Upstate Medical University
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Unveiling cell-type-specific transcriptome and genetic regulation in postmortem brains of schizophrenia patientsSchizophrenia is a complex psychiatric disorder with a poorly understood etiology. This dissertation addresses three critical questions in schizophrenia research: identifying involved cell types, characterizing their transcriptomic changes, and elucidating how these changes mediate genetic risk. After rigorous evaluation, we conducted a comprehensive analysis of cell-type-specific gene expression in postmortem brains of schizophrenia patients and controls with single-cell RNA sequencing and cell deconvolution methods. Our findings provide compelling evidence for the involvement of upper-layer neurons and multiple non-neuronal cell types in schizophrenia. We observed significant alterations in synaptic function, neurodevelopment, immune response, and vascular transport within their respective cell types. Notably, we demonstrate that genetic risk for schizophrenia is predominantly enriched in neurons, particularly upper-layer neurons, with partial enrichment in oligodendrocyte precursor cells and vascular cells. This cell-type-specific approach offers novel insights into the molecular underpinnings of schizophrenia, potentially bridging the gap between genetic risk factors and clinical manifestations. By highlighting key genes and pathways, our study establishes a robust foundation for future research and opens avenues for innovative preventive and therapeutic approaches.
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Sleep disturbances and racial-ethnic disparities in 10-year dementia risk among a national sample of older adults in the USARace/ethnicity and sleep disturbances are associated with dementia risk.
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Mucosal Innate Immunity of Human Surfactant Protein A Genetic Variants against SARS-CoV-2 InfectionMore than 7 million people have died of the coronavirus disease-2019 (COVID-19) since first reported in December of 2019. Infection in some patients manifests as life-threatening ALI/ARDS, multi-organ dysfunctions, and/or death characterized by active viral replication and profound inflammatory cell influx into tissues/organs. SARS coronavirus-2 (SARS-CoV-2) infects human angiotensin-converting enzyme 2 (hACE2)-expressing cells through its spike protein (S protein). The S protein is highly glycosylated and could be a target for lectins. Surfactant protein A (SP-A) is a collectin, expressed by lung alveolar type II cells and other mucosal epithelial cells; it plays a crucial role in innate immunity and inflammatory regulation. SP-A modulates pathogenic infection and disease severity by binding to microbial and host glycoproteins to alter infectivity and regulate host inflammation. The human SP-A gene is located on chromosome 10q22-23, which contains two functional genes SP-A1 and SP-A2 (gene names: SFTPA1 and SFTPA2), and a pseudogene. SP-A1 and SP-A2 are highly polymorphic and consist of several genetic variants, such as SP-A1 (variants 6A2, 6A4) and SP-A2 (variants 1A0, 1A3). It has been demonstrated that these variants have differential antiviral and immunoregulatory capacities in response to various viral infections. The goal of this study was to investigate the mechanistic role of human SP-A variants in response to SARS-CoV-2 infection and COVID-19 susceptibility and severity. The results from this study showed that native human SP-A can bind SARS-CoV-2 S protein, receptor-binding domain (RBD), and hACE2 in a dose-dependent manner. A decrease in S protein and RBD binding was observed in the presence of EDTA and sugars, indicating that the SP-A carbohydrate-recognition domain (CRD) mediates S protein binding in a calcium-dependent manner. We further showed that human SP-A can attenuate viral infectivity in susceptible host cells, evidenced by the dose-dependent reduction in viral load in infected cells. These results suggest that human SP-A can bind SARS-CoV-2 S protein, RBD, and hACE2 to attenuate SARS-CoV-2 infectivity in susceptible host cells. Next, we examined the variations in antiviral and immunoregulatory roles of human SP-A variants in response to SARS-CoV-2 infection. The binding studies showed that in vitro-expressed SP-A variants differentially interact with S protein. Moreover, cells inoculated with SARS-CoV-2 pretreated with the 1A0 variant had a more reduced virus titer than those pretreated with the 6A2 variant, indicative of their differential antiviral capacities. These findings from in vitro studies demonstrated that human SP-A and their genetic variants directly interact with viral S protein to differentially modulate SARS-CoV-2 infectivity. To perform in vivo study, six genetically modified double-hTG mouse lines, expressing both hACE2 and the respective SP-A variants: (hACE2/6A2 (6A2), hACE2/6A4 (6A4), hACE2/1A0 (1A0), and hACE2/1A3 (1A3), one SP-A knockout (hACE2/SP-A KO (KO) and one hACE2/mouse SP-A (K18) mice, were generated and challenged intranasally with 103 PFU SARS-CoV-2 (Delta) or saline (Sham). We observed that these infected mice had differential COVID-19 severity. Infected KO and 1A0 mice had more mortality and lung injury compared to other mouse lines, and disease severity correlated with enhanced upregulations of inflammatory genes that play vital roles in host immunity such as MyD88 and Stat3 in the lungs of KO and 1A0 mice. Furthermore, pathway analysis identified several important signaling pathways involved in lung defense, including pathogen-induced cytokine storm, NOD1/2, toll-like receptor, neuroinflammation, and Trem1 signaling pathways. Consistent with the transcriptomic data, expressions of inflammatory mediators such as G-CSF, IL-6, and IL-1β were comparatively higher in the lungs and sera of KO and 1A0 mice with the highest mortality rate. We further examined other organ injuries (kidney, intestine, and brain) in the infected mice; we found a more severe acute kidney injury (AKI) and intestinal damage in KO and 6A4 mice compared to other double-hTG mice. Viral titers were generally lower in the kidneys and brains of infected double-hTG mice relative to KO mice. Inflammatory mediators like TNF-α, IL-6, IL-1β, and MCP-1 were comparatively higher in KO and 6A4 mice with the most severe AKI. High virus presence and inflammatory markers were also observed in the brain and hippocampus of all infected mice. The results from in vivo studies suggest that SP-A variants differentially protect against severe COVID-19. Furthermore, the human COVID-19 patient studies revealed increased SP-A levels in the saliva of COVID-19 patients compared to healthy controls and highlighted the potential use of SP-A levels as a biomarker for COVID-19 severity. Collectively, these findings underscore the importance of host innate immune collectins and contribute to our understanding of the roles of host genetic variations in the observed population-level differences in COVID-19 susceptibility and severity.
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Deciphering cellular dynamics and crosstalk of trabecular meshwork and Schlemm's canal cells in a bioengineered 3D extracellular matrix hydrogel microenvironmentIn the conventional outflow pathway, Schlemm's canal (SC) inner wall endothelium interfaces with the trabecular meshwork (TM). Biomechanical changes in this microenvironment contribute to increased resistance to aqueous outflow, a characteristic of ocular hypertensive glaucoma. Notably, TM undergoes fibrotic-like remodeling and stiffening. Existing in vitro TM/SC models fail to accurately replicate native cell-cell and cell-extracellular matrix (ECM) interactions, limiting their use for studying glaucomatous outflow pathobiology. In this dissertation, we utilized a biomimetic ECM hydrogel system made from natural polymers resembling native tissue proteins. This ECM hydrogel can be (i) used to encapsulate donor-derived primary human TM cells or (ii) employed as a substrate for culturing donor-derived primary human SC cells on top. As ECM hydrogels gradually emerge as a preferred model in diverse research laboratories, a standardized fabrication method is essential to improve accessibility and consistency across experimental protocols. Thus, a detailed methodology for producing these ECM hydrogels is provided in Chapter 2. In Chapter 3, using the 3D TM hydrogel system, we demonstrated that simvastatin-mediated inactivation of Yes-associated protein (YAP) and transcriptional coactivator with PDZ binding motif (TAZ) attenuates pathological changes in TM cells. YAP/TAZ are key mechanotransducers involved in glaucoma pathogenesis and are shown to be regulated by the mevalonate pathway. By inhibiting this pathway, we hypothesized that statins could potentially improve TM cell pathobiology by modulating YAP/TAZ activity. Thus, targeting the mevalonate pathway with statins may offer therapeutic potential for glaucoma. Despite significant progress in understanding TM and SC cells individually, the dynamic interactions between them and their role in glaucoma pathogenesis remain poorly understood. These interactions are crucial in the pathogenesis of glaucoma, yet no effective model exists to study them. Therefore, in Chapter 4, we developed a novel co-culture hydrogel system to explore TM-SC interactions and assess how glaucomatous TM cells affect SC behavior. Our findings show that glaucomatous TM cells alone can induce pathological changes in SC cells, underscoring the critical role of cell-cell and cell-ECM interactions in glaucoma progression. Collectively, these biomimetic ECM hydrogels provide a unique platform for investigating glaucomatous outflow mechanisms and offering insights into disease pathogenesis.
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HIV-1 has a sweet tooth: glucose metabolism drives the multistep process of HIV-1 latency reversalThe major barrier to a cure for HIV-1 is the establishment of latency in long-lived CD4+ T cells within lymphoid tissues which readily fuel viral rebound upon antiretroviral therapy (ART) interruption. Therapeutic approaches aimed at eliminating these HIV reservoirs with latency reversal agents (LRAs) have hitherto yielded underwhelming results in clinical trials owing to our incomplete understanding of the exact determinants of meaningful latency reversal in vivo. While previous studies have associated glycolysis with HIV productive replication and latency reversal, the exact role and mechanistic link of glycolysis to HIV latency reversal remains undefined. Furthermore, few studies have investigated HIV latency under physiologically relevant metabolic conditions found in the anatomical reservoirs of HIV in vivo. The studies in this thesis reveal that glycolysis is a metabolic determinant of HIV latency reversal, particularly during physiological hypoxia. We show that the capacity of LRAs to modulate glycolysis determines their efficacies over a physiological range of glucose and oxygen availabilities as found across tissues in vivo. Mechanistically, glycolysis fuels histone lactylation, a novel post-translational modification (PTM) which we show is a stronger predictor xviii of latency reversal than the canonically recognized acetylation marks, and promotes chromatin accessibility at the HIV LTR. Beyond histone PTM modulation, glycolysis also modulates HIV RNA splicing, a critical post-transcriptional step in HIV latency reversal. Specifically, multiple splicing of rev, an HIV regulatory gene, is significantly downmodulated by glycolytic restriction in a hypoxia-dependent fashion. Finally, we show that glucose and oxygen availability impact the phosphorylation and lactylation of splicing factor 3B subunit 1 (SF3B1), a core component of the U2 spliceosome complex and HIV dependency factor which provides preliminary mechanistic insight to how glycolysis and hypoxia modulate HIV RNA splicing. Collectively, our findings uncover glucose and oxygen availability as critical metabolic determinants of HIV-1 latency reversal and support the rationale that physiologically relevant experimental conditions should be utilized in studies aimed at identifying therapeutic agents that effectively target the latent reservoir in vivo.
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Racial and ethnic disparities in social isolation and 11-year dementia risk among older adults in the United States.Social isolation has been implicated in the development of cognitive impairment, but research on this association remains limited among racial-ethnic minoritized populations. Our study examined the interplay between social isolation, race-ethnicity and dementia.
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Kohlschütter-Tönz protein ROGDI is the homolog of yeast Rav2 and a novel Rabconnectin-3 subunitV-ATPases are rotary proton pumps that are extraordinarily well-conserved among eukaryotes. V-ATPases function primarily to acidify intracellular compartments, critical to maintaining cellular homeostasis. The V-ATPase-generated proton gradient provides the optimal environment for lysosomal catabolism and drives intracellular protein trafficking. V-ATPases serve important functions throughout the human body. For example, V-ATPase activity energizes the active transport of neurotransmitters into synaptic vesicles, regulates the acid/base balance in the kidney, and helps the immune system recognize invading pathogens. However, when V-ATPase activity is inappropriately increased or decreased, these processes are affected, and disease can result. V-ATPases are composed of peripheral V₁ and integral membrane V₀ subcomplexes; V₁ hydrolyzes ATP and transmits rotation to V₀, which moves protons across a membrane. V-ATPase activity is regulated in part through the reversible association of the V₁ subcomplex and V₁C subunit from V₀. Upon disassembly, both V₁ and V₀ are catalytically inactivated. In yeast, the RAVE complex catalyzes the efficient reassembly of V-ATPases. Rabconnectin-3 is the human homolog of the RAVE complex and functions similarly. Mutations in the Rabconnectin-3 complex can reduce V-ATPase activity through decreased assembly, which leads to disease. Both Rabconnectin-3 subunits share substantial homology with the RAVE subunit Rav1. We have identified the poorly characterized protein ROGDI as the mammalian homolog of the yeast RAVE subunit, Rav2. ROGDI shares strong functional and structural homology with yeast Rav2. Expression of ROGDI in a rav2Δ yeast strain partially rescues the growth phenotype characteristic of RAVE mutants. ROGDI binds to the structurally conserved N-terminal β-sheet rich domain. AlphaFold3 modeling predicts that ROGDI binds between the Rabconnectin-3 subunits. ROGDI coimmunoprecipitates with Rabconnectin-3 and V-ATPase subunits. Additionally, ROGDI is present alongside V-ATPase and Rabconnectin-3 subunits on lysosomal membranes. This indicates that, like RAVE and Rav2, Rabconnectin-3 and ROGDI localize intracellular regions rich in V-ATPases. Identifying ROGDI as a novel Rabconnectin-3 subunit is a substantial step forward in our understanding of Rabconnectin-3 and how it influences V-ATPase activity.
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Research on Health Topics Communicated through TikTok: A Systematic Review of the LiteratureTikTok has more than 1.5 billion users globally. Health and wellness content on the application increased by more than 600% in 2021. This systematic review seeks to summarize which fields within medicine have embraced researching health communication on the TikTok platform and the most common measures reported within this literature. Research questions include what categories of health topics on TikTok are investigated in the literature, trends in topics by year, and types of outcomes reported. Embase, CINAHL, Scopus, and Ovid MEDLINE databases were searched in March 2024. Eligible studies met four criteria: (1) investigated human health topics on TikTok; (2) conducted in the United States; (3) published in English; and (4) published in a peer-reviewed journal. Of the 101 included studies, 50.5% (N = 51) discussed non-surgical specialties, 9.9% (N = 10) discussed topics within surgery, and 11.9% (N = 12) discussed COVID-19. The number of papers referencing non-surgical topics spiked in 2023, and no increase was seen in the number of COVID-19 papers over time. Most papers reported a number of interactions, and papers about mental health were least likely to report accuracy. Our findings highlight several health topics with a wide breadth of research dedicated to them, such as dermatology and COVID-19, and highlight areas for future research, such as the intersection of cancer and TikTok. Findings may be influential in the fields of medicine and healthcare research by informing health policy and targeted prevention efforts. This review reveals the need for future policies that focus on the role and expectations of the healthcare worker in health communication on social media. Implications for clinical practice include the need for providers to consider an individual's perception of health and illness, given the wide variety of information available on social media applications such as TikTok. This review was pre-registered on PROSPERO (CRD42024529182).
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Role of Cytoreductive Nephrectomy in Metastatic Clear Cell Renal cell Carcinoma in the Era of immunotherapy: An Analysis of the National Cancer DatabaseBackground: The effectiveness of the clinical outcome of CN (Cytoreductive Nephrectomy) in cases of mccRCC (Metastatic Clear Cell Renal cell Carcinoma) is still uncertain despite two trials, SURTIME and CARMENA. These trials, conducted with Sunitinib as the standard treatment, did not provide evidence supporting the use of CN. Methods: We queried the NCDB for stage IV mccRCC patients between the years of 2004 to 2020, who received (immunotherapy) IO with or without nephrectomy. Overall survival (OS) was calculated among three groups of IO alone, IO followed by CN (IOCN), CN followed by IO (CNIO). Cox models compared OS by treatment group after adjusting for sociodemographic, health, and facility variables. Results: From 1,549,101 renal cancer cases, 7983 clear and nonclear cell renal cell carcinoma cases were identified. After adjusting for sociodemographic and health covariates, patients who received IO followed by CN or CN followed by IO had a respective 64% (adjusted Hazard Ratio [aHR] = 0.36, 95% CI = 0.30-0.43, P = .006] and 47% (aHR = 0.53, 95% CI = 0.49-0.56, P = .001) mortality risk reduction respectively compared to patients who received IO alone. Compared to White adults, individuals who identified as Black exhibited 17% higher risk mortality (aHR = 1.17, 95% CI = 1.06-1.30, P = .002). Patients who received CN prior to IO had a 59% associated mortality risk compared to patients who received IO followed by CN who had a lower risk, 35.7% (P < .001). Conclusions: Patients receiving CN regardless of sequence with IO did better than IO alone in this national registry-based adjusted analysis for mccRCC. Presently available data indicates that the combination of CN and IO holds promise for enhancing clinical results in patients with mRCC.
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Imbalanced specialty representation of USMLE and NBME test writersPurpose: The United States Medical Licensing Examination (USMLE) is an examination series required for allopathic physician licensure in the United States (US). USMLE content is created and maintained by the National Board of Medical Examinations (NBME). The specialty composition of the USMLE and NBME taskforce members involved in the creation of examination content is currently unknown. Methods: Using the 2021 USMLE and 2021 NBME Committees and Task Forces documents, we determined each member's board-certified primary specialty and involvement in test material development committees who we dubbed "test writers". Total active physicians by primary specialty were recorded from the 2020 Physician Specialty Data Report published by the Association of American Medical Colleges (AAMC). Descriptive statistics and chi-square analysis were used to analyze the cohorts. Results: The USMLE and NBME test writer primary specialty composition was found to be significantly different compared to the US active physician population (USMLE χ2 [32]=172, p<.001 and NBME χ2 [32]=200, p<.001). Only nineteen specialties were represented within USMLE test writers, with three specialties being proportionally represented. Two specialties were represented within NBME test writers. Obstetrics and Gynecology physicians were proportionally represented in USMLE but not within NBME test writers. Internal Medicine (IM) accounts for the largest percentage of all USMLE test writers (60/197, 30%) with an excess representation of 31 individuals. Conclusions: There is an imbalance in the specialty representation of USMLE and NBME test writers compared to the US active physician population. These findings may have implications for the unbiased and accurate portrayal of topics in such national examinations; thus, future investigation is warranted.
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Human cytomegalovirus (HCMV) exploits heat-shock transcription factor 1 (HSF1) to promote viral replication: a potential novel antiviral target to combat HCMV infectionHuman cytomegalovirus (HCMV) is a highly prevalent beta-Herpesviridae virus infecting almost 80-90 % of the world population. Though HCMV infection is typically asymptomatic, it can cause significant morbidity and mortality among immunocompromised individuals. Because of its obligate intracellular nature, HCMV modulates the cellular environment to promote infection. HCMV activates different cellular responses and signaling pathways to facilitate a favorable state for viral replication. During the lytic cycle of HCMV infection, viral entry, and replication inside the cell initiate stress response due to nutrient deficiency, energy depletion, hypoxia, and proteotoxic stress. Stress responses are designed to sense the damage, initiating a cascade of events to survive the stress. Several studies showed that HCMV usurps components of heat shock-stress response (HSR) to mitigate stress-associated damage and promote viral gene expression and replication. In this study, we found that HCMV infection in fibroblast cells induces a unique biphasic activation of heat shock transcription factor 1 (HSF1), a master transcription factor that is activated in response to heat-induced proteotoxic stress. HCMV binding to the integrin-ᵝ receptor activates HSF1 through Src- kinases. Importantly, HCMV infection drives the translocation of HSF1 into the infected cell nucleus. During canonical activation of HSF1, nuclear HSF1 binds to the specific sequence on the genome called heat shock element (HSE) and initiates transcription of a wide variety of stress-related genes. Interestingly, HCMV also utilizes this master transcription factor by harboring HSEs on major immediate early promoter (MIEP) to regulate viral immediate early (IE) gene expression. We found inhibition of HSF1 with a novel anti-HSF1 targeting drug SISU102 (Direct Targeted HSF1 InhiBitor) attenuated IE protein expression, indicating that the HSF1 regulates HCMV lytic replication. Additionally, inhibition of HSF1 reduced late (L) gene expression and subsequent viral progeny production. To explore HSF1 as a potential in vivo anti-HCMV target, we employed a murine model involving the subcutaneous transplantation of human skin into athymic nude mice. Treatment with SISU102 significantly diminishes viral replication in skin xenografts compared to the vehicle-treated group, indicating HSF1 as a possible cellular protein target for HCMV antiviral therapy. Overall, our data suggest that HCMV infection rapidly activates HSF1 during viral binding and entry, driving nuclear localization to promote lytic replication, which can be exploited as an antiviral strategy.
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Effectiveness of Artificial Intelligence Technologies in Cancer Treatment for Older Adults: A Systematic ReviewBackground: Aging is a multifaceted process that may lead to an increased risk of developing cancer. Artificial intelligence (AI) applications in clinical cancer research may optimize cancer treatments, improve patient care, and minimize risks, prompting AI to receive high levels of attention in clinical medicine. This systematic review aims to synthesize current articles about the effectiveness of artificial intelligence in cancer treatments for older adults. Methods: We conducted a systematic review by searching CINAHL, PsycINFO, and MEDLINE via EBSCO. We also conducted forward and backward hand searching for a comprehensive search. Eligible studies included a study population of older adults (60 and older) with cancer, used AI technology to treat cancer, and were published in a peer-reviewed journal in English. This study was registered on PROSPERO (CRD42024529270). Results: This systematic review identified seven articles focusing on lung, breast, and gastrointestinal cancers. They were predominantly conducted in the USA (42.9%), with others from India, China, and Germany. The measures of overall and progression-free survival, local control, and treatment plan concordance suggested that AI interventions were equally or less effective than standard care in treating older adult cancer patients. Conclusions: Despite promising initial findings, the utility of AI technologies in cancer treatment for older adults remains in its early stages, as further developments are necessary to enhance accuracy, consistency, and reliability for broader clinical use.
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Microbiota Colonization Dynamics Dictate Systemic IgAEvolution of the mammalian gut is intimately linked with the microbes that inhabit this space. Immunological development of gastrointestinal and systemic tissues is fundamentally dependent on stimulation by symbiotic microorganisms. In some cases, the same species that are critical for host immunity display pathogenic qualities when homeostasis is disrupted. Bacteroides fragilis is one such species with numerous symbiotic and pathogenic characteristics. This thesis explores the generation of B. fragilis-specific systemic IgA and the role of this response in protecting the host from B. fragilis pathogenicity. Induction of systemic IgA specific to B. fragilis requires exposure of this bacterium to small intestinal Peyer's patches and results in migration of newly generated IgA plasma cells to systemic tissues. Colonization dynamics of B. fragilis in mouse models with endogenous gut microbiota revealed that the magnitude of systemic IgA responses occurs in a dose-dependent fashion. Finally, a framework for establishing B. fragilis colonization and subsequent immune modulation within a highly diverse intestinal ecosystem was developed.
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Investigating the role of formin FHOD3 during myofibrillogenesis in embryonic chick cardiomyocytesFormins are major actin polymerizing proteins which act via the FH2 domain to promote actin nucleation and polymerization, as well as the FH1 domain to accelerate FH2 mediated actin elongation. FHOD3 is a formin that has been shown to be expressed predominantly in the heart and is critical for myofibril maturation during development in mice. FHOD3 has been shown to localize where actin filaments overlap myosin filaments within the sarcomeres of mice, rat, and human induced pluripotent stem-cell derived cardiomyocytes, flanking both sides of the M-line in the sarcomere. However, the role of FHOD3 in the myofibrillogenesis and the timing of FHOD3's activity in myofibrils has yet to be determined. Using RT-PCR, I successfully identified expression of at least two different isoforms of FHOD3 within heart tissue, matching to predicted isoforms X5 and X6. I also identified two chemically conserved regions within the FHOD3 amino acid sequence that are related to the cardiac FHOD3 isoform's localization to myofibrils. Using immunofluorescence microscopy and western blotting I found that FHOD3 is present within embryonic chick cardiomyocytes and that the localization of FHOD3 matches prior reports. FHOD3 was determined to be transiently expressed at significantly higher rates on Days 3 and 4 of culture in cardiomyocyte myofibrils. 90% of measured sarcomeres containing FHOD3 had a Z-line to Z-line length ranging from 1.4-1.9 µm, suggesting not only a length-dependent role of FHOD3, but a myofibril maturity dependent localization of FHOD3. These observations illustrate that FHOD3 likely does not have a function in the initiation of myofibrillogenesis but may instead have a role in the maturation and elongation of sarcomeres. The transient nature observed also suggests that FHOD3 may be localized within the sarcomere only as needed. Knockdowns of FHOD3 performed with shRNAs showed no indication of knockdown causing myofibrillar disruption. Knockdowns of FHOD3 using DsiRNAs were statistically inconclusive for knockdown occurring but did have an upwards nonsignificant trend in the percentage of myofibril disruption in cardiomyocytes.
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HSV-1 targets a novel antiviral response of the STING pathwayIn order to establish a successful infection, herpes simplex virus-1 (HSV-1), a ubiquitous virus with high seropositivity in the human population, must undermine a multitude of host innate and intrinsic immune defense mechanisms, including key players of the stimulator of interferon genes (STING) pathway. Recently it was discovered that not only de novo produced intracellular 2'-3'cGAMP, but also extracellular 2'-3'cGAMP can activate the STING pathway by being transported across the cell membrane via the folate transporter, SLC19A1, the first identified extracellular antiporter of this critical signaling molecule in cancer cells. We hypothesized that the import of exogenous 2'-3'cGAMP would function to establish an antiviral state similar to that seen with the paracrine antiviral activities of interferon. Further, to establish a successful infection, viruses, such as HSV-1, must undermine this induction of the STING pathway by inhibiting the biological functions of SLC19A1. Herein, we report that treatment of the monocytic cell line, THP-1 cells and SH-SY5Y neuronal cell line with exogenous 2'-3'cGAMP induces interferon production and establishes an antiviral state. Using either pharmaceutical inhibition or genetic knockout of SLC19A1 blocks the 2'-3'cGAMP-induced inhibition of viral replication. Additionally, HSV-1 infection results in the reduction of SLC19A1 transcription, translation, and importantly, the rapid removal of SLC19A1 from the cell surface of infected cells. Our data indicate SLC19A1 functions as a newly identified antiviral mediator for extracellular 2'-3'cGAMP which is undermined by HSV-1 protein ICP27. This work presents novel and important findings about how HSV-1 manipulates the host's immune environment for viral replication and discovers details about an antiviral mechanism which information could aid in the development of better antiviral drugs in the future.
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Structural insights of the histone H3 tail and its role in the mechanism of histone H3 lysine-4 methylationStructural insights of the histone H3 tail and its role in the mechanism of H3 lysine-4 methylation Gene expression relies on the proper chromatin structure to provide the necessary access to the DNA for the large transcription complexes to carry out their tasks. If the chromatin is tightly condensed, transcription is unable to occur. To regulate and initiate access to the DNA, an elaborate network of histone modifying enzymes, chromatin remodeling complexes, and other supporting proteins must coordinate the writing, reading, and erasing of histone post-translational modifications (PTMs). One such PTM, methylation of histone H3 on the lysine-4 (H3K4) residue, is critically important for maintenance of gene expression states. This is done in a spatiotemporal manner, which is influenced by the number of methyl groups that are present. However, an understanding of how the degree of H3K4 methylation is regulated remains elusive. In this dissertation, we demonstrate the remarkable conservation of length and composition in the flexible N-terminal tails of histone proteins across evolution. Recent structural studies indicate several methyltransferase complexes bind to the nucleosome core, often leaving the N-terminal tails unbound. Research from our lab has also demonstrated that non-processive buildup of lysine-4 methyl groups takes place at multiple active sites. Based on these observations, we propose a hypothesis whereby the histone H3 tail acts as a swinging arm substrate, delivering residue side chains to different active sites to facilitate the progressive establishment of these epigenetic states. To investigate this hypothesis, we employed the CRISPR/Cas9 system in Saccharomyces cerevisiae to systematically modify the length of the H3 tail. We monitored histone H3 lysine 4 (H3K4) methylation, mediated by SET1, the primary H3K4 methyltransferase in budding yeast. Our findings demonstrate that altering the length of the H3 tail has varying effects on the extent of H3K4 methylation, in accordance with the swinging arm model. We also demonstrate that three proline residues are responsible for providing a segmented, tripartite structure with hinge-like joints that likely influence the tail's range of motion. Furthermore, the results support the proposed multiple active-site model, where mono-, di-, and trimethylation occur at distinct active sites within the COMPASS or MLL Core Complexes.
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Healthcare staff perceptions of an electronic hand hygiene monitoring system within a large university systemObjective: The acceptability of an electronic HH monitoring system (EHHMS) was evaluated among hospital staff members. Design: An electronic HH monitoring system was implemented in June 2020 at a large, academic medical center. An interdisciplinary team developed a cross-sectional survey to gather staff perceptions of the EHHMS. Setting: The survey was conducted at a public, tertiary acute care hospital. Participants: The survey included current employees and staff. 1,273 participants responded. The mean age was 44.9 years (SD = 13.5). Most of the samples were female (71%) and non-Hispanic white (83%). Methods: A survey was conducted between June and July 2021. Responses were analyzed using Stata statistical software. Multiple logistic regression models were constructed to examine factors associated with negative perceptions of the EHHMS and its radiofrequency identification (RFID) badge. Supporting qualitative analyses were performed using Atlas.ti version 9. Results: Three-quarters (75%) of respondents reported neutral to negative perceptions of the EHHMS and its associated badge. Respondents reported limited influence on HH practices. Age, campus location, length of employment, job role, and opinion on data sharing were associated with negative perceptions of the EHHMS and RFID badge. Position in a direct patient care role was associated with negative perceptions of the RFID badge. Conclusions: Perceptions of the EHHMS aligned with previous research. Identified associations provide opportunities for targeted education, outreach, and intervention to increase acceptability and uptake. Lack of acceptance is explained by poorly perceived ease of use and usefulness, as well as challenges in implementation.
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Pulse Pressure as a Hemodynamic Parameter in Preeclampsia with Severe Features Accompanied by Fetal Growth RestrictionBackground: Modern management of preeclampsia can be optimized by tailoring the targeted treatment of hypertension to an individual's hemodynamic profile. Growing evidence suggests different phenotypes of preeclampsia, including those with a hyperdynamic profile and those complicated by uteroplacental insufficiency. Fetal growth restriction (FGR) is believed to be a result of uteroplacental insufficiency. There is a paucity of research examining the characteristics of patients with severe preeclampsia who do and who do not develop FGR.We aimed to elucidate which hemodynamic parameters differed between these two groups. Methods: All patients admitted to a single referral center with severe preeclampsia were identified. Patients were included if they had a live birth at 23 weeks of gestation or higher. Multiple gestations and pregnancies complicated by fetal congenital anomalies and/or HELLP syndrome were excluded. FGR was defined as a sonographic estimation of fetal weight (EFW) < 10th percentile or abdominal circumference (AC) < 10th percentile. Results: There were 76% significantly lower odds of overall pulse pressure upon admission for those with severe preeclampsia comorbid with FGR (aOR = 0.24, 95% CI = 0.07-0.83). Advanced gestational age on admission was associated with lower odds of severely abnormal labs and severely elevated diastolic blood pressure in preeclampsia also complicated by FGR. Conclusions: Subtypes of preeclampsia with and without FGR may be hemodynamically evaluated by assessing pulse pressure on admission.
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Correlates of Loneliness and Social Isolation among Older Adults during the COVID-19 Outbreak: A Comprehensive Assessment from a National United States SampleThis study examined the correlates of loneliness and social isolation among older adults in the United States (U.S.) during the COVID-19 outbreak. We analyzed data from the 2020 National Health and Aging Trends Study, a nationally representative sample of 3257 U.S. older adults aged 65 years and older. We analyzed and identified the sociodemographic, health, social support, and community correlates of loneliness, higher loneliness during versus before the COVID-19 outbreak, and social isolation using weighted multiple logistic regression models. About 35.2% of U.S. older adults reported loneliness during the COVID-19 outbreak, 21.9% reported higher loneliness compared to before the COVID-19 outbreak, and 32.8% were socially isolated during the outbreak. Correlates for increased odds of loneliness included female gender, higher education, physical activity, depression, anxiety, functional limitations, and virtual communication access (only for higher loneliness during COVID-19 outbreak). Correlates for increased odds of social isolation included higher age, non- Hispanic Black, Hispanic, higher number of household children, and metropolitan residence. Our findings provide insights into evidence-based approaches to address social disconnection among U.S. older adults. The wide range of sociodemographic, health, social support, and community correlates identified in this study warrants multifaceted interventions that traverse individual, community, and societal levels to address the loneliness and social isolation epidemic.
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Role of TLDc Proteins Oxr1 and Rtc5 in Yeast V-ATPase Reversible DisassemblyThe vacuolar H+-ATPase (V-ATPase; V1Vo-ATPase) is a highly conserved, ATP hydrolysis-driven dedicated proton pump found on the membranes of intracellular organelles in virtually all eukaryotic cells and on the plasma membrane of specialized cell types. Regulation of V-ATPase activity is key to maintaining normal physiological functions, as aberrations in its activity are associated with several pathophysiological conditions. V-ATPase activity is mainly regulated by a mechanism called reversible disassembly, in which the assembly state - and hence the activity - of the enzyme is controlled by nutrient availability and extracellular cues. During the process, V-ATPase activity becomes either turned off by dissociation of the V1-ATPase from the Vo proton channel, or turned on by reassembling the two subcomplexes into an active enzyme. While the process is well-characterized at the cellular level, the molecular mechanism at the level of the enzyme remains elusive. Here, we show that two TLDc proteins, Oxr1p and Rtc5p, control the assembly state of yeast V-ATPase, with the former promoting disassembly, and the latter (re)assembly of the enzyme. Based on cryoEM analysis and in vitro and in vivo approaches, we discovered that Oxr1p is a V-ATPase disassembly factor. Oxr1p binding to V-ATPase results in autoinhibited V1 in two steps - first producing a disassembly intermediate, which, upon ATP hydrolysis, gets converted into autoinhibited V1. From in vitro experiments, we find that the second TLDc protein, Rtc5p, primes autoinhibited V1 for (re)assembly with Vo. CryoEM structures of Rtc5p bound V1 show Rtc5p's C-terminal ⍺ helix inserted into the catalytic core of the enzyme, thereby opening a second catalytic site, a conformational change that may facilitate (re)assembly of V1Vo. In vivo experiments, however, suggest that Rtc5p is not essential for V-ATPase reassembly in the cell, suggesting redundancy and/or alternative pathways. Overall, this study enhances our understanding of the molecular basis for the regulation of V-ATPase activity by reversible disassembly.