Now showing items 1-20 of 2007

    • Characterizing enhancer-driven transcriptional networks in schizophrenia

      Casella, Alex; Ament, Seth A. (2021)
      Genetic studies of schizophrenia have demonstrated that more than 90% of genetic risk is confined to non-coding portions of the genome. Advances in chromatin state prediction and chromatin accessibility assays have enabled us to better characterize the genomic features making up these regions and annotate genetic risk to these elements. The focus of this dissertation is to understand the role that tissue- and cell type- specific regulatory elements and the transcription factors that bind them play in risk for schizophrenia. I hypothesized that enhancer-based transcription factor-target networks that direct neuronal development are disrupted in schizophrenia. To test this hypothesis, I used high-quality chromatin state predictions in both the developing and the adult brain to develop a framework for testing enhancer properties for association with genetic risk. Any enhancer-level annotation can be used in this type of test, including transcription factor binding counts and chromosomal contact information. I first described and validated an atlas of transcription factor binding sites across multiple human tissue, including the brain. I then used this atlas to show that neurodevelopmental transcription factors and target genes are most associated with risk for developing schizophrenia.
    • Crossing the Chasm: A Pilot Study for Preparing PT’s for Telehealth through IPE and Simulation

      Gordes, Karen L.; Retener, Norman F.; Lee, Mei Ching W.; Horn, Linda B. (2021-10-23)
    • Altering Mechanisms of Frailty in Persons Living with HIV

      Nelson, Amy; Klinedinst, N. Jennifer (2021)
      Background: People with HIV experience frailty more often and earlier than others. Little is known about mechanisms driving early frailty in HIV. There are a lack of effective interventions for frailty in HIV. This study explored the mechanisms of musculoskeletal frailty in people living with HIV and the influence of baseline activity after a six-week aerobic exercise intervention. Methods: A literature review developed an adapted conceptual model for musculoskeletal frailty in HIV for the first manuscript. Due to COVID-19 restrictions, a secondary data analysis utilized the baseline activity measure (Yale Physical Activity Survey) from 11 healthy participants who completed a six-week moderate paced walking program, aged 50 to 65. Cellular energy production and inflammation markers were available pre- and post-intervention. Correlation with baseline activity was assessed using Kendall’s tau-b. Results: Mechanisms of musculoskeletal frailty in people living with HIV include increased inflammation, dysregulated energy metabolism, immune activation, and endocrine alterations. Aerobic exercise has the potential to moderate each of these. The relationship between baseline activity and changes in cellular energy metabolism was not statistically significant. However, strong positive associations were noted between body mass index and change in platelet spare respiratory capacity, the ability of mitochondria to produce more energy upon demand. In examining the effect of baseline activity on inflammatory markers, no significant relationships were found, and no markers showed significant change. Conclusion: Moderate walking did not make significant changes in inflammation after a six-week moderate paced walking intervention. Baseline activity levels did not play a significant role in the change of either inflammation or cellular energy production. This may be because healthy participants did not have impaired levels of inflammation or cellular energy metabolism at baseline. This study should be repeated in people living with HIV who have altered inflammation or cellular energy metabolism.
    • Genomic Medicine in Diabetes: Improving the Diagnostic Rate of Monogenic diabetes

      Zhang, Haichen; Pollin, Toni; 0000-0002-0615-2836 (2021)
      Monogenic diabetes is an uncommon type of diabetes caused by genetic defects in one of several genes, and it accounts for 1-2% of all diabetes. The primary subtypes are Maturity Onset Diabetes of the Young (MODY), neonatal diabetes, and syndromic diabetes. The correct treatment of each subtype of monogenic diabetes depends on the corresponding disease etiology that can only be confirmed by genetic testing. However, the diagnostic rate of monogenic diabetes is inadequate, mainly due to the overlapping phenotype of monogenic diabetes with type 1 diabetes and type 2 diabetes and lack of awareness among patients and physicians. To improve the diagnostic rate of monogenic diabetes, this project focuses on three aspects: 1) systematically screening of patients for genetic testing; 2) comprehensively re-analyzing next-generation sequencing (NGS) data from multiple diabetes cohorts; 3) assessing the ability of Direct-to-Consumer Genetic Testing (DTC-GT) raw data in detecting GCK-MODY variants. The Personalized Diabetes Medicine Program (PDMP) screened 2,522 patients with diabetes with a simple questionnaire, assigned patients to different algorithm criteria groups based on clinical features, and performed genetic testing on suspected patients. Overall, 38 of 313 patients suspected of monogenic diabetes were tested positive for causative variants. The group of patients diagnosed before age 30 who were not treated with insulin had the highest pick-up rate. The comprehensive re-analysis of NGS panel data in PDMP, including re-classification and updating variant calling algorithm, improved the diagnostic rate from 11.82% to 13.10%. Also, the comparison between exome sequencing (ES) and NGS panel or Sanger sequencing of the Progress for Diabetes Genetics in Youth samples showed ES failed to identify all MODY-causing variants, but re-analysis of ES unfiltered data discovered the missing variants. By analyzing the GCK variants in the 23andMe DTC-GT raw data from 3,044 anonymous volunteers and calculating the ancestry-specific allele frequency of GCK-MODY variants, some of the variants showed higher-than-expected minor allele frequency compared with the large population database. Such inconsistency suggests customers should not use DTC-GT as a supplementary method of clinical genetic testing for GCK-MODY. In a summary, these studies provide practical approaches to improve the diagnostic rate of monogenic diabetes.
    • Role of the Parabrachial Complex in Rodent Models of Pain

      Uddin, Olivia; Keller, Asaf; 0000-0001-7035-6836 (2021)
      The parabrachial complex (PB) is a midbrain structure that is vital to survival-related functions. This region receives and integrates incoming sensory information, communicating these signals to higher brain regions that are key in shaping behavior and affect. PB responds to painful somatosensory input, however, its most compelling role is in the development and maintenance of persistent pain. Pain persisting beyond the duration of a threat or tissue injury no longer serves a physiological purpose. Therefore, this type of maladaptive pain has a severe impact on health and quality of life. Current therapies for maladaptive pain are not optimal: it is necessary to better understand the neural circuitry underlying persistent pain so that we can design more effective therapies. The work presented here aims to outline PB’s role in four rodent models of pain. I approach this by combining anatomical tracing, electrophysiology, and behavioral studies. First, I show that in a model of orofacial neuropathic pain, PB neural activity is amplified. Next, I describe a novel and direct anatomical connection between the meninges and PB, via the trigeminal ganglion; this pathway is poised to underlie migraine headache-associated pain. Finally, I conduct behavioral studies aiming to hone rodent models of pain in the context of aging and amyloid accumulation, and pain during opioid withdrawal. These findings together confirm that PB is a crucial node in maladaptive pain processing and provide direction for further work clarifying PB’s role in new behavioral contexts.
    • Individual, Caregiver, and Family Characteristics Associated with Obesity in Preschool-age Children

      Rahmaty, Zahra; Johantgen, Mary E.; 0000-0001-6165-0881 (2021)
      Background and Objectives: Eating habits start from early childhood and may contribute to the development of obesity. Food neophobia (FN) occurs among 50% of preschoolers and has shown inconsistent associations with obesity. Caregiver feeding practices (FPs) influence eating habits but have limited evidence about how they employ together and how they associate with childhood obesity. The first paper examines the relationship between FN and preschooler's obesity/overweight. The second paper assesses patterns of FPs and their associated factors. The third paper examines how the patterns of FP relate to preschoolers' Body Mass Index (BMIz), an objective measure of obesity. Method: Data from the Creating Healthy Habits Among Maryland Preschoolers (CHAMP) study including preschoolers (N=500) and caregivers from 50 Maryland childcare centers were examined. Children's weight and height were measured, and BMI percentile and z-score were calculated. Caregivers reported demographics, weight and height, FN, FPs, child temperament via an online survey. Mixed models, factor analysis, latent profile analysis, and structural equation models were used. Results: A quarter of children were obese/overweight; caregiver-reported FN was not associated with preschoolers' obesity/overweight, although children were more likely to be obese/overweight if their caregiver was overweight (aOR=2.6) or obese (aOR=3.9). Three patterns of FP were found. Controlling class had high coercive control and low autonomy practices (69%), Regulating class had high coercive control, but moderate structural and autonomy practices (16%), and Balancing class were moderate in all practices (15%). Caregivers who desired their child to be heavier (aOR=0.40, 95%CI=0.22-0.72), had higher poverty levels (aOR=0.80, 95%CI=0.65-0.98), were single (aOR=0.38, 95% CI=0.18-0.80), and were less likely to be in the Balanced versus Controlling class. Children’s difficult temperament (b=0.09, p=0.008), caregiver’s BMI (b= 0.26, p<0.001), desire for the child to be thinner (b=0.23, p<0.001), desire for child to be heavier (b=-0.37, p<0.001), and Regulating versus Controlling FP (b=-0.09, p=0.03) were associated with child BMIz. Conclusion: Childhood obesity is a multifactorial phenomenon, with interactive effects among the child, family, and environment. FP are associated with preschooler’s weight and should be assessed comprehensively. Caregivers’ perceptions of child size and temperament may also provide insight into FP and obesity.
    • Staff-resident Interactions in Assisted Living: Optimizing the Quality of Daily Care Interactions

      Paudel, Anju; Galik, Elizabeth; Resnick, Barbara; 0000-0002-1784-5427 (2021)
      Background: A considerable amount of research has focused on understanding and improving staff-resident interactions in long-term care. Much of this work has focused on social communications between staff and residents in nursing home settings. Attention to care interactions in assisted living (AL) is lacking. Purpose: The purpose of this dissertation was to: (1) describe the staff-resident interactions in AL; (2) explore the resident and facility factors associated with the care interactions in AL; and (3) test the feasibility and preliminary efficacy of the Promoting Positive Care Interactions (PPCI)—a four-step intervention designed to establish positive care interactions between the staff and residents with cognitive impairment or dementia in AL. Methods: Utilizing baseline data in a randomized trial that included 379 residents from 59 AL facilities, aim 1 used descriptive statistics to describe the quality of staff-resident interactions in AL and aim 2 used stepwise regression to examine factors influencing interactions. Aim 3 involved pilot-testing of PPCI intervention in one AL community in Maryland using a single group pretest-posttest design. Feasibility was demonstrated with the evidence of delivery, receipt, and enactment of PPCI. Preliminary efficacy was evaluated with repeated measures ANOVA for staff outcomes and descriptive change in summary scores for facility outcomes. Results: Although majority of the interactions observed were positive, almost 25% were negative and neutral suggesting a need to improve the interactions in ALs. Factors influencing interactions included resident agitation and facility ownership which accounted for 8.2% of variance. Additionally, PPCI was implemented as intended with 100% staff exposure to education and considerable staff engagement in mentoring sessions. While there was an improvement in AL environment and policy, no significant changes were observed in staff outcomes post PPCI. Conclusions: Understanding the quality of staff-resident interactions in AL and the factors that influenced these interactions guided the development of PPCI. Pilot testing supported the feasibility and preliminary staff adoption of PPCI in ALs. PPCI will be further tested with a randomized trial, and a hybrid model with both online education and in-person mentoring and coaching of staff to improve staff knowledge and behavior related to care interactions.
    • Development of New Formulations of nCaF2 Dental Nanocomposites with Antibacterial and Remineralizing Properties

      Mitwalli, Heba; Weir, Michael D.; 0000-0003-3877-6632 (2021)
      It is desirable to use a minimally invasive approach in dentistry through conservative techniques in order to prevent destruction of the tooth structure. Since resin-based dental materials are increasingly used, the occurrence of re-infections is also increasing. Recurrent caries and secondary infections are major problems in the restorative dentistry field. The prevalence of recurrent caries associated with resin-based restorative materials was previously shown to reach 60%. It is the most common reason recognized for composite resin restorations replacement and failure. Many efforts have been made to incorporate antibacterial agents into restorative materials. However, the majority of these materials act by releasing these agents into the surrounding environment, leading to their depletion over time. There is a clinical need for durable bioactive composite restorations that resist the formation of secondary caries for an extended period of time. Dimethylaminohexadecyl methacrylate (DMAHDM) is an antibacterial agent that is immobilized in resin and not lost or released with time. Therefore, this dissertation aims to develop new composite resin formulations containing DMAHDM antibacterial, 2-methacryloyloxyethyl phosphorylcholine (MPC) protein repellent, and nanoparticles of calcium fluoride (nCaF2) remineralizing modalities which could be a promising approach for management of recurrent caries around or under restoration margins. This dissertation incorporated DMAHDM, MPC and nCaF2 into composite resin restorations to achieve potent, long-lasting antibacterial, protein repellent, and Ca and F ion release and recharge/re-release capabilities. Mechanical testing was performed for all composite formulations. To determine biofilm properties, a human salivary microcosm biofilm model was used. Biofilm colony-forming units (CFU), minimum inhibitory concentration, lactic acid production, and metabolic activity of biofilm were investigated. Fluoride (F) and calcium (Ca) initial ion releasing, recharging and re-releasing capabilities were tested. The majority of nCaF2 nanocomposites show matching mechanical properties to the commercial control composite. The nCaF2-DMAHDM nanocomposites have potent antibacterial effects that substantially reduce biofilm activities in all biofilm experiments. Similarly, all nCaF2 nanocomposites have higher values of F and Ca ion release- recharge, and re-release when compared to the commercial control composite. Therefore, these new composite resin formulations may potentially lead to a fundamental contribution in restorative techniques that can be used to fight this most common limitation of composite restorations - recurrent caries - and contribute to the longevity of composite restorations through long-lasting antibacterial and protein repellent properties and remineralization capabilities.
    • Role of Adipose Lipolysis in Development of Alcoholic Liver Disease

      Mathur, Mallika; Yu, Liqing (2021)
      Alcoholic liver disease (ALD) pathologies include hepatic steatosis, inflammation and liver injury. The liver receives ~60% of fatty acids from adipose tissue. We examined the role of adipose lipolysis in ALD pathogenesis using adipose-specific CGI-58 knockout (FAT-KO) mice, a model of impaired adipose lipolysis. FAT-KO versus control mice were almost completely protected against ethanol-induced hepatic steatosis and lipid peroxidation when subjected to the 15-day NIAAA chronic-binge ethanol diet. This was unlikely from reduced lipid synthesis because ethanol feeding downregulated hepatic expression of lipogenic genes similarly in both genotypes. On a control diet, FAT-KO relative to control mice had increased hepatocyte injury, neutrophil infiltration, and activation of transcription factor STAT3 in the liver, none exacerbated by ethanol. This was associated with increased hepatic leptin receptor mRNA and adipose inflammatory cell infiltration. These findings identify a critical role for adipose lipolysis in hepatic steatosis and oxidative stress during ALD development.
    • Neuronal Intrinsic Apoptosis Mechanisms and Their Modulation by Sp1 Inhibition

      Makarevich, Oleg; Stoica, Bogdan A.; 0000-0003-2061-2285 (2021)
      Drug-induced DNA damage, reactive oxygen species (ROS), inflammatory mediators and central neurotoxicity have all been shown to play a role in cognitive impairments after chemotherapy, after CNS injury or in neurodegenerative diseases. Intrinsic apoptosis is a regulated cell death pathway implicated in many of these conditions. This pathway proceeds sequentially through DNA damage responses, including phosphorylation of ATM, H2AX and Tumor Protein 53 (p53), transcriptional activation of pro-apoptotic BH3-only proteins, and mitochondrial outer membrane permeabilization (MOMP), activating caspase-dependent and caspase-independent apoptosis. Previous work has indicated that Sp1 may regulate p53’s transcriptional profile and ability to promote apoptosis after DNA damage. To examine the role of Sp1 in DNA-damage-induced apoptosis, my work has utilized Mithramycin, a drug that binds G-C rich DNA to compete with Sp1 chromatin binding. Our hypothesis is that Mithramycin competes with Sp1 chromatin binding and thus protects neurons from DNA damage-induced, p53-dependent intrinsic apoptosis. However, it is vital to recognize that neurons increase their resistance to cytotoxic stimuli as they mature, leading to decreased levels of apoptosis. Thus, we additionally characterized the changes in the intrinsic apoptosis pathway upon neuronal maturation. Despite significant attenuation of the intrinsic apoptosis pathway in mature neurons, we found evidence that Mithramycin can still attenuate DNA-damage dependent intrinsic apoptosis in this paradigm to the extent that it is induced by neurons. Additional experiments showed that mature animal cortices or hippocampi do not demonstrate evidence of end-stage apoptosis (caspase activity) in response to DNA damage, unlike those in immature animals. However, in vivo administration of Mithramycin was able to attenuate cortical expression of p53-dependent genes. Thus, our studies provide evidence that: 1) Mithramycin attenuates activation of the DNA damage-dependent intrinsic apoptosis pathway via indirect inhibition of p53-dependent transcription and 2) Mature neurons restrict intrinsic apoptosis pathway activation in response to DNA damage. Therefore, Sp1 is likely to play a significant role in DNA damage-dependent neuronal intrinsic apoptosis, and Mithramycin treatment is protective when this pathway is activated. However, mature neurons significantly downregulate this pathway, thus limiting the potential therapeutic effects of Mithramycin in adult animals.
    • Modulation of Inflammation and Stromal Remodeling Processes in Autoimmune Arthritis by Microbial Indole Derivatives

      Langan, David; Moudgil, Kamal; 0000-0002-4525-9767 (2021)
      Rheumatoid arthritis (RA) is an autoimmune disease characterized by chronic inflammation of the synovial tissue that can lead to joint damage and deformities. Inflammation, new blood vessel formation (angiogenesis), and bone resorption (osteoclastogenesis) are three key processes of the pathophysiology of RA. ‘Dysbiosis’ of the gut microbiota is implicated in RA pathogenesis because it can cause an imbalance in the microbial metabolites that regulate host health and disease. However, there is little information about the impact of two such indole derivatives, indole-3-aldehyde (IAld) and indole-3-acetic acid (I3AA), on arthritis-related processes. Using established cell-based models and the adjuvant-induced arthritis animal model, we conducted a comparative analysis of IAld and I3AA to understand how these metabolites might impact RA pathogenesis. To our surprise, despite their structural similarities, the bioactivities of these two metabolites were profoundly different. IAld, but not I3AA, altered the expression of genes encoding arthritis-associated cytokines (IL-1β, IL-6, VEGF) in RAW 264.7 (murine macrophage) cells stimulated with heat-killed M. tuberculosis. Further investigation of this anti-inflammatory activity of IAld suggested that inhibition of the MyD88-dependent activation of NF-κB and MAPK pathways was unlikely to be involved. IAld also exhibited pro-osteoclastogenic and pro-angiogenic activity. In contrast, I3AA exhibited only anti-angiogenic activity. Both IAld and I3AA are proposed agonists of the aryl hydrocarbon receptor (AhR). However, AhR inhibitor CH-223191 suppressed the anti-angiogenic activity of I3AA, but failed to mitigate any of the effects of IAld. There is a cross-talk between the AhR and Nrf2 pathways, and some plant-derived phytochemicals are multifunctional ligands of both pathways. Our findings show that IAld, unlike I3AA, can suppress the Nrf2-dependent antioxidant response of macrophages to an Nrf2 agonist, but that IAld also potentially reduces intracellular ROS levels during osteoclast differentiation. Furthermore, oral administration of IAld to rats resulted in the reduction of arthritis severity compared to the arthritic control group. Taken together, our findings suggest that the relative bioavailability of these microbial indole derivatives has the potential to influence their immunomodulatory effects in healthy individuals as well as patients with RA.
    • Monomeric IL-12p40 binds partner proteins to modulate immune cell function

      Gerber, Allison; Singh, Nevil; 0000-0001-9442-3255 (2021)
      Cytokines are critical mediators used by immune cells to communicate as well as protect. The IL-12 family of cytokines are made up of  and  subunits typically assembled within one cell and secreted as a heterodimer. IL-12p40 is the shared β-subunit for both IL-12 (paring with IL-12p35) and IL-23 (with IL-23p19). However, the IL-12p40 monomer is often secreted in excess during infections, but its biological role was not known. In this thesis we investigated the function of secreted IL-12p40 monomer in vivo with the central hypothesis that the monomer combines with multiple α-subunits in vivo to generate IL-12 as well as other heterodimeric cytokines. Consistent with this hypothesis, in chimeric mice containing mixtures of cells that can only express either IL-12p40 or IL-12p35, but not both together, we found that functionally active IL-12 was generated. This alternate two-cell pathway requires IL-12p40 from hematopoietic cells to extracellularly associate with IL-12p35 from radiation-resistant cells. The two-cell mechanism was sufficient to influence local T cell differentiation in sites distal to the initial infection and helped control systemic dissemination of a pathogen although not parasite burden at the site of infection. Broadly, this suggests that early secretion of IL-12p40 monomers by sentinel cells at the infection site may help prepare distal host tissues for potential pathogen arrival. In addition to this role in generating IL-12 through two-cell assembly, we found that IL-12p40 has a novel partner protein, CD5L. This novel heterodimer was present in the serum of uninfected mice, with differences in the basal levels between B6 and Balb/c animals, with Balb/c having higher amounts of p40-CD5L. Functionally, we found that treatment with p40-CD5L leads to IL-4 and IL-10 production by T cells. Taken together, this thesis offers at least two major fundamental advances in cytokine biology – one the concept of a two-cell assembled cytokine and second the identity of a novel TH2-promoting heterodimeric cytokine. The first has significance in immunotherapy and understanding immunity to tissue-specific modulation of immune responses. The second is expected to drive significant research on allergy, responses to parasites and immune deviation.
    • The Molecular Basis of IL-1 Family Signaling: Strategies to Modulate Inflammation

      Fields, James; Sundberg, Eric J.; 0000-0001-5923-108X (2021)
      Interleukin-1 (IL-1) family cytokines are potent signaling molecules that influence both innate and adaptive immune systems. The IL-1 family, composed of 11 cytokines and 10 receptors, mediate inflammation to a wide array of stimuli and act on myriad cell types for diverse immunological outcomes. Altogether, IL-1 family signaling is integral to a multitude of inflammatory responses and occurs in distinct steps. First, an agonist cytokine binds its cognate receptor at high affinity. Next, this cytokine-receptor complex recruits an often-shared co-receptor. As this cytokine/receptor/co-receptor complex forms, Toll/IL-1 Receptor (TIR) domains, residing cytoplasmically, oligomerize, initiating a potent signaling cascade that results in prototypical NF-κB signal transduction. Due to the strong nature of IL-1 family signaling, multiple physiological mechanisms exist to stem this inflammatory signal, including antagonist cytokines and decoy receptors. Within the IL-1 family, the cytokines and receptors can be further divided into four subfamilies dependent on their secondary receptors. The IL-1 subfamily contains IL-1, IL-33, and IL-36 as they all share IL-1RAcP as their secondary receptor; the IL-18 subfamily is distinct as it utilizes IL-18Rβ as its secondary receptor. Here, we describe how structural biology has guided our understanding of IL-1 family signaling and how that knowledge can be leveraged for the design of therapeutics to stem aberrant cytokine signaling. In our first study, we demonstrate the feasibility of targeting a shared co-receptor, IL-1RAcP, for selective cytokine inhibition. Indeed, dependent on the specific epitope targeted on IL-1RAcP, differential cytokine signaling inhibition can be achieved. In addition, we developed our own IL-33 therapeutics by leveraging the high affinity IL-33 has for its primary receptor, the stability imparted by the secondary receptor, and the extended half-life gained through an Fc-fused receptor. Two of these molecules inhibit IL-33 signaling better than the natural antagonist sST2. Altogether, structural biology has informed our understanding of IL-1 family signaling, generated approaches to improve existing therapeutics, namely antibody epitope targeting, and led to the creation of additional IL-33 target therapeutics in the form of our “cytokine traps.”
    • The Association of Antiretroviral Treatment and Early Menopause in Women Aging with the Human Immunodeficiency Virus

      Bozzi, Laura; dosReis, Susan; 0000-0001-5326-6521 (2021)
      Women living with HIV (WLWH) have irregular menses with several periods of prolonged amenorrhea but their risk of early menopause, clinically defined as before age 45 years, is unknown. This is largely because there is no gold standard method to confirm menopause. Antimullerian hormone (AMH) is a biomarker indicative of ovarian reserve; however, no prior study has used this to confirm menopause. This study aimed to 1) confirm menopause using AMH; 2) determine if WLWH are at an increased risk of early menopause compared to at-risk, uninfected women; and 3) evaluate the relationship between time-varying ART use with early menopause in WLWH. Data were derived from the Women’s Interagency HIV Study, which had four enrollment waves from 1994 through 2016 across 11 US clinic sites. Women were followed prospectively from their baseline visit until menopause confirmation, loss to follow up, or end of study (12/31/2018), whichever came first. The study cohort was women ages 18 or older with no history of: menopause; hysterectomy/uterine cancer/double oophorectomy; any type of cancer, except skin cancer. Women were censored if they experienced any aforementioned events in follow-up. The study measures confirm menopause were at least 12 months of amenorrhea without resumption of menses and an undetectable AMH (<0.10ng/mL). Age at menopause was determined upon confirmation of final menstrual period. A Cox Proportional Hazards model determined the risk of early menopause among WLWH relative to at-risk uninfected women. Marginal Structural Cox Proportional Hazards models with stabilized weights estimated the effect of >75% adherence to ART, modeled as a time-varying covariate, on the risk of early menopause. Age at confirmed menopause with undetectable AMH was 48.6±4.3 years as compared to 41.2±5.6 years for women with amenorrhea without menses resumption and detectable AMH. WLWH reached menopause at significantly earlier ages and had a two-fold increased risk of experiencing early menopause than at-risk, uninfected women. There was a non-significant protective effect of ≥75% ART adherence on early menopause. AMH can improve the precision in determining age of menopause, which is an integral part of understanding the risk for early menopausal and future planning for postmenopausal care in WLWH.
    • Behavioral symptoms associated with dementia and inappropriate psychotropic medication use in U.S nursing homes

      Yoon, Jung Min; Trinkoff, Alison M. (2021)
      Background: Behavioral symptoms associated with dementia often occur concurrently. Psychotropic medications are used to treat behavioral symptoms in nursing homes (NHs) despite limited efficacy and the risk of adverse effects. Psychotropics are considered an easier solution for behavioral symptoms with fewer nursing staff. Inappropriate psychotropic medication use has been the focus of policy attention due to safety issues. A NH deficiency of care can be cited for inappropriate psychotropics use (F-758 tag). Purpose: The purposes of this dissertation are to examine factors of co-occurring behavioral symptoms of dementia (Aim 1), the occurrence of F-758 tag citations in relation to nurse staffing (Aim 2) and to explore how NH deficiency citations describe inappropriate psychotropics use to manage behaviors (Aim 3). Methods: For aim 1, general linear mixed models were used to explore co-occurring behavior symptoms in relation to cognitive status, physical function and analgesics use among 336 NH older adults diagnosed with dementia. For aim 2, generalized linear mixed models estimated associations between the occurrence of F-758 tags and nurse staffing levels among 13,614 NHs from December 2017 to 2018. Aim 3 used a mixed-method study design that combined descriptive and content analysis of F-758 deficiency reports (n=444 NHs) during January to March 2018. Results: Having multiple behavioral symptoms was negatively associated with better cognitive status and regular analgesics use (p<.001 and p=.009, respectively) (Aim 1). NHs with greater hours per resident day for RNs (OR=0.54, 95% CI=0.44-0.67), certified nurse assistant (OR=0.87, 95% CI=0.77-0.99), total nurse staff (OR= 0.87, 95% CI= 0.79-0.96), and greater RN skill-mix (OR=0.10, 95% CI=0.04-0.26) had significantly lower odds of F-758 tags (Aim 2). Common reasons for inappropriate psychotropic medication use included failure to monitor behavioral symptoms (178 NHs), attempt gradual drug reduction (131 NHs) and maintain14 day limits on PRN psychotropic medication orders (121 NHs) (Aim 3). Conclusions: Consideration of cognitive function and pain management are important for multiple behavioral symptoms (Aim 1). NHs need to have adequate nurse staffing levels to reduce inappropriate psychotropic medication use (Aim 2). Aim 3 analysis suggests areas for improvement, that could potentially reduce F-758 citations.
    • Mechanisms of Beta Cell Metabolic Coordination

      Rao, Vishnu; Rizzo, Megan A.; 0000-0002-4396-1579 (2021)
      Glucose homeostasis is predominantly regulated by pancreatic hormones. Insulin, which is secreted by pancreatic beta cells, is vital for maintaining normoglycemia; insulin secretory failure is a prime contributor to diabetes progression. The insulin secreting beta cells are a heterogeneous population that are organized into islets, which display coordinated responses to glucose. Although the electrical coupling of beta cells is well described, upstream metabolic coupling has not been sufficiently explored. The goal of this research was to (1) determine if beta cells are metabolically coordinated prior to reaching electrical threshold potential, (2) confirm evidence of metabolic heterogeneity, and (3) examine the importance of this heterogeneity. To accomplish this goal, we used a variety of novel techniques and models. We restricted application of glucose to portions of the islet to show beta cells are metabolically coordinated by gap junctional diffusion of metabolites. Then, using mice expressing a plasma membrane ChR2, we demonstrated that beta cell metabolism is differentially regulated by calcium. Finally, we generated a novel mouse model expressing an anisotropic reporter of glucokinase activity to illustrate that differences in glucokinase activity are averaged by metabolic coupling. These results highlight the importance of metabolic coordination in evening out differences between cells. Our findings may explain how insulin secretion is enhanced in the presence of glucose but contained in its absence.
    • The Role of Testisin in Endothelial Cells and Angiogenesis

      Peroutka, Raymond; Antalis, Toni M.; 0000-0002-8639-5773 (2021)
      Testisin (PRSS21) is a membrane anchored serine protease, which is tethered to the cell surface via a glycosylphosphatidylinositol (GPI)-anchor. Testisin expression has been documented in eosinophiles, ovarian cancers, endothelial cells, and spermatozoa where its expression is highest. Although two substrates of testisin have so far been identified, protease activated receptor 2 (PAR2) and protein inhibitor C (PCI), little is known of the biological, physiological, and pathophysiologic characteristics of testisin. Thus far, there are no published data identifying an activator or inhibitor of testisin. To better characterize the biochemistry of testisin we produced the zymogen as inclusion bodies in E. coli and refolded using the insoluble cellular fraction. To better characterize the cellular functions of testisin, hybridomas producing anti-testisin monoclonal antibodies were acquired, antibodies purified, and then characterized. In an investigation of testisin’s function in endothelial cells we identified testisin as a novel regulator of physiological hormone-induced angiogenesis and microvascular endothelial permeability. Using a murine model of rapid physiological angiogenesis during corpus luteal development in the ovary, we found that mice genetically deficient in testisin (Prss21-/-) show a substantially increased incidence of hemorrhages which are significantly more severe than in littermate control Prss21+/+ mice. This phenotype was associated with increased vascular leakiness, demonstrated by a greater accumulation of extravasated Evans blue dye in Prss21-/- ovaries. Live cell imaging of in vitro cultured microvascular endothelial cells depleted of testisin by siRNA knockdown revealed that loss of testisin markedly impaired reorganization and tubule-like formation on Matrigel. Moreover, testisin siRNA knockdown increased the paracellular permeability to FITC-albumin across endothelial cell monolayers, which was associated with decreased expression of the adherens junction protein VE-cadherin and increased levels of phospho-(Tyr658)-VE-cadherin, without affecting the levels of the tight junction proteins occludin, claudin-5, or ZO-1. Decreased expression of VE-cadherin in the neovasculature of Prss21-/- ovaries was also observed without marked differences in endothelial cell content, vascular claudin-5 expression or pericyte recruitment. Together, these data identify testisin as a novel regulator of VE-cadherin adhesions during angiogenesis and indicate a potential new target for regulating neovascular integrity and associated pathologies.
    • Mitochondrial Dysfunction is Linked to Pathogenesis in the P497S UBQLN2 Mouse Model of ALS/FTD

      Lin, Brian; Monteiro, Mervyn J.; 0000-0003-2944-3051 (2021)
      Ubiquilin-2 (UBQLN2) mutations cause amyotrophic lateral sclerosis (ALS) with frontotemporal dementia (FTD), but the mechanisms that drive disease pathogenesis remain unclear. Neurons have especially high energy requirements and consume copious amounts of ATP to support synaptic transmission and other complex processes. As such, mitochondrial dysfunction has been thought to play a pathogenic role in ALS. Recently, UBQLNs have been implicated in mitochondrial protein quality control whereby their inactivation in cells leads to the accumulation of cytostatic mitochondrial precursors. However, it is unclear what specific role UBQLN2 plays in maintaining mitochondrial proteostasis and how UBQLN2 mutations impact mitochondria physiology. In this thesis, I tested my hypothesis that the ALS-linked UBQLN2 P497S mutation causes mitochondrial dysfunction through loss of UBQLN2 chaperone function and impaired mitochondrial import. Our lab previously generated proteomic profiles of early-stage (8 weeks) hippocampal and spinal cord (SC) tissues isolated from non-transgenic (Non-Tg), wild-type (WT356), and P497S UBQLN2 mutant mice, whereby the mutant animal closely models human ALS/FTD. Gene ontology analysis revealed “mitochondrial proteins” as a major category altered in P497S animals. I immunoblotted SC lysates of Non-Tg, WT356 and P497S UBQLN2 animals for various mitochondrial proteins, and found decreased levels of many mitochondrial proteins, including those involved in oxidative phosphorylation (OXPHOS), network dynamics and import. I discovered through Seahorse respiration assays that mitochondria purified from the SC of P497S mice have age-dependent respiration deficits unlike those of age-matched Non-Tg and WT356 animals. Electron microscopy of spinal motor neurons in the P497S animals revealed distortions to mitochondria cristae. I demonstrated that mitochondrial alterations found in P497S mutant animals are recapitulated in UBQLN2 knock-out cells, suggesting loss of UBQLN2 function may underlie the mutation’s effects. Additionally, inactivation of UBQLN2 compromised proper targeting and processing of the mitochondrial import factor, TIMM44, which subsequently could be rescued by reexpression of WT UBQLN2, but not by mutant UBQLN2 proteins. ALS/FTD UBQLN2 mutants bind weaker to TIMM44 compared to WT UBQLN2, providing a possible mechanism for the mitochondrial import defects. Overall, these studies highlight a potential key role of UBQLN2 in maintaining mitochondrial health, and how its function is impaired by mutations in UBQLN2.
    • BECC Adjuvanted Vaccine Provides Broad Protection from Homologous and Heterologous Influenza A Virus Infections

      Haupt, Robert; Frieman, Matthew B.; 0000-0001-8300-7184 (2021)
      Influenza A virus (IAV) is a leading cause of mild to severe respiratory disease worldwide with significant infections resulting in hospitalization or death, especially in older individuals, young children, and people with comorbidities. Because the influenza virus has a high rate of mutation, this allows the virus to evade the host immune system against new variants, new vaccines are produced annually to match circulating strains. FluAd, is currently the only approved influenza vaccine formulated with the adjuvant MF59, a squalene oil emulsion, to create a stronger immune response to the vaccination and is limited for use in people 65 years and older. Classic adjuvants such as oil emulsions and alum are not universally effective and potentiate toward a T helper 2 (Th2) response (effective versus extracellular pathogens), while another often-used vaccine adjuvant, monophosphoryl lipid A, skews toward a Th1 response (effective versus intracellular pathogens). Unique adjuvants have been created using the Bacterial Enzymatic Combinatorial Chemistry (BECC) system to produce novel Toll like receptor 4 (TLR4) immunostimulatory molecules. These molecules drive a balanced Th1/Th2 response and enhanced immunogenicity in the context of an influenza virus vaccine candidate as well as other vaccine platforms. These BECC adjuvanted vaccines display superior protection to challenge from a homologous IAV strain in a BALB/c mouse model, even in aged mice with a single prime vaccination. More important, these vaccines stimulate the production of broadly reactive antibody to the hemagglutinin stem and protection against heterologous IAV strain infection, possibly skirting the need to formulate new influenza vaccine annually.
    • Patient and Intimate Partner (IP) Illness Appraisals in Cancer: A Multi-Methods Study

      Francis, Martha Eileen; Johantgen, Mary E. (2021)
      Background: Psychological distress for patients with serious illness has been associated with increased physical and spiritual distress, decreased quality of life, and increased medical expenses at end of life (EOL). For both cancer patients and their intimate partners (IPs), appraising the illness and communicating about it can be challenging. Yet, there is little evidence on how best to support them. Objectives: Bodenmann’s Systemic-Transactional Model (STM) of dyadic coping provided foundation to study how living with cancer impacts communication at primary appraisal level in patient/ IP dyad. The purpose of this exploratory multi-methods study was to understand patient/IP illness appraisals. This was investigated through following aims: 1. Describe patient/IP perceptions of dyadic communication before and after diagnosis of advanced cancer (including barriers and facilitators to sharing emotionally vulnerable content); 2. Describe patterns (incongruent/congruent) of patient and IP communication during advanced cancer; and 3. Explore relationships between experiential suffering (Suffering Pictogram) and communication congruency (CCAT-PF measure). Methods: Descriptive phenomenology was used for the qualitative phase. In-depth, semi-structured interviews with dyads were done, followed by individual interviews. The quantitative phase assessed cancer communication and suffering using established measures. Results: The main findings from qualitative analyses included: 1) Vulnerable communication is complicated by balancing two opposing worlds: Hope/Positivity and Uncertainty/Fear of Death; and 2) Vulnerable communication about EOL and hospice is emotional and unfamiliar. Hearing the word ‘hospice’ ends dyad’s uncertainty, confirming death from cancer is definite. Patients and IPs articulated feeling unprepared and needing guidance about skills to cross this vulnerable environment toward open communication. Quantitative data from communication measures showed low to medium discrepancy between dyads yet, patients displayed more discrepant communication behavior than their IP counterparts. IPs consistently exhibited higher suffering scores than patients across Overall Suffering and in 6/8 suffering domains. Worry and Fear were identified as highest domains of suffering for both patients/IPs. Conclusion: For IP dyads to articulate preferences for care with providers they must first be provided external support to facilitate vulnerable conversations within the dyad itself. These dyadic conversations must be initiated early after diagnosis to strengthen available supports during illness and EOL.