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NEW TOOLS FOR IMAGE-INFORMED BONE TISSUE ENGINEERING
No full textCritical-sized calvarial bone defects pose complex clinical challenges due to complicated biological processes involved in bone healing. Among these, angiogenesis is essential for delivering oxygen, nutrients, and key cell types to the defect site, yet the detailed role of vascular function on bone growth and repair over the healing cycle remains insufficiently understood. Current clinical and preclinical studies mainly focus on structural imaging and lack insight into vascular functionality and the spatial-temporal effects of angiogenesis on bone regeneration at microvascular resolution. Furthermore, the successful integration of tissue-engineered constructs in calvarial defects requires effective oxygen delivery from the vasculature to sustain construct viability and promote long-term tissue repair. Recent advancements in tissue engineering, such as proangiogenic factors and oxygen-generating biomaterials, underscore the need for continuous monitoring of intravascular and tissue oxygenation to enhance our understanding of the bone healing microenvironment.
This thesis addresses these challenges by proposing (i): a novel imaging pipeline to facilitate longitudinal, in vivo characterization of calvarial bone healing, (ii): application of this imaging framework to study the impact of tissue-engineered treatments on bone regeneration in vivo, and (iii): the development of oxygen-sensing scaffolds capable of monitoring in vivo oxygenation in conjunction with the imaging platform. This pipeline will integrate high-resolution imaging techniques to provide quantitative data on vascular perfusion and tissue oxygenation throughout the bone healing process. Through detailed oxygen monitoring, the framework aims to reveal the role of oxygen gradients in cellular processes critical to bone repair.
The results of this study are expected to yield new insights into the in vivo dynamics of oxygen delivery and vascular function in bone healing, allowing quantitative analysis and improving the design of more effective treatment and tissue-engineering strategies. By combining scaffold design with real-time oxygen sensing and imaging, this research offers the potential for significant advancement in understanding and optimizing the conditions necessary for efficient calvarial bone defect repair. These findings may guide future therapeutic approaches, improving outcomes for patients with critical-sized bone defects
INVESTIGATING THE EXACT EFFECTIVENESS OF CUTTING PLANES OVER BRANCH-AND-BOUND IN INTEGER PROGRAMMING
Get PDFThis thesis investigates the exact effectiveness of cutting plane (CP) methods relative to branch-and-cut (BC) techniques in convex 0/1 integer programming under variable disjunctions, with a focus on the three-dimensional setting.
We prove that for a range of three-dimensional convex 0/1 integer programming problems, any valid inequality established by a BC proof tree can be derived by a corresponding pure CP proof tree with zero slack. Our approach combines structural induction and detailed geometric case analysis to systematically transform mixed BC proofs into CP-only proofs. In contrast to earlier ideas based on cut rotation or local approximation, our method fully replaces branching without increasing the overall proof size.
Although our results are currently limited to the three-dimensional case, they provide the first evidence that the finite substitution result may hold in low dimensions. Extending this to higher dimensions remains an open challenge
DEVELOPING AN INSTITUTIONAL REVIEW BOARD (IRB) HANDBOOK: A REGIONAL STUDY OF RESEARCH ETHICS PROCEDURES
Get PDFThis capstone project provides a comparative analysis of Institutional Review Board (IRB) handbooks and manuals of 16 universities within the Arab region, which led to the creation of an Institutional Review Board (IRB) handbook for Carnegie Mellon
University in Qatar (CMU-Q).
Currently the faculty, staff, and students have been conducting social behavioral research at CMU-Q with no central resource, where they can access the information that would guide them through the IRB process and provide necessary regulatory requirements. The purpose of this project is to understand the best practices for these institutions and the gaps in their current research policies. What was observed was the different level of details among these institutions; some provided clear ethical guidance and followed international or national regulations, while others did not include important sections like review types, definitions, principal investigator responsibilities, post-approval monitoring, and the IRB review process.
This comparison provides a set of recommendations to help design an IRB handbook for our university. These include things like clearly defining the ethical standards, whether international or national, the informed consent process, training requirements and principal investigator responsibilities, and the IRB composition and responsibilities. The goal of this IRB handbook is not only to provide CMU-Q researchers with an all-in one place to get all the necessary information they require to conduct ethical human subjects research, but also to be used by other institutions in the region as a model that they can follow to improve their IRB guidelines
DEVELOPING MULTIPLE R/BIOCONDUCTOR PACKAGES FOR: 1) HuBMAPR DATA RETRIEVAL, 2) DATA STORAGE, AND 3) BIASED FEATURE IDENTIFICATION IN SPATIALLY-RESOLVED TRANSCRIPTOMICS DATA
Get PDFGenomics data science has revolutionized biomedical research by providing unprecedented opportunities to investigate complex biological questions. Large-scale, high-throughput data, including single-cell RNA sequencing (scRNA-seq) and spatially resolved transcriptomics (SRT), offer novel perspectives on human disease, cellular composition, and tissue organization. These data enable researchers to dissect molecular mechanisms, facilitating deeper insights into human biology. Open-source software, particularly within the R/Bioconductor ecosystem, plays a critical role in bridging statistical methodologies with practical data analysis, fostering collaboration, reproducibility, and innovation in genomics. This thesis focuses on the development and implementation of open-source R/Bioconductor packages to enhance the accessibility and analysis of genomics data. Specifically, three packages were developed, each addressing a key challenge in genomics research.
First, the HuBMAPR package provides an efficient, programmatic interface for accessing and retrieving Human BioMolecular Atlas Program (HuBMAP) data. HuBMAPR facilitates the integration of HuBMAP resources into broader research workflows.
Second, the humanHippocampus2024 package, an ExperimentHub Data package within Bioconductor, enables convenient access to processed single-nucleus RNA sequencing (snRNA-seq) and SRT data from the Spatial Human Hippocampus Project (HPC) at the Lieber Institute for Brain Development (LIBD).
Third, the BatchSVG package introduces a binomial-deviance model-based quality control method for identifying biased spatially variable genes (SVGs) in the presence of batch effects. By examining per-gene deviance and rank values when the model was run with and without batch effects on SVGs, we developed a data-driven thresholding approach using the number of Standard Deviations for relative change in deviance and rank difference metrics
The Humanitarian-Development Nexus and Health Interventions in Fragile Settings
No full textObjectives
The intersection of humanitarian and development assistance in complex settings has been termed the humanitarian-development nexus (HDN), which focuses on the need to address the vulnerability of people before, during, and after crisis. The humanitarian-development-peace nexus (HDpN) expands the concept one step further to incorporate peace efforts. This dissertation research aims to make HDN/HDpN guidance more actionable and applicable for humanitarian and development actors, among other key stakeholders involved in health service delivery, by examining specific health and sexual, reproductive, maternal, newborn, child and adolescent health (SRMNCAH) interventions in fragile settings.
Methods
This dissertation consists of three manuscripts. The first manuscript is a scoping review that maps the existing literature related to the HDpN and its operationalization for SRMNCAH interventions in fragile settings. The second and third manuscripts are qualitative case studies from South Sudan and Mali, respectively, drawing from key informant interviews and a document review.
Results
The scoping review identified 45 publications that met inclusion criteria across 39 countries and addressed three aspects of HDN operationalization, including SRH service prioritization at different phases of an emergency; transitions between minimum and comprehensive health services; and health systems strengthening. In South Sudan, limited investment by the government in the health sector has perpetuated reliance on international assistance, and barriers to engagement with government counterparts have restricted coordination at the national level. In Mali, the government has worked to align humanitarian intervention packages with national health development plans. Attempts to shift from a reliance on humanitarian assistance have occurred, including the implementation of different health financing models. Investment in human resources for health was cited as a critical area of focus for the nexus in both case studies.
Conclusions
This dissertation research advances the literature on the HDN and health interventions in fragile settings, including SRMNCAH. This research is timely and relevant given the current global discourse on the HDN and the need to translate conceptual aspects of the nexus into practice for health. This research is relevant for all stakeholders involved in health service delivery in fragile contexts where both humanitarian and development health assistance are being delivered
MECHANISMS OF SMALL RNA REGULATION IN C. ELEGANS: CHARACTERIZING A NOVEL, NONCANONICAL miRNA BIOGENESIS PATHWAY AND INVESTIGATING miRNA ARGONAUTE ISOFORM DIVERSITY
No full textMicroRNAs (miRNAs) are short RNAs that post-transcriptionally regulate gene expression and play critical roles in development and differentiation. Despite their biological importance, the biogenesis of miRNAs is not completely understood. In canonical miRNA biogenesis, primary miRNAs are transcribed from intergenic loci or intronic regions by RNA polymerase II (Pol II). These transcripts are then processed by the Microprocessor, an enzyme complex comprised of Drosha and RNA-binding protein DGCR8 (known as PASH-1 in C. elegans). Subsequent processing by Dicer produces the miRNA that is loaded into Argonaute to repress target mRNAs.
This dissertation describes a newly identified, noncanonical miRNA biogenesis pathway in C. elegans. Analysis of a temperature-sensitive allele of pash-1 revealed a unique family of PASH-1-independent miRNAs, the mir-1829 family. Members of this germline-enriched miRNA family reside in the unusually long introns of three host genes that have no apparent overlapping functions. Based on 5′ RACE, promoter bashing experiments, and Pol III depletion assays, we determined that the mir-1829 family is derived from independent transcripts that are solely transcribed by Pol III. Although the biogenesis of the mir-1829 family bypasses the Microprocessor, we determined that it is Dicer-dependent using Northern blot, RT-qPCR, and small RNA-sequencing. Thus, we have delineated a novel biogenesis pathway involving Pol III and Dicer, but not Microprocessor. Future work will examine the biological function of these miRNAs.
Regulation of miRNAs also includes Argonaute loading. Argonautes, the core miRNA effector proteins, can exhibit isoform-specific functions, as shown by prior studies on CSR-1 isoforms in C. elegans. We investigate the isoform-specific roles of ALG-1/2, two miRNA Argonautes that each encode two isoforms that differ at their N-terminus. Using fluorescently tagged isoforms and isoform-specific knockout strains, we identified distinct expression patterns and functional roles for ALG-1 isoforms. When only the long isoform is expressed, developmental abnormalities such as protruding vulvas are observed, suggesting isoform-specific miRNA loading profiles. Ongoing studies aim to determine whether similar isoform-specific functions exist for ALG-2 and how these roles impact miRNA activity and development.
Together, these findings provide new insights into the complexity of miRNA biogenesis and regulation, advancing our understanding of small RNA biology
Onsager Theory of Wall-Bounded Turbulence: A Study of Dissipative Anomalies and Boundary Effects
Get PDFDissipation anomaly is a phenomenon in turbulent flow that apparently occurs when dissipation rate of kinetic energy asymptotically tends to a non-zero constant instead of vanishing as viscosity goes to zero. This phenomenon is a fundamental property of turbulence and is sometimes called the "zeroth law of turbulence''. Motivated by this, Lars Onsager, in late 1940s, conducted an exact analysis of the zero-viscosity limit of turbulent flow modeled by incompressible Navier-Stokes equations. His theory describes turbulent energy cascade through weak solutions of limiting Euler equations, providing a foundational understanding of this dissipative anomaly. The presence of boundaries, common in real-world turbulence, such as channel walls or the surface of an airplane wing, further complicates turbulent flows, leading to phenomena like drag, complex exchange processes for momentum, energy and vorticity, and possible breakdown of weak-strong uniqueness of weak Euler solutions. This dissertation provides a detailed and rigorous analysis of these aspects, extending Onsager's theoretical framework to wall-bounded domains. In particular:
Chapter 2: We extend Onsager-type analysis to wall bounded turbulence to study momentum dissipation anomaly hypothesized by Taylor and prove that anomalous skin friction is related to spatial momentum cascade for weak Euler solutions. This relation is analogous to the ``Onsager-Duchon-Robert'' relation between viscous energy dissipation anomaly and energy cascade.
Chapter 3: As an application of the results in Chapter 2, we show that the Josephson-Anderson (JA) relation for drag, valid for Navier-Stokes solutions for all , is also valid for weak Euler solutions obtained in the inviscid limit. By showing this connection between the JA relation and the Onsager-Duchon-Robert relation, we provide a novel resolution of d’Alembert paradox.
Chapter 4: We prove weak-strong uniqueness for suitable weak solutions of the Euler equations in both interior and exterior domains, establishing conditions under which uniqueness holds in the presence of solid boundaries. This analysis reveals significant implications for drag, dissipation, and extreme wall-stress fluctuations in turbulent flows.
Chapter 5: We extend Onsager-type analysis to finite Reynolds numbers, studying again momentum dissipation anomaly. We derive an upper bound on wall friction as a function of Reynolds number, providing a deterministic version of Prandtl's relation between drag laws and velocity profiles
INVESTIGATING THE MOLECULAR MECHANISMS OF GENETIC REGULATORY ELEMENTS USING SINGLE-MOLECULE MICROSCOPY
No full textGenetic regulatory elements are fundamental in controlling the timing, location, and levels of gene expression. Despite their importance, many regulatory mechanisms remain unclear. A key challenge lies in the nanometer-scale, highly dynamic interactions among these elements, which are difficult to capture with traditional biochemical methods. To address these challenges, my doctoral research has centered on using single-molecule microscopy to uncover the molecular mechanisms of three essential regulatory elements across prokaryotic, yeast, and human systems, providing detailed insights into their regulatory functions.
In the first half of my research, I collaborated with David Lilley's RNA Structure Lab to study riboswitch folding dynamics using single-molecule FRET (sm-FRET). The research explored how riboswitches control conformational changes to achieve translation inhibition, revealing that nascent RNA folding dynamics significantly impact their conformational switching capabilities. Chapter 2 will discuss these findings in detail, highlighting the intricate regulatory role of riboswitches.
The second half of my research shifted focus to understanding the molecular mechanisms underlying 3D chromosomal structure organization, employing high-speed AFM (HS-AFM) to capture dynamic structural changes at the nanoscale. Chapter 3 details the molecular mechanism by which pioneer transcription factors link multiple tether elements through site-specific multimerization. Chapter 4 delves into the chromosomal protein SMCHD1, revealing highly dynamic conformational changes that can be modulated by ATP and DNA binding. Collaborations with Toshio Ando, Carl Wu, and James Berger's lab were instrumental in investigating these complex interactions using HS-AFM. While my graduate research spans diverse topics, the central theme has been establishing connections between molecular structures, conformations, and functions with temporal resolution. I believe these single-molecule mechanistic studies offer essential foundations for future molecular manipulation strategies and potential therapeutic applications
Development of Computational Tools and Pipelines for Inferring and Modeling Gene Regulatory Networks
No full textGene regulatory networks (GRNs) are complex systems of interacting transcription factors (TFs) and genes that govern gene expression within a cell. Understanding GRNs is essential for studying cellular identities and functions. However, accurately inferring and constructing functional GRNs is not a trivial task. This dissertation presents methods developed for the inference of GRNs. The first part of the dissertation focuses on identifying the direct functional targets of CrebA in Drosophila salivary gland cells during organogenesis through the integration of ChIP-seq and scRNA-seq data. We first curated a single-cell atlas of Drosophila embryos by characterizing the embryonic tissues and identifying the transcription programs activated during organ specialization. This resource provides a baseline expression profile for the wild-type salivary gland cells, which we compared with CrebA mutant salivary gland cells to identify the differentially expressed genes affected by CrebA knockout. By integrating the differentially expressed genes with the CrebA binding locations identified using ChIP-seq, we determined the direct functional targets of CrebA and found that CrebA directly regulates genes involved in secretion. The second part of my dissertation focuses on the development of a novel tool called OneSC. OneSC is a computational method that infers functional GRNs and simulates synthetic single-cell expression profiles that recapitulate the cell state transitions using the inferred functional GRNs. We showed that OneSC can infer GRNs with higher simulation fidelity than current state-of-the-art methods. By applying OneSC to real scRNA-seq datasets, we showed that OneSC can accurately recapitulate cell state transitions and predict cell fate decision biases resulting from TF perturbations. Lastly, we applied OneSC to infer the core GRN driving osteosarcoma metastasis. In summary, this work presents a computational pipeline that integrates scRNA-seq with ChIP-seq to infer the direct functional target genes of CrebA in Drosophila salivary gland cells, and it introduces a computational tool designed to infer and simulate functional GRNs, enabling the simulation of cell state transitions and the prediction of the consequences of TF perturbations
Taming Interactions in Genomics and Clinical Trials
Get PDFThis dissertation develops statistical methods to address two challenges in contemporary biomedical research.
The first is understanding shared mechanisms underlying related diseases using high- throughput molecular data, motivated by a study of gene expression changes in a group of rare genetic disorders called Mendelian Disorders of the Epigenetic Machinery or MDEMs (Luperchio et al., 2021). We develop a simple approach to assessing the degree of overlap among the disorders based on rescaled differences in ranks of each gene across conditions, which we call “rank-gap” statistics. Rank-gap statistics have p-value-like properties that highlight concordance among genes ranked highly in at least one condition, and we argue that the number of smaller than expected rank-gap statistics estimates a meaningful overlap metric in the limit of small measurement error. Rank-gap statistics also possess three- and higher-way analogues. We compare our proposal with other rank-based overlap methods in the literature, and evaluate a diverse group of related methods in a simulation study. Our approach provides a simple complement to the predominant practice in genomics of forming Venn diagrams of statistically significant genes.
The second challenge is understanding how treatment effects vary among different types of patients using data from clinical trials. The standard approach—one-at-a-time subgroup or interaction analysis of candidate covariates—suffers from difficulties of inference and interpretation. We develop the proportional interaction model, a parsimonious extension of additive regression adjustment also considered by Follmann and Proschan (1999) and Kovalchik, Varadhan, and Weiss (2013) in which benefits and harms from treatment vary as a function of a “score” that also predicts outcomes in each treatment group. An asymptotic analysis enables us to assess the sample sizes required to detect proportional interactions in practice, and we propose a symmetrical re-parametrization of the model that allows for stable single-step inference. We re-analyze a clinical trial of treatments for schizophrenia using our approach, showing how proportional interactions can be incorporated into a traditional regression model-building framework