrichment of missense variants implicated in schizophrenia, autism and neurodevelopmental disorders. To investigate the structural basis of this signal, we developed a 3D enrichment framework that leverages the AlphaFold 3-predicted structure to pinpoint structurally constrained mutational hotspots with likely functional impact. We did this by testing for an excess of case-derived variants within 15Å spherical neighborhoods around each residue, and identify compelling candidates for downstream mechanistic interrogation. We identified an enrichment of case-derived
variants localized in close spatial proximity to both the Ca2+ permeation tunnel and binding site and the ATP:Mg2+ coordination site, suggesting two distinct mechanisms of ATP2B2 perturbation. In the Ca2+ binding neighborhood, substitution of the Ca2+-coordinating residue E457 with lysine (E457K) introduces a charge-reversal, suggesting disrupted binding as a focal mechanism of pathogenic variation in ATP2B2. We used AlphaFold 3 to simulate ATP2B2 and Ca2+ with and without E457K, and found it markedly reduced Ca2+ contact probabilities relative to wildtype, supporting an LoF effect.
We validated E457K’s impact in two orthogonal assays. The variant abolished ATPase activity in recombinant ATP2B2 and in a cellular context it impaired Ca2+ extrusion in HEK293 cells using a GCaMP6s-based imaging assay - both consistent with a LoF mechanism aligned with the direction of genetic risk. This suggests that case variants in ATP2B2 very likely compromise its function and disrupt intracellular Ca2+ homeostatic equilibrium. Our study constitutes a significant contribution to the neurobiological elucidation of etiological genetic risk and advances mechanistic insight into the pathogenesis of neuropsychiatric disorders.
First, in a biochemical assay, the variant completely abolished ATPase activity in recombinant ATP2B2. Second, in a cellular model, GCaMP6s-based Ca2+ imaging in HEK293 cells revealed a marked impairment in Ca2+ extrusion, indicative of disrupted Ca2+ clearance. Both findings converge on a loss-of-function mechanism, consistent with the direction of genetic risk observed in neuropsychiatric cases as well as the downregulation seen in functional genomics datasets. These results strongly suggest that pathogenic variants in ATP2B2 compromise its physiological role in maintaining intracellular Ca2+ homeostasis. Our study constitutes a significant contribution to the neurobiological elucidation of etiological genetic risk, and provides a mechanistic link between rare genetic variation and disrupted neuronal Ca2+ signaling, offering novel insight into the molecular pathogenesis of neuropsychiatric disordersBiological and Biomedical Science
Accelerating the development and application of genetic medicines
No full textPrecision gene editing offers the promise of permanently correcting alleles that cause disease in the genomes of living individuals with one-time treatments. Making this promise a reality has been a major objective of molecular biology and genetics since the descriptions of the mutations that cause cystic fibrosis—the first for any genetic disease—were reported in 1989. My doctoral work has focused on developing and applying precision gene editing technologies to correct cystic fibrosis-causing pathogenic mutations at ever increasing scales.
In Chapter 1, I discuss cystic fibrosis, an autosomal recessive disease arising from loss-of-function mutations in the CFTR gene. I summarize the pathogenesis and genetic epidemiology of the disease, and I describe the profound need for highly effective medicines that correct cystic fibrosis-causing alleles in the age of highly effective CFTR modulator therapies. I also illustrate how recent advances in gene editing technologies enable us to explore the development of genetic medicines at scale in different ways.
In Chapter 2, I describe the systematic optimization of prime editing to efficiently correct CFTR.F508∆, a three-nucleotide deletion that is the predominant cause of cystic fibrosis. By combining six recent advances in prime editing—epegRNAs, the PEmax architecture, MLH1dn, strategic silent edits, PE6, and dsgRNAs—we increased CFTR.F508∆ correction efficiency from .5% in model HEK293T cell lines to 58% in therapeutically relevant immortalized bronchial epithelial cells. In primary airway epithelial cells derived from people with cystic fibrosis, the optimized prime editing strategy enabled 25% precise correction of CFTR.F508∆, a 140-fold improvement over initial prime editing systems, with a 3.4-fold higher edit-to-indel ratio than nuclease-mediated homology directed repair approaches, and minimal off-target editing. Editing primary airway cell cultures restored CFTR ion channel function to >50% of wild-type levels, comparable to treatment with the modulator drug combination elexacaftor/tezacaftor/ivacaftor. Direct and efficient correction of CFTR.F508∆ suggests a durable one-time treatment for cystic fibrosis and provides a blueprint for optimizing prime editing to correct other pathogenic gene variants, including the >1000 other cystic fibrosis-causing alleles that have been described to date.
In Chapter 3, I present work that explores a variety of therapeutically viable viral and nonviral strategies for the efficient delivery of CFTR-correcting prime editors to the airway. Building on the high-potency CFTR.F508∆ gene editing strategies we devised in Chapter 2, we formulate helper-dependent adenoviral vectors (HD-Ad), engineered virus-like particles (eVLPs), and lipid nanoparticles (LNPs) for the delivery of prime editors to both differentiated and undifferentiated primary airway cells as well as to human bronchial epithelial cell lines. We demonstrate functional restoration of CFTR-mediated channel currents that approaches the efficacy of existing small molecule modulator drugs by delivering CFTR.F508∆-correcting prime editors via HD-Ad and eVLPs. We also develop a prime editing formulation for the modulator-ineligible allele CFTR.G542X that achieves 40% correction efficiency when delivered to 16HBEge cells via LNPs. To enable efficient packaging of CFTR-correcting prime editors into dual adeno-associated viral (AAV) vectors, we assess the ability of size-minimized SpCas9 domains to support prime editing. Our work provides insight into the relationship between editing efficiency and functional correction for CFTR, suggesting that modest (~3%) editing efficiencies can restore CFTR function to therapeutic levels but that 15-20% correction is likely needed to match the functional levels of current standard-of-care modulators. These proof-of-concept in vitro experiments lay a foundation for the evaluation of diverse delivery modalities to correct CFTR via prime editing in the airways of animal models.
In Chapter 4, I summarize efforts to leverage self-targeting lentiviral screens to accelerate the optimization of prime editing formulations for therapeutic applications. We describe LVPrime, a computational toolkit that facilitates the design and analysis of self-targeting lentiviral screens for optimizing therapeutic pegRNAs. We use these tools to screen thousands of pegRNAs for 17 cystic fibrosis-causing alleles that do not respond to modulator therapies in multiple cell types, including ~3200 pegRNAs to correct W1282X, ~2300 pegRNAs to correct c.489+1G>T, and ~1100 pegRNAs to correct R1162X. We also describe adaptive triaging, an active learning method that enables iterative optimization of pegRNA silent edit strategies in an allele- and cell type-agnostic manner. Adaptive triaging facilitates identification of high efficiency pegRNAs without the need to comprehensively screen multiple pegRNA parameters simultaneously and shows efficacy across a diversity of experimental scales. The tools and findings from this work could be applied in future translational studies that endeavor to correct disease-causing alleles via prime editing in preclinical settings.
In Chapter 5, I describe a series of studies to evolve and characterize botulinum neurotoxin (BoNT) proteases to cleave therapeutically relevant protein targets. We used phage-assisted evolution to reprogram BoNTs to cleave procaspase-1, a mediator of programmed inflammatory cell death, and NaV1.7, a voltage-gated sodium channel implicated in the sensation of acute pain. We also developed an efficient platform to broadly characterize the substrate specificity of both wild-type and evolved BoNT protease variants. Substrate profiling of evolved procaspase-1 cleaving BoNT/X variants demonstrated that evolved proteases acquired enhanced substrate sequence specificity over their wild-type BoNT/X starting point and enabled the nomination of proteome-wide off-target cleavage activity. We leveraged substrate profiling to inform evolutionary trajectories for a NaV1.7-cleaving BoNT/E, demonstrating that information on the substrate preferences of proteases can be used for the forward design of reprogrammed proteases. Together, these results expand the repertoire of experimental tools available for therapeutic protease development.Biophysic
Characterization of endogenous cereblon substrates with new chemical biology methods
No full textC-Terminal cyclic imides are posttranslational modifications on proteins that are recognized and removed by the E3 ligase substrate adapter cereblon (CRBN). Despite the observation of these modifications across the proteome by mass spectrometry-based proteomics, an orthogonal and generalizable method to visualize the C-terminal cyclic imide would enhance detection, sensitivity, and throughput of endogenous CRBN substrate characterization. I therefore developed an antibody-like reagent, termed “cerebody,” for visualizing and enriching CRBN substrates. I describe my protein engineering campaign to identify cerebody and showcase cerebody’s utility in identifying CRBN substrates by Western blot and enrichment from whole cell and tissue lysates. Furthermore, CRBN substrates identified by cerebody enrichment are mapped, validated, and further characterized for dependence on the C-terminal cyclic imide modification. These methods will accelerate the characterization of endogenous CRBN substrates and their regulation.
In Chapter 1, I introduce the E3 ubiquitin ligase substrate adaptor CRBN and provide an overview of its clinical use, structure, substrates it recognizes, and recognition motifs. I then discuss methods to recognize CRBN substrates through the C-terminal cyclic imide modification, and criteria for improved methods for detection of CRBN substrates through this modification. In Chapter 2, I discuss the directed evolution of CRBN’s thalidomide binding domain through random mutagenesis phage display in pursuit of a tool protein for detection of CRBN substrates. In Chapter 3, I move from the thalidomide binding domain scaffold to CRBNmidi and describe the development of the tool protein called cerebody. I then detail its use for detection and enrichment of endogenous CRBN substrates. In Chapter 4, I describe the validation of UROD as a PCMT1-dependent CRBN substrate, a discovery which was enabled and accelerated by cerebody enrichment. In Chapter 5, I discuss the utility of cerebody beyond endogenous substrates, including its use for identifying CRBN neosubstrates and its potential alterations to also provide structural insight into the interface between CRBN and its substrates and neosubstrates. I also provide detailed protocols for the implementation of cerebody methods. I conclude by discussing the broad applicability for cerebody and derived methods to illuminate CRBN’s biology, and what happens when these pathways are altered.Chemistry and Chemical Biolog
On-chip generation and manipulation of quantum states of light in thin-film lithium niobate
No full textWithin the last decade, thin-film lithium niobate (TFLN) has emerged as a leading integrated photonics platform with immediate applications in classical communications. Concurrently, TFLN components tailored to the requirements of photonic quantum information processing have also garnered considerable interest. Chief among these is the quasi-phase matched (QPM) nonlinear
frequency mixer—implemented via periodic domain inversion in ferroelectric lithium niobate—which can be used to generate photon pairs, squeezed states of light, and to perform single photon frequency conversion. Here, we demonstrate progress towards realizing a spectrally separable photon pair source in lithium niobate via waveguide dispersion engineering—a technique uniquely enabled by sub-wavelength optical mode confinement in the thin-film platform. Subsequently, we design optimize a scalable fabrication process to produce QPM TFLN devices for applications that require strict adherence to a specified operating wavelength, such as quantum frequency conversion in a quantum communications network. Finally, we explore how high-performance electro-optic devices can be combined with these nonlinear optical devices to realize a multi-functional platform in which quantum states of light can be generated and manipulated within a single, compact photonic integrated circuit.Engineering and Applied Sciences - Applied Physic
Applications and Host Responses to Adeno-Associated Virus (AAV) as a Gene Therapy Vector in the Eye
No full textAdeno-associated virus (AAV) vectors are state-of-the-art delivery vectors to treat genetic diseases. In 2017, the first AAV gene therapy was approved by the Food and Drug Administration. Called Luxturna, this AAV gene therapy helps restore vision in patients suffering from an inherited form of blindness called Leber’s Congenital Amaurosis Type 2 (LCA2). Beyond LCA2, there are many blinding diseases, such as retinitis pigmentosa (RP) and age-related macular degeneration (AMD), for which it may be possible to slow vision loss via gene agnostic therapies leveraging AAV delivery vectors.
Dry AMD is the most frequent cause of visual impairment in individuals over age 50 in developed countries. It is characterized by subretinal deposits of oxidized proteins and lipids and results in progressive loss of high acuity vision. One major risk factor is smoking, which causes oxidative stress in many tissues, including the eye. We previously showed that an adeno-associated viral vector expressing human NRF2 (AAV8/Best1-Nrf2), a transcription factor that regulates responses to oxidative damage, slowed degeneration in mouse models of another blinding disorder, RP, which also includes oxidative stress. Here, our AAV8/Best1-Nrf2 vector was tested in a model of oxidative stress wherein sodium iodate was injected systemically, as this is often used to model dry AMD. Sodium iodate causes acute oxidative damage to supporting cells of the retina, the retinal pigment epithelial cells, and ultimately leads to photoreceptor death. Subretinal injection of AAV8/Best1-Nrf2 led to protection of the retinal pigment epithelium and photoreceptors, as well as preservation of visual function, in rat and mouse sodium iodate models. AAV8/Best1-Nrf2 may serve as an effective gene-agnostic therapy for diseases with oxidative stress, including dry AMD.
As AAVs have also been reported to elicit ocular toxicity in the clinic, we additionally sought to elucidate mechanisms of toxicity upon subretinal injection of toxic or nontoxic AAVs in wild type and immune KO mice. Several transgenes, self and non-self, were tested for toxicity, with no clear correlation for this variable. Bulk RPE RNA-sequencing revealed upregulation of translational processes, cell stress, cytokine release, antiviral responses, and leukocyte infiltration pathways. Possible toxicity-inducing pathways were explored for causality by injecting toxic AAVs into mice deficient for intrinsic, innate, or adaptive immune pathways. Knocking out the interferon receptor, IFNAR1, partially alleviated toxicity. RNA-sequencing revealed >200 interferon stimulated genes upregulated in toxic vs. non-toxic AAV-injected RPE samples. In situ hybridization of interferon pathway transcripts (IFNB1, IFNAR1) revealed that the RPE and retina can produce and potentially respond to interferon. These data suggest that transgene-induced cell stress responses and host interferon responses contribute to toxicity following subretinal injection of AAVs encoding particular transgenes. This work elucidates mechanisms of toxicity following ocular AAV administration.Virolog
Kant’s Free Play and Aesthetic Judgment in Architecture: A New Interpretation as Visual Calculating
No full textFollowing Kant’s view of drawing or shape as the “proper object” of aesthetic judgments in architecture, I present an interpretation of a central concept in his theory of aesthetic judgment, viz., the free play of imagination and understanding, as visual calculating in shape grammars. Calculating with identity rules formalizes Kant’s reflective judgments in free play, which he explains as imagination sustaining a “lively engagement” with form. This interpretation departs from determining judgments, which underlie twentieth-century mathematical and computational approaches to aesthetics. With this interpretation in place, I address a central issue concerning computation and aesthetic intelligence, engaging Kant’s concept of “adherent beauty”: How are we to employ computation as a practical method for value judgment while preserving free play’s reflective property that refreshes aesthetic experience, especially when creative work must meet defined functions and end-goals?Computer ScienceHistory of Art and ArchitecturePhilosophyArt, Film, and Visual StudiesAccepted Manuscrip
Investigating PD-1 regulation of CD8+ T cell fate following acute influenza infection
No full textThe cornerstone of the adaptive immune system is the capacity to remember and rapidly respond to previously encountered pathogens. Tissue-resident memory (TRM) CD8+ T cells represent a specialized subset of memory T cells that maintain permanent residence at common barrier tissues and enable rapid response to tissue perturbation upon re-encounter with antigen. Checkpoint receptors such as PD-1 have been heavily studied in the context of chronic antigen settings but are also known to be upregulated constitutively on TRM. However, the impact of PD-1 on the differentiation and function of CD8+ TRM cells following acute contexts is not well understood.
This work explores how PD-1 signaling regulates the differentiation of CD8+ TRM cells following acute influenza infection. Genetic deletion of PD-1 in influenza-specific CD8+ T cells impaired acquisition of CD69 and CD103 – canonical TRM markers – at both effector and memory time points. Targeted deletion of an exhaustion-associated PD-1 enhancer region attenuated PD-1 expression in CD8+ T cells in flu-infected tissues, reducing tissue residency marker acquisition. Mouse models with germline or CD8-specific PD-1 loss had reduced tissue-residency marker expression within antigen-experienced CD8+ T cells. However, antibody blockade of the PD-1 pathway during priming did not recapitulate the TRM defects observed with genetic deletion. Our findings highlight PD-1's complex, context-dependent effects on CD8+ T cell fate decisions and establish its critical role in balancing protective immunity with tissue damage following respiratory viral infection.Graduate Educatio
Mimi and the Jackalope
No full text“Mimi and the Jackalope” is a midgrade portal fantasy set in West Texas during
the early years of the Operation Iraqi Freedom (OIF) and Operation Enduring Freedom
(OEF) campaigns in the Middle East. In this story, a young girl—Mimi—is raised by her
older half-sister, a veteran of OIF/OEF. Mimi has difficulty understanding the way her
sister’s PTSD manifests and seeks escape by having imaginary adventures in the rugged
environment of West Texas. During one of these adventures, Mimi encounters a mythical
Jackalope that is entirely real. She follows this Jackalope to a world populated by elves,
fairies, dragons, and other fantastic beings. As she explores, she realizes that the citizens
of this world are on the brink of their own war. Mimi ultimately must find a way to
communicate with her older sister in order to protect the fairy world she has come to
love.
“Mimi and the Jackalope” is a portal fantasy, but it is also a war story. It is not a
story about fighting wars, but a story about the human cost of fighting wars. This story
pre-supposes (and argues) that war has a lasting traumatic effect not only on the fighters,
but also on all the friends and relatives of the dead, wounded, and witnesses. Even though
this story deals with mature themes such as war, PTSD, substance dependency, death,
and child neglect, these themes are handled in an age-appropriate way that is suitable for
midgrade readers.Extension Studie