Lipid Metabolism

Perilipin 2, also known as Adipose differentiation-relation protein or PLIN2, is a lipid droplet-binding protein present in almost every tissue. The absence of PLIN2 upregulates hepatic very low-density lipoprotein secretion, relieves hepatosteatosis, and improves whole body insulin resistance in mice. Despite of the importance in mediating lipid metabolism, the regulation of PLIN2 itself remains largely unknown. Previous reports have shown that X-box binding protein 1 (XBP1) is an important regulator of lipogenesis. XBP1 is a transcription factor that recognizes and binds to a consensus sequence, 5’-TGACGTGG-3’. Interestingly, when we looked through the promoter region of mouse Plin2 gene, we found that the consensus sequence is present in the Plin2 promoter. Therefore, we hypothesize that XBP1 might directly bind to Plin2 promoter and regulate the Plin2 expression. To test our hypothesis, we will perform the luciferase assay to examine whether the Plin2 promoter activity is regulated by XBP1. We first designed forward and reverse PCR primers, which include BglII and BamHI restriction enzyme sites respectively, to amplify the Plin2 promoter region (from -1100 to +40). We performed PCR and cloned the Plin2 promoter to a TA vector. The TA vector was then sequenced to exclude any point mutations. After sequencing, we sub cloned the Plin2 promoter into a vector containing a luciferase reporter. In the future, we will transfect 293T, a human embryonic kidney cell line, with the Plin2 promoter-luciferase vector we generated. We will compare the Plin2 promoter activity by measuring the luminescence in the presence or absence of XBP1

Impact of Continuity in Nursing Care on Patient Outcomes in the Pediatric Intensive Care Unit

Background: Nursing care is known to improve patient outcomes during hospitalization, but the mechanisms by which outcomes are improved have not been fully explicated. Continuity in nursing care (CINC) may be an important characteristic of nursing care delivery that impacts patient outcomes. However, evidence linking CINC to patient outcomes is limited. Purpose: The first aim of this study was to examine the relationship between CINC and patient outcomes - length of intensive care unit (ICU) stay, duration of mechanical ventilation, adverse events, and ICU-acquired infections - in a pediatric ICU. The second aim was to examine whether the match of nursing expertise to mortality risk enhances the relationship between CINC and patient outcomes. Methods: This cross-sectional study was a secondary data analysis of prospectively collected data that were merged from multiple databases from one pediatric ICU. The analytical database was a combination of four databases: the Nightingale Metrics database, the Virtual Pediatric Intensive Care Unit Performance System database, the Medical/Surgical Intensive Care Unit-Acquired Infection database, and the Safety Errors Reporting System database. The relationships between CINC and patient outcomes were assessed using a proportional hazard regression model and a logistic regression model. The final sample included 332 pediatric ICU subjects. Results: In multivariable regression analyses, more CINC was associated with a longer ICU stay and a longer duration of mechanical ventilation. CINC was not significantly associated with adverse events and ICU-acquired infections. A match of nursing expertise and mortality risk did not have a significant effect on the relationship between CINC and any of the four patient outcomes. However, the moderating effect of the match variable on the negative association between CINC and nurse-sensitive adverse event was significantly less for the matched group; specifically fewer different experienced nurses created a safer environment, than the mismatched group. Conclusion: This study provides preliminary data evaluating the relationship between CINC and pediatric ICU patient outcomes. Additional studies in other settings are needed to better understand these findings. Future research should focus on refining the measurement of CINC and exploring links between CINC and other outcomes such as patient/family satisfaction and being well-cared-for

Are systematic reviews up-to-date at the time of publication?

BACKGROUND: Systematic reviews provide a synthesis of evidence for practitioners, for clinical practice guideline developers, and for those designing and justifying primary research. Having an up-to-date and comprehensive review is therefore important. Our main objective was to determine the recency of systematic reviews at the time of their publication, as measured by the time from last search date to publication. We also wanted to study the time from search date to acceptance, and from acceptance to publication, and measure the proportion of systematic reviews with recorded information on search dates and information sources in the abstract and full text of the review. METHODS: A descriptive analysis of published systematic reviews indexed in Medline in 2009, 2010 and 2011 by three reviewers, independently extracting data. RESULTS: Of the 300 systematic reviews included, 271 (90%) provided the date of search in the full-text article, but only 141 (47%) stated this in the abstract. The median (standard error; minimum to maximum) survival time from last search to acceptance was 5.1 (0.58; 0 to 43.8) months (95% confidence interval = 3.9 to 6.2) and from last search to first publication time was 8.0 (0.35; 0 to 46.7) months (95% confidence interval = 7.3 to 8.7), respectively. Of the 300 reviews, 295 (98%) stated which databases had been searched, but only 181 (60%) stated the databases in the abstract. Most researchers searched three (35%) or four (21%) databases. The top-three most used databases were MEDLINE (79%), Cochrane library (76%), and EMBASE (64%). CONCLUSIONS: Being able to identify comprehensive, up-to-date reviews is important to clinicians, guideline groups, and those designing clinical trials. This study demonstrates that some reviews have a considerable delay between search and publication, but only 47% of systematic review abstracts stated the last search date and 60% stated the databases that had been searched. Improvements in the quality of abstracts of systematic reviews and ways to shorten the review and revision processes to make review publication more rapid are needed

Assessment of Cross-protective Responses to Emerging Variant Viruses

Viruses pose a great threat to human health. Outbreaks and pandemics due to different viruses occur in all areas of the world. Two viruses, which emerged globally in the 1980s and in 2020, that are still causing ongoing pandemics in 2021 are dengue virus (DENV) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), respectively. These viruses circulate widely and currently infect hundreds of millions of people. Immunity from a previous infection may confer protection against subsequent infection of a related virus. Thus, understanding the degree of cross-protection can help define correlates of protection that are necessary to determine which people are at greatest risk of infection. DENV is a mosquito-transmitted flavivirus that causes the most common arthropod-borne viral disease in the world. Infection with DENV causes a range of disease manifestations, with the most severe being hemorrhage, vascular leakage leading to organ impairment and circulatory collapse or shock, and consequently death. Historical observations have classified DENV into four serotypes due to lack of cross-immunity and protection between each group. Instead, immunity from one DENV serotype infection can enhance disease upon secondary infection with a different serotype. Therefore, establishing how immunity against one DENV impacts infection of similar and divergent DENV strains is essential to mitigate disease. As more variant DENV strains have been isolated in humans and inducing immunity in this small but expanding population over the last several years, it remains to be determined how this level of viral variation will impact global DENV immunity. In 2007, the most divergent DENV to date was isolated from a Dengue patient in Malaysia. Sequencing results determined it had a level of genetic divergence encroaching on serotype level status, and in 2015, this virus, DKE-121, was unveiled as a novel DENV-5 serotype. Immediately, questions arose of whether immunity from other DENV serotypes or tetravalent vaccines would remain protective or become enhancing in relation to DKE-121 infection. Herein, I examined the relationship of DKE-121 to its closest circulating DENV serotype, DENV-4, for cross-neutralization and cross-protection. My experiments show that DENV-4 immune sera can neutralize DENV-4 and DKE-121 similarly, whereas DKE-121 immune sera preferentially neutralized DKE-121 compared to DENV-4. This directionality in neutralization suggests that DENV-4 and DKE-121 are antigenically related, yet distinct. In passive transfer experiments, neither DENV-4 nor DKE-121 immune sera prevented infection of DKE-121 or DENV-4, respectively, suggesting that humoral immunity alone does not confer protection against the related virus. However, tetravalent vaccination in humans induced antibody responses that neutralized highly variant DKE-121 and DENV-4 vaccination was protective upon DKE-121 challenge in non-human primates. Thus, these findings highlight the complexity of DENV genetic variation and challenge the canonical classification of distinct serotypes. In 2021, in the context of the COVID-19 pandemic, another situation arose with urgent questions of how pre-existing immunity against one virus protects against newly emerging variant viruses. As therapeutics and vaccines were rapidly developed and widely distributed in hopes of ending the COVID-19 pandemic, SARS-CoV-2 variants of concern (VOC) emerged. These VOC contain mutations within the viral spike protein receptor binding domain (RBD), which SARS-CoV-2 uses to enter host cells. Since many vaccines and antibodies were developed using historical spike and/or RBD sequences to interfere with this interaction, it was unknown whether these VOC mutations would enable virus escape. Herein, I assessed therapeutic monoclonal antibodies and vaccine or infection-elicited polyclonal antibodies for cross-neutralization and cross-protection against the SARS-CoV-2 variant viruses. To understand whether monoclonal antibodies (mAbs) targeting historical spike protein would neutralize SARS-CoV-2 VOC, I tested a panel of mAbs against multiple viruses including those with spike protein point mutations corresponding to emerging variants. Most mAbs retained neutralizing potency to these variants compared to historical SARS-CoV-2. However, some mAbs showed reduced or completely abrogated neutralizing activity to viruses containing mutations at position 484 of the spike protein. Remarkably, serum polyclonal antibodies induced against historical spike from vaccination or natural infection showed reduced potency against E484K mutation-containing SARS-CoV-2 variants. This was true for mouse, hamster, and non-human primate sera from adenoviral vectored (ChAd-SARS-CoV-2-S) vaccinated animals in addition to human sera from Pfizer BNT162b2 mRNA vaccinated or previously SARS-CoV-2 infected individuals. Thus, SARS-CoV-2 variants containing a mutation at position 484 (e.g., B.1.351 (Beta) and B.1.1.28 (or P.1, Gamma) lineages, are of particular concern for antibody-mediated protection. As some therapeutic mAbs were given Emergency Use Authorization (EUA) or under development in advanced clinical trial stages, it was essential to understand whether they retain in vivo efficacy against emerging SARS-CoV-2 variants. These mAbs included monotherapy from Lilly under EUA (at the time) and combination mAb therapy from AstraZeneca, AbbVie, and Vir Biotechnology under development and from Regeneron and Lilly under EUA. When tested for neutralization, most mAbs retained potency against VOC including those from the B.1.351 and B.1.1.28 lineages. However, mAbs that had contact residues at position 484 (those from Lilly and AbbVie) showed reduced potency or complete loss of neutralizing activity against B.1.351 and B.1.1.28. When tested for therapeutic efficacy in K18-hACE2 mice, monotherapy and combination therapy with the Lilly mAbs failed to confer protection. The other combination therapy (from Regeneron) approved for EUA use, in addition to two others (AstraZeneca and Vir Biotechnology) were highly protective. While one mAb completely lost neutralizing activity in AbbVie’s cocktail against Beta and Gamma, the other retained activity and conferred protection against weight loss in mice. Thus, neutralizing sensitivity correlated with in vivo therapeutic efficacy, which has utility in screening mAbs against any emerging SARS-CoV-2 variants. Herein, I describe two viruses that have emerged globally within the last few decades or year and how pre-existing immunity may influence subsequent infection. For DENV, these interactions are complex and can be protective or enhancing between different viruses within this family. As more divergent DENV strains are found, the canonical serotype definitions become blurred, with unclear impacts on infection and immunization outcome. For SARS-CoV-2, many therapeutics and vaccines were generated using 2019 virus spike sequences and may be susceptible to escape from emerging variants. However, combination use of multiple therapeutic mAbs targeting different areas on the spike protein may help prevent additional viral escape. In addition, rapid development of COVID-19 vaccines has provided a framework, which can be quickly adapted to create boosters against the variants. Hence, the studies I describe here expand our understanding of the cross-protective immunity that exists against DENV and SARS-CoV-2 and with this knowledge, we are better equipped to end these pandemics and hopefully prevent future ones

White Teachers' Racial Identities, Perceptions of Student' Behaviors, and Symptoms of Burnout

Thesis advisor: Janet E. HelmsEducational research has examined factors contributing to teachers' burnout symptoms, including their perceptions of student behaviors (Ingersoll, 2003). Interestingly, teacher and students' races have been differentially related to teachers' perceptions of student behavior (Downey and Pribesh, 2004); this disparity in perceptions has been associated with teachers making more negative recommendations for African American students than for White students (Tenenbaum and Ruck, 2007). However, racial categories are not psychological constructs and offer little room for designing interventions to restructure teachers' perceptions of student behavior as a strategy to prevent teacher burnout. Since most teachers are White, using Helms's (1995) White racial identity model could offer a conceptual framework for examining different perspectives by which teachers understand their students' racial dynamics, which in turn, might affect how teachers feel, think, and act. Thus, if teachers' racial identity relates to their burnout symptoms, perceptions of student behavior, and recommendations, educational researchers could investigate more effective means of preventing teacher burnout symptoms and affect teachers' reactions to racially diverse students. White teachers (N = 237) completed an on-line survey containing an abbreviated White Racial Identity Attitudes Scale (Helms, 2011), behavior subscales of the Conners' Comprehensive Behavior Rating Scale - Teacher Form (Conners, 2008), Maslach Burnout Inventory - Educators' Survey (Maslach, Jackson, and Leiter, 1996), two teacher recommendations measures, and a demographic questionnaire. Results from Multivariate Analyses of Covariance suggested that teachers did not react differently to students' ethnic names or pictures with respect to their perceptions of students' behavior or teachers' likelihood of using specific recommendations. However, canonical correlations suggested that teachers' levels of burnout symptoms were related to their perceptions of students' Defiant Aggressive and Conduct Disorder symptoms. Moreover, teachers' biased and confused racial identity perspectives were strongly positively related to teachers' (a) burnout symptoms; (b) perceptions of angry, argumentative, and defiant behaviors; and (c) likelihood of using negative behavior management strategies with their students. Discussion included recommendations for educational training programs, methodological limitations, and implications of the results.Thesis (PhD) — Boston College, 2013.Submitted to: Boston College. Lynch School of Education.Discipline: Counseling, Developmental, and Educational Psychology

Emergency admissions and subsequent inpatient care through an emergency oncology service at a tertiary cancer centre: service users’ experiences and views

Purpose Avoiding unnecessary emergency admissions and managing those that are admitted more effectively is a major concern for both patients and health services. To generate evidence useful for improving services for direct patient benefit, this study explores service users’ views and experiences of emergency admissions and subsequent inpatient care.Methods Participants were recruited during a cancer-related emergency admission from a tertiary cancer centre with an emergency oncology service and emergency department. Semi-structured interviews were conducted with fifteen patients and twelve carers post hospital discharge. Interview transcripts were analyzed using framework analysis.Results Twenty patients experienced 43 emergency admissions over six months. Most admissions (35/43) followed patients presenting acutely or as emergencies with cancer treatment side-effects. Most admissions (35/43) were directly to an oncology ward following specialist advice, review and triage and thus unavoidable. Participants experienced outstanding inpatient care because of: prompt and effective symptom control and stabilization of acute conditions; continuity of cancer care and coordination between acute and long-term treatment; satisfactory professional-patient communication and information sharing; responsive, motivated and competent staff; and less restrictive visiting times. Gaps in care were identified. Conclusions Many emergency admissions are necessary for people with cancer. Future work should focus on: improving easy access to specialist advice and triage, and the process of admission; providing rapid palliation of symptoms and prompt stabilization of acute conditions, and satisfactory inpatient care; closing the circle of care for patients by actively involving primary care and palliative/end-of-life care services to address the complex needs of patients and carers

Convergent antibody responses to the SARS-CoV-2 spike protein in convalescent and vaccinated individuals

Unrelated individuals can produce genetically similar clones of antibodies, known as public clonotypes, which have been seen in responses to different infectious diseases, as well as healthy individuals. Here we identify 37 public clonotypes in memory B cells from convalescent survivors of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection or in plasmablasts from an individual after vaccination with mRNA-encoded spike protein. We identify 29 public clonotypes, including clones recognizing the receptor-binding domain (RBD) in the spike protein S1 subunit (including a neutralizing, angiotensin-converting enzyme 2 [ACE2]-blocking clone that protects in vivo) and others recognizing non-RBD epitopes that bind the S2 domain. Germline-revertant forms of some public clonotypes bind efficiently to spike protein, suggesting these common germline-encoded antibodies are preconfigured for avid recognition. Identification of large numbers of public clonotypes provides insight into the molecular basis of efficacy of SARS-CoV-2 vaccines and sheds light on the immune pressures driving the selection of common viral escape mutants

Immunochemistry of Elastotic Material in Sun-Damaged Skin

The nature of elastotic material in sun-damaged human skin was investigated by indirect immunofluorescence. Antibodies were used against the following components of the dermis: type I and type VI collagens, aminopropeptide of type I and type III procollagens, fibronectin, elastin, microfibrillar proteins, and basement membrane represented by the 7S domain of type IV collagen, laminin, and nidogen. The elastotic material exhibited marked fluorescence for elastin and microfibrillar proteins which codistributed with fibronectin. The presence of type I and VI collagens and procollagen type III were demonstrated to a lesser extent within the elastotic material. These results suggest that solar elastosis is primarily derived from elastic fibers and not from preexisting or newly synthesized collagens