Heart disease is the leading cause of death in the United States, contributing to 1 in every 4 deaths. One of the most common causes of heart failure is dilated cardiomyopathy (DCM), and approximately 35 percent of these cases are caused by mutations in genes encoding for a variety of sarcomeric, cytoskeletal and nuclear envelope proteins. There are nearly 1,500 separate mutations in sarcomere protein genes that can lead to DCM, but patients are not currently treated differently based on these unique mutations. In this study, I use two mouse models of genetic DCM, D230N and I61Q, which have mutations that occur in cardiac Tropomyosin (Tm) and cardiac Troponin C (cTnC), respectively. These mutations result in desensitization of the cardiac myofilament, leading to negative health consequences such as left ventricular dilation and reduced ejection fraction. I am testing the hypothesis that the ionotropic Levosimendan will be more effective in the I61Q cTnC DCM model because the drug binds directly to the N-terminal domain of cTnC, increasing its affinity for Ca2+ which results in increased force generated. At 3-4 months of age, cardiomyocytes are isolated and suspended in Tyrode’s buffer, an isotonic physiological solution. Cell shortening and relaxation was measured using video microscopy (IonOptix, Milton, MA) with pacing at 1.0 Hz and 37°C in presence of Levosimendan ( 1 µM) or control (DMSO). Compared to DMSO, 1 µM Levosimenden increased myocyte shortening in NTG mice (9.36 to 13.02 %, n=5) and I61Q cTnC (7.3 to 8.4 %, n=3) but had no impact on D230N Tm mice (10.4 to 8.2 %, n=5). My results suggests that Levosimendan is more effective in I61Q cTnC compared to D230N Tm model. In future studies I plan to make similar measurements using the myosin activator Omecamtiv Mecarbil.

Hemoglobinopathies, a hereditary condition involving abnormalities in the structure of hemoglobin, currently require expensive and highly sophisticated medical facilities to treat using gene therapy. The purpose of this study is to provide a highly portable and scalable approach using in vivo hematopoietic stem cell (HSC) gene therapy to potentially overcome these limitations. The main idea of our in vivo HSC gene therapy approach is to mobilize HSCs from the bone marrow, and while they circulate at high numbers in the periphery, transduce them with an intravenously injected HSC-tropic, helper-dependent adenovirus HDad5/35++ gene transfer vector system. The transduced cells return to the bone marrow where they remain long-term. This study followed one animal for 22 weeks after in vivo HSC transduction with a human-γ-globin expressing HDAd5/35++ vector using Sleeping Beauty transposase 100X for integration. Treatment with G-CSF/AMD3100 (a bone marrow stimulant) resulted in efficient HSC mobilization into the periphery blood circulation. The treatment was well tolerated and after in vivo selection, gamma-globin marking in peripheral red blood cells rose to ~90% and was stable during the duration of the study. Enrichment of gene-modified cells by in vivo selection was also reflected by an increase in mgtm and gamma-globin mRNA levels. Our data suggest that in vivo gene therapy with HDAd5/35++ is feasible and side effects can be minimized or prevented with appropriate pretreatment. This is the first proof-of-concept study that in vivo HSC gene therapy could be feasible in humans and provide the necessary portability and accessibility to reach patients in places with limited medical resources. Future studies will involve the optimization of HSC mobilization, gene transfer vectors and in vivo selection.

Heart disease is a major health pandemic, and the myocardial infarction (MI), also known as a heart attack, is the leading cause of death. This is because adult cardiomyocytes (CMs) present in the heart cannot divide and proliferate, preventing the heart from regenerating itself and repairing tissue damage. From developmental studies in rodents, we know the Notch signaling pathway is crucial in mediating expansion and proliferation of CMs during development, as impairments in Notch lead to cardiac defects. Notch is inactive in adult CMs, which suggests it plays a role in CM renewal; however, this mechanism is still unknown. Thus, our goal is to investigate the role of Notch in CM proliferation and cell cycle regulation. For the purpose of this study, we are using CMs differentiated from a human embryonic stem cell line, RUES2. Differentiation was completed over a 17-day period by culturing the cells as a monolayer. On Day 0, Chiron 99021 is added to activate Wnt signaling, promoting mesoderm formation. Wnt-C59 is added at Day 2 to inhibit Wnt signaling and differentiate the cells into progenitors, and B27 supplement at Day 6 promotes full differentiation into CMs. This adherent protocol recapitulates every step of natural heart development in vitro. Purity of the cell populations, as assessed by flow cytometry staining for cardiac troponin T (cTnT), was 97.1 ± 0.9% cTnT+. We then determined the proliferative capabilities of CMs in the presence of a Notch inhibitor, DAPT. DAPT inhibits gamma-secretase, a transmembrane protein that normally proteolytically cleaves Notch during signaling, thus inactivating the Notch pathway. Treatment with DAPT significantly decreased cell proliferation by about 50%, confirming that Notch directly affects CM proliferation. The results of this study increase our knowledge of CM physiology and the mechanisms behind cell cycle withdrawal, and provide new insights into improving CM renewal.

In 2018, tuberculosis (TB) was the leading cause of death by a single infectious disease, causing 10 million new cases and 1.5 million deaths. Upon Mycobacterium tuberculosis (Mtb) infection most people develop non-transmissible latent TB, and some develop active TB disease. Healthy hosts have a 5-15% lifetime risk of progressing from latent to active TB, and prior studies have demonstrated that a type I interferon-stimulated gene (ISG) signature can predict progression to active disease. Type I interferon (IFN) and ISG expression is induced by DNA-sensing pathways and has anti-viral functions. However, the innate immune mechanisms and genetics controlling type I IFN responses following Mtb infection are not well understood. We hypothesize that genetically regulated higher type 1 IFN responses are associated with lower anti-microbial responses and higher Mtb replication in macrophages, as well as increased risk of TB disease. To test this hypothesis, blood was collected from 40 healthy donors. Donors were genotyped using Illumina MEGA^EX SNP Array. Monocytes were isolated, differentiated into monocyte-derived macrophages (MDMs), and stimulated with 3 ligands (supercoiled plasmid DNA, cyclic guanosine monophosphate–adenosine monophosphate, and sheared calf thymus DNA) to activate DNA-sensing pathways and induce a type I IFN response. RNA was isolated at 4 and 24 hours. Interferon-beta (IFN-β) and interleukin-6 (IL-6) gene expression were quantified using Real-Time PCR. Donors had highly variable IFN-β induction upon ligand stimulation. Donors’ genotypes will be linked to these in vitro phenotypes to identify expression quantitative trait loci (eQTLs) that regulate IFN-β expression. We will assess if these functional polymorphisms in genes of interest are associated with TB disease using patient samples from a Brazilian cohort. The results of this investigation will identify novel pathways that control TB progression that can inform vaccine development and host-directed therapeutic approaches.

Of the estimated 38 million HIV infections globally, approximately 1-2 million are people living with HIV-2 (PLHIV-2). Infection with HIV-2, which is endemic in West Africa, is characterized by lower viral loads (VL) and slower disease progression to AIDS than HIV-1. However, many PLHIV-2 eventually progress to AIDS and death if left untreated. Antiretroviral therapy (ART) for HIV-2 is complicated by fewer effective drugs and significant challenges in drug resistance testing to identify appropriate therapy. This study aims to validate a novel method of HIV-2 drug resistance testing using nucleic acid from dried blood spots (DBS). One hundred and fifty DBS have been collected as part of two clinical studies of ART for HIV-2 in Senegal. HIV viral nucleic acid (DNA/RNA) is extracted and the regions encoding protease (PR) and reverse transcriptase (RT), which are the most common drug targets, are amplified by PCR, then sequenced. Sequence data are examined for evidence of drug resistance-associated mutations. To date, we have tested 115 samples with a median viral load of 84 (range: 0-24,000) and obtained drug resistance data from 43 (37.3%). Testing was most commonly successful from samples with higher viral loads; samples with viral loads >250 copies/ml were successful 75.7% of the time. We observed several drug resistance mutations, including PR V47A (5 patients), I50V (3 patients), and L90M (2 patients), and RT K65R (4 patients), Q151M (2 patients), and M184V (10 patients). Eight patients had no known resistance mutations. Once DBS testing is complete, we will select a subset of sequences to compare to genotypic resistance testing from corresponding plasma samples to evaluate the sensitivity of DBS-based testing vs. standard methods. DBS-based resistance testing has the potential to revolutionize ART for HIV-2 by allowing faster, easier, and cheaper assessment of drug resistance to optimize second-line therapy for HIV-2-infected patients.

Chronic myeloid leukemia is among myeloproliferative neoplasms that cause an increase of myeloid cells. All CML cases have a reciprocal translocation causing the novel fusion BCR-ABL gene causing the disease. BCR-ABL protein is a drug target and BCR-ABL mRNA is a diagnostic target. The polymerase chain reaction (PCR) is a technique used to diagnose and monitor treatment response in CML. Tyrosine kinase inhibitors (TKIs) have raised the survival rate of CML patients from an average lifespan of 6 years to a normal lifespan. However, monitoring and diagnosing CML patients requires expensive equipment and technical expertise. Medical testing and therapy of CML in lower to middle income countries is difficult, because drugs are expensive and access to sophistical equipment rare. However, when the disease is confirmed by testing, a non-profit organization (the Max Foundation) can provide TKIs free for life through partnerships with pharmaceutical companies. Dried blood spots (DBS) can potential stabilize nucleic acid for long periods of time. This approach has facilitated retrieval of the genetic material on the DBS that can be used to analyze more genetic analysis related to CML response and progression and has reduced transportation costs. In this study, we isolated DNA & RNA from DBS, quantified the genetic material, performed quality control, and carried out DNA & RNA sequencing-based on gene mutation assays. Tapestation and qPCR were performed to check the quality of the genetic material. DNA samples showed a higher DNA integrity number with an average of 7.7. However, the RNA samples demonstrated a lower RNA integrity number with an average of 1.8. The DNA samples had excellent quality whereas the RNA samples had poor quality but were acceptable enough to carry on with our study. These results will help us determine if genetic material on DBS can be used for more genetic analysis.

Infection is the most common cause of mortality among patients with multiple myeloma undergoing autologous stem-cell transplant. Understanding the risk factors that lead to higher probability of infection is instrumental in their prevention. Although some risk factors are known, there are no robust predictive models for infection among myeloma patients receiving a hematopoietic stem-cell transplant. In this research study, we compared a variety of machine learning algorithms to reveal clinical features pre-transplant that are most predictive of infection post-transplant. Using features extracted from the electronic health record by means of natural language processing and manual abstraction, we trained our model using patients diagnosed with multiple myeloma who received an autologous stem-cell transplant at the Seattle Cancer Care Alliance. Along with being robust to missing data, the model is capable of processing diverse, heterogeneous data and identifying a class of predictive factors that can guide treatment decisions. Preliminary results point to lab tests such as lymphocyte, neutrophil, and platelet counts as the strongest predictive features for infection, and further exploration is underway to understand the full impact of these and other features.

Skin cancer is the most prevalent cancer in the U.S., with an annual incidence of 5.5 million, exceeding all other cancers combined. Given the extremely high incidence of skin cancer, preventive measures are important. Intriguingly, multiple human epidemiological studies have demonstrated that caffeinated coffee, which is widely consumed in the U.S., is associated with decreased risk of developing skin cancer in a dose-dependent manner. Importantly, decaffeinated coffee had no such preventive effect. To determine the per-cup effect of caffeinated coffee consumption on skin cancer prevention, we performed linear regression analyses of previously published data. Males showed 3.0% reduced risk per cup of caffeinated coffee, whereas females showed 4.4% reduced risk. To determine the average reduced risk among males and females in the U.S. population, we extracted coffee consumption data from the National Health and Nutrition Examination Survey (NHANES) 2015–2016. Subsequently, the average number of cups of caffeinated coffee consumed was combined with the per-cup effect on skin cancer prevention. Among caffeinated coffee drinkers, males and females consumed on average 2.9 and 2.0 cups, respectively. However, both male and female caffeinated coffee drinkers coincided with 9% reduced risk of skin cancer incidence even though coffee consumption and the per-cup effect on risk reduction were different between males and females. The cancer-preventive effects of caffeinated coffee per cup appear to be stronger in females than in males. However, greater coffee consumption in males with weaker per-cup effect on skin cancer prevention leads to the net effect in males being comparable to that in females. For both genders, current levels of caffeinated coffee consumption may be preventing thousands of skin cancers annually.

Human immunodeficiency virus (HIV) infection is a significant global health issue, with approximately 75 million infections, and over 35 million deaths, since the beginning of the AIDS pandemic. The majority of these are attributable to HIV type 1 (HIV-1). A second form of HIV – HIV type 2 (HIV-2) – is endemic in West Africa and has spread to other areas with socioeconomic ties to the region. Historically, regimens for first-line treatment of HIV-2 have differed from those used in HIV-1-infected patients due to the intrinsic resistance of HIV-2 to nonnucleoside reverse transcriptase inhibitors. This distinction is coming to an end, as countries throughout West Africa are implementing a new WHO-recommended treatment regimen for first-line treatment of all HIV-infected patients, including those with HIV-2. The regimen, known as TLD, is comprised of the nucleoside reverse transcriptase inhibitors tenofovir and lamivudine and the integrase inhibitor, dolutegravir that has potent activity against both HIV-1 and HIV-2. Although treatment-emergent drug resistance has been well characterized for HIV-2 patients receiving tenofovir and lamivudine, there are few data regarding resistance mechanisms in patients receiving the third component of TLD, dolutegravir. The two objectives of my project are: (1) to construct a system for generating recombinant HIV-2 clones that encode and express integrase sequences from TLD-treated HIV-2 patients, and (2) to determine the in vitro susceptibility of viruses produced from the patient-derived clones to the integrase inhibitor dolutegravir. Specifically, I am engineering a plasmid vector into which patient-derived integrase sequences can be ligated for virus production and drug resistance testing in culture. The plasmid vector produced in this study will be used to characterize novel genetic pathways to dolutegravir resistance in HIV-2 and will help identify patients who are failing TLD treatment due to drug resistance. This information is crucial for improving treatment outcomes in HIV-2-infected individuals worldwide.

Human Immunodeficiency Virus (HIV) remains on the forefront of research due to the ongoing global epidemic. HIV is comprised of two genetically different types, HIV-1 and HIV-2. HIV-2 is inherently resistant to some classes of antiretroviral drugs, and many HIV-2 patients develop drug resistance to first-line and subsequent regimens. HIV-2 can further be divided into two distinct genetic groups: A and B. While both are endemic to West Africa, group A accounts for the majority of infections and remains the most studied of the two groups. In-depth knowledge of drug resistance in HIV-2 group B is lacking, as only a few patients with drug-resistant virus are described in the literature and there have been no systematic efforts to characterize the drug resistance patterns of HIV-2 group B isolates in cell culture. My project's goal is to build drug resistance mutations, documented in literature, for HIV-2 group A into a full-length HIV-2 group B infectious molecular clone. Those results are used to compare the relative drug resistance conferred by those mutations to the phenotypes observed for equivalent mutants of HIV-2 group A. More specifically, common drug resistance mutations are introduced into the pol gene of a group B clone, individual mutant clones are isolated, and these are used to transfect replication-competent cells for virus production and drug susceptibility testing. Inhibitors targeting the reverse transcriptase, protease and integrase targets of HIV-2 are evaluated. The resultant drug resistance profiles are then compared to those found in published datasets for HIV-2 group A to determine how HIV-2 group A and group B mutants differ in terms of the magnitude and/or scope of drug resistance. These data are essential for developing evidence-based treatment guidelines for HIV-2–infected patients that harbor drug-resistant group B strains.

The Toll-interacting protein (TOLLIP) is an adaptor protein involved in the signaling pathways of interleukin-1 (IL-1) and Toll-like receptors (TLRs) in innate immunity. Evidence in published literature suggests that TOLLIP acts as a negative regulator of IL-1 and TLR-mediated immune responses by inhibiting the activity of IL-1 receptor-associated kinase (IRAK1), a serine/threonine kinase in the IL-1 and TLR signaling pathway. Lipopolysaccharide (LPS) is a major component of the gram-negative bacteria cell wall that activates host immune response upon recognition by TLR4. We hypothesize that TOLLIP deficiency leads to impaired inhibition of the innate immune response, resulting in increased inflammation in LPS-induced lung injury. We treated wild type (WT) and TOLLIP knockout (KO) mice with LPS through intratracheal instillation and bronchial alveolar lavage fluid (BALF) was collected at 3 days post-injury. Lung inflammation was measured by BALF total white blood cell (WBC) count and cell differential, BALF total protein, and BALF cytokine levels. Contrary to our hypothesis, TOLLIP deficiency was associated with decreased inflammation in LPS-induced lung injury as demonstrated by lower polymorphonuclear (PMN) cell count and significantly lower levels of cytokines in KO mice. In future studies, we will examine the mechanisms by which TOLLIP positively regulates inflammation in the LPS model of lung injury.

Prostate cancer remains the second leading cause of cancer-related deaths of men in the US. Currently, the major challenge is to prevent the tumor cell from gaining the resistance to androgen deprivation therapy which almost inevitably leads to lethal castration-resistant prostate cancer. We are repositioning antiparasitic agents to develop a novel therapy to target androgen receptor (AR) mediated metastatic castration-resistant prostate cancer (mCRPC). Among them, bumped kinase inhibitor (BKI) 1553, which was originally designed to inhibit Toxoplasma calcium-dependent protein kinase 1, has shown to efficiently inhibit AR-dependent prostate cancer growth and AR signaling. Upon binding to its cognate ligand androgen, AR undergoes a conformational change to translocate from the cytoplasm into the nucleus to act as a transcriptional factor. The aim of my project is to identify the target site of BKI 1553 on AR transcription activation pathway. I examined the subcellular localization of AR by using biochemical subcellular fractionation and immunofluorescence assay. I also observed the phosphorylation status of AR via Western blot, and evaluated the level of AR signaling through a luciferase-based reporter assay and quantitative PCR assay of several AR target genes, including PSA, FKBP5, and Nkx3.1. The results of the study reveal the potential site of action of BKI-1553 in AR signaling. This study will provide a better understanding of the mechanism of BKI-1553 on prostate cancer and contribute to the development of a new therapy to mCRPC.

The blood-brain barrier (BBB) is a layer of tight-junction endothelial cells that make up the capillaries in the brain and strictly regulate what molecules can pass from the blood into the brain. Many molecules, including insulin, cannot passively cross this barrier but require an active transport system at the surface of the BBB. Once in the brain, insulin plays a role in memory and cognition. Indeed, Alzheimer’s disease is characterized by decreased sensitivity to insulin, which could be explained by a malfunctioning insulin receptor (IR) or impaired transport at the BBB. However, before we can begin to investigate the IR under disease conditions, we must first understand its standard regulation and function in a healthy system. Specifically, we aim to determine what factors mediate the endocytosis of insulin into the endothelial cells of the BBB. To do this, we focused on clathrin and caveolin, two proteins involved in different endocytic pathways. We performed cardiac perfusions on mice, where we first administered a drug to inhibit either clathrin or caveolin, and then we perfused with radiolabeled insulin. Afterwards, brains were collected and dissected into regions. Radioactivity was measured in the hypothalamus, olfactory bulbs, and whole brain, and the data was graphed over time to determine if there were changes in insulin binding or transport rates. Our results help elucidate the molecular processes necessary for insulin transport and binding at the BBB, which can ultimately help us understand how IR uptake and insulin transport may go awry in Alzheimer’s disease.

The field of regenerative medicine is approaching the goal of using stem cell therapy to replace part of an organ that has been damaged irreversibly. Our laboratory differentiates kidney organoids, 3D multicellular structures that functionally and compositionally resemble the respective organ they model, from human iPSCs (induced pluripotent stem cells). However, organoids we work with are largely avascular, whilst organs in vivo are highly vascularized. Our goal for this project is to build a microfluidic, vascular platform in which organoids can grow. To accomplish this, we adopted a microfluidic chip which was fabricated using soft lithography. Consequently, PDMS (polydimethylsiloxane), a polymer commonly used in soft lithography, was molded and bound to a glass coverslip using plasma binding. With this platform, we successfully engineered microvascular networks through vasculogenesis and angiogenesis and optimized the protocol of vascularization to sustain the cells by submerging the microfluidic chip in cell culture medium. Human umbilical vein endothelial cells and human lung fibroblasts were suspended in fibrinogen ECM (extracellular matrix), seeded into the microfluidic chips with micropillars to contain the cells within their respective channels, and developed into 3D microvascular networks with visible lumen. We then stained the vasculature with endothelial cell markers (i.e. CD31, CD54, VWF) and tested the perfusability by flowing polysterene beads through the microfluidic chip, observing the retention of polysterene beads within the vessels. Finally, we altered our design, specifically the height and width of the channels, to incorporate kidney organoids. Currently, we are using this platform for vascularizing kidney organoids and simultaneously implementing a flow system to induce shear stress on the microvasculature to attain physiological parameters. Ultimately, we aim to vascularize a kidney organoid to demonstrate the vascularization of stem cell tissue in vitro and see growth of tissue within our system, which would further our process in the translation pathway from bench to bedside for kidney regenerative medicine.

In the US, the number of children with obesity has reached a staggering 13.7 million. Though there are diets to assist weight loss, recent research suggests a neurobiological basis of obesity specifically related to the mediobasal hypothalamus (MBH), a critical brain structure involved in energy homeostasis, metabolism, and appetite. However, proliferation of hypothalamic gliosis, a cellular inflammatory response, disrupts the function and is shown, in rodents, as a key component in diet-induced obesity. Further conclusions reveal that highly caloric and high-fat diets, in rodents, can cause MBH gliosis. This project seeks to investigate the relationship between diet and hypothalamic gliosis in children, the latter assessed by magnetic resonance imaging (MRI). We expect children with an unhealthy diet to have an increased BMI z-score and greater evidence of MBH gliosis. Participants (N=192) were recruited as part of the longitudinal NIH Adolescent Brain Cognitive Development study. Anthropometric and demographic data were collected along with brain MRI T2-weighted images at the baseline visit. MBH gliosis was measured by using the signal ratio for T2 intensity of the mean bilateral MBH/Amygdala signal ratio; Putamen/Amygdala was used as control ratio. At the one year follow-up, the child’s habitual diet in the past year was assessed using a parent-report food frequency questionnaire. Higher total points represented a healthier overall diet. Additionally, follow-up anthropometric data was obtained to determine the child’s adiposity change over time. At baseline, mean age was 9.9±0.6 and 49% were males. Mean BMI z-score was 0.76±1.05, 19% were overweight and 23% with obesity. Preliminary results in a subset of participants (N=60) revealed a trend for an association between an unhealthy diet and evidence of MBH gliosis (t=1.59, P=0.117). By emphasizing the neurobiological basis of obesity, potential insights can inform targeted diet-related treatments of childhood obesity; thus, furthering the understanding of child obesity pathogenesis.

CRISPR is a powerful gene-editing tool with several advantages over state-of-the-art viral vectors used for gene therapy. However current methods to deliver CRISPR to human cells requires electroporation, which is cytotoxic and not viable for in vivo delivery. The Adair lab previously demonstrated gold nanoparticles (AuNPs) to passively deliver CRISPR gene editing into blood stem/progenitor cells, with Cas12a (Cpf1) nuclease resulting in high levels of homology-directed repair (HDR) compared to Cas9 when co-delivered with a single-stranded, homology-directed DNA template (HDT). Here we tested this AuNP-mediated delivery system in primary human T cells. We hypothesized that AuNP delivering Cpf1+HDT would also mediate high levels of HDR in T cells, compared to Cas9+HDT. To facilitate HDR readouts, the HDT encoded an 8 bp NotI restriction enzyme site. Both guide RNAs targeted the same C-C chemokine receptor type 5 (CCR5) gene locus. A mutation at this locus confers resistance most human immunodeficiency virus (HIV) strains. Briefly, CD3+ T cells from 8 different human donors (n=8 biological replicates) were either cultured overnight and exposed to AuNP-CRISPRs, or activated with CD3/CD28 beads prior to AuNP-CRISPR treatment. Test conditions for each donor included Mock-treated cells (negative control), naked AuNP, AuNP-CRISPR/Cpf1, AuNP-CRISPR/Cpf1+HDT, AuNP-CRISPR/Cas9, or AuNP-CRISPR/Cas9+HDT at identical cell and AUNP concentrations for 48 hours. Cell viability and counting by trypan blue dye exclusion assay demonstrated >70% viability and yield across all conditions tested. Gene editing analysis by tracking of indels by decomposition (TIDE), demonstrated the highest levels of gene editing with AuNP-CRISPR/Cpf1+HDT, with >90% of all editing being successful HDR. In non-activated T cells, mean gene editing levels were 19.7% across all donors (n=4). Final results for activated T cells are currently pending. These data demonstrate AuNP-mediated CRISPR/Cpf1 delivery as an efficient method for HDR in primary human T cells and suggest utility in human clinical applications.

There is evidence that the decision to admit patients to the hospital is not always correlated with their medical acuity. Emerging evidence suggests that a patient’s environment or psychosocial support may affect their likelihood of admission. However, there has been no comprehensive description of this body of literature. Given the breadth of this field, completion of a scoping review to map the current literature is warranted. The research question which this scoping review explores is: what factors other than medical acuity have been studied in relation to the physician’s decision to admit adult patients into the hospital? The scoping review method used in this review involves the following steps: identifying the research question, identifying and selecting studies that were relevant, charting and summarising the results extracted from these studies, and lastly consultation with stakeholders in order to add valuable insight. We collaborated with a research librarian to perform a comprehensive search of seven databases (PubMed, CINAHL, PsycINFO, EMBASE, Web of Science, Sociological Abstracts, and Social Science Abstracts) that resulted in 15,571 unique abstracts. A two-step screening process is currently underway. First, each abstract was reviewed independently by two research team members. From this, 1,368 abstracts were selected and are now being reviewed in full-text to confirm appropriateness for inclusion in the final analysis. We will next begin creating a narrative summary to describe the study designs used, factors studied, the terminology used, and outcomes identified. This research will provide a comprehensive view of trends that affect hospitalization and can be used to build new policies for treatment practices to improve healthcare in all communities.

Randomized controlled trials form the basis of translating research data into clinical practice. Adequately powered trials are essential to draw a precise and accurate conclusion. Our study aims to determine the proportion of randomized controlled trials published in the field of emergency medicine that were sufficiently powered to detect a true 25% difference in outcomes between study groups. We conducted a PubMed search to identify randomized trials related to emergency care published in 5 top-ranked general medical and emergency medicine journals in the last 10 years. Standard statistical techniques were used to calculate the sample size required to have at least a 90% probability of detecting a 25% difference in the primary outcome between study groups. Adequate power was defined as a planned sample size larger than the sample size required to detect this difference. We found that approximately half of the studies that met inclusion criteria reported no significant difference between study groups. 36.3% of these “negative” studies had adequate power to detect a 25% difference between study groups. When grouped by study setting, 26.4% of Emergency Medical Services (EMS) based studies as compared to 22.5% of Emergency Department (ED) studies had adequate power to have at least a 90% chance of detecting a 25% difference between study groups (p=0.11). Therefore, we concluded that a large proportion of randomized trials in the medical literature had inadequate power to detect a clinically significant difference between study groups. Our study would help to strengthen research practice in the field of emergency medicine and to advance knowledge in this field. 

Though there has been significant attention to preventing unnecessary hospital admissions, emerging evidence indicates that some patients who lack acuity of illness warranting admission continue to be hospitalized. In this study, we use a qualitative approach to further explore the hospitalist perspective and determine what factors influence the disposition decision for patients thought not to be appropriate for inpatient hospitalization. All admission calls to the acute care Medicine service at our hospital are logged into a Triage Database on an ongoing basis. This database allows the assessing physician to record a brief written narrative about the case and issues with making admission decisions. Entries were included in the study if admission calls originated from the Emergency Department and if the hospitalist assessed that the degree of patient’s medical acuity did not warrant hospitalization. Three hundred entries met inclusion criteria and were exported to Dedoose for qualitative analysis. A comprehensive codebook was iteratively created through independent coding accompanied by weekly meetings, with a final inter-rater reliability kappa score of 0.79. Using a grounded theory approach, a conceptual model of the admission decision-making process was constructed. We found that many factors are considered during admission assessment, both within and beyond physician control. The decision appears to occur in two distinct steps. First, an initial assessment considers patient factors, unmet medical and social needs, assessment of risk for discharge, and/or system-level barriers. A final admission decision is made after considering the influence of outside pressures such as: emergency department activity, coordination of care, resource constraint, and/or physician disagreements. Some of these factors may strongly outweigh the lack of medical acuity. Future work should investigate how these factors may vary at institutions with differing patient demographics and the effectiveness of possible interventions to alleviate the pressures of non-acute admission assessment factors.

Adolescence alcohol use and opioid addiction in adults are systemic issues afflicting the world, and thus, it is important to elucidate the long-term individual and relational consequences of both substance abuse disorders. The purpose of this experiment was to examine the long-term consequences of voluntary adolescent alcohol use on morphine tolerance, fentanyl self-administration, and the effects of previous opioid exposure on fentanyl self-administration in adulthood. Using a preclinical model to examine this hypothesis, adolescent rats had access to alcohol in gelatin form for twenty days, after which a three week withdrawal period occured. Morphine was then administered intraperitoneally for five days and morphine tolerance was measured by a tail-flick test for those five days. Finally, fentanyl self-administration occured in an operant chamber and self-administration will be measure by the amount of fentanyl consumed. My expected results were that adolescent alcohol use will increase morphine tolerance as evidenced by decreased tail-flick time, and fentanyl self-administration will also be increased. Additionally, I expect that previous opioid exposure will increase fentanyl self-administration. Future research should examine the neurobiological mechanisms by which adolescent alcohol use increases morphine tolerance and fentanyl self-administration and how previous opioid exposure increases fentanyl self-administration since these biological mechanism are not well understood.