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Predicting Time-to-Event Incidence of Neurological Diseases and Mortality Using Objective Physical Activity Measures in the UK Biobank
Get PDFWearable accelerometers are instruments that can measure human activity in free-living environments in an objective and reliable way. Accelerometer-derived physical activity measures have been shown to be powerful predictors of many diseases, as well as mortality. This thesis presents three case studies that use data from the largest objective physical activity study to date, the UK Biobank, to predict time-to-event outcomes of: (i) Parkinson's disease; (ii) Alzheimer's disease; and (iii) mortality. The study of physical activity data is complicated by the fact that it can be processed and examined at different resolutions. For example, scalar summary statistics or vectors of high-resolution 1440 minute-level data points. Different outcomes and data resolutions present unique challenges that are addressed in this thesis with reproducible solutions
AUTOPHAGY AND NEURODEGENERATION: DISCOVERING CRITICAL INSIGHTS AND INNOVATIVE THERAPEUTIC STRATEGIES USING A NOVEL HUMAN NEURON MODEL OF BPAN
Get PDFBPAN (β-propellor protein-associated neurodegeneration) is a rare, X-linked, incurable, and devastating neurodegenerative disease, characterized by childhood static encephalopathy, subsequent dementia and movement disorders in adulthood, brain atrophy, axonal swellings, neurofibrillary tangles, and iron accumulation in the basal ganglia. BPAN is caused by loss-of-function mutation in the WDR45 gene, which encodes WIPI4, a critical adaptor for protein complexes involved in the maturation of vesicles in the degradative pathway of autophagy. Mounting but incomplete evidence attests to essential roles of autophagy in neuronal health and neurodegeneration, but there is great potential for resolving the remaining knowledge gaps about autophagy impairment in neurodegenerative diseases by investigating the pathophysiology of BPAN. However, cells and mice with WDR45 mutations or knock-out have thus far proven to be imperfect models of BPAN due to incomplete recapitulation of essential disease phenotypes, limited in part by a lack of studies specifically in human neurons. In this study, we successfully establish a novel human neuron model system for BPAN, consisting of patient-derived human iPSC cell line expressing the WDR45 mutation c.235 G>A (p.Ala79Thr, or A79T). We engineered these iPSCs to differentiate into cortical neurons (iNeurons), offering a platform to explore disease mechanisms and validate innovative rescue strategies rapidly and robustly. A79T iNeurons demonstrate abnormal levels of autophagy markers LC3 and p62, indicating impaired autophagic flux. We also found decreased cell viability in A79T iNeurons, which could not be mitigated by pharmacologic induction or inhibition of autophagy. Overall, our results confirm the usability of a novel human neuron model system for BPAN, which faithfully captures several essential disease phenotypes. Our experimental platform will serve as a pivotal foundation for exploring molecular mechanisms that are targetable for therapeutic intervention in both BPAN and other neurodegenerative diseases characterized by impaired autophagy
Characterization of TEAD1 Biomolecular Condensates in Renal Cell Carcinomas
No full textTEAD1 (TEA domain transcription factor 1) is a transcription factor in response to Hippo pathway signaling. In Renal Cell Carcinoma (RCC), a particularly lethal form of kidney cancer with poor treatment options, TEAD1 and its coactivators YAP1 are often up-regulated and indicate poor prognosis. TEAD-family transcription factors can bind YAP1 and form phase-separated biomolecular condensates, specifically transcriptional hubs, in response to hyperosmolarity and other cellular and environmental cues. We have found that in certain patient-derived RCC cell lines with high levels of YAP1 and TEAD1 expression, TEAD1 forms atypical large nuclear foci, distinct from normal kidney cells and other subtypes of RCC. These few TEAD1 foci co-occupy with the repressive histone mark H3K9me3 and are generally located near the nuclear periphery or nucleolus. This contrasts with TEAD1 patterns both in normal cells and within other regions of the same nuclei, where TEAD1 is distributed into many sub-resolution foci and associates with markers of active chromatin. As YAP1-TEAD complexes are important mediators of chromatin architecture and genome organization, these findings provide insight into how those mechanisms can be hijacked in cancer. We also investigate the roles of small-molecule YAP1-TEAD inhibitors as tools to manipulate chromatin landscapes. We have used a combination of high-resolution microscopy, ChIP-seq, and biochemical methods to identify the unique chromatin architecture of these cancer-associated TEAD1 foci and propose that they represent a novel subtype of biomolecular condensate where normally activating transcription factors are recruited into repressive heterochromatin
SHIPWORM ENDOSYMBIONT, TEREDINIBACTER TURNERAE, AS A PROMISING MODEL FOR LIGNOCELLULOSE DEGRADATION
Get PDFLignocellulose, the primary component of wood and plant biomass, is the most abundant renewable carbon source on earth. Deconstructing lignocellulose is an essential ecological process and holds significant potential for biotechnological applications in renewable energy and chemical production through biomass conversion. However, the complex structure of lignocellulose creates obstacles to bioconversion in natural and engineered systems. We used a combination of genomic, transcriptomic, and proteomic analyses to investigate strategies for lignocellulose degradation in Teredinibacter turnerae, a cultivable gammaproteobacterium that exists both as a free-living bacterium in the marine environment and as an intracellular endosymbiont in shipworms, wood-eating marine bivalves. From the pan-genome analysis of 11 strains, including two newly sequenced strains, we identified a large genetic repertoire containing diverse clade- and strain-specific carbohydrate-active enzymes (CAZymes). This suggests that populations may be specialized in their abilities and contributions to lignocellulose degradation. Transcriptional profiles via RNAseq of T. tunerae cells in culture and in host indicate a regulation of expression of CAZyme-encoding genes that includes both substrate-specific control and host-control. Importantly, we learned that CAZymes represent the highest expressed genes while in the host, indicating a complete transcriptional reprogramming towards lignocellulose degradation. Understanding this ability of control could be used in future systems-biology approaches for enzyme production. Additionally, we show that T. turnerae cells produce outer membrane vesicles (OMVs) when grown with cellulose as the sole carbon source. Proteomic analysis reveals OMVs contain a diverse set of CAZymes, and enzymatic assays further show that OMVs are capable of cellulose hydrolysis with specific activities comparable to or greater than that observed for an industrial standard enzyme mix. The packaging of enzymes within or bound to OMVs can have distinct advantages over free enzymes, including protection from degradation and simultaneous delivery of specific combinations, concentrations, and spatial organization of enzymes that could significantly enhance degradation efficiency. Our results support T. turnerae as a promising model that will improve the understanding of mechanisms of lignocellulose degradation and provide insight into the design of engineered lignocellulose deconstruction systems
BONE MORPHOGENETIC PROTEIN 2: A NEW PLAYER IN MELANOMA DORMANCY
Get PDFMelanoma, the most aggressive form of skin cancer, is heavily impacted by age, notably contributing to skin cancer-related deaths in the US. Despite being typically associated with aging, melanoma poses a significant risk to adolescents and young adults, emerging as the second most prevalent cancer in this demographic. The microenvironment plays a critical role, facilitating melanoma cell escape from primary tumors and subsequent growth in the lungs. While melanoma cells in young individuals' lungs remain dormant, they activate in aged lung tissue, fostering metastatic tumor outgrowth. This process is orchestrated by distinct fibroblast secretomes of the lung microenvironment., We have previously reported that BMP2 secreted by the aged melanoma tumor microenvironment (TME) triggers the development of a slow-cycling, invasive behavior in melanoma cells. This secretion of BMP2 by aged fibroblasts promoted intrinsic resistance and eventually by melanoma cells themselves increased acquired resistance to BRAFi/MEKi therapy, respectively. Furthermore, as melanoma cells acquire resistance to therapy, they increase their production of BMP2, which primes them for invasion and metastasis to distal sites such as the lung. At the distant sites, they can remain dormant for extended periods before initiating metastatic growth. Our study demonstrates that melanoma cells overexpressing BMP2 exhibit reduced metastatic growth in the lungs of mice compared to control cells, suggesting a potential role for BMP2 in promoting dormancy
Multifaceted Investigation into Cryptococcus neoformans Biomolecules
Get PDFCryptococcus neoformans is an environmental encapsulated yeast-like Basidiomycete and a major opportunistic fungal pathogen, ranked as the first on the WHO fungal priority pathogen list. As a major virulence factor, capsule, formed by polysaccharides (capsular-PS) deposited at the cell surface, helps the initiation of infection by protecting the fungal cells from the host immune defense mechanisms. Meanwhile, C. neoformans secretes additional polysaccharides to the extracellular environment (exo-PS), and exo-PS interferes with host immunity and exacerbates disease manifestation. Therefore, both capsular-PS and exo-PS are targets for vaccine design and antibody treatments. Such initiatives require a thorough understanding of fungal PS structures, yet recent studies have shown the difficulty of preserving PS natural structure. To better preserve PS structural integrity, we pioneered the use of neutron scattering techniques to study native capsular-PS and exo-PS structures. Our PS structure analysis of neutron scattering data collected at the National Institute of Standards and Technology provides solid experimental evidence supporting previous computational modeling studies and has important implications for other PS-related studies.
Melanin, a unique class of pigments, produced by C. neoformans is another important virulence factor since it protects fungal cells from multiple host’s immune responses and reduces the efficacy of antifungal drugs. To gain further insight into the melanin structure, which remains largely an enigma, we extended the neutron scattering technique to probe the structural information. Here we also present the first neutron scattering study on the extracted fungal melanin. Other than reporting the values of some crucial parameters and structure-based modeling attempts, we have provided instructions on how to conduct neutron scattering studies on those complicated particles.
The multifunctionality makes melanin promising for incorporation into numerous biotechnologies, that desire melanin production in large quantities. Therefore, we attempted to develop a method to isolate fungal cells containing more melanin so that melanization enhancement approaches might be found. We find the fluorescence intensity of the cell wall dye UviTex can be an excellent parameter to show melanization levels. Meanwhile, we have been focusing on the supplement optimization of N-acetylglucosamine, which has been identified as a promising ingredient in the culture media to increase melanization. This study has provided an implication for future advancement of melanin biosynthetic production
ASSOCIATION OF CORONARY ARTERY AND EXTRA-CORONARY CALCIFICATION WITH ANKLE BRACHIAL INDEX IN THE ATHEROSCLEROSIS RISK IN COMMUNITIES STUDY
Get PDFBackground: Traditionally, atherosclerosis is considered a systemic condition with similar risk factors across different vascular beds. However, contemporary insights suggest distinctive pathophysiological processes across different vascular beds. We aimed to explore potential differential associations of ankle brachial index (ABI), a marker of subclinical atherosclerosis, with calcification in different vascular beds and cardiac valves.
Methods: We studied 1,420 community-dwelling older adults (mean age 80.2 [SD 4.1] years, 60.2% female, and 16.6% Blacks) from the Atherosclerosis Risk in Communities (ARIC) Study. ABI was measured using an oscillometric device, and coronary artery calcification (CAC) and extra-coronary calcification (thoracic aorta, aortic valve, and mitral valve) were assessed through non-contrast cardiac-gated computed tomography. We ran multivariable logistic regression models, with any calcification (Agatston score >0) and high calcification (Agatston score ≥75th percentile) as outcome variables.
Results: Using any calcification as an outcome, the odds ratio (OR) for ABI ≤0.9 was largest for CAC (OR 7.05 [95%CI 1.03, 53.22]), followed by ascending aorta (OR 3.03 [95% CI; 1.80, 5.11]). The associations of low ABI were weakest for aortic valve and mitral valve/annular calcification. Using high calcification as an outcome, ABI ≤0.9 was significantly associated with all vascular and valvular calcification tested, but again its association was strongest for CAC. High ABI >1.3 tended to be more strongly associated with valvular calcification than vascular calcification regardless of any or high calcification as an outcome.
Conclusions: Likely reflecting their nature as measures of subclinical atherosclerosis, low ABI was most robustly associated with CAC. Its association was weaker for thoracic aorta calcification and weakest for valvular calcification. These findings further support distinct pathophysiology of calcification across vascular beds and cardiac valves
MEASURING MALADJUSTMENT: PRAGMATIST PSYCHIATRY, SYMPTOM SCALES, AND THE ECONOMIZATION OF MENTAL HEALTH, 1925-1975
Get PDFMeasuring Maladjustment explores the influence of Pragmatist philosophy on twentieth-century U.S. psychiatry, with a particular focus on the ergasiology of Adolf Meyer (1866-1950) and his colleagues. Drawing from historians of psychiatry like Susan Lamb, as well as an interdisciplinary literature on paper technologies and tools, it departs from conventional narratives that revolve around intellectual shifts within the field, in order to high-light the broader continuity and impact of Meyer’s practices. Each chapter profiles a series of research projects conducted by his trainees, with special emphasis on the careers of Thomas A.C. Rennie (1904-1956) and Alexander H. Leighton (1908-2007). By closely examining the forms and procedures used to conduct these studies, they collectively demonstrate how ergasiological concepts and methods were operationalized in various settings, and combined with those of other fields to analyze interactions between individuals and their environments.
This dissertation also explores the intersection of mental healthcare and governance, particularly during World War II, which catalyzed various collaborations between psychiatrists and the state. Such initiatives would eventually inspire the creation of symptom scales, standardized assessment tools which inadvertently perpetuated reductive metrics of psychological adjustment – often by emphasizing vocational capacity over broader sociocultural factors. By describing their development and dissemination throughout the postwar period, later chapters show how these instruments shaped not only academic research but also psychiatric services and welfare policy, contributing to what Michelle Murphy has called “the economization of life.” Ultimately, Measuring Maladjustment aims to inform re-cent efforts toward structural competency in medicine, and demonstrate the utility of historical methods in the critical evaluation of assessment tools
Isolation and Characterization of Enterovirus D68 Clinical Specimens From the 2022 Maryland Outbreak
Get PDFEnterovirus D-68 (EV-D68) is a respiratory pathogen that is most commonly associated with circulation in pediatric populations and has been linked to cases of acute flaccid myelitis (AFM). Since 2014, outbreaks have followed a biennial pattern, which was interrupted in 2020, potentially by COVID-19 respiratory precautions. While EV-D68 respiratory cases spiked in 2022, cases of AFM did not increase proportionally, in contrast to other recent outbreaks.
The Johns Hopkins Health System (JHHS) observed an increase in Influenza-like illness in September 2022 that correlated with an increase in general enterovirus/rhinovirus positivity, with EV-D68 comprising approximately one-third of the positive samples. Whole-genome sequencing of the clinical specimens showed they were highly similar to 2018 circulating strains.
We attempted to isolate and characterize EV-D68 from specimens collected during 2022 at the JHHS. Thirty-five RTPCR confirmed clinical samples were received, and virus isolation was attempted on rhabdomyosarcoma (RD) cells at 33 and 37°C. No infectious virus was detected by TCID50 or plaque assay. Isolation and quantification methods, including TCID50, plaque assay, and RTqPCR, were optimized using 2014 and 2018 outbreak isolates. Eighteen clinical samples were isolated in human nasal epithelial cell (hNEC) primary culture at 33°C with a 2018 outbreak isolate, MD/18-23209, used as a positive control. Five isolates have been characterized by RTqPCR and plaque assay. All isolates were negative for infectious virus by plaque assay on RD cells, but three were positive by RTqPCR. The positive control MD/18-23209 showed consistent viral RNA and infectious virus titers for early time points before infectious viral titers decreased but viral RNA remained high. An infectious clone model for a 2022 outbreak isolate also shows an inability to be tittered by plaque assay but has high RNA titers. These results suggest that the 2022 isolates may have lost the ability to replicate on RD cells but retain replication capacity in a primary respiratory epithelial cell culture model. Further research is needed to determine if this change in cell tropism could be linked to the lack of AFM cases observed during this outbreak
MULTI-OMIC AND INTERDISCIPLINARY ASSESSMENT OF THE ROLE OF THE AXON GUIDANCE GENE FAMILY IN PANCREATIC CANCER IMMUNE INFILTRATION
No full textBackground: Pancreatic ductal adenocarcinoma (PDAC) is characterized by a dismal prognosis and high metastatic potential. Previous research has highlighted the importance of axon guidance pathway genes within the SEMA-PLXN pathway (SEMA3A, SEMA3B, SEMA3C, SEMA3D, SEMA3E, SEMA3F, SEMA3G, NRP1, NRP2, PLXNA1, PLXNA2, PLXNB1, PLXNB2, PLXNC1, and PLXND1) in PDAC, as well as the connection between neuronal development and the tumor microenvironment (TME). However, the relationship between these 16 genes and tumor immune cell infiltration has not been explored. Additionally, a recent study identified 20 new neuronal development genes (DLG2, NRCAM, NRXN3, MAPK10, PDGFD, PRKCE, KCNMA1, PKHD1, NCAM1, NRG1, ZNF667, CFTR, ACSM3, C6, PTPRM, HIF1A, ADCY5, AJAP1, NBEA, and SCN9A) associated with perineural invasion and poor prognosis in PDAC. The association between genetic alterations in these 20 genes and the TME has not been investigated previously.
Methods: To explore this relationship, we utilized quantitative sequential multiplex immunohistochemistry (IHC), whole genome sequencing, and whole exome sequencing results from biopsy specimens obtained from 63 patients with PDAC.
Results: Our findings indicate that except for PLXNA1, PLXND1, PTPRM, and NBEA, genetic alterations involving these 36 genes are linked to significant changes in the densities of major immune cell subtypes. Furthermore, except for SEMA3A, PLXNA2, and AJAP1, copy number losses involving these two panels of neuronal development genes are significantly associated with alterations in immune cell infiltrates. Conversely, copy number gains in a much fewer subset of genes, including SEMA3A, SEMA3C, SEMA3D, SEMA3E, NRP2, PLXNA2, NRXN3, ZNF667, ACSM3, C6, ADCY5, SCN9A, and PRKCE, are significantly correlated with changes in immune cell infiltrates.
Conclusions: Our study suggests that neuronal development genes play a role in modulating immune composition in the TME of PDA