UTMB Health SHARED (Univ. of Texas Medical Branch at Galveston)
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MicroRNA-122 and Poly(C)-Binding Protein 2 Bind to the Hepatitis C Virus Genome, Regulating Viral Replication and Genome Structure
No full textHepatitis C virus (HCV) has a positive-sense RNA genome which is used as the template for both viral translation and negative-strand RNA synthesis. As both processes work in opposite directions on the genome the virus requires a mechanism to coordinate what process is acting on a given copy of genome. It has been suggested that the host factors, microRNA-122 (miR-122) and poly-(C) binding protein 2 (PCBP2), may regulate these two processes. These host factors promote HCV replication and compete for overlapping binding sites on the 5’ untranslated region (UTR) of the HCV genome. This work investigates how miR-122 and PCBP2 bind the 5’ UTR of the HCV genome, and how the competition between the two factors regulates viral translation and replication.
The binding competition of miR-122 and PCBP2 was characterized and the impacts on HCV replication identified. Near the 5’ terminus of the HCV genome there are two miR-122 binding sites, S1 and S2. PCBP2 binds a sequence that overlaps the second miR-122 binding site (S2) causing direct competition with miR-122 for binding. Steady-state binding assays determined that PCBP2 and miR-122 have similar affinities for the overlapping binding sites. Electron microscopy shows the competition of these two factors modulates the PCBP2-mediated circularization of the HCV genome. Additionally, a novel interaction between the viral polymerase, NS5B, and the 5’ UTR was shown. The affinity of this interaction is increased by miR-122 binding to HCV, potentially promoting RNA synthesis. The modulation of genome structure and polymerase affinity may form a mechanism by which the competition of miR-122 and PCBP2 regulates genome templating for translation versus RNA synthesis.
The structure of the miR-122 and HCV genome was investigated. The first miR-122 binding site (S1) is the higher affinity of the two sites and is not affected by PCBP2 binding. To determine the structure of the RNA complex via cryo-EM, a proheadRNA-Assisted RNA Imaging Scheme (pARIS) was developed. This novel method can be applied to other small RNA structures that otherwise cannot be reconstructed via cryo-EM. We found that the HCV:miR-122 complex is flexible and accessible in the absence of the argonaut-2 protein
Ehrlichia chaffeensis Activates Notch Signaling Through SLiM Mimicry to Inhibit Apoptosis
No full textEhrlichia chaffeensis is a small, obligately intracellular gram-negative bacterium, and the etiological agent of human monocytotropic ehrlichiosis (HME), an emerging, life threatening tick-borne zoonosis. E. chaffeensis infects mononuclear phagocytes and has evolved molecular strategies to reprogram the host cell involving secreted effectors that interact directly with the host cell targets. Recently, we have shown E. chaffeensis evasion of innate defenses of the macrophage involve activation of Wnt, Hedeghog and Notch signaling pathways. Interestingly, the E. chaffeensis tandem repeat effector,
TRP120, has been shown to interact with host proteins important for activation and regulation of conserved signaling pathways including Wnt, Notch and Sonic Hedgehog. In this study, we investigated the molecular interactions and functional implications of Notch activation during E. chaffeensis infection.
The objective of this research project is to identify the SLiM ligand mimetic in E. chaffeensis TRP120 and the functional implications of TRP120 Notch activation. Two aims were originally proposed to investigate this hypothesis. Aim 1 was to elucidate the molecular interactions required for TRP120 Notch activation. Aim 2 was to investigate the role of E. chaffeensis Notch stabilization of XIAP and inhibition of caspase activation. Based on the evidence collected during my research, I have concluded that a TRP120-
TR Notch SLiM memetic motif directly binds Notch-1 at a region containing the LBD to activate Notch signaling. TRP120 Notch activation results in an anti-apoptotic program involving inhibitor of apoptosis proteins (IAPs) that inhibits caspase activation for intracellular survival. We demonstrate sequence homology between TRP120 and Notch ligands and determined that the TRP120-TR shares significant identity with known Notch ligands. We determined direct interactions between TRP120 and NECD recombinant protein containing ligand interaction domain, EGFs 1-13. We further defined the TRP120-TR domain as being capable of Notch activation and have defined the TRP120 Notch SLiM memetic motif required for Notch activation. Furthermore, we determined a direct correlation between Notch activation and inhibition of apoptosis linked to an increase in XIAP expression during E. chaffeensis infection. Increased XIAP levels correlated with increased NICD levels during E. chaffeensis infection and after TRP120 Notch ligand mimetic peptide treatment. Additionally, increased XIAP expression was consistent with
increased pro-caspase levels. siRNA knockdown or inhibition of XIAP with small
molecule inhibitor significantly increased apoptosis and Caspase-3, -7 and -9 levels and decreased ehrlichial load. This investigation reveals a mechanism whereby E. chaffeensis repurposes Notch signaling to stabilize XIAP and inhibit apoptosis. Understanding the molecular basis of Ehrlichia-Notch ligand mimicry is important for understanding the survival strategies of intracellular pathogenesis. Defining such interactions may lead to the development of novel therapeutics that target host-pathogen protein-protein interactions.
The proposed study is highly significant in revealing a molecular mechanism
whereby obligately intracellular pathogens, with small genomes and limited protein effectors, have evolved moonlighting proteins and molecular mimicry to rewire conserved signaling pathways and cellular functions to ensure persistent infection and survival. The identification of a short linear motif found within a non-canonical Notch ligand gives more insight into the underlying molecular mechanisms of aberrant Notch activation and may therefore lead to therapeutic approaches for diseases by which constitutively activated Notch signaling leads to disease onset and progression. Understanding Notch activation during E. chaffeensis infection will allow for the development of agents targeting critical steps of Notch signaling to inhibit infection and survival in the macrophage
Location, Location, Location: Assessing the Role of Context in Moderating the Relationship Between Childhood Adversity and Mental Health Burden
No full textAdverse childhood experiences (ACEs) have profound mental health implications throughout the life course. While literature has focused on individual risk factors for ACEs and their burden on health, there are social and physical environmental contextual factors outside the individual that have been overlooked in studies of childhood adversity. This dissertation sought to understand how environmental context (social and physical environment) moderates the relationship between prior ACE exposure and mental health by using the 2015 Behavioral Risk Factor Surveillance System (BRFSS) and County Health Rankings. By using the BRFSS and the County Health Rankings, this dissertation assessed ACE burden and mental health as well as examined participants’ context at the county level. The goals of this dissertation are to: (1) explore how to define context using social and physical contextual variables, (2) evaluate if context can act as an effect modifier in the relationship between childhood adversity on mental health, and (3) assess if there are racial/ethnic differences on how context moderates ACE burden on mental health. The results of this proposal shed light on how certain aspects of context can buffer the burden of ACEs on mental health, and these results can be used as evidence for allocation of funds to help alleviate the ACE burden at the county level
Aging Skeletal Muscle Plasticity in Exercise and Injury
No full textSarcopenia—progressive loss of muscle mass and strength—diminishes quality of life and longevity. In addition to progressive atrophy, older adults exhibit impaired regeneration after muscular injury. Activity of muscle stem cells, termed satellite cells, is dysregulated with aging which impairs skeletal muscle remodeling and limits plasticity of aged muscle. Aim 1 (Chapter 2) of this dissertation seeks to clarify the debated requirement of satellite cells for overload-induced hypertrophy in aging muscle via a common surgical overload model, with the original hypothesis that while aging satellite cells contribute to overload-induced hypertrophy to mitigate sarcopenia, growth can occur in the absence of satellite cells via expansion of the myonuclear domain. In light of Aim 1 results demonstrating no overload-induced hypertrophy in aging skeletal muscle with surgical overload, Aim 2 (Chapter 3) examines the efficacy of a novel and translatable murine resistance exercise model to elicit satellite cell expansion with myonuclear accretion and hypertrophy in aging skeletal muscle, with the hypothesis that this novel exercise model would result in expansion of the satellite cell pool along with hypertrophy not observed in old mice with the surgical overload model. Indeed, PoWeR (Progressive Weighted wheel Running) elicited hypertrophy in old mice, likely supported by a robust angiogenic response in hind limb muscles. Lastly, Aim 3 (Chapter 4) examines the efficacy of a novel NNMT inhibitor to enhance regeneration of aging skeletal muscle after injury by enhancement of satellite cell proliferation and fusion to myofibers, confirming the hypothesis that NNMT inhibition would rescue age-related deficits in satellite cell activity to promote superior regeneration of muscular injury of old mice
Yellow Fever: Role of Heterologous Flavivirus Immunity on Urban Emergence
No full textDuring the recent yellow fever (YF) epidemics in Brazil, human cases were attributed only to spillover infections via sylvatic mosquito transmission. Despite YF virus (YFV) transmission in major urban centers with insufficient vaccination coverage and abundant populations of the domestic vector, Aedes aegypti, there was no evidence of human-amplified transmission. Furthermore, the complete historic absence of YF in Asia, despite abundant Ae. aegypti and a completely immunologically naive human population represents a longstanding epidemiological enigma. We tested the hypothesis that pre-existing, heterologous flavivirus immunity, specifically from dengue (DENV) and Zika (ZIKV) viruses, limits YFV viremia and transmission by Ae. aegypti. We infected cynomolgus macaques with DENV or ZIKV, then challenged them 6-9 months later with YFV. We then measured viremia and disease, and allowed Ae. aegypti mosquitoes to feed during peak macaque viremia. Although prior heterologous immunity had variable effects on disease, DENV and ZIKV immunity consistently suppressed YFV viremia, leading to a significant reduction in Ae. aegypti infection and a lack of transmission potential. Next, we utilized an interferon α/β receptor knock-out mouse model to determine the role of pre-existing DENV-2 and ZIKV immunity in YF virus infection, and to determine mechanisms of cross-protection. We utilized African and Brazilian YF strains and found that DENV-2 and ZIKV immunity significantly suppresses YFV viremia in mice but did not consistently protect against disease outcome. Cross-protection appears to be mediated mainly by humoral immune responses. These studies underscore the importance to re-assess the risk of YF outbreak accounting for prior immunity from flaviviruses that are endemic
A Naturalistic Inquiry of the Experiences of Women Diagnosed with Gestational Diabetes
No full textGestational diabetes, hyperglycemia that develops in pregnant women with no prior history of diabetes, affects 2% to 10% of all pregnancies in the United States (Centers for Disease Control and Prevention [CDC], 2022). About 50% of women with GDM go on to develop T2DM, and babies of women diagnosed with GDM also are at increased risk of developing T2DM later in life (CDC, 2022). Effective management of GDM requires women to change their health behaviors related to diet, exercise, and medication adherence. Although some studies have examined the experiences of women who have been diagnosed with GDM in other parts of the world, there is a need for research that explores the experiences of women with GDM living in the United States. The current study utilized Naturalistic Inquiry [NI] (Erlandson et al., 1993; Lincoln & Guba, 1985) to explore and describe the experiences of women diagnosed with gestational diabetes [GDM] in a previous pregnancy and were living in the United States.
Data collection took place in the form of one-on-one interviews via Zoom Video Conference with women diagnosed with GDM in a previous pregnancy currently living in the United States. Study data consisted of interview data and participants’ demographic data. Interview data was analyzed using Erlandson et al.’s (1993) interpretation of Lincoln and Guba’s (1985) approach to inductive data analysis. Data analysis revealed three major categories: 1) Finding Out About the Gestational Diabetes Diagnosis, 2) Mastering GDM, and 3) Life After GDM. The implications of the study’s findings pertain to nurses and other healthcare providers who help care for pregnant women diagnosed with GDM. Women with GDM need more information about GDM risk factors, how to incorporate GDM recommendations into their daily lives, mental health resources, and whether having had GDM posed long-term risks for themselves and their children
Searching for their Path: Understanding Parents of Children with Emotional Disturbance
No full textBackground: Children with Emotional Disturbance (ED) have unpredictable reactions and behaviors, which cause significant problems for themselves, their families and society (Zionts et al., 2016). The child’s reactions are highly disproportionate, chronic and frequent (Gage, 2013). These children have significant risks related to their ED, including school drop-out, incarceration, unemployment and substance abuse (Lipscomb et al., 2018).
Purpose: The purpose of this study was to explore participants’ experiences related to raising a child with ED, participants’ response to situations related to their child’s ED, and how a child with ED impacted the family.
Method: This study explored the parenting experiences of mothers of children with ED, utilizing Naturalistic Inquiry as described by Lincoln and Guba (1985) and Erlandson et al. (1993). After IRB approval, eight mothers were recruited nationally through social media.
Results: Three overarching constructs emerged in the study. The first construct, “concerns and suspicions,” was the evolution of mothers’ concerns, rationalizations, escalations, decision and eventual confirmation that their child was “different.” “Actions and adaptations” depicted the actions used to manage their child with ED, along with actions for self-care and siblings’ care. “Altered families and advice for others” described how every family member was impacted by the child with ED and participants’ advice on how to ease the difficulties for other families. Study findings suggested needed changes to nursing practice, education and policy
The Balance of Interferon-γ and Interleukin-10 during Influenza A Virus Infection Complicated by Methicillin-Resistant Staphylococcus aureus Superinfection
No full textImmune activation is necessary to mount a protective immune response against viral and bacterial infections, but an overzealous response can lead to the development of acute respiratory distress syndrome (ARDS). ARDS is a highly lethal inflammatory lung injury that is associated with dysregulated cytokine production and decreased lung compliance. No specific treatment exists for ARDS due to the limited understanding of its pathogenesis. We have developed an influenza A virus (IAV) and methicillin-resistant Staphylococcus aureus (MRSA) superinfection model with antibiotic therapy that resembles severe secondary bacterial pneumonia in patients that often progresses to ARDS. Despite antibiotic therapy, mice still succumb to superinfection-induced inflammatory lung damage. Using single-cell RNA sequencing, we reveal significant transcriptome alterations prompted by interferon (IFN)-γ. Transgenic mouse studies demonstrate that IFN-γ receptor (IFN-γR) signaling in mononuclear phagocytes induces tumor necrosis factor (TNF)-α hyperproduction and lethal inflammatory lung damage, with no detectable benefit to viral or bacterial clearance. In contrast, we show that interleukin (IL)-10 is crucial to counteract the lethal IFN-γ-induced cytokine storm and preserve lung function. Transgenic mice with ablation of the IL-10 receptor α gene in mononuclear phagocytes have significantly higher levels of IFN-γ and TNF-α, and a higher mortality rate. This reveals the importance of mononuclear phagocytes in shifting the balance between immunopathology and protective immunity. By omitting antibiotic therapy from the superinfection model, we show that IL-10 also impairs bacterial clearance, specifically when the IL-10Rα gene is ablated in interferon-I-responsive monocytes. Collectively, this body of evidence demonstrates the dominant role played by hypercytokinemia and acute lung damage during post-influenza bacterial pneumonia and the balance of IFN-γ and IL-10 that shifts the balance between immunopathogenesis and bacterial outgrowth
Normalizing the Abnormal: Self-Identified Traumatic Events and Critical Care Nurses, A Focused Ethnography
No full textAccording to The National Institute for Occupational Safety and Health (2022), 20 million healthcare workers in the United States (U.S.) are at risk for symptoms of mental unhealthiness directly related to their work environment and experiences. Despite nurses reporting higher levels of burnout and other adverse effects within their work environment, approximately two-thirds of nurses indicate they are not receiving supportive resources for mental health, which is known to impact physical health (Berlin et al., 2023). The pandemic has shed light on pre-existing conditions for nurses, such as the workplace culture and understaffing, leading to unmanageable patient loads, which all minimize nurses' mental health (Bowie, 2022). Critical care nurses (CCNs) encounter singular and often repetitive traumatic events (TEs) within the work environment, which lead to adverse effects such as anxiety, depression, post-traumatic stress disorder, secondary traumatic stress disorder, burnout, compassion fatigue, moral distress, substance use, and suicidal behaviors. A TE is described as an extreme event causing an individual’s ability to cope to be threatened, resulting in unusual and strong cognitive, emotional, or behavioral reactions (Kleim et al., 2015).
A limited number of studies have directly addressed the experiences of self-identified TEs among CCNs in the clinical work environment. This study aims to describe the CCN’s TE, explore potential psychological impacts on the CCN, explore work culture impacts on the CCN’s response, and examine resources offered to the CCN. A focused ethnography (Roper and Shapira, 2000) methodology was used for the study and was guided by the research question, “What are CCNs’ experiences and responses to a self-identified traumatic event(s) in the emergency department or intensive-care unit settings that caused them distress?”
Purposive and snowball sampling was used for recruitment. CCNs (n=11) representing various regions in the U.S. participated in semi-structured interviews. Study data included demographic data, transcribed interviews, field notes, and methodological journals. Collected data was analyzed using the constant comparative method (Glaser, 1998; Glaser and Strauss, 1967) until redundancy and data saturation occurred. The study utilized Beck’s (1993) criteria to demonstrate scientific rigor. Data analysis revealed four themes of normalizing the abnormal, suffering in silence, badge of honor, and resilience: we are CCNs
Life-Space Mobility, Falls, and Healthcare Use Among Community Dwelling Mexican-American Older Adults
No full textPhysical mobility affects all aspects of daily life and is a crucial part of independent living. Life-space mobility (LSM) is a valuable concept for measuring both mobility and independence of older adults in their day-to-day lives. Aims are: 1) to determine the threshold values of restricted LSM among Mexican American older adults in comparison to the original life-space assessment (LSA) validation on measures of disability, depressive symptoms, physical function, and mortality; 2) to evaluate LSM as a predictor of falls and fear of falling with and without visiting a hospital or Emergency Department (ED) among Mexican American older adults over time; and 3) to determine the association between LSM and healthcare use of Mexican American older adults over time. We included 799 respondents aged 80 and older from the Hispanic Established Population for the Epidemiologic study of the Elderly survey (2010/11-2016). The independent variable was restricted life-space mobility. Outcome variables included falls/fear of falling, and acute/post-acute healthcare use. Covariates were age, sex, education, marital status, nativity, living arrangement, pain, cognitive and physical function, comorbidities, depressive symptoms, and sensory deficits. Receiver operating characteristic curve analysis showed that the new threshold for LSM restriction in this population was a LSA of ≤35 points for disability, physical function, depressive symptoms, and mortality which was lower than the initial score of ≤60 points found in the original LSA validation cohort. Generalized Estimating Equations models showed that the new restricted LSM (LSA ≤35) is associated with falls with visits to the hospital or ED [odds ratio (OR)=1.58 95% Confidence Interval (CI)=1.03-2.44], fear of falling (OR=1.88, 95% CI=1.36-2.60), and hospital admissions (OR=1.57, 95%CI=1.13-2.19) after controlling for all covariates. No significant association was found between restricted LSM and post-acute care. In conclusion, there is a lower threshold for restricted LSM among Mexican American older adults and that the lower threshold more accurately predicts falls that required a hospital or ED visits, fear of falling, and acute healthcare use. This suggests that the new LSA threshold may be a better assessment tool for health outcomes among Mexican American older adults