Now showing items 1-20 of 1921

    • Mapping Expanded Prostate Cancer Index Composite (EPIC) Questionnaire to EuroQoL-5D (EQ-5D) Utility Weights to Inform Economic Evaluations for Prostate Cancer

      Khairnar, Rahul; Palumbo, Francis Bernard, 1945- (2020)
      OBJECTIVES: To develop a mapping algorithm to obtain EuroQoL-5D-3L (EQ5D) health utilities from Expanded Prostate Cancer Index Composite (EPIC) questionnaire. METHODS: This mapping study utilized baseline data from an international, multicenter, randomized controlled trial (NCT00331773) of patients with low-risk prostate cancer. Patient health-related quality-of-life (HRQoL) data were collected using EPIC and health utilities were obtained using EQ5D. Data were divided into an estimation sample (n=765, 70%) and a validation sample (n=327, 30%). The relationship between the instruments was estimated using ordinary least squares (OLS), Tobit, and two-part models. Five-fold cross-validation (in-sample) was used to compare the predictive performance of the estimated models. Final models were selected based on root mean square error (RMSE). OLS models using baseline cross-sectional data, combined data from all assessment periods, and random effects (RE) models that explicitly model the longitudinal nature of the data were estimated to compare predictive ability of algorithms derived from cross-sectional and longitudinal data. Longitudinal predictive performance of OLS models derived using baseline data was examined in the post-intervention data. RESULTS: A total of 565 patients in the estimation sample had complete information on both EPIC and EQ5D questionnaires at baseline. Mean observed EQ5D utility was 0.90±0.13 (range: 0.28-1) with 55% of patients in full health. Low to moderate correlations were found between EQ5D utility and urinary (r=0.38), bowel (r=0.34) and hormonal (r=0.55) domains of EPIC; sexual domain was weakly correlated (r=0.18) with EQ5D utility. OLS models outperformed their counterpart models for all pre-determined model specifications. The best model fit was: “EQ5D utility = 0.248541 + 0.000748*(Urinary Function) + 0.001134*(Urinary Bother) + 0.000968*(Hormonal Function) + 0.004404*(Hormonal Bother) – 0.376487*(Zubrod) + 0.003562*(Urinary Function*Zubrod)”; RMSE was 0.10462. When comparing cross-sectional vs. longitudinal data, a mapping algorithm obtained using combined EPIC subdomain data outperformed other model types. Mean absolute differences (MDs) between reported and predicted were low in general and decreased as the time of assessment increased. CONCLUSIONS: This study identified mapping algorithms to generate EQ5D utilities from EPIC domain or sub-domain scores, with satisfactory longitudinal predictive performance. The study results will help estimate quality-adjusted life-years in future economic evaluations of prostate cancer treatments.
    • Evaluating Antidepressant Use in Nursing Home Residents with Moderate to Severe Cognitive Impairment

      Nalls, Victoria; Galik, Elizabeth; Resnick, Barbara (2020)
      Background: Antidepressants are commonly prescribed medications among nursing home residents and used to treat symptoms of dementia. Concerns have been raised, however, about disparities and potential inappropriate use of these medications within this population. Purpose: The purpose of this dissertation was to: (1) Describe factors associated with antidepressant use in nursing home residents with moderate to severe cognitive impairment; (2) Describe differences in antidepressant use between white and black nursing home residents with moderate to severe cognitive impairment; (3) Evaluate trends in antidepressants and antipsychotics prescribing among nursing home residents with moderate to severe cognitive impairment. Methods: This secondary data analysis used data from the Function and Behavior Focused Care (FBFC) for Nursing Home Residents with Dementia randomized control trial. A total of 336 residents were included in the study, who were mostly white, female, severely cognitively impaired (MMSE=7.8, SD=5.1) and the average age was 82. Data collection was done at baseline and 12 months and based on chart reviews, input from staff, and observation of residents. Descriptive statistics and logistic regression were used to address aims 1 and 2. Generalized linear mixed modeling with a binary distribution and logit link function was used for aim 3. Results: At baseline, 59% of the sample was taking an antidepressant. Race was significantly associated with antidepressant use (β=0.51; p=0.01). Black residents were half as likely to be on antidepressants compared to white residents (OR=0.499 CI=0.305-0.817) and received lower dosages of sertraline (t=2.68; p=0.01). There was no significant change in antidepressant or antipsychotic use at 12 months. Conclusions: Black nursing home residents with moderate to severe cognitive impairment were significantly less likely to be on antidepressants and when treated, were likely to be on lower dosages of some antidepressants. It is unknown if this is due to misdiagnoses and disparities in treatment or lack of need for antidepressants or differences in responses to specific drug classes. Further research is needed to explore these differences and evaluate the influence of resident, prescriber, and facility factors on the use of antidepressants among nursing home residents with moderate to severe dementia.
    • Early Symptom Improvement as a Predictor of Antidepressant Response in Children and Adolescents Diagnosed with Depression: Translating Evidence from Randomized Controlled Trials to Community Practice

      Spence, O'Mareen; dosReis, Susan (2020)
      Statement of the Problem: A common problem among children and adolescents diagnosed with depression who receive care in community settings is that antidepressant regimen changes such as psychotropic augmentation may occur soon after starting treatment. This raises the question as to whether such changes are implemented among youth who would otherwise respond to the antidepressant. Thus, the overarching objectives of this dissertation were to 1) distinguish early in treatment children and adolescents who are likely to respond, and 2) empirically evaluate the association between predicted response and psychotropic augmentation or switching in real world settings. Summary of Methods: Using randomized clinical trial (RCT) data, this research applied a Bayesian approach to predict the likelihood of initial (12 week) and sustained (18 week) response to treatment as a function of early changes in depressive symptoms (i.e. mood, somatic, subjective and behavioral) and other demographic and clinical factors. An innovative application of combined sample multiple imputations (CSMI) was used to estimate the 12-week predicted probability of response among commercially insured adolescents who received care in real-world settings. Each adolescent received a probability of treatment response, which was then used to compare the odds of psychotropic augmentation or switch. Results: Early changes in mood and somatic symptoms within the first six weeks of treatment are primary predictors of initial (at 12 weeks) and sustained (at 18 weeks) response to an antidepressant. Baseline depression severity is an important prognostic factor for initial response, and additional, though minimal improvement, in somatic symptoms from weeks 6 to 12 is indicative of sustained response. In a highly selected cohort of adolescents receiving care in community settings, an augmentation or switch occurred similarly among adolescents with a high versus low likelihood of responding to fluoxetine. Conclusion: The results suggest that other factors beyond expected antidepressant response (or lack thereof) might influence current treatment practices. Our findings have clinical and public health implications that support measurement-based care in pediatric depression. Our application of CSMI highlights several key areas of consideration for future pharmacoepidemiologic research aimed at translating RCT evidence to real world data to better understand clinical practices patterns.
    • Informing the role of RIFINs in malaria pathogenesis, natural immunity, and design of a severe malaria vaccine

      Zhou, Albert; Travassos, Mark A.; Laufer, Miriam K. (2020)
      Plasmodium falciparum is a eukaryotic parasite that causes severe malaria and contributed to 405,000 deaths worldwide in 2018. Victims of severe malaria are predominantly sub-Saharan African children, who typically present with symptoms of severe anemia or unarousable coma. The pathogenesis of severe malaria is poorly understood but mediated by the expression of adhesive variant surface antigens (VSAs) on infected red blood cells. VSAs are involved in sequestration and rosetting, unique virulence processes that allow the parasite to evade host immune responses and prevent clearance in the spleen. A relatively unstudied family of VSAs, the repetitive interspersed family (RIFIN) proteins, have recently been found to be important in rosetting and host immune suppression. RIFINs also appear to be targets for protective immunity; humoral immune responses against RIFINs have been correlated with asymptomatic infections. In this dissertation, I applied a multi-faceted approach using protein and peptide microarrays, transcriptomics, and reverse vaccinology to identify appealing RIFIN candidates for inclusion in a future severe malaria vaccine. I show that serological responses against epitopes within the semi-conserved domain of RIFINs associated with severe malaria reflected age-related malaria exposure. Sequencing and identifying specific rif genes expressed in clinical infections have not been feasible. I have addressed these challenges by adapting a novel bioinformatic pipeline and developing an HMM-based tool to process, assemble, classify, and subtype RIFIN sequences from peripheral blood samples. This takes advantage of a targeted probe capture method that I determined yields more abundant, full-length RIFIN sequences than other library enrichment approaches. Finally, I performed a comprehensive genomic survey of RIFIN gene repertoires using publicly available whole genome data of sixteen P. falciparum isolates to identify highly conserved, strain-transcendent sequences. Together, these results provide insights and powerful tools that can advance our understanding of the role RIFINs play in severe malaria pathogenesis and the development of naturally-acquired immunity to severe malaria. This work will aid efforts to determine targets for vaccines to protect children from the deadliest consequences of malaria.
    • Inhibition of Traumatic Brain Injury (TBI)-Induced Neuroinflammation Using Pharmacological Modulators of Metabotropic Glutamate Receptors

      Vinueza, Gelareh; Faden, A. I.; 0000-0002-1036-6378 (2020)
      Chronic dysregulated microglial activation is a major hallmark of persistent inflammation and progressive neurodegeneration following traumatic brain injury (TBI). Thus, research has focused on strategies to inhibit chronically activated microglial responses following TBI. Metabotropic glutamate receptors (mGluRs) 4 and 5 are expressed on microglia and can modulate microglial activity; therefore, they may serve as potential therapeutic targets for inhibition of microglial-dependent neuroinflammation. In the first of these studies, based on its reported neuroprotective roles, we examined the effects of the mGluR5 positive allosteric modulator (PAM) VU0360172 in an established fluid percussion injury (FPI) rat model of TBI plus hypobaria (HB). Systemic administration of VU0360172 significantly reduced pro-inflammatory cytokines, chemokines and microRNAs (miRs) at 1- and 7- days following FP+HB. However, VU0360172 did not alter injury-induced behavioral deficits examined over the following 28 days. In order to assess potential mechanisms underlying the inflammatory changes, we used Nanostring analysis to identify miRs that modulate neuroinflammation and compared plasma changes for selected miRs with brain tissue changes. The pro-inflammatory miR-223 showed the strongest correlation between plasma and brain tissue expression levels at the 7d time-point in TBI+HB experimental rodent models. An additional series of studies addressed the purported anti-inflammatory effects of mGluR4 PAMs. We employed in vitro models of immortalized microglia cell lines and primary microglia to elucidate the molecular mechanisms responsible for the modulation of inflammation by ADX88178 and other mGluR4 PAMs. ADX88178 downregulated lipopolysaccharide (LPS)-induced expression of pro-inflammatory mediators in BV2 cells and primary microglia. However, ADX88178 anti-inflammatory effects appeared to be mGluR4-independent as mGluR4 expression in our in vitro models was very low and its actions were not altered by pharmacological or molecular inhibition of mGluR4. Moreover, we showed that putative mGluR4 PAMs attenuate pro-inflammatory pathways in BV2 microglia through mGluR4/Gi-independent mechanisms involving activation of cAMP-response element binding protein (CREB) and inhibition of NFkB. Overall, these studies show that mGluR4 and mGluR5 PAMs can significantly attenuate microglial activation. Therefore, further studies should examine their potential therapeutic effectiveness after TBI.
    • Programmed Death Ligand 1 in Oral Potentially Malignant Epithelium and Implications of Regulation by Interferon Gamma

      Elnaggar, Manar; Younis, Rania H. (2020)
      Programmed death ligand 1 (PD-L1) is an immune-checkpoint regulator. Expression of PD-L1 in a subset of head and neck squamous cell carcinoma (HNSCC) was linked to improved response to immunotherapy. Oral potentially malignant disorders (OPMDs) are characterized by increased risk for malignant transformation. We investigated the expression of PD-L1 in immune cells (ICs) and epithelial cells of OPMD, which is essential to prevent progression to malignancy. Immunohistochemistry analyses in HNSCC whole excision tissue sections (104 patients) indeed demonstrated predominant expression of PD-L1 in the epithelial margins (~86%). This directly correlated with PD-L1 expression in underlying ICs (P=0.0172) with a predominating lichenoid pattern of IC infiltrate. Immunoblotting analyses demonstrated the role of human recombinant IFN-γ in the upregulation of PD-L1 expression in the oral premalignant cell line (DOK) with implications of downstream activation of pS6. Our work suggests a role for IFN-γ/PD-L1 in the immune escape of OPMDs.
    • Transient mechanical stimuli elicit rapid mechano-chemical signal transduction in non-tumorigenic and malignant mammary epithelial cells

      Pratt, Stephen JP; Martin, Stuart S.; 0000-0003-3627-7258 (2020)
      Changes in are observed during breast tumor formation and progression, which promote malignant phenotypes in both normal mammary epithelial cells and breast cancer cells. In this context, understanding the molecular details of mechanotransduction signaling may provide unique therapeutic targets. This work applied time-lapse confocal microscopy and quantitative methods to define the rapid mechanically-stimulated calcium signaling mechanisms that occur in breast epithelial cells. While most tumor cell studies focus on long-term effects of mechanical stimulation (>24h), the current approach detected an immediate initiation of cytosolic calcium signals within 2 seconds after transient mechanical stimulation. Two novel methods were developed to describe this response and underlying mechanisms: a) the real-time scratch assay and b) scratch on low elastic dishes (SLED). The real-time scratch assay revealed an ATP/P2Y2/Ca2+ signaling axis in response to scratch with implications as a path toward EMT. The second method developed was SLED, a non-damaging application of mechanical stress to breast epithelial cells with physiologic implications as the elasticity of the cell substrate better matched that of the mammary gland in vivo than the typical glass or plastic experimental dishes. New data obtained using this novel approach revealed that normal breast epithelial cells are mechanically sensitive, responding to mechanical stimuli through a two-part calcium signaling mechanism. An immediate, robust rise in intracellular calcium (within seconds) was observed followed by a persistent extracellular calcium influx (up to 30 minutes). This persistent calcium was sustained via microtubule-dependent mechano-activation of NADPH oxidase 2 (NOX2)-generated reactive oxygen species (ROS), which acted on TRPM8 channels to prolong calcium signaling. Disruption of this conserved mechanobiology mechanism was possible through oncogenic activation such that, among common oncogenic mutations, constitutively-active KRas suppressed this signaling pathway. Therefore, certain oncogenes render cells mechanically unresponsive which could have interesting implications for the evolution of cancer cell mechanosensing in the tumor microenvironment as cells acquire new mutations. In addition, altered expression of the ROS generator (NOX2) and the ROS-responsive channel (TRPM8) are indicators of reduced overall survival in ER-negative breast cancer, suggesting that this mechano-pathway identified in breast epithelial cells may also be modified in patients in vivo.
    • FOXN2 Expression: from Pluripotent Stem Cells to Neural Progenitor Cells

      Saleh, Wissam; Ament, Seth A. (2020)
      Several transcription factors (TFs) have been demonstrated as risk genes for schizophrenia (SCZ) based on genome-wide association studies (GWAS), fine-mapping, and functional studies. In this project, we prioritized TFs that have multiple lines of evidence supporting their likelihood of being causal risk loci for SCZ. We integrated results from four published studies, which used GWAS, gene expression and chromatin conformation studies to identify genes and gene networks associated with SCZ. These analyses revealed that the TF FOXN2 is a strong candidate risk gene for SCZ. Next, we characterized the dynamic expression profile of FOXN2 in our stem-cell based cortical neurogenesis model by qPCR. This study helped in establishing FOXN2 as a risk gene for SCZ potentially involved in cortical neurogenesis and building a strong base for the future genetic editing experiments designed for functional characterization of FOXN2 in cortical neurogenesis, that might relate to the pathophysiology of SCZ.
    • Delineating the Role of NKT cell Activation in B cell Lymphoma

      Lee, Michael; Webb, Tonya J. (2020)
      Natural Killer T (NKT) cells play an important role in cancer surveillance and can reduce lymphoma burden in vivo; however, a hallmark of cancer is its ability to evade immune surveillance. Our goals were to elucidate novel mechanisms utilized by B cell lymphoma to evade NKT cell-mediated immune surveillance and determine the prognostic potential of assessing NKT cell function in lymphoma patients. We found that knockdown of sphingosine kinase 1 (SK1) in human lymphoma cells results in a significant increase in CD1d-mediated NKT cell activation. Lipidomic and co-culture studies identified cardiolipin as being upregulated in SK1 knockdown cells and implicated cardiolipin as an NKT cell-specific cancer neoantigen. We also sought to determine the efficacy of NKT cell-based therapy on survival and the induction of anti-tumor immune responses in a mouse model of B cell lymphoma. We found that activation of NKT cells via early administration of α-galactosylceramide (α-GalCer) only provided modest protection. Our data suggest that the lack of protection is due, at least in part, to the expansion of myeloid-derived suppressor cells in α-GalCer-treated tumor bearing mice. Lastly, we sought to identify novel immunological biomarkers in lymphoma patients. It was found that lymphoma patients have a reduction in NKT cell function compared to healthy donors. Furthermore, lymphoma patients have significantly higher levels of both pro- and anti-inflammatory cytokines in their sera compared to healthy donors. In addition, lymphoma patients who experience relapse have significantly reduced NKT cell function in the blood, compared to lymphoma patients who did not relapse. Collectively, our studies demonstrate the multifaceted role NKT cells play in immune responses to B cell lymphoma and will help inform the next generation of cancer immunotherapy.
    • Burden and Mental Health of Family Caregivers of Cancer Patients: The Impact of Spirituality

      La, In Seo; Johantgen, Mary E. (2020)
      Background: As the primary source of care for individuals with cancer, family caregivers are relied on for treatment support and emotional care during the cancer trajectory. Studies on caregiver burden and psychological sequelae among cancer caregivers have been conducted cross-sectionally. Spirituality has been suggested as a potential buffer between burden and sequelae. Yet, there have been very few longitudinal studies addressing burden, depression, and spirituality, and there is limited information on psychometric properties of the spirituality measures in cancer caregivers. Purpose: The aims of this study were to: 1) evaluate validity of the Spiritual Perspective Scale (SPS) and explore differences in spirituality across caregiver and patient characteristics, 2) describe caregiver burden during active cancer treatment and explore caregiver and patient factors influencing caregiver burden, and 3) examine changes in caregiver burden, spirituality, and depression and explore the moderating effect of spirituality on burden-depression relationship over time. Methods: A secondary analysis of data from a longitudinal study of cancer caregivers from the NIH Clinical Center was conducted. Caregivers completed measures, including the Spiritual Perspective Scale (SPS), Caregiver Reaction Assessment (CRA), and NIH Toolbox and PROMIS® measures. Structural equation modeling and linear mixed modeling were used for testing study aims. Results: The SPS was found to have satisfactory psychometric properties in cancer caregivers. Adjusting for a direct effect of race did not alter the pattern of results, and caregivers who were older, female, racial/ethnic minorities, less educated, affiliated with a religion, and who provided care to anyone other than the patient reported higher levels of spirituality. Baseline mutuality between the caregiver and patient was negatively associated with initial burden. Changes in caregiver burden were related to being spouse caregivers, sole caregivers, and income. Scores on total burden, spirituality, and depression remained stable over time. Caregivers’ spirituality moderated the link between burden and depression (-1.26, p = .025). Conclusions: Higher levels of spirituality may act as a protective factor in the relationship between burden and depression during active cancer treatment. Identified factors related to burden and strategies to strengthen spirituality should be considered to improve caregiver mental health.
    • Dimensional Stability of CAD/CAM Patterns: A Longitudinal Study

      Byun, Shane S.; Masri, Radi, 1975- (2020)
      The goal of this study was to investigate the accuracy, over time, of computer-aided design and computer-aided manufactured (CAD/CAM) dental patterns in two different materials (resin and wax) using two different fabrication methods (subtractive and additive manufacturing). The intaglio surface of the patterns (n=48/time period) were evaluated at five predetermined time periods (zero minutes, 20 minutes, 24 hours, one week, and two weeks) relative to fabrication time. Intaglio surface scans of the generated samples were aligned with the Best Fit alignment to the design file (.stl) and compared with 3D Compare on Geomagic Control X to obtain the deviations as a root mean square (RMS). Trueness of the patterns were compared at all time points using three-way Analysis of Variance (ANOVA) (=.05). Accuracy of dental patterns deteriorated over time. When materials were considered, wax had better dimensional stability than resin. When fabrication method was considered, milled patterns had better dimensional stability than printed patterns. Time, material type, fabrication method, and all their interactions, showed a significant effect, however, the differences were very small (ranging from <1 m to 20 m). Thus, both resin and wax CAD/CAM patterns fabricated by additive and subtractive manufacturing can be used to produce dental restorations with acceptable accuracy.
    • Follicular Lymphoma Stage at Diagnosis: Determinants, Prediction from Administrative Claims Data and Impact on Healthcare Costs

      Albarmawi, Husam; Onukwugha, Eberechukwu (2020)
      Introduction: Follicular lymphoma (FL) stage is an important determinant of survival, treatment options and treatment outcomes. However, the determinants of advanced FL, defined as Ann Arbor stages III and IV, and its impact on the economic burden of FL are unknown. Moreover, for studies that rely on administrative claims data, it is not clear if advanced FL can be accurately predicted from this data source. Methods: Using the linked Surveillance, Epidemiology, and End Results-Medicare database we identified patients newly diagnosed with FL. We estimated a modified Poisson regression to explore the effect of pre-diagnosis healthcare resource utilization patterns and baseline county-level factors on FL stage. We estimated the 1-year and 5-year incremental costs of stages II-IV compared to stage I using generalized linear models. To predict FL stage from claims data, we developed and tested two random forests models. Results: We identified 11,078 patients diagnosed in 2000-2013. Half of the sample had advanced FL. Higher counts of specialist physician visits in the 3 years pre-diagnosis were associated with lower risk of advanced FL (4th quartile vs. 1st quartile: Relative Risk [RR]=0.92; 95% CI=0.86–0.99). The risk of advanced FL was 8% lower among women receiving screening mammography compared to men (RR=0.92; 95% CI=0.88–0.97). Living in counties designated as health professional shortage areas (HPSA) was associated with 7% increased risk of advanced FL (RR=1.07; 95% CI=1.00–1.14, p=0.049). In 2004-2009, FL patients with stages II, III and IV had statistically higher 1-year ($14,911; $15,106; $24,639, respectively, p<0.01) and 5-year costs ($21,590; $23,599; $34,968, respectively, p<0.01) compared to stage I patients. The random forests models exhibited poor accuracy of classifying limited and advanced FL from Medicare claims data (accuracy: ≤64%; sensitivity: ≤72%; specificity: ≤57%). Conclusions: Higher frequencies of specialist physician visits and living in counties with no HPSA can reduce the risk of presenting with advanced FL. Patients with stages II-IV incur significantly higher costs compared to stage I patients. The incremental cost increases with higher FL stage. Predicting advanced FL from claims data may not be feasible and researchers may need to rely on datasets with existing clinical information.
    • Effect of Transient Heat Exposure on Drug Delivery from Transdermal and Topical Products

      Thomas, Sherin; Stinchcomb, Audra L. (2020)
      Heating pads and electric blankets are widely used for relief from pain and to provide warmth, respectively. Their unintentional application simultaneously with a transdermal or topical system can result in unexpected drug levels in systemic circulation. Designing well-characterized in vitro and in vivo methods are vital to understanding the effect of heat and hence can aid in the development and evaluation of these products. The objective of this work was to evaluate the effect of heat on products with the same active pharmaceutical ingredient (API) but different inactive ingredients. Four drug molecules with different physicochemical properties were chosen. For each drug, formulations with different excipients were selected. In vivo serum drug profile and in vitro flux profile data can provide mechanistic understanding of heat effect on these formulations. Four topical diclofenac formulations were evaluated for heat effect in vitro under continuous heat exposure. Their flux profiles demonstrated the influence of formulation design and excipients on drug permeation at elevated skin temperature. Serum profiles of two different oxybutynin formulations evaluated under heat exposure showed very different magnitude of enhancement in serum levels under similar heat exposure conditions. Another objective of this work was establishing an in vitro - in vivo correlation (IVIVC) of heat effect on topical and transdermal formulations. This will help in characterizing and predicting heat effect minimizing the need of clinical trials and support the regulatory evaluation of these dosage forms. For buprenorphine patch, study design for in vitro permeation testing (IVPT) using human skin was well characterized to align with and mimic in vivo conditions of heat exposure. Level A and Level C IVIVC were established under normal as well as elevated temperature conditions. For lidocaine patches, IVIVC was observed for early heat effect. However, poor correlation was observed for late heat effect. The findings from this work determined IVPT studies can correlate with and be predictive of in vivo results under normal temperature conditions. But under suboptimal conditions like heat exposure, IVPT may have limitations and should be used in addition to other methods to evaluate heat effect.
    • Post-translational Regulation of Glucokinase in Hypothalamic Neurons

      McFarland, Jennifer; Rizzo, Megan A. (2020)
      Glucose-sensing tissues utilize glucokinase (GCK), the activity of which is rate-limiting for glucose metabolism, to sense and, consequently, counteract deviations from glucose homeostasis. Post-translational regulation of GCK is well defined in the liver and the pancreas, and is critical for the maintenance of glucose homeostasis; yet, post- translational regulation of GCK in hypothalamic neurons, which play a central role in maintaining glucose homeostasis, remains relatively unexplored. Here, we use a hypothalamically-derived, glucose-sensing GT1-7 neuronal cell line to provide evidence of a receptor-driven, ER Ca2+-mediated S-nitrosylation and activation of GCK. Strategic pharmacological manipulations were paired with the assessment of GCK activity, done by either measuring NAD(P)H autofluorescence while raising extracellular glucose, or through expression of a homotransfer FRET GCK biosensor. Further, a biotin-switch assay was used to confirm the presence of GCK S-nitrosylation. This work illustrates a central mechanism of post-translational GCK regulation, which may underlie metabolic signal integration in the hypothalamus and may contribute to the pathology of diabetes.
    • A clinical evaluation of the ability of finishing files to supplement the removal of bacteria and endotoxin from primarily infected root canals (Part I- Initial evaluation)

      Kim, Eunice; Martinho, Frederico C. (2020)
      Aim: The aim of this study was to evaluate the ability of XP-endo Finisher (XPF) to supplement the removal of bacteria and endotoxin from primary infected root canals after instrumentation. Methodology: This randomized and blinded controlled trial included eight subjects. Instrumentation was performed using Vortex Blue or XP-endo Shaper, followed by supplemental instrumentation with XPF. All canals were irrigated with 2.5% sodium hypochlorite. Bacterial and endotoxin samples were taken using sterile paper points. Samples were collected before and after instrumentation and after XPF. Results: Bacteria was present in all root canals. After XPF, bacterial mean was reduced from 255 ± 311.82 CFU/mL to 2.5 ± 7.07 CFU/mL (p= .056). Endotoxin was detected in all root canals by the LAL method (KQCL test). After XPF, endotoxin mean was significantly reduced from .85 ± .26 EU/mL to .03 ± .01 EU/mL (p= .00004). Conclusion: The findings of this study showed that the supplemental use of the XP-endo Finisher after root canal instrumentation was effective in significantly reducing endotoxin but not bacteria present in primary endodontic infections.
    • Novel PC2 regulation of ezrin in renal epithelia reveals insights into ADPKD cystogenesis

      Dixon, Eryn; Woodward, Owen Maxwell (2020)
      Autosomal dominant polycystic kidney disease is caused by the loss of function of either two transmembrane proteins, polycystin-1 or polycystin-2. In renal epithelia, the consequence of polycystin loss is the formation of progressive, focal, fluid-filled cysts. However, the function and associated downstream signaling pathways specific to the polycystins have not been defined. Therefore, a new in vitro tubuloid model was designed to investigate the proximate cellular changes in renal epithelial cells following inactivation of Pkd2, the gene that encodes for polycystin-2. This model system reinforced the relevance of proteins associated with cell junctions, adhesions, and matrix in the cyst mechanism. The impact of this model was further supported through morphometrical analysis of epithelial compartmentalization in human ADPKD tissue, demonstrating an altered apical compartment in emerging cysts compared to noncystic tubules. Seeking connection between the junctions and disrupted apical compartment led to investigation of ezrin, a master scaffold in the apical compartment in renal epithelial cells. Ezrin plays a critical role in regulation of polarity, cytoskeleton organization, and protein trafficking, and the downstream consequences of its disruption have not been elucidated. Investigation into the initiating events of cystogenesis in ADPKD revealed a dramatic change in ezrin, following loss of PC2 in our tubuloid model, cystic mouse model, and pathological human ADPKD tissue. Based on this novel regulatory relationship between PC2 and ezrin, as well as the antecedent loss of ezrin to cyst formation in mice, ezrin was overexpressed in the pkd2 morpholino zebrafish model. Increased expression of ezrin diminished the formation of pronephric cysts. This lead to the design of a cyst rescue mouse model, which has exhibited promising preliminary data for cyst area reduction with additional ezrin. The disruption of ezrin in Pkd2 inducible in vitro and in vivo model systems, changes in ADPKD patient tissue, and rescue of pronephric cysts in the pkd2 MO suggest there is a role of ezrin in renal cystogenesis. Understanding the relationship of ezrin, with PC2 in renal epithelial cells will help elucidate the mechanism of ADPKD cystogenesis and define important downstream pathways necessary for epithelial functions.
    • Evaluation of the Equivalency of Generic Drugs

      Das, Sharmila; Polli, James E. (2020)
      The objective of this dissertation is to assess the bioequivalence of generic drugs. Patients with epilepsy complain about more seizures and side effects after brand-generic or generic-generic switching of an anti-epileptic drug (AED). Generic brittleness (GB) is the familiar notion that, upon switching between AEDs of pharmaceutical equivalents, a patient experiences negative outcome. Aim 1 is to probe the individual patient attributes thought to predispose a patient to generic brittleness. At the University of Maryland Medical Center, 148 patients from the outpatient epilepsy clinic were recruited for an observational case-control study. An algorithm for being GB (40% of patients) and not GB was devised. A patient with epilepsy was categorized as GB if the patient negatively opined about generics and was taking brand of their most problematic AED when generic was available. Two demographic factors that increased the odds of being GB were a patient currently taking a problem AED and increasing total number of current medications. Interestingly, taking lamotrigine increased and taking any one of six “protective” anti-epileptic drugs decreased the odds of being GB, respectively. Furthermore, no genetic, clinical laboratory or neuropsychiatry tests or their sub-elements differentiated GB patients from not GB patients. Aim 2 involves a comparative pharmacokinetic (PK) analysis upon challenging sixteen GB patients to brand-generic or generic-generic switch of an AED that they are currently on, using a four-way crossover replicate design. For each patient, test and reference PK profiles were the same, despite patients being GB. Aim 3 is to assess the noninferiority of the generic sodium ferric gluconate (SFG) against the reference product Ferrlecit with respect to drug bound iron (DBI), after single dose intravenous administration of brand and generic SFG in 44 healthy volunteers. Using a two-way crossover replicate design, plasma PK profiles of SFG to Ferrlecit were the same across two iron species (e.g. DBI and NTBI), although adverse event rates differed. In conclusion generics of AEDs and intravenous sodium ferric gluconate are bioequivalent to the brand-name drugs. Results support FDA criteria for bioequivalence in regards to AEDs and complex iron products.
    • The Role of Type I and III Interferons in the Pathogenesis of Bordetella pertussis Infection and Disease.

      Ardanuy, Jeremy; Carbonetti, Nicholas H. (2020)
      Bordetella pertussis is a Gram-negative bacterial pathogen that infects human respiratory tracts and is the causative agent for the disease pertussis, otherwise known as whooping cough. In 2012, there were 48,277 reported cases in the United States, the most since 1955. Symptoms build up to severe paroxysmal coughing, often for 10+ weeks after onset. For infants, pertussis can be fatal due to complications including pulmonary hypertension, pneumothorax, high-level circulating lymphocytosis, and pneumonia. In adults, the disease is severe too, with long-lasting cough, lung damage, and serious symptoms resulting in hospitalization. A difficulty in treatment/prevention of pertussis is a suboptimal vaccine that confers waning immunity, and a lack of effective treatments available. Antibiotics are administered to patients to prevent transmission, but usually don’t change the clinical course of disease. Host directed therapeutics treating pertussis could benefit individuals with severe cough and save the lives of infected infants. Using RNA-sequencing transcriptomics we investigated lung gene expression responses to Bordetella pertussis infection in adult mice, revealing that type I and III interferon (IFN) responses and signaling pathways may play an important role in promoting inflammatory responses. In B. pertussis infected mice, lung type I/III IFN responses correlated with increased proinflammatory cytokine expression and lung inflammatory pathology. In mutant mouse models with increased type I IFN signaling, B. pertussis exacerbated lung inflammatory pathology, whereas knockout mice with deficiencies in type I/III IFN signaling had reduced lung inflammation compared to wild-type mice. In direct contrast, infant mice didn’t upregulate type I/III IFNs in response to B. pertussis infection and were protected from lethal infection by increased type I IFN signaling, indicating age dependent effects of type I/III IFN signaling during B. pertussis infection. The induction of type I/III IFNs was found to be MyD88 dependent, and TLR9 and STING were identified as DNA sensing pattern recognition receptors required for type I/III IFN responses, as well as for typical levels of lung inflammatory pathology. This observation, coupled with results showing DNase treatment of B. pertussis-infected mice causing reduced lung pathology, indicated a DNA dependent induction of type I/III IFNs, making these targets for therapeutic intervention.
    • Social Workers and Disproportionate Minority Contact: A Mixed Methods Study

      Afkinich, Jenny Lee; Bright, Charlotte Lyn (2020)
      Disproportionate minority contact (DMC) is the disproportionate representation of racial minority youth at all levels of the juvenile justice system. DMC is evident in rates of initial arrests, referrals to court, delinquency findings/ adjudications, out-of-home placements, and transfers to adult criminal court. Race remains a significant predictor of legal outcomes for youth even when factors such as prior legal history and current charge severity are considered despite White and minority youth reporting similar levels of offending. This mixed methods study examined the relationship between community social workers employed by the South Carolina Department of Juvenile Justice (SC DJJ) in the agency’s county offices and DMC. Administrative data from SC DJJ was utilized to determine the extent of DMC in the state, to compare legal outcomes (i.e., receiving confinement dispositions and being waived to adult court) for youth in counties with community social workers to youth in counties that do not employ community social workers, and to compare the legal outcomes for youth in counties with community social workers over time. The results indicate that DMC continues to exist in South Carolina when measured via relative rate indices. Overall, there was little evidence that employing community social workers is sufficient to reduce DMC at the disposition or waiver stage. Qualitative interviews with nine of the 11 community social workers were used to identify and understand the mechanisms, barriers, and facilitators for reducing DMC. The findings suggest multiple nuanced ways the social workers can play a role in reducing DMC. The social workers identified two stages in the juvenile justice process in which they can and have had an impact on increasing equity: (1) out-of-home placement decisions for youth on probation or parole and (2) determining probation requirements. The social workers described a need for hiring additional social workers. They also believe they could train police officers and school officials about alternatives to making a referral to SC DJJ to reduce inequitable decisions at the front-end of the juvenile justice system. Implications for the study include an expanded role for community social workers and new ways to examine DMC quantitatively.
    • A Bridge Between Integration Stations: Insula’s Connection with the ventral Bed Nucleus of the Stria Terminalis

      Girven, Kasey; Sparta, Dennis R. (2020)
      Individuals suffering from substance use disorder often experience relapse events that are attributed to drug craving. Insular cortex (IC) function is implicated in processing drug-predictive cues and is thought to be a critical substrate for drug craving. Here, we uncover the functional connectivity of a novel projection from the IC to the ventral bed nucleus of the stria terminalis (vBNST), a portion of the extended amygdala shown to modulate dopaminergic activity within the ventral tegmental area, and investigate the role of this pathway in establishing reward-predictive cues. We hypothesized that these cues activate IC projections that synapse onto projection neurons within the vBNST, which then activate the mesolimbic dopamine pathway, resulting in the acquisition of associations between exteroceptive stimuli and rewards. In addition, due to the BNST’s role in ethanol self-administration combined with IC’s role in processing interoceptive cues associated with addictive substances, we predicted exposure to ethanol would affect the characteristics of vBNST-projecting IC neurons. Here we utilized both ex vivo slice electrophysiology and in vivo optogenetics to examine the functional connectivity and bidirectionally control the IC-vBNST projection in various reward-related behavioral paradigms. We also examined the effect of alcohol consumption on the IC-vBNST projection in acute and repeated ethanol exposure as well as in withdrawal. This work provides a potential mechanism by which the IC processes exteroceptive triggers that are predictive of reward.