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The full abstract for this thesis is available in the body of the thesis, and will be available when the embargo expires.

Contact dependent inhibition (CDI) systems facilitate antagonistic toxin exchange between closely-related bacteria in a proximity-dependent manner. CDI+ bacteria use the large (~300 kDa) cell surface protein CdiA to intoxicate their neighbors by binding to an outer membrane (OM) receptor. Receptor binding occurs at a central 200-300 aminoacids (RBD) and initiates cleavage and translocation of the ~200 carboxy-terminal residues of CdiA (CdiA-CT). Delivery of CdiA-CT requires receptor proteins at each stage of membrane transport into the target cell. We have previously identified three OM receptors for CdiA in Enterobacteria: BamA, OmpC/F, and Tsx. Using the CDI locus from E. coli STEC_O31 as a model (cdiBCAI), in Chapter 2 I introduce a fourth class of CdiA (CdiA4) that exclusively binds to lipopolysaccharide to initiate CdiA-CT translocation. CdiA4 is also linked to an accessory gene, cdiC, which encodes a predicted lysine acyltransferase. Site-directed mutagenesis and HPLC-MS reveals CdiC and its close homologues form a subfamily of bacterial lysine acyltransferases within the toxin-activating acyltransferase (TAAT) family that modify their cognate CdiA proteins to promote LPS binding. In Chapter 3, I use this new CDI system to investigate the interaction between CdiA and O-antigen and find that O-antigen presents a barrier to toxin translocation at the outer membrane of target cells. Finally, in Ch. 4, I identify four new inner-membrane (IM) protein receptors that facilitate toxicity by two new members of the CDI ionophore family. I demonstrate that the native activity of these receptors is not required for CDI, but their presence in the IM is necessary for ionophore activity both from the periplasm or from the cytosol. Combined with a primary sequence analysis of these CdiA-CTs, the results in Chapter 4 offer a wealth of opportunities to explore the mechanism of CDI ionophore activity further. Together, the results of this work present a new physiological application for bacterial protein lipidation, introduce a new family of accessory acyltransferases, reveal that O-antigen can be a barrier to CdiA-CT delivery, and advance our understanding of the functional diversity of CDI toxins and how they exploit target cell proteins for activity.

Caregivers of persons with dementia (PWDs) were socially isolated with little support during the COVID-19 pandemic “Stay-At-Home” order in the United States. To enhance social and emotional connection for diverse caregivers, a culturally/linguistically appropriate telephone intervention provided compassionate listening, mindful breathing, and COVID-19 safety education. The study purpose was to understand caregiving challenges and to evaluate the intervention for caregivers during the early pandemic using a qualitative approach. Twenty-three caregivers participated in the intervention provided by bilingual research assistants for 3 months. Call logs were used to describe the caregivers’ dialogue. Thematic analysis identified (a) the challenges, including fear of coronavirus disease, providing around-the-clock care, and forced isolation and negative emotions; and (b) caregivers’ experience with the intervention, including connecting with the outside, relief from emotional stress, reliable COVID-19 information, and reinformed caregiving skills. Results suggest that the telephone support was of benefit to diverse caregivers of PWDs during the pandemic by promoting social connection and reducing emotional distress.

Recent advances in genome sequencing have greatly improved our ability to understand and identify the causes of genetic diseases. However, there remains an urgent need for innovative, safe, and effective treatments for these diseases. CRISPR-based genome editing systems have become important and powerful tools in the laboratory, and efforts are underway to translate these into patient therapies. Therapeutic base editing is one form of genome engineering that has gained much interest because of its simplicity, specificity, and effectiveness. Base editors are a fusion of a partially deactivated Cas9 enzyme with nickase function, together with a base-modifying enzyme. They are capable of precisely targeting and repairing a pathogenic mutation to restore the normal function of a gene, ideally without disturbing the rest of the genome. In the past year, research has identified new safety concerns of base editors and sparked new innovations to improve their safety. In this review, we provide an overview of the recent advances in the safety and effectiveness of therapeutic base editors and prime editing.

Indiana University-Purdue University Indianapolis (IUPUI), The occurrence of colorectal cancer among those over the age of 50 is decreasing; conversely, the rate of diagnosis for those under 50 years old is increasing. While medical researchers scramble to identify the cause for this increase, young onset colorectal cancer (YOCC) patients and survivors are left to navigate a new normal. This new normal often includes awkward and troublesome concerns such as scarring, colostomy bags, and bowel problems. Contrary to those diagnosed with colorectal cancer later in life, those that are diagnosed at a younger age are forced to deal with these issues for many years. The purpose of this exploratory study was to identify privacy issues surrounding YOCC. Because of the significant increase in diagnoses, YOCC is now being researched independently from colorectal cancer in general. The topic of privacy has been researched in academic disciplines, including medicine. Privacy issues surrounding cancer have been researched, as well. Yet, the topic of privacy concerns facing YOCC patients/survivors has been overlooked. It is important to identify privacy concerns specific to YOCC patients/survivors as the information could help health care providers, communication scholars, and caregivers. Patient narratives were analyzed employing thematic analysis to identify privacy concerns of YOCC patients/survivors through the lens of Communication Privacy Management theory (CPM theory). Results indicated that participants discussed disclosure of their YOCC journey as a process. Within this disclosure process, YOCC patients/survivors identified specific privacy issues that influenced the way they disclosed or concealed information specific to their illness. There is a growing need for more research into the YOCC community due to the increase in diagnosis rates and their unique privacy concerns. Potential topics for future research include the impact of COVID-19, patient desire to help others, social media influence on disclosure, how patient disclosure could impact provider training, dating with YOCC, and specific demographic research.

As the demand for healthcare workers fluent in foreign languages continues to grow, so does the necessity for successful educational programs that produce providers with the appropriate language skills and cultural humility to serve the target population. Given the interdisciplinary nature of teaching languages for specific purposes, we propose a model of collaboration between experts in the different fields involved to train well-rounded professionals. In the particular case of languages for healthcare, professionals in both the medical and language learning fields should combine their expertise to offer students a well-integrated training that builds on their medical education at the same time that it provides them with meaningful opportunities to practice and develop their language and culture skills. While this paper focuses on Spanish, the main points discussed in it can be applied to teaching any language in the healthcare setting.

Complex socio-economic, political and demographic factors have driven the increased conversion of Europe’s semi-natural grasslands to intensive pastures. This trend is particularly strong in some of the most biodiverse regions of the continent, such as Central and Eastern Europe. Intensive grazing is known to decrease species diversity and alter the composition of plant and insect communities. Comparatively little is known, however, about how intensive grazing influences plant functional traits related to pollination and the structure of plant-pollinator interactions. In traditional hay meadows and intensive pastures in Central Europe, we contrasted the taxonomic and functional group diversity and composition, the structure of plant-pollinator interactions and the roles of individual species in networks. We found mostly lower taxonomic and functional diversity of plants and insects in intensive pastures, as well as strong compositional differences among the two grassland management types. Intensive pastures were dominated by a single plant with a specialized flower structure that is only accessible to a few pollinator groups. As a result, intensive pastures have lower diversity and specificity of interactions, higher amount of resource overlap, more uniform interaction strength and lower network modularity. These findings stand in contrast to studies in which plants with more generalized flower traits dominated pastures. Our results thus highlight the importance of the functional traits of dominant species in mediating the consequences of intensive pasture management on plant-pollinator networks. These findings could further contribute to strategies aimed at mitigating the impact of intensive grazing on plant and pollinator communities.

While the advent of checkpoint blockade antibodies has provided a powerful means to reinvigorate pre-existing T cell responses in the treatment of cancer, therapies capable of priming or expanding endogenous antitumor immunity are notably lacking within the current clinical cancer immunotherapy “toolbox”. Neoantigen peptide vaccines, dendritic cell-based vaccines, small molecule adjuvants, oncolytic viruses, and GM-CSF secreting tumor cells have demonstrated striking efficacy in murine tumor models yet, thus far, have failed to broadly impact the clinical treatment of advanced cancer. Difficulties faced translating success from the lab to the clinic have highlighted the need for further development of strategies to ignite effective antitumor immunity in formats which are cheaper, more tumor agnostic, and readily available in off-the-shelf formulations. With this aim in mind, we engineered tAb-pManTLR7, comprised of tumor cell-binding antibodies (tAbs) covalently linked to our pManTLR7 agonist, to localize and sustain delivery of a powerful adjuvant to the tumor microenvironment while endowing existing anti-tumor antibodies with a heightened vaccinal effect. Upon intratumoral injection, pManTLR7-armed antibodies bind and opsonize tumor cells, increase the residence time of pManTLR7 in the tumor microenvironment, and functionally link the delivery of endogenous tumor antigens with strong TLR7 activation. Increased tumor-specific cellular responses and intratumoral T cell accumulation upon tAb-pManTLR treatment mirror the salient outcomes of a traditional subunit or peptide vaccine but here, the tumor itself provides an endogenous source of antigen. In aggressive ‘cold’ models of melanoma and triple-negative breast cancer, tAb-pManTLR treatment slowed or eradicated established tumors in a CD8+ T cell-dependent manner and generated antitumor memory which protected mice against subsequent abscopal tumor challenge. Our engineered approach exemplifies a modular, off-the-shelf immunotherapy capable of activating systemic antitumor immunity with the sufficient magnitude and functionality required for therapeutic efficacy. This work provides a strong proof-of-principle for the further development of tAb-pManTLR as a therapeutic platform to treat numerous malignancies.

Background: The ProtecT trial reported intention-to-treat analysis of men with localised prostate cancer (PCa) randomly allocated to active monitoring (AM), radical prostatectomy, and external beam radiotherapy. Objective: To determine report outcomes according to treatment received in men in randomised and treatment choice cohorts. Design, Setting, and Participants: This study focuses on secondary care. Men with clinically localised prostate cancer at one of nine UK centres were invited to participate in the treatment trial comparing AM, radical prostatectomy, and radiotherapy. Intervention: Two cohorts included 1643 men who agreed to be randomised; 997 declined randomisation and chose treatment. Outcome Measurements and Statistical Analysis: Health-related quality of life impacts on urinary, bowel, and sexual function were assessed using patient-reported outcome measures. Analysis was carried out based on treatment received for each cohort and on pooled estimates using meta-analysis. Differences were estimated with adjustment for known prognostic factors using propensity scores. Results and Limitations: According to treatment received, more men receiving AM died of PCa (AM 1.85%, surgery 0.67%, radiotherapy 0.73%), whilst this difference remained consistent with chance in the randomised cohort (p = 0.08); stronger evidence was found in the exploratory analyses (randomised plus choice cohort) when AM was compared with the combined radical treatment group (p = 0.003). There was also strong evidence that metastasis (AM 5.6%, surgery 2.4%, radiotherapy 2.7%) and disease progression (AM 20.35%, surgery 5.87%, radiotherapy 6.62%) were more common in the AM group. Compared with AM, there were higher risks of sexual dysfunction (95% at 6 mo) and urinary incontinence (55% at 6 mo) after surgery, and of sexual dysfunction (88% at 6 mo) and bowel dysfunction (5% at 6 mo) after radiotherapy. The key limitations are the potential for bias when comparing groups defined by treatment received and outdating of the interventions being evaluated during the lengthy follow-up required in trials of screen-detected PCa. Conclusions: Analyses according to treatment received showed increased rates of disease-related events and lower rates of patient-reported harms in men managed by AM compared with men managed by radical treatment, and stronger evidence of greater PCa mortality in the AM group. Patient Summary: More than 90 out of every 100 men with localised prostate cancer do not die of prostate cancer within 10 yr, irrespective of whether treatment is by means of monitoring, surgery, or radiotherapy. Side effects on sexual and bladder function are much better after active monitoring, but the risks of spreading of prostate cancer are more common.

This chapter provides an update on rat (Rattus norvegicus) embryology, including the steps of preimplantation, implantation, and gastrulation. It also describes the process of organogenesis in the rat, with emphasis placed on neural tube formation, craniofacial development, and limb development. Teratology is the study of abnormalities of physiological development. Experimental teratology gained its current relevance after the thalidomide tragedy, which led to the development and revision of specific regulatory guidelines in drug safety. In studies to evaluate effects on embryo–fetal development, usually the rat is used as the primary species and rabbits as the secondary species. The International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use guidelines state that the rat is the most often used rodent species for reasons of practicality, general knowledge of pharmacology in this species, and the large amount of historical background data. This chapter describes in detail the various aspects of embryo–fetal developmental study design using the rat as the primary species.

Keywords: multiple myeloma, bone disease, osteogenesis, adipogenesis, adipose-derived stem cells, bone marrow, senescence, Dickkopf-related protein 1, systemic disease

Outpatient practice forms an integral part of medical care for patients needing care who are not admitted into the hospital, but receive the needed care and are discharged the same day which also largely includes primary care service. The growing demand for primary care physicians added by the complexity in managing outpatient practice effectively demands an optimized practice management system to coordinate care between different departments and provide high quality care on a timely fashion. The advent of health information technology has rapidly changed the way how outpatient practice has been done especially with the use of Electronic Health Records (EHR). Efficient use of such EHR has always been challenging, and several commercial software applications are available to facilitate outpatient practice management such as Microsoft's Amalga. However, commercial software does not aptly fit the needs of dynamically changing requirements in a hospital setting. Hence, the outpatient practice management department in Mayo Clinic which consistently ranks at the top in providing high quality care developed a custom outpatient practice application called Cohort Knowledge Solutions (CKS) to optimize the practice workflow and hence needs of the patients. This paper describes the design and development of CKS system, and its application in CTP/palliative care department and heart clinic which improved workload management, reduced follow up visits and readmission of patients using CKS.

The requisites for effectively addressing the demands of modern work have become a clear interest of organizational scholars. In line with this need has come increased focus on resilience in the wo...

Objectives To present the baseline patient-reported outcome measures (PROMs) in the Prostate Testing for Cancer and Treatment (ProtecT) randomized trial comparing active monitoring, radical prostatectomy and external-beam conformal radiotherapy for localized prostate cancer and to compare results with other populations. Materials and Methods A total of 1643 randomized men, aged 50–69 years and diagnosed with clinically localized disease identified by prostate-specific antigen (PSA) testing, in nine UK cities in the period 1999–2009 were included. Validated PROMs for disease-specific (urinary, bowel and sexual function) and condition-specific impact on quality of life (Expanded Prostate Index Composite [EPIC], 2005 onwards; International Consultation on Incontinence Questionnaire- Urinary Incontinence [ICIQ-UI], 2001 onwards; the International Continence Society short-form male survey [ICSmaleSF]; anxiety and depression (Hospital Anxiety and Depression Scale [HADS]), generic mental and physical health (12-item short-form health survey [SF-12]; EuroQol quality-of-life survey, the EQ-5D-3L) were assessed at prostate biopsy clinics before randomization. Descriptive statistics are presented by treatment allocation and by men’s age at biopsy and PSA testing time points for selected measures. Results A total of 1438 participants completed biopsy questionnaires (88%) and 77–88% of these were analysed for individual PROMs. Fewer than 1% of participants were using pads daily (5/ 754). Storage lower urinary tract symptoms were frequent (e.g. nocturia 22%, 312/1423). Bowel symptoms were rare, except for loose stools (16%, 118/754). One third of participants reported erectile dysfunction (241/735) and for 16% (118/731) this was a moderate or large problem. Depression was infrequent (80/1399, 6%) but 20% of participants (278/1403) reported anxiety. Sexual function and bother were markedly worse in older men (65– 70 years), whilst urinary bother and physical health were somewhat worse than in younger men (49–54 years, all P < 0.001). Bowel health, urinary function and depression were unaltered by age, whilst mental health and anxiety were better in older men (P < 0.001). Only minor differences existed in mental or physical health, anxiety and depression between PSA testing and biopsy assessments. Conclusion The ProtecT trial baseline PROMs response rates were high. Symptom frequencies and generic quality of life were similar to those observed in populations screened for prostate cancer and control subjects without cancer.