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Single-Molecule Expansion Microscopy to Visualize Chromatin Structure and Heat Shock Response in Yeast
No full textExpansion microscopy (ExM) is a recently developed technique that enables super-resolution imaging using conventional widefield and confocal microscopes. This method involves anchoring specimens to a hydrogel, followed by physical expansion in water, achieving approximately four-fold isotropic enlargement. ExM has been widely applied to mammalian cells and organoid specimens; however, its application to Saccharomyces cerevisiae presents unique challenges due to the presence of a rigid cell wall, which restricts the diffusion of anchoring molecules and stains. In this study, we developed a robust protocol integrating ExM with stochastic optical reconstruction microscopy (STORM) to visualize chromatin and protein spatial organization in yeast. To overcome the barrier imposed by the yeast cell wall, we incorporated an enzymatic digestion step using zymolyase, allowing efficient permeability of anchoring chemicals and DNA stains. We further optimized staining by integrating in vivo protein labeling using Janelia Fluor HaloTag ligand dyes, to minimize non-specific labeling often associated with post-expansion staining techniques. Additionally, we refined DNA staining and washing steps to enhance signal specificity and resolution. Applying this method to investigate the yeast heat shock response, we observed distinct morphological changes in the nucleolus, which adopted a circular shape following heat shock. Interestingly, chromatin organization mirrored this structural rearrangement, relocalizing within the circular nucleolus. Furthermore, we confirmed that RNA Polymerase II (RNAPII) remains predominantly localized within the nucleoplasm, consistent with known literature describing its role in mRNA transcription. This work establishes a novel ExM-STORM protocol tailored for yeast, enabling high-resolution visualization of chromatin spatial organization
Investigating the Use of Dipeptides as Singular Amino Acid Replacements in Chinese Hamster Ovary Cell Cultures
No full textThe objective of this study was to identify alternative high-solubility formats of essential amino acids for supplementation in mammalian cell culture-based protein production workflows. Current methodologies for supplementation involve supplying these amino acids in their singular forms. However, the biopharmaceutical industry encounters challenges with some of these amino acids due to their relatively low solubilities when supplied to cells via nutrient feed solutions, complicating the formulation of nutrient feeds and hindering the efficient transfer of cultivation strategies to larger manufacturing scales. Diluting feed solutions to overcome solubility challenges is not always feasible because total feed volume is limited by the maximum capacity of the closed bioreactor system. Given that multiple feeds are required for culture growth and productivity, it is imperative to find high solubility nutrient substitutions to enhance overall process efficiency.
This study investigated dipeptides, which are linked groups of two amino acids, as a potential solution to key limitations in industrial production processes due to their enhanced solubility compared to single amino acids. The linear forms of cysteine, leucine, and phenylalanine are examples of amino acids with relatively low solubilities at neutral pH levels. Both dipeptide dimers and other dipeptide forms of these amino acids were tested in various concentrations and combinations to observe their influence on cell growth and productivity. The dipeptide dimers included alanyl-cystine and lysyl-cystine, while the dipeptide forms included leucine-glycine, leucine-alanine, and phenylalanine-alanine
PARALLEL TRANSPORT ON LANDMARK MANIFOLDS IN LDDMM: THEORY AND NUMERICAL SCHEMES
Full text linkParallel transport provides the canonical mechanism for comparing tangent-space quantities across points on a Riemannian manifold. In computational anatomy, particularly within the Large Deformation Diffeomorphic Metric Mapping (LDDMM) framework for landmark data, it facilitates the transfer of deformation momenta between anatomical configurations, essential for groupwise registration, longitudinal studies, and statistical shape modeling. This thesis derives the second-order Hamiltonian system governing landmark parallel transport under the Levi–Civita connection, induced by a reproducing-kernel Hilbert space of vector fields.
We implement and systematically compare five numerical schemes: explicit Euler, fourth-order Runge–Kutta (RK4), the RK23 integrator, and the geometric Schild and Pole ladder constructions, realized via LDDMM shooting and matching operations. Experiments conducted on five two-dimensional landmark models—circle–ellipse, star, cardioid, circle–star, and circle–smooth-square—assess each method’s accuracy, computational efficiency, and momentum conservation across varying kernel widths and matching tolerances.
Results identify RK4 as the most effective integrator, offering exact numerical transport (as the defined ground truth), minimal computational cost, and superior Hamiltonian preservation compared to all other methods. While RK23 achieves comparable accuracy, it does not outperform RK4 in speed or stability. Euler is efficient but incurs moderate errors, and ladder methods exhibit significant shape-dependent instability and large relative errors. Consequently, RK4 is strongly recommended as the default numerical scheme for reliable and efficient landmark parallel transport tasks in computational anatomy
DEVELOPMENT AND EVALUATION OF PEGYLATED POLY (ASPARTIC ACID) NANOPARTICLES FOR CISPLATIN DELIVERY IN GLIOBLASTOMA TREATMENT
No full textGlioblastoma (GBM) is one of the most aggressive and common malignant brain tumors. Treating GBM is a challenge due to its highly aggressive nature and the blood-brain barrier (BBB) restriction which needs to be passed to deliver drugs to the brain. Conventional chemotherapy drugs, such as cisplatin (CDDP), have significant drawbacks such as off-target toxicity, limited ability to cross the BBB and poor brain penetration once in the brain. Nanoparticle-based drug delivery systems, combined with MRI-guided focused ultrasound (FUS), offer a promising non-invasive strategy to enhance cisplatin delivery to glioblastoma tumors. In this study, poly(aspartic acid) (PAA) was selected to encapsulate CDDP into nanoparticles due to its biodegradable nature and high drug encapsulation capacity. The surface of the nanoparticles was coated with polyethylene glycol (PEG) to increase stability in vivo and facilitate more effective penetration into brain tissue. We identified a promising brain penetrating nanoparticles (BPN) formulation that has desired physiological characteristics (diameter＜70 nm, zeta potential between -5 mV to 5 mV), high glioblastoma tumor cell cytotoxicity, and stability in biological environments. This formulation has the potential to be combined with MRI-guided focused ultrasound for efficient and non-invasive glioblastoma treatment
CHANGES AND FACTORS AFFECTING INPATIENT SELF-REPORTED QUALITY OF CARE EXPERIENCE IN CHINESE HOSPITALS
Full text linkThis nationwide study investigates patient self-reported quality of care experience—a multidimensional construct integrating subjective perceptions and objective evaluations of healthcare services—within China’s evolving hospital system (2016–2023). Guided by three research aims about patient experience (1. longitudinal trends; 2. relationship with institutional & individual factors; 3. relationship with the People-Centered Care (PCC)), we analyzed 834,546 inpatient records from tertiary, secondary, and community hospitals using multivariate regression and proportional odds models to address hierarchical data structures.
Our key findings reveal the overall patient experience in China in the past few years(2016–2023) is improving (by satisfaction, recognition, recommendations). These improvements likely reflect systemic enhancements in care coordination and PCC implementation across hospital tiers. Our research find out that tertiary hospitals showing more stable improving trends, while secondary hospitals experienced a pre-2020 better performance, and post-2020 declines. General hospitals performed more stable than specialized hospitals. Private hospitals had significantly higher per than public hospitals pre-2020. However, the difference became insignificant(at 5% level) after 2020. Individual factors, especially age and income, have a significant impact on patient experience.
Our study reveals that institutional, personal and process factors collectively influence the patient experience outcome. These findings advocate for composite intervention strategies addressing both systemic constraints and population-specific care expectations.
The study operationalizes PCC through five attributes: continuity (coordinated care pathways), information sharing (transparent communication), enhanced access (reduced barriers), effectiveness (perceived outcomes), and respect (dignity preservation), and confirms the transformative potential of PCC in enhancing patient experience, with care continuity emerging as the most influential attribute.
The findings propose three strategic priorities to sustain care quality improvements: (1) Structural reinforcement of tiered systems through PCC-driven specialization of secondary hospitals in chronic care, supported by digital referral protocols and financial incentives for hierarchical care adherence; (2) Equity-focused resource allocation via tiered insurance reimbursement models and specialist outreach to rural facilities; (3) Technology-augmented process optimization leveraging AI-driven triage systems and real-time patient feedback platforms to standardize dignity-preserving practices. These priorities collectively address systemic fragmentation while operationalizing PCC’s five attributes—as interdependent levers for equitable health system transformation
A LIFE OF CURIOSITY
Full text linkI have always wanted to be a science writer; I just did not know it. Constant questions plagued my science teachers through grade school, and I could never settle on any one field to study, so decided to choose all of them by studying journalism. Now that I have committed to a career of learning, I have gone back through my life and pulled from the environment I grew up in, interests both as a child and adult, and some of the most impactful moments of my life to make up for lost time and answer a few of those ceaseless questions
INTERNALIZED MULTILEVEL VIOLENCE AMONG BLACK PEOPLE LIVING IN AREAS OF CONCENTRATED DISADVANTAGE IN BALTIMORE, MD
No full textBlack people living in areas of concentrated disadvantage (ACD) in the United States experience inequitable structural, collective, and interpersonal violence that may internalize as self-directed violence (SDV). However, there is relatively little research on the internalization of multilevel violence in Black communities. This research advances the state of the art by examining a process of multilevel violence internalization within a community-based sample of Black people living in ACD in Baltimore, MD, USA.
This study takes a gender approach using multiple groups structural equation modeling to evaluate the psychometric properties of a population-based measure of SDV risk. Additionally, this study examines structural differences between females and males in a longitudinal model of internalization from perceived neighborhood disorder (PND) through exposure to interpersonal violence (ETV) to SDV. Finally, the study uses the same approach to evaluate how support network characteristics of size, density, percentage in the family, and multiplexity influence this process. Results found the 2-factor 10-item population-based measure of SDV was reliable and displayed measurement invariance and concurrent validity for females and males. The study found evidence of violence internalization such that PND at baseline predicted later exposure to witnessing interpersonal violence (WV) directly correlated with experiencing interpersonal violence (EV) that then significantly associated with SDV risk.
Finally, the study found significant gender differences in the influence of network characteristics on violence internalization. Greater support network size associated with greater SDV risk through greater ETV. However, this effect was significantly stronger among females who reported less PND. Meanwhile, network density was significantly associated with less SDV risk only among males. Support multiplexity significantly decreased SDV risk among females, however this effect did not significantly differ from males, suggesting it may offer generalizable protective effects. These results suggest that community-based interventions on SDV are practicable and may offer distinct advantages over individual-level interventions. Results also suggest that SDV research and prevention among Black people living in ACD would be more effective when considering compositional and structural differences between females and males
IN-VITRO CHARACTERIZATION OF SEASONAL INFLUENZA A VIRUSES IN MARYLAND DURING THE 2023-2024 SEASON
Full text linkThe 2023–2024 influenza season in Baltimore, MD exhibited an atypical epidemiological pattern for Influenza A viruses, initially dominated by H1N1, later transitioning to H3N2 dominance, contrasting distinctly with the patterns observed in the previous two seasons. To understand the underlying reason of this unusual pattern, we investigated the antigenic characteristics, viral fitness, and cell tropism of circulating H1N1 and H3N2 viruses. Phylogenetic analysis revealed co-circulation of two distinct H1N1 clades (6B.1A.5a.2a and 6B.1A.5a.2a.1) early in the season. A representative H1N1 virus from clade 6B.1A.5a.2a, harboring mutations in HA antigenic sites (K160R, K169Q, R142K, R223Q), and a representative H3N2 virus from clade 3C.2a1b.2a.2a.3a.1, were selected to be compared on their viral fitness and cell tropism, and the representative H1N1 virus was also used for serum neutralization studies. H1N1 vaccine efficacy assessments using serum neutralization assays showed greater fold change in pre- and post-vaccination antibody titers, along with higher seroconversion rates for the vaccine virus compared to seasonal viruses, indicating that the vaccination provides stronger protections for the vaccine virus itself in contrast to the antigenically distinct seasonal virus. However, H1N1 seasonal virus showed higher neutralizing antibody titers at pre-vaccination level, suggesting the sample population may have pre-existing immunity against this virus strain. Replication kinetics in human nasal epithelial cell (hNEC) cultures revealed distinct growth dynamics at late times post infection, where H3N2 seasonal virus displayed a fitness advantage compared to H1N1 seasonal virus at 37oC. Cell tropism studies in hNEC cultures revealed H3N2 seasonal virus infected goblet cells more efficiently than H1N1 seasonal virus, likely due to its preference for elongated α2–6 sialic acid receptors. Further investigation into serological studies on H3N2 seasonal virus and H1N1 seasonal virus from the other clade (6B.1A.5a.2a.1) may provide more insights on the dynamics between the H1N1 and H3N2 circulating viruses observed this season
Phenomenological shift in Bioethics Methodology
No full textAs the most used tool, principlism is a valuable and efficient methodology for bioethics. The scientific progress in medical care should mean there needs to be more tools in bioethics to better address people’s issues and reevaluate our understanding of care.
In this paper, I argue that phenomenology can help enrich the use of principlism to analyze ethical issues. Specifically, phenomenology can help us understand people’s embodied experiences, see how embodied experience may be shaped by social structures, and help healthcare providers to problematize their patients’ statements, which helps discover the hidden values related to their ethical decisions and reflect upon our own values and assumptions. Those methods could potentially provide us with a new perspective on the form of ethical questions healthcare providers should focus on and develop better understanding between doctors and patients
FROM GWAS TO BIOLOGICAL MECHANISM: MAKING STRIDES TOWARDS UNDERSTANDING AND TREATING NEUROLOGICAL DISEASE
No full textParkinson’s disease (PD) and schizophrenia are common neurological disorders with complex genetic architecture. Preceding research efforts have been dedicated to unearthing the genetic loci and protein-coding variants associated with these diseases, as well as identifying symptom management strategies. However, it is increasingly acknowledged that the complex genetic architecture of these two disorders extend beyond coding regions of the genome. This dissertation investigates the role of non-coding variation in the regulation of gene expression, with a particular focus on how this contributes to the pathogenesis of PD and schizophrenia. By combining genome-wide association studies (GWAS) data with cutting edge molecular biology techniques and functional genomic data, the primary objective of this work is to prioritize non-coding variation associated with disease risk and query functional regulatory elements for their role in mediating disease pathology.
Specifically, for schizophrenia, I characterize regulatory variation in neural progenitor cells by studying how neuroanatomically relevant enhancer regions drive differential expression of genes implicated in neuronal function and psychiatric risk. These findings contribute to a more nuanced understanding of the regulatory networks governing neuronal identity and disease susceptibility for individuals at risk for schizophrenia.
In my studies of Parkinson’s disease, I characterize a regulatory element specific to midbrain dopaminergic neurons, the primary cell type affected by neurodegeneration in this disease. By validating an enhancer element harboring PD-associated variation, my work provides insight into the non-coding architecture of PD and reveals novel therapeutic targets. A key aspect of this work is the development and validation of a novel gene therapy strategy targeting this cell-specific cis-regulatory element. By disrupting a midbrain dopaminergic-specific enhancer using CRISPR-based techniques, this work represents a proof-of-concept for a targeted therapeutic intervention strategy that significantly slows PD progression in a well-characterized mouse model of PD.
Overall, this dissertation provides a comprehensive examination of non-coding regulatory mechanisms in PD and schizophrenia, offering an advancement in our understanding of these disorders, as well as new therapeutic targets and strategies for disease intervention. My findings highlight the importance of non-coding variation in neurological disorders and pave the way for future research into gene regulation as a therapeutic avenue