Enhancing Lipid Metabolism Promotes Cytokine-Induced Activation and IFN-γ Production in Naïve Human Natural Killer Cells

From the Washington University Senior Honors Thesis Abstracts (WUSHTA), Spring 2018. Published by the Office of Undergraduate Research. Joy Zalis Kiefer, Director of Undergraduate Research and Associate Dean in the College of Arts & Sciences; Lindsey Paunovich, Editor; Helen Human, Programs Manager and Assistant Dean in the College of Arts and Sciences Mentor: Megan A. Coope

The Localization and Function of Novel Tetrahymena thermophila Cytoskeletal Genes BBC29 and BBC39

Ciliary function is known to play an important role in many human conditions, including chronic sinus and pulmonary diseases and problems with infertility. Cilia are cytoskeletal structures that protrude from the cell body to facilitate movement. Ciliary structure is conserved throughout eukaryotes, from unicellular to multicellular organisms, including humans. A clear understanding the proteins that compose cilia and how they interact with one another will increase our knowledge about important cytoskeletal elements. Because cilia are difficult to study in multicellular organisms, the unicellular eukaryote Tetrahymena thermophila serves as a useful model for the study of cytoskeletal genes, due to their robust and longitudinally organized cilia. Here, we examine two novel kinetodesmal fiber proteins in T. thermophila, BBC29 and BBC39. Kinetodesmal fibers stabilize and organize cytoskeletal elements to ensure that all the cilia beat together and in the same orientation to maximize the effects of their beating. Genes BBC29 and BBC39 were isolated using PCR amplification, cloned into pICY-gtw vectors, and transformed into T. thermophila. The pICY vector included an inducible 3’ MTT cadmium promoter and a 5’ yellow fluorescent protein tag to allow visualization of the proteins’ location in live cells under UV light. Both genes localized to the kinetodesmal fiber structures, indicating their role to help stabilize and coordinate ciliary movement. We examined the interaction between the two proteins using co-immunoprecipitation to determine if the two proteins are dependent on each other for proper function. Our results demonstrate the complex interactions that occur between cytoskeletal proteins, such as those in the kinetodesmal fibers of cilia, and illuminate the need to further understand the relationship among the many proteins that allow for proper ciliary function. Because cytoskeletal proteins are often highly conserved between species, results gleaned through study with T. thermophila may be extended to elucidate the biochemical basis of human ciliary diseases

Putting a Face on Hunger: A Community-Academic Research Project

Food insecurity is a growing concern for Eau Claire County residents in Western Wisconsin. A community-academic partnership studied food insecurity through the voices of families struggling to access food and institutions that assist with hunger related problems. Data were collected through focus groups held in urban and rural parts of the county. Participants reported that food insecurity affected all aspects of daily life, increasing stress and reducing coping abilities. Results indicate that when Extension and campus-based staff partner with community groups, they can increase community awareness of and find innovative solutions to pressing community needs, such as food insecurity

Step width variability as a discriminator of age-related gait changes

Background There is scientific evidence that older adults aged 65 and over walk with increased step width variability which has been associated with risk of falling. However, there are presently no threshold levels that define the optimal reference range of step width variability. Thus, the purpose of our study was to estimate the optimal reference range for identifying older adults with normative and excessive step width variability. Methods We searched systematically the BMC, Cochrane Library, EBSCO, Frontiers, IEEE, PubMed, Scopus, SpringerLink, Web of Science, Wiley, and PROQUEST databases until September 2018, and included the studies that measured step width variability in both younger and older adults during walking at self-selected speed. Data were pooled in meta-analysis, and standardized mean differences (SMD) with 95% confidence intervals (CI) were calculated. A single-decision threshold method based on the Youden index, and a two-decision threshold method based on the uncertain interval method were used to identify the optimal threshold levels (PROSPERO registration: CRD42018107079). Results Ten studies were retrieved (older adults = 304; younger adults = 219). Step width variability was higher in older than in younger adults (SMD = 1.15, 95% CI = 0.60; 1.70; t = 4.72, p = 0.001). The single-decision method set the threshold level for excessive step width variability at 2.14 cm. For the two-decision method, step width variability values above the upper threshold level of 2.50 cm were considered excessive, while step width variability values below the lower threshold level of 1.97 cm were considered within the optimal reference range. Conclusion Step width variability is higher in older adults than in younger adults, with step width variability values above the upper threshold level of 2.50 cm to be considered as excessive. This information could potentially impact rehabilitation technology design for devices targeting lateral stability during walking

Exploring Health Insurance Status and Emergency Department Utilization

Emergency department (ED) use, by both insured and uninsured, leads to significant health care costs in the United States. While frequent ED use is often attributed to the uninsured, there is some evidence that insured populations also report utilizing the ED when otherwise preventable or nonurgent. We conducted in-person surveys of patients visiting the ED at a large research hospital and examined the differences in their characteristics based on the health insurance status. While less than the uninsured, insured individuals still report barriers to access to care outside the ED that include lack of access to another health care facility and unavailability of a doctor’s office or clinic

Glycolytic requirement for NK cell cytotoxicity and cytomegalovirus control

NK cell activation has been shown to be metabolically regulated in vitro; however, the role of metabolism during in vivo NK cell responses to infection is unknown. We examined the role of glycolysis in NK cell function during murine cytomegalovirus (MCMV) infection and the ability of IL-15 to prime NK cells during CMV infection. The glucose metabolism inhibitor 2-deoxy-ᴅ-glucose (2DG) impaired both mouse and human NK cell cytotoxicity following priming in vitro. Similarly, MCMV-infected mice treated with 2DG had impaired clearance of NK-specific targets in vivo, which was associated with higher viral burden and susceptibility to infection on the C57BL/6 background. IL-15 priming is known to alter NK cell metabolism and metabolic requirements for activation. Treatment with the IL-15 superagonist ALT-803 rescued mice from otherwise lethal infection in an NK-dependent manner. Consistent with this, treatment of a patient with ALT-803 for recurrent CMV reactivation after hematopoietic cell transplant was associated with clearance of viremia. These studies demonstrate that NK cell-mediated control of viral infection requires glucose metabolism and that IL-15 treatment in vivo can reduce this requirement and may be effective as an antiviral therapy

Beta cell extracellular vesicle miR-21-5p cargo is increased in response to inflammatory cytokines and serves as a biomarker of type 1 diabetes

AIMS/HYPOTHESIS: Improved biomarkers are acutely needed for the detection of developing type 1 diabetes, prior to critical loss of beta cell mass. We previously demonstrated that elevated beta cell microRNA 21-5p (miR-21-5p) in rodent and human models of type 1 diabetes increased beta cell apoptosis. We hypothesised that the inflammatory milieu of developing diabetes may also increase miR-21-5p in beta cell extracellular vesicle (EV) cargo and that circulating EV miR-21-5p would be increased during type 1 diabetes development. METHODS: MIN6 and EndoC-βH1 beta cell lines and human islets were treated with IL-1β, IFN-γ and TNF-α to mimic the inflammatory milieu of early type 1 diabetes. Serum was collected weekly from 8-week-old female NOD mice until diabetes onset. Sera from a cross-section of 19 children at the time of type 1 diabetes diagnosis and 16 healthy children were also analysed. EVs were isolated from cell culture media or serum using sequential ultracentrifugation or ExoQuick precipitation and EV miRNAs were assayed. RESULTS: Cytokine treatment in beta cell lines and human islets resulted in a 1.5- to threefold increase in miR-21-5p. However, corresponding EVs were further enriched for this miRNA, with a three- to sixfold EV miR-21-5p increase in response to cytokine treatment. This difference was only partially reduced by pre-treatment of beta cells with Z-VAD-FMK to inhibit cytokine-induced caspase activity. Nanoparticle tracking analysis showed cytokines to have no effect on the number of EVs, implicating specific changes within EV cargo as being responsible for the increase in beta cell EV miR-21-5p. Sequential ultracentrifugation to separate EVs by size suggested that this effect was mostly due to cytokine-induced increases in exosome miR-21-5p. Longitudinal serum collections from NOD mice showed that EVs displayed progressive increases in miR-21-5p beginning 3 weeks prior to diabetes onset. To validate the relevance to human diabetes, we assayed serum from children with new-onset type 1 diabetes compared with healthy children. While total serum miR-21-5p and total serum EVs were reduced in diabetic participants, serum EV miR-21-5p was increased threefold compared with non-diabetic individuals. By contrast, both serum and EV miR-375-5p were increased in parallel among diabetic participants. CONCLUSIONS/INTERPRETATION: We propose that circulating EV miR-21-5p may be a promising marker of developing type 1 diabetes. Additionally, our findings highlight that, for certain miRNAs, total circulating miRNA levels are distinct from circulating EV miRNA content

Methods for identifying surgical wound infection after discharge from hospital: a systematic review.

Background: Wound infections are a common complication of surgery that add significantly to the morbidity of patients and costs of treatment. The global trend towards reducing length of hospital stay post-surgery and the increase in day case surgery means that surgical site infections (SSI) will increasingly occur after hospital discharge. Surveillance of SSIs is important because rates of SSI are viewed as a measure of hospital performance, however accurate detection of SSIs post-hospital discharge is not straightforward. Methods: We conducted a systematic review of methods of post discharge surveillance for surgical wound infection and undertook a national audit of methods of post-discharge surveillance for surgical site infection currently used within United Kingdom NHS Trusts. Results: Seven reports of six comparative studies which examined the validity of post-discharge surveillance methods were located; these involved different comparisons and some had methodological limitations, making it difficult to identify an optimal method. Several studies evaluated automated screening of electronic records and found this to be a useful strategy for the identification of SSIs that occurred post discharge. The audit identified a wide range of relevant post-discharge surveillance programmes in England, Scotland and Wales and Northern Ireland; however, these programmes used varying approaches for which there is little supporting evidence of validity and/or reliability. Conclusion: In order to establish robust methods of surveillance for those surgical site infections that occur post discharge, there is a need to develop a method of case ascertainment that is valid and reliable post discharge. Existing research has not identified a valid and reliable method. A standardised definition of wound infection ( e. g. that of the Centres for Disease Control) should be used as a basis for developing a feasible, valid and reliable approach to defining post discharge SSI. At a local level, the method used to ascertain post discharge SSI will depend upon the purpose of the surveillance, the nature of available routine data and the resources available

Sixteen years of Collaborative Learning through Active Sense-making in Physics (CLASP) at UC Davis

This paper describes our large reformed introductory physics course at UC Davis, which bioscience students have been taking since 1996. The central feature of this course is a focus on sense-making by the students during the five hours per week discussion/labs in which the students take part in activities emphasizing peer-peer discussions, argumentation, and presentations of ideas. The course differs in many fundamental ways from traditionally taught introductory physics courses. After discussing the unique features of CLASP and its implementation at UC Davis, various student outcome measures are presented showing increased performance by students who took the CLASP course compared to students who took a traditionally taught introductory physics course. Measures we use include upper-division GPAs, MCAT scores, FCI gains, and MPEX-II scores.Comment: Also submitted to American Journal of Physic