|Corinne Praska||Vitamin D inhibition of a T cell pro-inflammatory cytokine by an epigenetic mechanism
Vitamin D reduces autoimmune-mediated pathology by inhibiting T cell production of interferon-gamma (IFNg), a pro-inflammatory cytokine, through an unknown mechanism. I hypothesize that the vitamin D hormone, calcitriol, promotes methylation of the Ifng gene promoter to silence gene transcription in T cells. I will test the hypothesis by activating myelin-specific T cells in vitro in the presence and absence of calcitriol and quantifying IFNg secretion by ELISA and Ifng gene promoter methylation using the methylation-sensitive restriction enzyme SnaBI. If the calcitriol samples show less IFNg promoter fragmentation by SnaBI and reduced IFNg secretion compared to the placebo samples, the data would support an epigenetic mechanism. Understanding calcitriol mechanisms of IFNg regulation will facilitate prevention and therapeutic control of autoimmune disease.
|Haleigh Mulholland||Understanding the pathology of cognitive deficits of Parkinson’s disease using a PINK1 -/- rat model
Parkinson’s disease (PD) is a neurodegenerative disorder characterized by progressive deficits in motor function over time. PD also has a preclinical period during which patients experience deficits in mood, arousal, learning and memory. The underlying pathology of these deficits is unknown; however, there is evidence that aggregation of alpha-synuclein protein in the brain regions associated with arousal/memory (the amygdala, locus coeruleus, hippocampus, and dorsal raphe nuclei) may be associated with these deficits. In order to characterize the pathology in an animal model of PD, the PINK1 -/- rat, 10 month old tissue was stained for alpha-synuclein aggregation in the regions of interest. This research will lead to a better understanding of cognitive decline in PD patients and potentially serve as a biomarker for early diagnosis.
|Brittany Derynda||THE RELATIONSHIP BETWEEN SLEEP QUALITY AND THE ACCUMULATION OF AMYLOID PROTEIN IN THE BRAIN
Background: Sleep dysfunction is a risk factor for Alzheimer’s disease (AD). Given the lack of effective treatments, strategies to delay or prevent AD are needed. Despite several studies linking sleep dysfunction with development of AD pathology in rodent models, the extent to which sleep dysfunction contributes to AD pathology (especially amyloid accumulation) in cognitively normal humans is not known.
Objective: The purpose of this study is to determine whether sleep dysfunction is associated with the accumulation of amyloid protein (Aß) in the brain, a hallmark of AD.
Methods: 18 participants from the Wisconsin Alzheimer’s Disease Research Center (mean age 59.8+/- 4.5 years, 79.2% female, 75% with family history of AD) underwent an overnight sleep study using polysomnography. Objective sleep quality was measured using Wake After Sleep Onset (WASO). AD pathology was measured using cerebrospinal fluid samples analyzed for Aß42 and Aß40 levels (expressed as a ratio Aß42/Aß40). Multiple regression was used to test the association between Aß42 levels and sleep measures.
Results: Worse sleep quality (higher WASO) was significantly associated with lower Aß42 levels (p<.05).
Conclusion: Poor sleep quality may contribute to amyloid accumulation, although it is also possible that amyloid accumulation contributes to poor sleep. Additional longitudinal studies are needed to determine the causation. Poor sleep may be a modifiable risk factor for AD.
|Mikayla Gallenberger||FCGR3A VARIABILITY: COPY NUMBER, SINGLE NUCLEOTIDE POLYMORPHISMS, AND EFFICACY OF IMMUNOTHERAPY
Immunotherapeutic treatment options for cancer continue to rise, but not everyone will benefit from this therapy. To improve the efficacy of immunotherapy, personalized treatment strategies based on one’s genotype can be determined. My project focused on genotypic differences in the Fc gamma receptor (FCGR) gene, FCGR3A (expressed on NK cells). FCGR3A is subject to genetic variability in both copy number variation (CNV) and in single nucleotide polymorphisms (SNP), both of which affect the ability of the FCGR3A protein to interact with immunotherapies. We adapted the reference-query pyrosequencing (RQPS) assay to simultaneously genotype the SNP and CNV of FCGR3A. We utilized RQPS to genotype metastatic renal cell carcinoma patients to delineate the relationship between patient response to immunotherapy and their FCGR3A CNV/SNP genotypes.
|Adam Kaufmann||Chemical Mutagenesis and Isolation of Persistence mutants in B. subtilis
Staphylococcus aureus, Streptococcus pneumonia, and Listeria monocytogenes, are all harmful and even fatal Gram-positive bacteria, whose treatment is made difficult by the occurrence of a remarkably rare sub-populations known as persisters. These persisters are phenotypically different from normal cells in that they are able to survive antibiotic treatment, which allows for lingering infections and promotion of antibiotic resistance selection. Persisters differ from resistors in that they do not grow in the presence of antibiotics, but instead remain in a state of dormancy, resuming growth once the concentration of antibiotics decreases sufficiently. Despite recent progress in the understanding of persister cells in gram-negative bacteria, persistence in gram-positive bacteria remains largely unknown. The goal of this study is to examine the genetic cause of this phenomenon in gram-positive bacterial model B. subtilis, subjecting cultures to chemical mutagenesis and selecting for mutants with elevated levels of persistence. Analysis of the genetic and physiological variations of these high-persistence (Hip) mutants will allow for further research into more effective treatment options of bacterial infections.
|Katlynn Rodgers||Effects of Traumatic Brain Injury on Drosophila Hemolymph
Traumatic brain injury (TBI) affects millions of individuals each year and is a contributing factor to one-third of all injury-related deaths in the United States. At present, there is no precise medical test to diagnose or measure the severity of TBI. Methods to diagnose TBI are needed to instruct treatment options as well as evaluate the success of treatments. Thus, the goal of my research is to identify proteins that reflect biological responses to TBI and can be detected non-invasively, quantitatively, and rapidly. To achieve this goal, I will use a fruit fly (Drosophila melanogaster) model of TBI to determine changes in protein level in the hemolymph, the circulatory fluid of the fly, that occur following TBI. My data indicate that at least one protein changes expression following TBI and that higher resolution studies will uncover other protein changes. These studies may provide insight into the relationship between protein expression and TBI as well as lay the groundwork for identifying biomarkers associated with TBI in humans that can be used as diagnostic and therapeutic tools.
|Adam Slavick||Improving activity of Adenylation domains from Myxococcus xanthusNonribosomal Peptide Synthetase
Nonribosomal Peptide Synthetases (NRPSs) are multi-modular enzyme complexes that produce natural products in many bacterium and fungi. Some of these notable products produced by NRPSs include siderophores and medically relevant metabolites like antibiotics. An NRPS may contain any number of modules and within each module there are three core catalytic domains: adenylation (A), condensation (C), and peptidyl carrier protein (PCP). A single module is responsible for recognition, activation, and incorporation of a particular substrate; specifically, the A domain recognizes and activates the substrate. Until recently it was thought that all A domains functioned in a similar manner. That view changed when it was determined that some A domains require an accessory protein for solubility and function. These assessor proteins are called MbtH-like proteins (MLPs). While we know MLPs are essential for some NRPSs, the reason for why they are essential is still unknown. To investigate the role of MLPs play in NRPS enzymology, we are studying an MLP in the bacterium Myxococcus xanthus. This MLP is unusual in that the coding gene it is not directly associated with any NRPS-encoding gene cluster. Using comparative genomics, a partner NRPS for this MLP was identified and I am involved in the in vitro characterization of the NRPS/MLP interactions. In particular, I am examining whether an alternative method for protein purifications would improve detectable NRPS/MLP interactions. To date, work has focused on characterizing an isolated A domain with the MLP, but this has had mixed results. We hypothesize that purification of a complete NRPS module (C-A-PCP) will improve A domain activity because of possible interactions with adjacent domains that might be required for protein stability, which can lead to an active protein. To test this hypothesis, I have constructed a recombinant plasmid containing module DNA, overexpressed the associated gene, and purified the NRPS module when co-produced with the MLP. I will then test the solubility and activity of the A domain of this new construct, enabling me to fully test my hypothesis.
|Samantha Blanke||Novel Treatment for Dysphagia in Patients with Cognitive Impairment
Dysphagia, or swallowing dysfunction, resulting in entry of food or liquid into the airway, frequently leads to development of pneumonia in patients with Alzheimer’s disease; yet there are currently no effective treatments for dysphagia in patients with AD. The purpose of this study was to examine the effects of a novel 8 week lingual strengthening treatment for dysphagia on swallowing-related outcomes in four patients with dysphagia and cognitive impairment (Mini-mental status exam scores<24). Preliminary results show increases in lingual pressures (3/4 patients), lower ratings of swallowing-related effort (2/4 patients) and improvements in several subscales of the SWAL-QOL. These findings suggest that lingual strengthening therapy may be a feasible treatment option for patients with dementia and supports the need for future research.
|Madeleine Jones||An Economic Case Study of Farmers’ Markets and Food Deserts in the District of Columbia
I examined price differences between farmers’ markets (FM), a grocery store (GS), and a supercenter (SC) in wards with low food access in Washington DC. Prices for farmers’ market produce in low-access wards were also compared with those in farmers’ markets located in areas with higher access to fresh, healthy food. Average prices were lowest in low-access farmers’ markets and highest in high-access farmers’ markets. This was the only statistically significant result, however, descriptive results indicate that prices may be slightly higher at supercenters than farmers’ markets in low-access areas of DC, and higher still in low-access grocery stores. The discount achieved at low access farmers’ markets as opposed to other types of markets varied by product. The cost-competitive prices at farmers’ markets compared to other food retailers in low access areas may have
important implications for policy measures aimed at ameliorating food deserts.
|Ranveer Vasdev||Enhanced Rapid Prototyping of Skull CT via Novel Thresholding and Binary Smoothing Algorithm
3D printing of skull computed tomography (CT) scans can provide insight for cranial surgical interventions. However, segmentation and production of organ prototypes are often a rate limiting steps for both surgical interventions and educational purposes. We hypothesize a semi-automated algorithm for segmentation and surface mesh transformation of human skull CT scans to improve the speed of rapid prototyping. Image processing utilized a weighted otsu algorithm thresholding and combined Gaussian and Laplacian surface mesh smoothing to reduce image acquisition errors of a single male skull. Life sized 3D prototypes are under review by the Royal College of Surgeons in London, England. Minimal image acquisition errors are still resistant to binary smoothing, however current prototypes are valid models for educational purposes. Modification of this smoothing procedure is necessary to reduce CT acquisition error and minimize surface mesh shrinkage. However, preliminary prototypes are valid models of target CT. Following optimization and extended preliminary studies, this semi automated algorithm can be extended to fields such as archeology, surgical interventions, prosthetic construction, and education.
|Michael Diny||Investigating the Role of Kdm3b, a Histone Demethylase, in the Acquisition of Pluripotency
Induced Pluripotent Stem Cells (iPSCs) are somatic cells that have been reprogrammed to acquire the properties of embryonic stem cells by the overexpression of a small set of proteins. iPSCs have the unique ability to self-renew and maintain pluripotency, allowing them to differentiate into any of the three primary germ layers. This makes them suitable tools for regenerative therapy and disease modeling. Since somatic cells and their corresponding iPSCs share the same genome, iPSCs have acquired pluripotency by modifying their epigenome, the chemical marks on their DNA and histone tails. These epigenetic modifications are performed by various epigenetic enzymes. These enzymes have shown to be highly specific in their activity, but the mechanism by which they operate is not well understood. In order for safe, controllable and efficient iPSC therapy to be achieved, the molecular mechanism underlying the transition to the pluripotent state must be better understood. The goal of this proposal is to help delineate the molecular mechanism of an important epigenetic enzyme, Kdm3b, during the reprogramming process.
|Julia Loosen||Understanding the experiences of older adults with frequent hospitalizations: A qualitative study
Five percent of Americans account for almost half of all health care spending, yet patients with “high-utilization” patterns are underrepresented in research studies. Understanding the goals, values, and perceptions of patients with “high-utilization” patterns is critical to designing effective interventions. The purpose of this grounded theory study was to develop a theoretical model of factors contributing to frequent hospitalizations, grounded in lived experiences of patients and their families. We conducted in-depth interviews with frequently hospitalized patients and any caregivers they identified as important in their health care. Management strategies of patients and caregivers fell into “late reactive”, “early reactive”, and “proactive” categories. Use of these strategies depended on personal thresholds for accessing health care and on expectations of “being listened to” by health care providers.
|Joshua Michael Ferraro||The Pluripotency Marker Sox-2 is Elevated in the Alzheimer’s Disease Brain
Alzheimer’s disease (AD) is the most common form of dementia, and is characterized by a general decline in cognitive function. There are endless theories regarding what actually causes this disease and how it can potentially be cured. One current theory regarding AD focuses on post mitotic neurons re-entering the cell cycle. It is believed that this re-entry causes the neuronal degeneration and the progressive cognitive decline observed in Alzheimer’s patients. Previous research has shown that certain genes/transcription factors contribute to the regulation and renewal of embryonic stem cells, and could be causing post-mitotic neurons to re-enter the cell cycle. There are two main families of transcription factors that contribute to embryonic development: the Octamer family and the Sox family. Therefore, one specific factor being focused on is the presence of Sox-2, a member of the Sox family, in human adult brain tissue sections (AD and non-AD). Considering that Sox-2 is involved almost exclusively in development, it is hypothesized that in healthy/non-AD tissues, gene expression would be turned off after the organism has reached complete development. Under this hypothesis Sox-2 is expected to be expressed in the AD brain tissue sections but not in the healthy-non-AD tissue sections. This hypothesis was tested using immunohistochemistry (IHC) and analyzed using a fluorescent microscope. Experimental results found a Sox-2 expression level of 45% in AD tissue samples and 9% in Non-AD tissue samples. These results further support the hypothesis that the cause of neuronal death in Alzheimer’s disease is caused by neurons re-entering the cell cycle, and that Sox-2 is a contributing factor to Alzheimer’s disease.
|Theoren Loo||Muddied Waters: An Interdisciplinary Assessment of Water Security in Mmangweni Village, South Africa
By the end of 2014, 19 million people in South Africa lived in rural communities. Many of these areas depend on ground springs for drinking water and lack the capacity to transport and protect these sources. This results in the prevalence and transmission of waterborne diseases in low-resource communities in South Africa. We analyzed the current factors contributing to water stress in Mmangweni village, a rural community on the Eastern Cape of South Africa, by employing an interdisciplinary approach to recommend an implementation of a suitable water security system. Surveys were conducted to assess water accessibility, quality, and usage; photographs of springs were geo-tagged and tested for fecal coliforms; and a map of the village’s living dwellings, springs, and community centers was created. We found that all springs in the village were contaminated with fecal coliforms and that 61% of community members suffer from gastrointestinal symptoms. Villagers also complained about poor water quality and collection time (109 minutes per day on average). We recommend that national programs implement a water security system to prevent the transmission of waterborne diseases and to increase water accessibility to those living in neglected, rural communities in South Africa.
|Emily Forster||Inhibition of the Inflammasome to Improve Listeria monocytogenes as a Cancer Immunotherapeutic Platform
Listeria monocytogenes is a genetically tractable intracellular bacterium that induces strong innate and adaptive immune responses in its hosts. These characteristics make L. monocytogenes a promising immunotherapeutic platform for delivery of tumor antigens directly to host cells. However, the current L. monocytogenes-based immunotherapy is limited in that the bacteria can only secrete unmodified, soluble, simple antigens. To bypass this limitation, L. monocytogenes has been engineered to lyse within the host cytosol to deliver DNA encoding complex tumor antigens for production by host cells. We have shown that lysis of L. monocytogenes triggers a cytosolic innate immune response, the inflammasome. Surprisingly, we have found that inflammasome activation impairs the host adaptive immune response. Novel inflammasome inhibitors, PYRIN-only proteins (POPs), block inflammasome activation. We hypothesize that POP-mediated inflammasome inhibition will prevent the detrimental consequences of inflammasome activation due to bacteriolysis for antigen-encoding DNA delivery. This strategy will likely improve the host adaptive immune response, making L. monocytogenes more successful as a cancer immunotherapeutic platform.
|Emily Sheely||Designing individualized nutrition plans for preterm infants using urine metabolomics
The purpose of this research is to determine if metabolomic differences exist in urine between preterm infants being fed parenteral TrophAmine, an intravenous amino acid supplement, and infants on other feeding regimens. For this pilot study, we analyzed and compared urine samples from twenty-five subjects, taken on their third day of life. Three components of parenteral nutrition—tyrosine, tryptophan, and homolanthionine—increased significantly in infants fed TrophAmine in comparison to those who were not. These metabolites are vital for the synthesis of neurotransmitters, bone and cardiovascular health, or the formation of some proteins that are necessary for life. This demonstrates that we can identify significant metabolic differences between infants exposed to different conditions. It lays the ground work for future studies to explore metabolic development in the preterm infant.
|Sadie Gugel||Low-dose antibiotic treatment in early life impacts gut microbiota of active and hibernating ground squirrels
The annual hibernation cycle modifies the gut microbiota of the 13-lined ground squirrel (Ictidomys tridecemlineatus); while the total abundance of microbes gradually decreases over the winter fasting period, the taxonomic composition of the microbiota changes more rapidly. To further investigate these changes in the gut microbiota during hibernation, early life antibiotic treatments were used. Pregnant ground squirrels were given low dose penicillin (PEN) in drinking water for 1-2 wk prior to parturition and during the 4-wk lactation period (control mothers drank untreated water). Cecal contents collected from squirrel pup sampled in summer and ~3 months of hibernation were analyzed for bacterial abundance through qPCR using universal primers for all bacteria and specific primers for the mucin-degrading bacterium Akkermansia muciniphila. PEN treatment of mothers slightly reduced abundance of all bacteria in summer pups (2.3-fold) but the effect was absent in hibernating pups. However, PEN induced a large increase in abundance of Akkermansia in summer pups (39-fold) and there was a similar trend for hibernating pups. These results demonstrate that administration of a low dose antibiotic to ground squirrels in early life has modest effects on total abundance of gut bacteria but has substantial effects on composition of the bacterial community, including increasing abundance of Akkermansia muciniphila.
|Natalie Gallesr||Appropriate Statistical Modeling Sheds Light on the Complex Relationship Between Diet and Inflammatory Physiology
A typical Western Diet is characterized by food consumption simultaneously high in calories, fats, and carbohydrates. Since the adoption of highly processed foods in American culture after WWII and the publication of various dietary guidelines since the 1940s, macronutrient balances have shifted away from fats in favor of processed grains and sugars. The incidence of obesity, metabolic syndrome and metabolism-associated diseases has drastically increased, currently reaching epidemic proportions. Scientific literature and media outlets still present conflicting studies and controversial reports regarding the role of dietary styles on health and disease today. In the present study, we sought to elucidate the associations between macronutrient balance and micronutrient intake on cardiovascular health by considering glucoregulatory and lipid mediators, as well as the constrictions of metabolic regulation. We are utilizing sophisticated statistical approaches to distinguish among the complex relationships governing diet, metabolic regulation, and inflammatory physiology in a large sample of adults assessed by the National Health and Nutrition Examination Survey (NHANES) between 2003 and 2004. We hypothesized that diets rich in refined sugars and starches and poor in healthy fats contribute to the incidence of obesity and cardiovascular inflammation. We also hypothesized multiple interactions between macronutrients in moderating metabolic and inflammatory outcomes. In light of the many conflicting dietary recommendations popularly available, we hope to provide a more thorough examination of the relationships between nutrition and health through statistical models that carefully adhere to known physiological processes.
|Kennedy Ringelberg||Unnatural amino acid incorporation and covalent crosslinking as a means to identify specific protein interactions
The C-terminal domain (CTD) of RNA Polymerase II (Pol II) is a repeating heptapeptide of the consensus sequence YSPTSPS that plays a crucial role in orchestrating the events of gene transcription. Pol II’s activity is modulated by post-transcriptional modification of the CTD which serves to recruit various proteins responsible for efficient processing of the RNA transcript. One such factor is mRNA capping enzyme, Ceg1 in S. cerevisiae, which stabilizes the 5’ end of nascent transcripts. While phosphorylation at the Serine-5 position of the heptapeptide is necessary for Ceg1 recruitment to transcriptional promoters, the specific spatial organization of Ceg1 binding is unknown. Here we employ a photo-crosslinking system to uncover specific Ceg1 targets through incorporation of the photoreactive, unnatural amino acid, p-Benzoyl phenylalanine (pBpa), into the CTD binding site of the capping enzyme. When excited with UV radiation, pBpa has the capacity to covalently bind cellular factors within a 3.1Å radius. We show successful incorporation of pBpa into the Ceg1 enzyme via amber stop codon suppression, as well as successful covalent crosslinking to Pol II and other novel species. Also interesting is the finding that Ceg1 is seemingly able to bind to two conformations of the CTD. Previously solved X-ray cocrystal structures for Ceg1 homologues bound to CTD peptides display two grooves in which CTD can bind. By incorporating pBpa in both grooves, it was determined that Ceg1 efficiently crosslinked to CTD in either groove. Furthermore, determining the spatial location along the CTD at which Ceg1 binds will answer a question that has evaded the scientific field to this point and offer understanding to the specificity with which transcriptional cofactors interact with RNAP II. This study serves to validate unnatural amino acid incorporation as a method for studying known protein interactions as well as discover novel ones.
|Wen Fu||Synthesis of a GlfT2 acceptor substrate analog for elucidating galactan biosynthesis
Tuberculosis is the second greatest cause of mortality due to infectious disease worldwide. The disease-causing agent of tuberculosis, Mycobacterium tuberculosis (Mtb), has a characteristically thick cell wall, which makes Mtb infections hard to treat. The mycobacterial cell wall consists of peptidoglycan connected to galactan, a polysaccharide of about 20-40 galactofuranose (galf) residues that is polymerized by the galactofuranosyltransferase GlfT2, and then arabinan, another polysaccharide, followed by the mycolic acid membrane. Since the cell wall is essential for mycobacterial survival, several inhibitors of cell wall biosynthesis have been developed into frontline anti-mycobacterial agents. Although several drugs have been developed to target mycolic acid and arabinan biosynthesis, no drug has been developed to target galactan biosynthesis. A previous study from the Kiessling group shows that 12-phenoxy-dodec-2-enyl-D-galactofuranosyl-beta-(1->6)-D-galactofuranoside (12PDEG) is an effective exogenous acceptor for GlfT2. I will synthesize 12PDEG using the route published by the Kiessling group. Accessing 12PDEG will allow us to continue characterizing the function of GlfT2 in this essential synthetic pathway in mycobacteria and other related pathogens. This basic research may promote the development of probes that inhibit GlfT2 and galactan biosynthesis.
|Chinar S Raul||DNMT Knockdown In Vitro Decreased Axon Regeneration in DRG Neurons From Spinal Cord Injured Untreated Progeny of Folic Acid Treated Rats
Folic acid, a dietary supplement and a key methylator of the mammalian CNS has been reported to be effective in preventing neural tube defects and also in CNS regeneration and repair after injury. Our recent studies have revealed that the effect of folic acid on regeneration of the injured axons is transgenerational. Thus in this study, we silenced (in vitro) various DNA methyltransferases (DNMT1, DNMT3a and DNMT3b) of DRG neurons isolated from untreated F6 progeny of rats, whose parents (F0) were treated with folic acid (80µg/kg body weight). Our results shows that treating DRG neurons from F6 generation with siRNA against DNMT1, DNMT3a and DNMT3b significantly decreases axon elongation in vitro. The observed effect is the highest at 48 hrs after treatment. If confirmed statistically, these results suggest that DNMTs have a predominant role in axon regeneration after spinal cord injury and that methylation mechanisms are key to transgenerational axonal regeneration with folic acid.
|Jayne-Norah Ntambi||Does the semi-direct pathway interfere with indirect pathway in the tvDTH bioassay?
The trans-vivo Delayed Type Hypersensitivity (tvDTH) linked suppression bioassay is a bioassay that detects one aspect of infectious tolerance, T regulatory function, by measuring linked suppression of responses to a recall antigen. Cell lysates used in all the tvDTH assays are prepared by sonication followed by 16,000 x g centrifugation to remove large cell fragments, but not nanovasicles. Anti-HLA alloresponses are mediated by both indirect and direct/semidirect recognition, whereas anti-minor H responses are exclusively peptide-focused. We have found that given a similar degree of minor H-mismatch between HLA-identical vs HLA-MM siblings, the regulatory response (linked suppression) is greatly diminished by an HLA mismatch. Ultra-centrifugation at 100,000 g of a sonicated-cell antigen preparation generates isolated nano-vesicles. We hypothesized that anti-HLA semi-direct response (due to exosomes in the antigen preparation) enhance the regulatory response through nano-vesicle-cross dressing of dendritic cells. Data from experiments designed to test this hypothesis will be presented.
|Atzie Sobotik||Collagen Content in Rats Exposed to Postnatal Hyperoxia
Premature birth, birth at fewer than 37 weeks of gestation, accounted for about 1 in 10 births in 2014. Advances in modern medicine such as postnatal oxygen therapy and synthetic surfactant allow babies born as early as 22 weeks to survive past infancy. These methods are essential to survival: the lungs of the preterm babies at 22-26 weeks do not sufficiently make surfactant and are severely underdeveloped, causing respiratory distress. Together, hyperoxia and synthetic surfactant assist in developing lung cross-sectional area and alveolarization, promoting efficient gas exchange at the time of treatment. Although these methods are vital, life-saving treatments during infancy, recent studies show that oxygen therapy (hyperoxia) induces inflammation, which causes long-term alterations including irregular lung development, such as impaired alveolarization, reductions in pulmonary vasculature cross-sectional area, and increased collagen content, leading to Bronchopulmonary Dysplasia (BPD). Previous studies using a BPD rodent model (hyperoxia exposure, HYP) demonstrate an increase in lung collagen content within the first 20 days of birth. These works revealed that the aberrant lung development in the BPD model is associated with pathological changes in the ECM (i.e. increased collagen content). Collagens are the major structural proteins in the lung. They are found in principal structures like airways, blood vessels, interstitial parenchyma, and basement membranes. Following injury, collagen production is upregulated in many cell types including fibroblasts, epithelial cells, and inflammatory cells, which can change collagen’s deposition pattern. The purpose of this study is to determine if 14 days of postnatal hyperoxia exposure affects collagen accumulation in the lungs of male and female rats.
|Courtney McCourt||Supplementation of choline increases very-low density lipoprotein export from bovine primary hepatocytes
Supplementation of choline in dairy cattle decreases liver lipid accumulation, potentially through increased very low-density lipoprotein (VLDL) export. The direct impact of choline supplementation on VLDL export has not been confirmed due to an inability to quantify bovine VLDL. In other species, VLDL is separated from low- and high-density lipoproteins based on size; however, bovine VLDL differs in density compared with nonruminant VLDL particles due to lipid profile. The objectives of this experiment were to validate a bovine-specific ELISA technique for use in quantification of bovine VLDL and determine the effect of increasing concentrations of choline chloride (CC) and dL-methionine (dLM) during fatty acid challenge on VLDL export in cell culture. Primary hepatocytes isolated from 4 Holstein calves were maintained as monolayer cultures for 24 h prior to treatment with CC (33, 100, 2000, 4500 µM) and dLM (16, 30, 100, 300 µM) within a factorial design, with 1 mM FA cocktail. Treatments mimicked expected physiological concentrations. After 24 h media was collected for VLDL quantification by ELISA. The ELISA (NeoBiolab, Cambridge, MA) is a competitive immunoassay that utilizes antibodies for apolipoprotein B100 and C, which in combination uniquely binds to VLDL particles. The assay was validated using cell culture and postpartum bovine serum samples with purified LDL and HDL spikes. Recovery of samples with spiked standards, compared with independently analyzed samples and standards, was 102 ± 1%. Cell culture data were analyzed using PROC MIXED of SAS 9.4 with linear and quadratic contrasts including fixed effect of treatment and random effect of calf. Interactions were not significant and therefore only main effects are discussed. Increasing concentrations of CC linearly increased (P = 0.02) VLDL export from hepatocytes. Treatment with dLM did not alter (P = 0.7) VLDL export. This research validates an ELISA for use in quantification of bovine VLDL. Furthermore, these data support the role of CC, but not dLM, supplementation in increasing VLDL export from hepatocytes.
|Sarah Laudon||THE USE OF A NATIONAL SURVEY TO UNDERSTAND PARENTS’ PERSPECTIVES OF COMMUNITY SERVICES AND PROGRAMMING FOR THEIR CHILDREN DIAGNOSED WITH FRAGILE X SYNDROME AND/OR AUTISM SPECTRUM DISORDER
Autism Spectrum Disorder (ASD), now found in 1 out of every 68 children, is characterized by lack of social reciprocity, communicative difficulties, and repetitive behaviors. Fragile X Syndrome (FXS) is the most common known cause of inherited intellectual disability; children diagnosed with FXS can present with learning disabilities and social and behavioral challenges. These diagnoses can occur together. This study will examine parent perspectives in the services provided to their child with ASD and/or FXS. This will include medical care, Birth to Three services, speech pathology services, as well as literacy benchmarks and goals, IEP planning by their school, and level of independence. Past research has indicated that no large scale survey covering a variety of treatment services and developmental milestones has been completed.
|Alyssa Marie Joachim||Early cellular response due to infection with human rhinoviruses A, B, and C in airway epithelial cells in vitro
Human rhinoviruses (HRVs) infect the respiratory system of humans and are the cause of the majority of common colds. There are over 100 types of rhinoviruses, divided into 3 families: A, B, and C. Past studies have investigated cellular responses at 24 hours post-infection when cells begin to release inflammatory factors; however, this study examines the differences between RV-A, RV-B, and RV-C infection of human epithelial cells in vitro at early time points of 4 and 8 hours as depicted by changes in RNA expression levels. We hypothesized that the genes commonly up or down regulated by all 3 viruses would be genes involved in signaling to uninfected cells as a defense mechanism. We infected three 12-well plates of primary epithelial cells, each plate from a different donor, with RV-A, RV-B, and RV-C. By sequencing the mRNA present within host cells we were able to observe how gene expression changed at 4 and 8 hours post-infection. We found that the most commonly up-regulated genes at early time points were genes involved in transcription regulation (FOS, FOSB, JUN), as well as genes involved in vascularization of tissue (PGF, VEGF, VEGFR2), which can lead to a runny nose and is often the first symptom observed upon rhinovirus infection. The results from this study provide an overview of what changes occur early on in cells infected by rhinoviruses and can be further studied to determine why the cells respond this way and how this response may affect the progression of the infection.
|Keighley Reisenauer||Measuring Mitochondrial Nonenzymatic Lysine Acetylation
Acetylation is a regulatory modification affecting numerous biochemical and cellular processes. Over half of the proteins in mitochondria have been identified as acetylated, however the mechanism of how this occurs has not been elucidated. The focus of this project is to understand the mechanism of mitochondrial protein acetylation, with the hypothesis that non-enzymatic acetylation is responsible for the majority of observed acetylation. In order to understand non-enzymatic acetylation, we must be able to quantify this reaction. Quantifying nonenzymatic acetylation is accomplished by incubating a protein with increasing concentrations of acetyl-CoA. The rate is determined by plotting the change in acetylation over concentration of acetyl-CoA. The slope of the line is the second order rate constant (lysine reactivity). This study focuses on measuring the second order rate constants of ACAT, PDH, HMGCS2, HMGCL1, and aKGDH, mitochondrial proteins which are reported to be highly acetylated and also because they contain substrate-binding sites for acetyl-CoA and other CoA derivatives.
|Ryan Prestil||Engineering the Cellular Microenvironment to Reveal Mechanisms of Human Development
Human pluripotent stem cells possess the unique ability to proliferate indefinitely and to differentiate into any cell type in the adult body, and somatic cells are now routinely reprogrammed to pluripotency via ectopic expression of the OSKM factors (Oct4/Sox2/Klf4/c-Myc). During both differentiation and reprogramming, gene expression changes dramatically along with physical changes to the shape, size, and organization of both the nucleus and the cell body as a whole. We have developed high-content microcontact-printed biomaterial platforms to control cell patterning to 10µm resolution and to create a stable gradient of soluble molecules, and we have derived a new multi-transgenic line of human induced pluripotent stem cells to combine doxycycline-inducible OSKM reprogramming factors with live, dynamic fluorescent labels on histones and actin filaments. Combining these capabilities has permitted unprecedented profiling of the processes underlying differentiation and reprogramming and characterization of intermediate cell states.
|Yunmiao Wang||Rubber Hand Illusion: Robotic Stroke vs. Manual Stroke
The Rubber Hand Illusion (RHI) is a procedure in which synchronized tactile and visual sensation, stroking the finger with a soft brush, causes a participant to perceive a rubber hand as being their actual hand. An ongoing study in the Davidson lab is using the RHI as a test of integration of bodily sense of self in relation to other psychological variables. This study developed automated robots for stroking in a controlled and repeatable way; however, it is unknown whether this is equally effective as the traditional manual stroking. To test this, I have conducted a sub-study of difference in response to robotic and manual stroking in the RHI. This sub-study uses the same procedure as the main study, but with a within-subjects comparison of the two stroking methods. Outcome measures are: self-report of the illusion, and facial electromyography (EMG) and skin conductance response (SCR). In addition, I have measured surface temperature of the hand subjected to the illusion. Preliminary data suggest that the temperature responds to the illusion and that the robotic stroke is less effective than the manual stroke.
|Maria Olaru||Exploring High Self-Identification Versus Low Self-Identification Motivators for Altruistic Behavior through Comparison of Behavioral-Based Tasks
Our lab’s Economic Decision-Making and Wellbeing (ECWELL) study has used the factor of Self-Identification (SI) to explore altruistic behavior. The ECWELL study implements a form of dictator game called the Self-Other (SO) game as a behavioral measure of altruism. My study will investigate the differential motivators of altruistic behavior in the context of the existing ECWELL study by pairing the Are You The Same (AYTS) questionnaire to measure high SI versus low SI motivated donations with the SO game. The AYTS questionnaire has been validated in a recent paper published by the ECWELL study. Then, I will have the participants complete the SO game twice, once with monetary gains received by a stranger, and once with monetary gains received by a friend, in order to measure for high and low SI motivators. I hypothesize that participants scoring high on SI will show a difference in altruistic behavior depending on the recipient of the monetary donations, while those scoring low on SI will not show differences in altruistic behavior between the conditions.
|Daniel Aaron Fox||Roles for Zic2 and Zic3 in the developing brain
The Hedgehog (Hh) signaling pathway plays a crucial role in modulating embryonic development. Malfunctions in the vertebrate Hh pathway involving Sonic Hedgehog Homolog (Shh) have been linked to cancers including basal cell carcinoma and developmental disorders like holoprosencephaly. Zic3, a zinc-finger transcription factor, is hypothesized to activate Shh-mediated Hh signaling. This is based on data demonstrating ZIC3 often binds to GLI consensus motif, and Zic3-depleted embryos express lower levels of Shh. To test this hypothesis, a line of zebrafish embryos carrying a nonsense mutation of Zic3 was examined for morphology of the forebrain and retina. Unexpectedly, no visible defects were found in embryos homozygous for the mutant allele through five days of development, suggesting Zic3 has redundant functions or gene compensation is occurring, in comparison to Zic3-depleted embryos. To test this hypothesis, expression of four Hh pathway genes (Shh, Shhb, Hhip, and Gli2a) will be assayed via in situ hybridization in zic3 mutants. If my hypothesis is correct, Zic3 mutants will show normal patterns of Hh target expression.
In a parallel approach, we are investigating Alx1, a candidate target of Zic2. Alx1 is strongly implicated in the development of neural crest (NC) cells, as is the Zic gene family. Zic3, Zic2 and possibly Alx1 activate transcription of FoxD3 and Sox10, which in turn promote specification of NC cells. Pericytes, a mural cell type important for vascular development, are one of many cell lineages derived from NC cells. Zic2 mutants exhibit hemorrhage indicative of aberrant vasculogenesis, which may be due to aberrant pericyte formation. To test this hypothesis, in situ hybridization and immunohistochemistry will be used to assay expression of Pdgfrb, a gene expressed in pericytes, in Zic2 mutant and Alx1 morpholino-injected embryos. My hypothesis predicts reduction and/or mispatterning of pericytes as a consequence of Alx1 or Zic2 depletion. This study will provide insight into the mechanism of Zic gene family function in early development and will potentially enhance understanding of diseases associated with it.