Now showing items 1-20 of 2221

    • Moving Beyond ‘A white Man’s Thing’: A Case Study of Urban Kenyan Youth Mental Health

      Katerere-Virima, Thuli; Shdaimah, Corey S. (2023)
      Background: Kenya is a lower middle-income country located in Eastern Africa with a population of over 54 million people and a median age of 20 (World Bank, 2020). Competing health emergencies, a healthcare infrastructure ill-prepared for crisis, and inconsistent framing of mental health in culturally relevant terms have all created a gap between mental health need and services in Kenya (Meyer & Ndetei, 2016). This study explores how 15–24-year-olds in Nairobi, Mombasa and Kisumu counties define their mental health and which resources and barriers impact their engagement with mental health services. This study was designed to contribute to the ongoing REACH-MH (Reaching, Engaging Adolescent and youth adults for Care Continuum in Health-Mental Health) project. Methods: I used an inductive approach to answer two research questions: 1) How do adolescents/young people (AYP) define their mental health? and 2) How do relevant stakeholders describe resources and barriers to AYP mental health? For this case study focused on LVCT Health’s One2One program, I used five sources of data: in-depth qualitative interviews with One2One hotline counsellors; One2One hotline data; youth focus group transcripts; stakeholder meeting notes; and government document review of the Mental Health Taskforce Report of 2020 and the Mental Health Amendment Act of 2022. Findings: Five themes emerged from the data regarding the universality of “stress” as a concern for youth, the common conflation of mental health and mental illness, and recommendations for youth-friendly provision of mental healthcare. Overwhelmingly, study participants defined “mental health” in ways that captured broader social determinants of health, along with descriptions of “emotional, psychological and social wellbeing”. Barriers to mental health included cost and a lack of trust in mental health professionals, while youth’s capacity for coping and knowledge of the few, but existent, community services available were reported as facilitating factors. Conclusions: Though challenges abound, also numerous are the strengths and resources possessed by Kenya’s people who continue to solve problems and utilize ways old and new to strive toward a uniquely Kenyan conceptualization of mental health.
    • The Impact of Simulation-based Ethical Education on Prelicensure Nursing Students' Levels of Moral Distress

      Dalton, Jennifer; Gordes, Karen L. (2023)
      Research demonstrates the prevalence of moral distress in licensed nurses often begins during undergraduate training. While the concept of moral distress is evolving, this study used a conceptual definition of moral distress defined as negative self-directed emotions in acknowledgement of involvement in morally difficult situations. The purpose of this study is to determine whether integrating an ethical simulation-based education (SBE) module into fourth semester undergraduate nursing students’ curriculum would affect moral distress levels as measured by a validated Moral Distress Scale for Nursing Students. Using a longitudinal embedded mixed methods design in a required final-semester undergraduate nursing leadership course, students participated in an ethical SBE module with debriefing and completed the It-ESMEE as a pretest. During weeks two and three of the semester participants completed the SBE module using role play to practice speaking up with integrity. The closing of the module included a recorded debriefing to collect qualitative data related to the module and feelings of moral distress and the It-ESMEE for a second time. The final It-ESMEE was completed 13 weeks after the SBE module the final week of the semester. The recorded debriefings were transcribed and thematically analyzed. A repeated measures analysis of variance tested the within-subject differences of mean total It-ESMEE moral distress scores across all time points. A total of 48 students were included in the study. There was no statistically significant difference among mean moral distress scores, (F (2, 94) = 1.369, p = .259). Analysis of recorded debriefings revealed four primary themes: 1) powerlessness as a student, 2) students have a basic moral understanding, 3) there is discomfort in speaking up with integrity, and 4) students understand the importance of emotional intelligence. In conclusion, the measurement of overall moral distress levels over three time points did not show a statistically significant difference. However, the mean 13-week post-intervention survey score (M = 173.85) was the lowest of all three perceived moral distress scores. Though not statistically significant, this data in conjunction with the results of the thematic analysis could indicate that integrating meaningful simulation-based ethical training throughout undergraduate nursing curriculum may affect perceived moral distress levels. Future studies should address integration of ethical SBE throughout curricula.
    • The Economic Burden of Chronic Obstructive Pulmonary Disease and Comparative Effectiveness of Maintenance Inhaler Medications in the United States

      Shah, Chintal; Zafari, Zafar (2023)
      Introduction: Chronic Obstructive Pulmonary Disease (COPD) is a highly prevalent condition in the United States (US). Among individuals with moderate to very severe COPD, inhalation therapy is the mainstay of disease management, with the goal to reduce COPD exacerbations. Maintenance medications, especially combinations of long-acting beta2 agonist (LABA)/long-acting muscarinic antagonist (LAMA) or LABA/inhaled corticosteroids (ICS), are commonly used. This dissertation aimed to examine the (i) economic burden of COPD, (ii) comparative effectiveness of LABA/LAMA and LABA/ICS fixed dose combination (FDC) single inhaler therapy across various subgroups, and (iii) comparative effectiveness of LABA/LAMA combinations with different ingredients and inhaler types, vilanterol/umeclidinium (VI/UMEC) and olodaterol/tiotropium (OLO/TIO). Methods: Medical Expenditure Panel Survey data was used to estimate the economic burden of COPD (Aim 1). COPD-specific (adjusted) costs were determined for various service categories using a regression-based weighted two-part model among patients aged 45 years and older. Medicare Chronic Conditions Warehouse data was used for the comparative effectiveness studies (Aim 2, Aim 3). A new user active comparator retrospective cohort study design was utilized, and the outcome of interest was time to first COPD exacerbation. To ensure comparability between groups, they were matched based on their high-dimensional propensity scores. Results: The total COPD-specific direct medical cost was 2018 US $4,322 (Standard Error (SE): US $577) per patient per year with prescription drugs contributing US $1,887 (SE: US $216). The resultant overall annual total COPD-specific cost was US $24.0 billion, with prescription drugs contributing US $10.5 billion. For the interclass comparative effectiveness analysis, the hazard ratio (HR) of time to COPD exacerbation was 0.846 (95% Confidence interval (CI): 0.776-0.923) for LABA/ICS compared to LABA/LAMA initiators. Among LABA/LAMA FDCs, the HR of time to first COPD exacerbation was 0.948 (95% CI: 0.813-1.105) for individuals initiating OLO/TIO versus VI/UMEC. Conclusion: This dissertation found that COPD poses a significant economic burden on the US healthcare system, with prescription drugs being a major contributor. Optimizing therapy can help reduce this burden. While a statistically significant interclass difference was observed between LABA/LAMA and LABA/ICS initiators, no statistically significant intraclass difference was observed between initiators of LABA/LAMA FDCs: VI/UMEC and OLO/TIO.
    • Altered Gene Expression Profiles and Immune Responses in a Murine Model of a Non-lethal Flame Burn with Pseudomonas aeruginosa Infection

      Kambouris, Adrienne Renee; Cross, Alan S. (2023)
      Humanity has lived with fires for millennia, but combat, domestic use, and recent wildfires have increased the risk of burn injuries. Worldwide, over 100,000 deaths occur each year due to burns. If these patients survive the burn wound itself, the most common causes of death are multiorgan failure and sepsis, often caused by infection by Pseudomonas aeruginosa (PA). Utilizing a 10% total body surface area (TBSA) non-lethal flame burn model in mice, a superimposed infection of PA caused 100% mortality within 36 hours post-burn. This effect was transient, as infection 72 hours post-burn resulted in survival. The hypothesis was that this mortality could be linked to changes in gene expression that altered host-pathogen interaction. NanoString™, a system that allowed us to develop a custom panel of probes, was utilized to measure Mus musculus and PA gene transcripts simultaneously in each sample. Sampling from the blood, spleen, liver, and skin, gene expression in the burn and infection condition (B/I) was significantly different in each tissue when compared to mice that were burned alone, infected alone, and neither burned nor infected (Sham). The expression of the anti-inflammatory gene, Il10 is significantly increased over time in the spleen; administering anti-IL-10 antibodies delayed mortality by one day. While Arg1 and Nos2 gene expression were not significantly altered, administering arginine concurrently with PA restored survival in our mouse model, likely due to an inhibition of both PA motility and growth. We also hypothesized that burn-induced neutrophil dysfunction allowed for PA proliferation. Neutrophils isolated from the seroma of burned mice had a decreased ability to produce antibacterial reactive oxygen species (ROS) compared to neutrophils in the circulation of the same mice. Surprisingly, naïve neutrophils in the circulation of burned mice had a decreased ability to kill PA, possibly due to their premature ROS production induced by a burn-generated DAMP, HMGB1, present in the serum of burned but not Sham mice. In conclusion, a non-lethal burn injury is sufficient to induce multi-faceted changes in the murine immune system that results in an increased susceptibility to lethal PA superinfection.
    • Pim kinase inhibitor enhances FLT3 inhibitor gilteritinib efficacy through GSK-3β activation and GSK-3β-mediated c-Myc and Mcl-1 proteasomal degradation

      Lee, Jonelle; Baer, Maria R. (2023)
      Acute myeloid leukemia (AML) with fms-like tyrosine kinase 3 internal tandem duplication (FLT3-ITD) has poor outcomes. FLT3-ITD drives constitutive and aberrant FLT3 signaling, activating STAT5 and upregulating the downstream oncogenic serine/threonine kinase Pim-1. FLT3 inhibitors have limited clinical efficacy. We previously showed that concurrent Pim and FLT3 inhibition increases apoptosis induction in FLT3-ITD-expressing cells through post-translational downregulation of Mcl-1. Here we further elucidate the mechanisms of action of this dual targeting strategy. Protein expression and turnover, cytotoxicity and apoptosis were measured in FLT3-ITD-expressing cell lines and AML blasts treated with FLT3 inhibitor gilteritinib and/or Pim inhibitors AZD1208 or TP-3654. Pim and FLT3 inhibitor co-treatment decreased c-Myc protein, prior to Mcl-1, increased turnover of both proteins, rescued by proteasome inhibition, dephosphorylated (activated) GSK-3β, and increased apoptosis and in vivo efficacy. GSK-3β inhibition prevented c-Myc and Mcl-1 downregulation and apoptosis. Pim and FLT3 inhibitor co-treatment of Ba/F3-ITD cells infected with T58A c-Myc or S159A Mcl-1 plasmids, preventing phosphorylation at these sites, did not downregulate these proteins, increase their turnover or induce apoptosis, consistent with GSK-3β activation and c-Myc T58 and Mcl-1 S159 phosphorylation as the mechanism of combination treatment. These data further support GSK-3β activation as a therapeutic strategy in FLT3-ITD AML.
    • Role of Social Determinants of Health on the HIV Testing and Treatment Cascade in Nigeria

      Mohanty, Kareshma; Stafford, Kristen (2023)
      Introduction: The Joint United Nations Programme on HIV and AIDS proposed that to achieve epidemic control of HIV by 2025; 95% of all people living with HIV are aware of their status, 95% of people diagnosed with HIV receive sustained antiretroviral therapy (ART), and 95% of all people on ART are virally suppressed (VLS). The 2018 Nigeria HIV/AIDS Indicator and Impact Survey (NAIIS) found that in Nigeria, while only 47% knew their status, 96% had received ART, and 81% had achieved VLS. Social determinants of health (SDH), like wealth index (WI), have been shown to play a significant role in HIV in western countries, but the evidence has been limited and mixed in Nigeria. Identifying political, social, cultural, demographic, economic, and behavioral indicators of SDH, can better explain and address the disparities in the HIV epidemic, especially in the testing and treatment cascade, that are preventing the UNAIDS targets from being met in Nigeria. Objective: Examine the role of singular and composite indicators of SDH on the 95-95-95 targets: HIV testing, receipts of ART, and VLS in people living with HIV in Nigeria. Additionally, examine if wealth modifies the relationship between SDH indicators and the three 95 targets. Methods: Using the World Health Organization-SDH framework and Factor Analysis, I constructed composite indicators of SDH for Nigeria from various population-level survey data sources. Scores from the sub-indices and Global Terrorism Index were categorized as low, medium, and high, and individual or states were assigned one of these categories. Subsequently, I examined the association of the composite and singular SDH factors with HIV testing, receipt of ART, and achievement of VLS through survey-weighted multivariable logistic regression. Additionally, I examined the significance of the SDH indicators with the testing and treatment outcomes, by each of the wealth quintiles. Results: Out of the seven sub-indices constructed, only Access to Public Services, Crime & Conflict, Government Corruption, and Government Performance met the internal reliability criterion (Cronbach alpha > 0.7). Global Terrorism Index was constructed based on the prescribed methodology. When examining HIV testing, the first target in the 95-95-95 UNAIDS strategy, medium levels of Government Corruption, lower/medium Government Performance, and high Terrorism was associated with lower testing. Unemployment, living in rural areas, and married before 18 years of age were significantly associated with lower odds of HIV testing. For receipt of ART, second 95-95-95 target, low/medium treatment coverage was associated with lower odds of being on treatment. Younger age, male sex, being single, and living in rural areas were the singular factors associated with lower receipt of ART. Finally, for the third 95-95-95 target, only singular SDH, like lack of condom usage during sex, CD4 count (<500), and ethnic languages were associated with lower VLS. Wealth modified the relationship between the social determinants and HIV testing and treatment, but the role was weak. Wealth may increase the gap between the lowest and highest wealth index strata; HIV-related disparities experienced might be more pronounced between the two ends. Conclusion: Understanding and addressing structural determinants like political stability, terrorism, gender equality, accessibility to public services, and treatment facility coverage, rather than individual-level behavioral factors, could help Nigeria achieve the 95-95-95 targets.
    • Smoking Cessation Among People With Severe Mental Illness

      Alghzawi, Hamzah Mohammad; Storr, Carla L. (2019)
      Introduction: People living with mental illnesses have a high rate of smoking and make up over half of those dependent on nicotine. A considerable body of research has shown that social support, stressful life events (SLE), receiving help for tobacco/nicotine use, intention to quit, and smoking use-related factors are associated with smoking cessation in the general population. Yet, little is known about these factors among people with severe mental illness (SMI). Purpose: This study aims to: 1) examine gender differences in the interrelations among social support, SLEs, and smoking cessation, 2) estimate the probability of remission from NUD by type of help/services received for tobacco/nicotine use (pharmacological, non-pharmacological, and both), and 3) estimate gender and racial/ethnic differences in the probability of smoking cessation among those with a history of intention to quit. Methods: A sample of 4610 people with SMI and a history of tobacco/nicotine use were identified in a public limited dataset of the 2012-2013 National Epidemiologic Survey on Alcohol and Related Conditions (NESARC-III). Four mediation and moderated mediation models were used in the first manuscript, whereas survival analyses were used in the second and third manuscripts. All analyses took into account the complex sampling design and controlled for possible confounders (i.e. sociodemographic characteristics) and covariates (i.e. comorbidity with another mental illness). Results: Total, appraisal, and tangible support in females exerted indirect effects on improving smoking cessation via decreased SLEs (total=.0094, appraisal=.0229, tangible=.0298; p<.05). The probability of remission from NUD was higher among those who received non-pharmacological services (28.5%, HR=1.95, p<.05) or those who received both services (19.6%, HR=1.52, p<.05) compared to those who only had pharmacological services (17.6%). Among those with a history of intention to quit, 31.7% had stopped. The probability of smoking cessation was highest for Hispanic females (HR=2.07, p<.05), non-Hispanic other females (HR=1.59, p<.05), non-Hispanic other males (HR=1.45, p<.05), Hispanic males (HR=1.40, p<.05), and non-Hispanic Black females (HR=1.35, p<.05) compared to non-Hispanic Black males. Conclusion: A greater understanding of subgroup differences and the correlates of smoking cessation among tobacco/nicotine users with SMI can enhance efforts to design and implement smoking cessation programs for people with SMI.
    • PA Education and Practice in Maryland: Current Status, Opportunities and Challenges

      PALLA (Physician Assistant Leadership and Learning Academy) (2024-01)
      Physician assistants/associates have become a well-established component of the US health care work force. In Maryland, the PA profession is flourishing with more than 3600 PAs in active clinical practice and 5 operational PA education programs. A growth rate of approximately 40% has been projected between 2018-2028. The Physician Assistant Leadership and Learning Academy (PALLA), based at the University of Maryland, Baltimore with funding from the state, was created in 2019 to advance PA education, research, policy, and practice in the state. Consistent with this mission, PALLA regularly conducts an environmental scan to ascertain current opportunities and challenges facing the PA profession in the state. In fall of 2023, PALLA undertook an examination of the status of the profession and the state’s PA educational programs. For this report, PALLA conducted a series of semi-structured interviews with Maryland PA program faculty, program leaders, and deans. PALLA also analyzed secondary data from both local and national agencies. Several themes consistently arose from this analysis; among them were a strong demand among applicants, PA graduates staying local, concerns regarding program leadership and faculty stability, challenges in the development of effective student remediation processes, and the issue of clinical site and preceptor shortages. Additional areas of concern identified by Maryland PA program leadership and faculty were issues related to accreditation, levels of institutional support, faculty turnover and workload, sufficiency of support, and lack of diversity among faculty and students. There was a desire among interviewees for increased collaboration between PA programs and other health professions. At a practice level, PAs are making a strong impact in various clinical settings and the demand currently outweighs the supply in Maryland. However, scope of practice regulations remains a major barrier in maximizing PA value in Maryland. Based on current opportunities and challenges identified, we present a series of specific and detailed recommendations that we hope will stimulate discussion among various stakeholders while advancing the quality of PA education, policy, and practice in Maryland
    • Impact of Host and Parasite Factors on Gametocyte Production in Plasmodium falciparum

      Vareta, Jimmy Amakwa; Laufer, Miriam K.; Takala-Harrison, Shannon (2023)
      Stalled progress in reducing the malaria burden over the past few years suggests the need to develop new interventions to augment existing ones, including interventions aimed at interrupting gametocyte transmission from humans to the mosquito vector. To develop and effectively apply interventions that target gametocytes, there is a need to understand patterns of gametocytemia observed between individuals. Gametocytemia varies by host age, season, symptom status, antimalarial drugs use, and complexity of infection; however, the underlying mechanisms of this variation are not fully understood. Complexity of infection may modulate gametocytemia in P. falciparum; however, mechanisms of how clone composition influences gametocyte production are not clear. Addressing this gap requires a genotyping assay that can detect and estimate relative clone frequency of gametocytes and asexual parasites in infections. The dissertation aims were two-fold: 1) To develop an amplicon sequencing assay to genotype P. falciparum mature gametocytes; and 2) To evaluate the impact of host and parasite factors and gametocyte production in P. falciparum infections. We identified a polymorphic region of the pfs230 gene as a marker to distinguish P. falciparum mature gametocyte clones. When evaluating the impact of host and parasite factors on gametocyte production and gametocytemia, we found that more complex and high parasite density infections were more likely to produce and harbor gametocytes. The proportion of infections that produced gametocytes were similar between age-groups and between symptomatic and asymptomatic individuals, but children and asymptomatic individuals were more likely to harbor gametocytes than adults and symptomatic individuals, respectively. These findings suggest that complexity of infection and parasite density may increase gametocyte production, but additional factors such as host immunity and duration of infection may contribute to the presence or absence of gametocytes after initiation of gametocyte production. Coupled with the development of the gametocyte genotyping assay which will be an important tool for studies aimed at understanding dynamics of gametocyte production in polyclonal infections, understanding the impact of host and parasite factors on gametocyte production and gametocytemia will help explain variation in gametocytemia observed between individuals. This knowledge could inform development and effective deployment of transmission interrupting interventions.
    • Attitudes Toward Medical Aid in Dying in a National Sample of Hospice Clinicians

      Becker, Todd D.; Becker, Todd; Cagle, John G. (2023)
      As of this writing, medical aid in dying (MAID) is available in 11 U.S. states and the District of Columbia, with further expansion projected. Legal protections for conscientious objection foreground clinician attitudes as substantial barriers or facilitators to MAID access for interested patients. Although upward of roughly three quarters of patients who use MAID are enrolled in hospice care, little is known about hospice clinicians’ attitudes toward MAID. The purpose of this three-paper dissertation was to examine attitudes toward MAID in a national sample of hospice clinicians. Participants were recruited from national hospice and palliative care membership associations representing the four core disciplines of the hospice interdisciplinary group (i.e., medicine, nursing, social work, chaplaincy) to complete a one-time, self-administered survey. Paper 1 examined the preliminary psychometric properties of a modified version of the only empirically evaluated scale on attitudes toward MAID. Confirmatory factor analysis results indicated that the Attitudes Toward Medical Aid in Dying Scale demonstrated factorial validity. Construct validity was established through correlation analyses targeting convergent validity (vis-à-vis a researcher-constructed measure of attitudes toward MAID) and discriminant validity (vis-à-vis a researcher-constructed measure of attitudes toward euthanasia and a scale assessing religiosity). High congeneric reliability estimates supported internal consistency reliability. Despite the favorability of these statistical results, conceptual mismatches between scale items and the U.S. practice context as defined by state laws caution against wider scale use. Further psychometric development is warranted. Paper 2 explored institutional factors tied to the hospice context of care as correlates of MAID attitudes. Using a 3-point version of Paper 1’s ordinal convergent validity item, results of a partial proportional odds model indicated that professional experience working in a state where MAID was legal and increased orientation toward patient-centeredness were both significantly associated with higher odds of more supportive MAID attitudes across each threshold of the dependent variable. Increased commitment to the hospice philosophy of care also was significantly associated with higher odds of more supportive MAID attitudes. Accounting for differing slopes across dependent variable thresholds, however, this association reached statistical significance only when estimating the odds of being in a category above the midpoint response option (neither support nor oppose). Findings support the assessment of ecological factors that drive hospice ethos and functioning when exploring attitudes toward MAID. Paper 3 explored attitudes toward being physically present throughout MAID in a hypothetical patient scenario governed by certain safeguards. The 74% of participants who indicated willingness to be present did so based on feelings of personally derived support for MAID, definitions of quality clinical care, and values from their professional training. This broad support, however, was conditioned by boundary setting though which participants described specific conditions required for their participation. In contrast, 15% of participants were unwilling to be present. These attitudes were attributed to objections to the concept of MAID, objections to participation in MAID, and perceptions that MAID is misaligned with health care. Merely 11% of participants were unsure, relating their hesitation to feelings of ambivalence and a lack of experience with MAID. The tensions that participants across samples reported experiencing with themselves, their profession, and broader society reflect a need for greater professional guidance on the safe and effective provision of MAID.
    • An Insight to Further Malaria Vaccine Development: PfSPZ Vaccine Correlates of Protection Appear to be Cross-Reactive Antibodies to Immunodominant Low-Complexity Epitopes

      Berry, Andrea; Takala-Harrison, Shannon (2023)
      Plasmodium falciparum circumsporozoite protein (PfCSP) coats the sporozoite surface and is the target of multiple malaria vaccines in development. Discovery of additional vaccine candidate antigens beyond PfCSP may lead to improved vaccines. To identify new antigens, we used peptide microarrays to map antibody responses to P. falciparum proteins in adults who received a whole organism sporozoite vaccine, PfSPZ Vaccine, and were protected or unprotected after controlled human malaria infection. We discovered antibody responses that correlate with protection, but further examination, including of two monoclonal antibodies derived from protected PfSPZ Vaccine recipients, suggests that some antibodies elicited by PfCSP cross-react with peptides representing non-CSP proteins, a demonstration of inter-protein cross-reactivity. This work provides evidence that PfSPZ Vaccine elicits inter-protein cross-reactivity, provides amino acid-specific detail of putative epitopes, and introduces opportunities for further exploration that may help to elucidate underexplored immunological mechanisms and inform development of next-generation malaria vaccines.
    • Nitric Oxide in Mucormycosis Pathogenesis

      Soare, Alexandra; Bruno, Vincent, Ph.D. (2023)
      Mucormycosis is classified by NIAID as an emerging disease and is caused by Mucorales fungi. The recent surge of mucormycosis cases among COVID-19 patients has thrust the disease and lack of available treatments into the spotlight. Clinical data suggests a lack of inflammatory responses during mucormycosis despite severe fungal angioinvasion and tissue necrosis. In this dissertation, I sought to characterize immune evasion mechanisms by Mucorales, focusing on the interaction between fungi and macrophages. Macrophages infected with Mucorales fungi block the production of nitric oxide, a free radical molecule with strong antimicrobial properties and an important signaling role in immunity. Despite the increased expression of Nos2 mRNA and inducible nitric oxide synthase (iNOS) protein in Mucorales-infected macrophages, these macrophages are unable to produce nitric oxide, even when stimulated with nitric oxide-producing stimuli (LPS and IFN-γ). My results suggest that Mucorales fungi prevent the accumulation of nitric oxide through at least 2 mechanisms: (1) removal of nitric oxide from the surrounding environment, and (2) depletion of nutrients required to make nitric oxide. Additionally, a potent nitric oxide-donor (DETA-NONOate) inhibits in vitro growth of Mucorales fungi indicating that nitric oxide may be have antifungal activity against Mucorales. At lower concentrations of DETA-NONOate that are unable to inhibit growth of Mucorales, I observed downregulation of mRNAs encoding Mucorales virulence proteins including Mucoricin, a ricin-like toxin that is critical for Mucorales pathogenesis. By downregulating these genes, nitric oxide could be attenuating the virulence potential of the fungus, rendering it less pathogenic. My research describes a new immune evasion mechanism by Mucorales fungi and presents nitric oxide as a potential therapeutic for mucormycosis.
    • Role of Galectin-3 in Airway Epithelial Barrier Integrity During Influenza A Viral Infection

      Iqbal, Muddassar; Vasta, Gerardo R. (2023)
      Influenza A virus (IAV) infects the airway and alveolar epithelia in humans, and causes seasonal influenza annually which is a major global health concern, along with COVID-19, besides exhibiting pandemic potential. In severe cases IAV infection causes acute respiratory distress syndrome (ARDS) resulting from increased alveolar permeability due to the disruption of cell-cell tight junctions. The detailed mechanisms involved, however, remain to be fully elucidated. Galectins are β-galactose-binding lectins implicated in diverse cellular functions as well as in pathogen infections, including those from respiratory viruses. In previous studies from our lab on a murine model, IAV infection enhanced galectin-3 (Gal-3) secretion in the bronchoalveolar fluid. In the present study, I investigated in vitro the potential role of the secreted Gal-3 in airway epithelial barrier function during IAV infection. The results thus acquired using the human airway epithelial A549 cells indicate that IAV infection leads to a significant desialylation of the cell surface, exposing the sub-terminal β-galactose ligands, with a concomitant increase in the binding of recombinant galectin-3 (rGal3). I detected potential Gal-3 receptors CD147, integrin-β1 and MUC1 on the surface of A549 cells, and observed an increase in the secretion of matrix metalloproteinases MMP2 and MMP9 after exposure of these cells to rGal3. Furthermore, I observed disruption of the A549 cell surface distribution of tight junction proteins occludin and ZO-1 upon both IAV infection and Gal-3 exposure, as well as significant increase in the monolayer permeability. Using rGal-3, I demonstrated its direct interaction with the A549 cell CD147 as well as integrin-β1 which possibly mediated the above-mentioned cellular effects and validated some of the key findings in A549 cells from this study to the primary small airway epithelial cells (SAECs). In the end by searching for the sequence variants in the Gal-3 encoding genes LGALS3 in two public genetic databases, I found three rare variants that might alter protein function and one that was associated with the phenotypes indicative of a role for Gal-3 in influenza outcome: including influenza vaccine, flu treatment, acute sinusitis, and emphysema.
    • Microtubules as a therapeutic target in Duchenne muscular dystrophy

      Vanegas, Camilo; Ward, Christopher, Ph.D. (2023)
      Microtubules (MT) are dynamic polymers of tubulin protein whose post-translational modifications regulate MT interactions with other proteins, including intermediate filaments and actin. This integrated cytoskeletal network performs a vast array of cellular functions, including cell movement, transport of cellular cargo, defining cell shape and stiffness, and transmitting mechanical information to proteins that generate biological signals (mechanotransduction). My work is in Duchenne muscular dystrophy, where the absence of dystrophin leads to the proliferation of microtubules marked by an increase in posttranslational (PTM) modifications to their tubulin including detyrosination (deTyr-tubulin) and acetylation (acetyl-tubulin). The consequences of this change are increased cytoskeletal mechanics (i.e., stiffness) of the muscle fiber and increased mechanotransduction through NADPH Oxidase 2 (Nox2) dependent reactive oxygen species (ROS) and calcium signals during contraction that together have been implicated in the contraction injury that drives the pathology. My work addressed two open questions related to the pathobiology of these microtubule changes independent of dystrophic disease (Aim 1; Chapter 2) and the potential for targeting their reduction as a therapeutic option for halting or slowing disease progression (Aim 2; Chapter 3).
    • Biological, psychological, and sociocultural contributions to pain processing in the central nervous system: A whole-person approach to understanding chronic pain

      Cundiff-O'Sullivan, Rachel; Colloca, Luana (2023)
      Introduction: Chronic pain affects approximately 20% of the global population, yet effective treatments remain elusive. This study employs a biopsychosocial model, recognizing the importance of all facets of life, to investigate the complex interactions of biological, psychological, and sociocultural factors influencing pain outcomes and pain processing in the central nervous system. Methods: This project utilized functional magnetic resonance imaging to assess pain severity, interference, and endogenous pain modulation via placebo analgesia in a sample of participants with temporomandibular disorder. Brain-age difference was estimated via a pre-trained Gaussian Process Model using cortical thickness as a predictor of pain outcomes. Structural equation modeling and high dimensional multivariate mediation were employed to examine the influence of pain catastrophizing (PC) on pain outcomes. Differences in pain outcomes between religious/spiritual (R/S) and atheist participants were also explored. Results: TMD participants exhibited accelerated brain aging which was associated with pain severity and interference but not placebo analgesia. Structural equation modeling revealed that PC did not mediate pain outcomes or influence placebo analgesia. However, resting-state functional connectivity between the hippocampus and cuneal cortex mediated the relationship between PC and pain interference. Although there were no significant differences in behavioral outcomes between R/S and atheist participants, differential connectivity underlying these outcomes was found, such that atheist participants exhibited greater reliance on low-level sensory input compared to R/S participants who utilized higher cognitive regions for better emotional regulation of pain. Conclusion: This work provides valuable insights into the neurobiological mechanisms underlying how brain age, PC, and R/S factors influence chronic pain and placebo-induced pain reductions. By unraveling the complexities of pain processing using a whole-person approach, this research promotes a more holistic understanding of chronic pain and contributes to the development of more effective interventions for individuals suffering from chronic pain.
    • Immunomodulatory Nanoparticles as a Multimodal Approach to Attenuate Immune Dysregulation in Severe Inflammation and Sepsis

      Truong, Nhu; Pearson, Ryan M. (2023)
      Sepsis, a life-threatening condition triggered by an uncontrolled immune response to infection, currently lacks an FDA-approved therapeutic intervention to enhance patient survival. Severe inflammatory conditions can disrupt the balance of histone acetyltransferase (HAT)/histone deacetylase (HDAC) activity, leading to global cellular hypoacetylation. Histone deacetylase inhibitors (HDACi) restore acetylation profiles and reverse transcriptional silencing. Suberoylanilide hydroxamic acid (SAHA), a pan-HDACi, was modified by para-hydroxymethylation (termed SAHA-OH), which resulted in a favourable reduction in SAHA-associated toxicity under inflammatory lipopolysaccharide (LPS) challenge. SAHA-OH was incorporated into immunomodulatory nanoparticles (iNPs), previously developed by our lab, to form iNP-SAHA using a prodrug approach through the covalent modification with poly(lactic-co-glycolic acid) (PLGA). iNP-SAHA treatment significantly reduced proinflammatory cytokines in vitro and in vivo, improved the viability of LPS-stimulated primary macrophages, and enhanced survival of mice in an LPS-induced endotoxemia model. iNP-SAHA treatment did not significantly improved mice survival compared to the iNP treatment alone; however, the synergistic anti-inflammatory properties of iNP-SAHA are potentially promising for future exploration in alternative models of inflammatory disease. We evaluated the efficacy and cellular mechanism of iNP activity using a clinically relevant cecal ligation and puncture (CLP) murine model of polymicrobial sepsis. iNPs, when administered as an adjuvant to antibiotics, significantly improved survival compared to antibiotics alone. Interestingly, iNP treatment marginally affected local and systemic cytokine profiles, despite mitigating organ dysregulation. Minimal effects on immune cell proportions at local sites were observed, but iNP treatment normalized monocyte levels in peripheral blood and alveolar macrophages in lung tissues. Further studies enumerated that iNPs modulated cellular adhesion and migration surface marker expression as well as apoptotic levels on immune cells. These findings highlight the potential of iNPs as an adjunctive therapy for sepsis, particularly when combined with antibiotics, suggesting promising prospects for future clinical translation. Lastly, a high-throughput microfluidic approach for iNP formulation to enable facile scale-up was developed. We optimized the microfluidic method and the impact of polymer and surfactant concentrations, surfactant chemistry, flow rate ratio (FRR), and anti-inflammatory activity. This work demonstrated a controlled and reproducible microfluidic method for iNP formulation, showcasing their inherent anti-inflammatory properties and offering a promising avenue for inflammation management.
    • Conversion of Small-Molecule Inhibitors into Heterobifunctional Compounds in the Discovery of Novel Chemotherapeutics

      Chan, Alexandria; Fletcher, Steven (2023)
      Heterobifunctional polypharmacologic agents are compounds that have individual pharmacophores for at least two separate biological targets. Our work spans two distinct sets of heterobifunctional molecules: 1. Polypharmacologic agents that inhibit two proteins known to contribute to the disease state, and 2. Protein degraders: Proteolysis targeting chimeras (PROTACs) and molecular glues. Both types of protein degraders function through recruiting an E3 ligase to the protein of interest, resulting in a hijacking of the ubiquitin-proteasome system, and the subsequent destruction of the target protein. The use of type 1 compounds is rapidly growing as such polypharmacologic agents are postulated to exhibit distinct advantages over the monovalent, parent drug compounds from which they are constructed, including the ability to increase therapeutic effect, lower effective dosage, and circumvent treatment resistance. Type 2 compounds – the protein degraders – can eliminate a target of interest, requiring the cell to resynthesize the protein to regain its cellular function. These compounds may have a catalytic mechanism of action wherein the compounds are recycled after mediating the degradation of the target protein, thereby requiring non-stoichiometric amounts of drug while also directly countering resistance that manifests through target protein upregulation. Moreover, such degraders retain activity with resistant proteins where traditional, non-covalent small-molecule drugs fail. Due to these advantages, there is increasing enthusiasm that targeted protein degraders may herald a new class of anti-cancer therapeutics. Herein, our efforts towards the discovery of heterobifunctional pharmaceuticals for the treatment of drug-resistant hematological malignancies are described.
    • Novel Bioactive Low-Shrinkage-Stress Composite with Antibacterial and Remineralization Properties

      ALHUSSEIN, ABDULLAH; Xu, Huakun H. (2023)
      Methacrylate-based resin composites are frequently employed in dentistry for their aesthetic qualities, durability, and adhesive properties. Nevertheless, these restorations generally exhibit a lifespan of 5 to 10 years, with recurrent caries and tooth fractures being primary failure factors. Marginal integrity and the absence of bioactivity at the tooth-restoration junction contribute to recurrent caries development. Consequently, this dissertation endeavors to introduce a novel bioactive low-shrinkage-stress nanocomposite, featuring dimethylaminododecyl methacrylate (DMADDM) as an antibacterial agent, as well as remineralization nanoparticles of calcium fluoride (nCaF2) and nanoparticles of amorphous calcium phosphate (NACP), with the potential of increase the longevity of dental restoration and protect tooth structure. All novel formulations of low-shrinkage-stress composite were subjected to a series of mechanical, antibacterial, cytocompatibility, and ion release assessments. First, we investigated the optimum concentration of DMADDM that can be incorporated with a low-shrinkage-stress composite without compromising mechanical properties. We found that incorporation of up to 5% DMADDM into a low-shrinkage stress composite efficiently inhibited Streptococcus mutans (S. mutans) biofilm commonly associated with secondary caries. This potent antibacterial effect is achieved while maintaining excellent mechanical properties and minimizing polymerization shrinkage stress, potentially improving the long-term success of dental restorations. Next, we investigated the antibacterial and cytocompatibility of the incorporation of 3% DMADDM with 20% nCaF2 or 20% NCAP into a low-shrinkage-stress nanocomposite. We found that incorporating DMADDM with either nCaF2 or NACP into a low-shrinkage-stress nanocomposite provides a potent antibacterial effect against S. mutans biofilm while maintaining excellent mechanical properties. In addition, the novel formulations demonstrated excellent biocompatibility against human gingival fibroblasts and dental pulp stem cells. Lastly, we investigated the ions release and antibacterial properties against a salivary biofilm for our innovative formulations. The innovative mixture of DMADDM, NACP, and nCaF2 demonstrated strong antibiofilm effects on salivary biofilm, while concomitantly releasing a significant amount of remineralizing ions. This nanocomposite is a promising dental material with antibiofilm and remineralization capacities, with the potential to reduce polymerization-related microleakage and recurrent caries.
    • Utilizing Pharmacometrics to Facilitate Generic Drug Development of Orally Inhaled Products and Optimize Pharmacotherapy of Antifibrinolytics

      Li, Shuhui; Gobburu, Jogarao (2023)
      This thesis has two parts. The first part is related to the pharmacokinetic (PK) batch-to-batch variability of orally inhaled products, which may pose challenges for generic product development. I applied the techniques of pharmacometrics to propose and evaluate alternative PK bioequivalence (BE) study designs using Advair Diskus as an example product, aiming to facilitate generic development. First, population PK models for Advair Diskus were developed and qualified to simulate PK BE study. Next, the effect of batch-to-batch variability on the establishment of BE was evaluated using the developed models. Batch-to-batch variability substantially elevates the probability of reaching a false conclusion in a PK BE study for equivalent and inequivalent comparisons. Therefore, ignoring batch-to-batch variability when presenting will increase the risk of either patients being treated with an inequivalent formulation or pharmaceutical companies not obtaining approval for an equivalent formulation. This calls for alternative PK BE approaches to account for the batch-to-batch variability. I proposed and evaluated a two-phase study framework that uses a pilot study to select reference and test batches for the pivotal BE study. A parallel design with ≥ 12 patients per sequence or a crossover design with ≥ 6 patients per sequence is recommended for the pilot study design. The proposed criteria for selecting batches based on the pilot study results include (1) 0.9 ≤ T/R ≤ 1.11 and (2) higher conditional power. The two-phase study design offers the flexibility to select batches in a PK study to minimize the impact of batch-to-batch variability on the generics development. The two-phase framework might be applied to other products with similar characteristics and high batch-to-batch variability in the reference products. The second part of this thesis used pharmacometrics to optimize the pharmacotherapy of an anti-fibrinolytic, tranexamic acid (TXA), in special patient populations. The PK and pharmacodynamics (PD) of TXA in special populations are understudied; therefore, the PK/PD-driven optimal doses for them are unknown. First, I characterized the PK and PD of TXA in pregnancy and found that pregnant women have up to 30% higher clearance and volume of distribution than the general non-pregnant population. A dose of 650 mg maintains both PK and PD targets for > 1 hour in most patients, which is recommended as the postpartum prophylactic dose for future confirmatory clinical studies. In addition, I evaluated a current dosing regimen for cardiac surgery patients who use cardiopulmonary bypass (CPB) during their surgeries from a PK perspective. This dosing regimen consists of a long infusion of TXA at 100 mg/hr for 5 hours before CPB initiation, a 1 g bolus of TXA at CPB initiation, and another 1 g bolus at the end of CPB. While kidney function affects the clearance of TXA, and the CPB procedure increases the volume of distribution of TXA, the current dosing regimen was confirmed to provide sufficient TXA exposure (15 mg/L) from CPB initiation till 3 hours post-CPB, achieving the therapeutic goal. Both studies contribute to understanding how TXA dosing can be optimized in special patient populations.
    • Regulation of Epidermal Skin Immunity by a Tick Bite

      Marnin, Liron; Pedra, Joao H. F. (2023)
      Hard ticks are hematophagous arthropods of public health and veterinary importance. Following a tick bite, these arthropods take prolonged, continuous bloodmeals that facilitate pathogen transmission. Successful bloodmeals are attributed to components of the tick saliva that alter inflammation, inhibit hemostasis, and block pain and itch responses in the mammalian skin. Recent studies have reported that extracellular vesicles (EVs) in the saliva enable tick feeding and redirect skin immunity. Notably, the skin epidermis, which interfaces with the external environment, has been mostly neglected when studying the relationship between ticks and their mammalian hosts. Here, we report that Ixodes scapularis EVs enable tick feeding by affecting epidermal γδ T cells frequency and co-receptor expression at the skin site. Epidermal γδ T cells have a critical role in wound healing and interact with keratinocytes, which comprise 95% of the epidermal layer. We further coupled entomological approaches with flow cytometry, single cell RNA sequencing, and animal strains devoid of epidermal γδ T cells to demonstrate that tick EVs disrupt networks involved in keratinocyte proliferation, suggesting an effect on epithelium wound repair. Collectively, this work broadens our knowledge of ectoparasitology, vector-host interactions, and principles of immunology.