OBJECTIVES/GOALS: The history of immune suppression, especially CD4 nadir, has been shown to be a strong predictor of HIV-associated neurocognitive disorders (HAND). However, the potential mechanism of this association is not well understood. This study examined the relationship between CD4 nadir and brain atrophy. METHODS/STUDY POPULATION: Fifty-nine people with HIV participated in the cross-sectional study (mean age, 56.5 ± 5.8; age range, 41-69; 15 females; 46 African-Americans). High resolution structural MRI images were obtained using a 3T Siemens scanner. From a comprehensive 7-domain neuropsychological test battery, a global deficit score (GDS) and HAND diagnoses were determined for each participant. The correlation between CD4 nadir (the lowest ever lymphocyte CD4 count) and cortical thickness was investigated using a vertex-wise non-parametric approach with a conservative statistical threshold of p < 0.05 (FWE-corrected). RESULTS/ANTICIPATED RESULTS: Out of the 59 participants, 12 met standard Frascati criteria for asymptomatic neurocognitive impairment (ANI) and two met the criteria for mild neurocognitive disorder (MND). Across all participants, low CD4 nadir was associated with widespread cortical thinning, especially in the frontal and temporal regions. Higher GDS (indicating worse global neurocognitive function) was associated with bilateral frontal cortical thinning, and the association largely persisted in the subset of participants who did not meet HAND criteria. DISCUSSION/SIGNIFICANCE OF IMPACT: These results suggest that the low CD4 nadir may be associated with widespread neural injury in the brain, especially in the frontal and temporal regions. This spatial profile might contribute to the prevalence/phenotypes of HAND in the cART era, such as the frequently observed deficits in the executive domain.

ABSTRACT IMPACT: Small molecule readthrough compounds are a promising therapeutic with the potential to overcome nonsense mutations thereby enabling the production of functional ATM protein in patients with Ataxia Telangiectasia OBJECTIVES/GOALS: To generate a novel mouse model of Ataxia-Telangiectasia for testing small molecule readthrough compounds that both expresses a clinically relevant nonsense mutation and recapitulates the major symptoms of the disease, including a progressive loss of motor coordination not previously observed in prior A-T animal models. METHODS/STUDY POPULATION: Using a double-hit strategy to increase genotoxic stress, we generated a novel A-T mouse model that expresses a clinically relevant (c.103C>T) mutation in the Atm gene and a knockout of the functionally related Aptx gene. We then characterized the mouse across multiple domains related to the various symptoms related to the disease. This includes examination of survivability, immunologic function, cancer prevalence, and motor behavior and its associated cerebellar dysfunction and atrophy. Lastly, we tested the ability of small molecule readthrough compounds to enable production of ATM from tissue explants extracted from these ATM deficient mice. RESULTS/ANTICIPATED RESULTS: The double mutant mice display reduced survivability compared to control mice (53% vs. 97%; p<0.0001), dying at a clinically relevant rate of about 30% from thymomas. At postnatal day 400 (P400), only AtmR35X/R35X; Aptx-/- mice, and none of the controls expressing at least one wildtype Atm or Aptx gene develop a motor behavioral deficits that are associate with reduced Purkinje neuron diameter (8.0 ±0.4 µm vs. 9.92 ±0.5; p<0.01) and density (4.3 ±0.2 vs. 6.0 ±0.3 per 100 µm; p<0.05) as well as cerebellar atrophy (cerebellum/forebrain area 0.26 ±0.01 vs. 0.31 ±0.01; p<0.001). ATM deficient mice also display disrupted thymocyte development and metabolic function. When exposed to small molecular readthrough compounds, greater than 50% of the ATM protein is restored. DISCUSSION/SIGNIFICANCE OF FINDINGS: We have created a novel, clinically relevant A-T mouse model that develops a severe ataxia associated with changes in cerebellar function and atrophy as well as demonstrate the potential of SMRT compounds as an A-T therapeutic.

OBJECTIVES/SPECIFIC AIMS: Objectives: To determine genes that are shared between human and obese Zucker rat hypertrophic hearts, in order to identify potential early biomarkers and drug target for heart failure. METHODS/STUDY POPULATION: Four age-paired lean and obese Zucker rats were used. The human data are derived from doi:10.1152/physiolgenomics.00122.2016. RESULTS/ANTICIPATED RESULTS: We expect to find genes that are upregulated and downregulated in Zucker rats and humans that present cardiac hypertrophy. DISCUSSION/SIGNIFICANCE OF IMPACT: The genes and proteins determined from this study will provide future directions in order to determine whether obese Zucker rats are a valid model organism for the development of cardiac hypertrophy.

OBJECTIVES/SPECIFIC AIMS: Describe (1) the features of the first release of Duke’s myRESEARCHhome portal for researchers, and 2) the methods and results of adoption strategies METHODS/STUDY POPULATION: Through methods described previously (cite ACTS poster, 2016), the myRESEARCHhome portal team conducted a needs assessment to determine priorities for inclusion in the tool. Based on results of that assessment, the “minimal viable product” launched in June 2016 included the following features, organized into 9 distinct widgets: Access to all web-based research applications; ability to find and request research services; at-a-glance view of financial, protocol, and salary distribution information; access to financial and personnel reports; access to status of agreements and patents; access to CTSA-supported navigation services; visibility into required training and expiration dates; listing of announcements relevant to researchers; customized links area; ability to customize portal. The portal was developed using Ruby on Rails™, with a REACT grid framework. The development team, internal to Duke University, followed industry-standard best practices for development. After the initial release, the team employed several strategies to ensure awareness and adoption. Although written communications were an important factor for awareness, the presentations and hands-on studios proved most important. RESULTS/ANTICIPATED RESULTS: Use of the portal was directly related to in-person outreach efforts. There were small spikes after written communications, but strategies such as presentations, hands-on demonstrations, training sessions, and faculty meetings garnered the steadiest adoption rates. As of early January, 2017, almost 3000 users have interacted with the portal, with numbers rising steadily. There are an estimated 10,000+ faculty, staff, and trainees engaged in research at Duke. DISCUSSION/SIGNIFICANCE OF IMPACT: To maintain high adoption rates with the research community, engagement strategies must be ongoing. In addition to frequent in-person demonstrations, updates via written communications, and attendance at events, the portal team will employ a key adopt strategy—engaging the researchers in ongoing needs assessments. By maintaining the portal’s relevance to the needs of the research community, the tool can better improve the efficiency of research at a large academic medical center.

OBJECTIVES/GOALS: This study aimed to determine the prevalence of cardiovascular risk factors in a group of victims of CSA in Puerto Rico and determine the impact of both the offender and the number of victimizations on the presentation of cardiovascular risk factors. METHODS/STUDY POPULATION: A study design of a retrospective chart review at a clinical forensic service in Puerto Rico. The demographic variables were age, sex, and health plan; the CV risk factors were family health history, level of physical activity, blood pressure, BMI, and lipid profile. Sexual violence experience variables were sexual assault, sexual molestation, the relationship with the offender, and the number of victimizations. Medical records were used to identify cardiovascular risk factors and variables associated with child sexual abuse victimization. Central tendency and frequencies were used to describe the risk factors and victimization. The Mann–Whitney and Fisher exact tests were used to determine the differences between the type of victimization and the risk factors for cardiovascular health. RESULTS/ANTICIPATED RESULTS: Most of the victims were female (81%), with an average age of 10 (SD 3.8). According to the 31 reviewed charts, 55% were victims of sexual assault, the offender was a family member (84%), and the assault had occurred more than once (81%). The study also found that systolic blood pressure, diastolic blood pressure, total cholesterol, and body mass index (BMI) were at unhealthy levels (based on age and sex); when the victim reported sexual assault, the offender was a family member, and more than one assault occurred. Systolic blood pressure, diastolic blood pressure, and fasting blood sugar were statistically significant among victims who reported being either sexually assaulted or sexually molested when the offender was a family member, and the victimization occurred more than once. DISCUSSION/SIGNIFICANCE: This study indicated a higher prevalence of CVD risk factors in children victims of sexual assault. The blood pressure, lipid profiles, and BMIs were much higher than the standards. Early childhood screening is crucial in alerting health professionals to a child’s exposure to trauma.

OBJECTIVES/SPECIFIC AIMS: Abnormalities in sensorimotor behavior are present in the majority of individuals with ASD and associated with core symptoms. Cortico-cerebellar networks that control sensorimotor behavior have been implicated in ASD, but little is known about their function during sensorimotor actions. The purpose of this functional magnetic resonance imaging (fMRI) study was to examine cortical-cerebellar function during feedback-guided motor behavior in ASD. METHODS/STUDY POPULATION: Individuals with ASD (11-30 years; N = 18) and age-matched controls (N = 15) completed a visuomotor task of feedback-guided precision gripping during fMRI. Participants pressed with their right thumb and forefinger on a force transducer while viewing a green FORCE bar on a screen that moved upwards with increased force toward a fixed white TARGET bar. Individuals were instructed to maintain the FORCE bar at the level of the TARGET bar for 24 seconds. Target force levels were set at 20% and 60% of each participant’s maximum voluntary contraction (MVC). Force variability was characterized as the coefficient of variation (i.e., standard deviation of the force time series / mean force output; CoV). RESULTS/ANTICIPATED RESULTS: Mean force did not differ between groups indicating participants were able to follow task demands. Participants with ASD showed increased force variability (F(1,30) = 5.214, p = 0.03) at both 20% (d = .45) and 60% (d = .77) MVC compared to controls. Compared to controls, individuals with ASD showed decreased activation in left angular gyrus during the visuomotor task compared to rest (AG; maximum t = 4.31). Individuals with ASD also showed greater visuomotor activation compared to controls in ipsilateral ventral M1, extending anteriorly into posterior ventral pre-motor cortex (PMv; maximum t = −4.06, cluster size = 38 voxels). This difference reflected the finding that control participants showed a selective deactivation of ipsilateral M1/PMv during visuomotor behavior, whereas individuals with ASD did not show this pattern. A significant group x force interaction was observed for contralateral Crus I activation (maximum t = −2.42) that was driven by an increase in activity during 60% compared to 20% MVC in control participants, while individuals with ASD showed no change in Crus I activation between force levels. DISCUSSION/SIGNIFICANCE OF IMPACT: Increased force variability in individuals with ASD suggests impaired processing of sensory feedback to guide precision motor behaviors. Individuals with ASD did not show deactivation of right motor cortex during visuomotor behavior relative to rest, suggesting reduced ability to selectively modulate motor cortical output. Reduced activation in left AG may reflect an inability to integrate visual, haptic, and proprioceptive inputs to reactively adjust ongoing motor output. Failure to show force-dependent scaling of Crus I in ASD suggests lateral cerebellar circuits do not adapt sensory prediction and error processes to maintain precision motor output during more demanding conditions. Together, our results demonstrate multiple cortical-cerebellar mechanisms associated with sensorimotor imprecision in ASD.

OBJECTIVES/GOALS: The COVID-19 pandemic disrupted HIV care, though it prompted preventive measures for respiratory pathogens, particularly among PWH. We therefore quantified trends in respiratory ADE incidence during vs. before the COVID-19 pandemic to assess effects of these measures on non-COVID-19 illnesses. METHODS/STUDY POPULATION: We included PWH aged ≥18 years in care at the Vanderbilt Comprehensive Care Clinic in Nashville, Tennessee from 2017-2023. Individuals contributed time from the last of March 31, 2017 or clinic enrollment until the first of death, March 31, 2023 (study close), or final clinic visit (if there was no visit ≤12 months before study close). We described respiratory ADE incidences (per 1,000 person-years) in each year of the study; we used Poisson regression with robust variance to estimate the incidence rate ratio (IRR) and 95% confidence interval (CI) for respiratory ADEs in the three years following vs. before the World Health Organization’s pandemic designation for COVID-19 (March 2020). RESULTS/ANTICIPATED RESULTS: Among 4,880 persons contributing 19,510 person-years, 69 (1.4%) developed ≥1 respiratory ADE. Median age at cohort entry was 42.6 (interquartile range [IQR]: 32.1, 52.3) years and at first respiratory ADE was 43.6 (IQR: 36.1, 51.2) years. The overall average respiratory ADE incidence in the pre-pandemic period (March 2017-March 2020) was 4.5 (95% CI: 3.3-6.3) per 1,000 person-years and during the post-pandemic period (April 2020-March 2023) was 4.1 (95% CI: 1.8-9.0) per 1,000 person-years. When accounting for repeated outcomes and annual variation, the modeled respiratory ADE incidence was 10% lower (IRR=0.9, 95% CI: 0.6-1.4) during vs. before the COVID-19 pandemic. DISCUSSION/SIGNIFICANCE: Respiratory ADE incidence dropped 10% following the COVID-19 pandemic declaration, though the confidence interval for this change contains the null. It is plausible that nonpharmaceutical COVID-19 mitigation measures drove a brief but impermanent decline, though further research is needed to assess whether diagnostic biases also played a role.

OBJECTIVES/GOALS: Our goal is to explore the complex, the non-linear interplay among chronic conditions collectively contributing to a greater detrimental impact on the progression of Alzheimer’s disease (AD) than a single chronic condition alone in individuals with normal cognition, MCI, and AD. METHODS/STUDY POPULATION: We used longitudinal data from National Alzheimer Coordinating Center (n = 41,437) and focused on individuals with normal cognition (n =16,884, mean age (SD) = 70.72 (9.7)). Random forest models were used to predict newly developed MCI or AD from baseline to the most recent visits. We used self-reported baseline data on 50 chronic conditions and comprehensive clinical and demographic information (e.g., age, sex, APOE status, mini-mental status exam (MMSE) scores, education, BMI, and depressive symptoms). A binomial random forest was used to identify significant interactions (with p-values RESULTS/ANTICIPATED RESULTS: Our model demonstrated an AUC of 0.708 and a classification error rate of 25.4%. Variables of importance for predicting MCI or dementia were age, coronary artery bypass, depression, APOE status, smoking, and depressive symptoms. Two-way interactions, such as age X MMSE score, age X depressive symptoms, and age X BMI, were significant. Three-way interactions, including age X depressive symptoms X MMSE score, or depressive symptoms X BMI X MMSE score, were significant. However, when we explored the random forest model using only the chronic condition data, we found an AUC of 0.602 and an error rate of 27.15%. We found that depression, anxiety, hypercholesterolemia, stroke, and the interaction between BMI and anxiety were significant. DISCUSSION/SIGNIFICANCE: Random Forest models indicate that not only known factors including age, baseline cognitive status, and APOE status, but also chronic conditions like depression, anxiety, hypercholesterolemia, and stroke may predict cognitive impairment.

OBJECTIVES/GOALS: The HIV acquisition rate is higher for cisgender black women in the United States. This population has a much lower use of FDA-approved pre-exposure prophylaxis drug (PrEP) for HIV prevention. This systematic literature review examines factors that hinder PrEP use among cisgender black women. METHODS/STUDY POPULATION: PubMed, CINAHL, and Cochrane were used to conduct a systematic literature search. Inclusion and exclusion criteria were predetermined to narrow down the search and to identify the appropriate studies. The retrieved literature was exported to EndNote for deduplication. The literature was then uploaded into Rayyan for title and abstract screening. The two collaborators were blinded to limit bias. Literature characterized as conflict was resolved by an independent, third collaborator. Full-text screening of the selected studies was done and a PRISMA flow diagram was developed for identification, screening, eligibility, and to detail included literature. RESULTS/ANTICIPATED RESULTS: 185 articles were obtained from the systematic literature search. 172 were excluded and 13 full-text articles were accessed for eligibility. One was removed. 12 studies involving 974 study participants met inclusion criteria for analysis. Despite the disproportionally high risk of HIV infection, black women only represented 29% of the study participants. Top three barriers to low PrEP use were stigma, socio-economic factors, and personal factors. Stigma included concerns about HIV and sexuality. Socio economic barriers were related to homelessness and low income. Personal factors included substance abuse, incarceration, and commercial sex work. The data confirms that these risk factors were not always rigid but changed based on patient characteristics and external factors. DISCUSSION/SIGNIFICANCE: The study outcomes confirmed a multitude of risk factors that serve as impediments in the use of PrEP among cisgender black women. When these risk factors are identified and addressed, the decision to use PrEP among cisgender black women may increase, therefore this warrants further investigation.

OBJECTIVES/GOALS: Our goal is to develop a non-invasive stimulation technique using magneto-electric nanoparticles (MENs) for inducing and enhancing neuronal activity with high spatial and temporal resolutions and minimal toxicity, which can potentially be used as a more effective approach to brain stimulation. METHODS/STUDY POPULATION: MENs compose of core-shell structures that are attracted to strong external magnetic field (~5000 Gauss) but produces electric currents with weaker magnetic field (~450 Gauss). MENs were IV treated into mice and drawn to the brain cortex with a strong magnetic field. We then stimulate MENs with a weaker magnetic field via electro magnet. With two photon calcium imaging, we investigated both the temporal and spatial effects of MENs on neuronal activity both in vivo and in vitro. We performed mesoscopic whole brain calcium imaging on awake animal to assess the MENs effects. Furthermore, we investigated the temporal profile of MENs in the vasculatures post-treatment and its toxicities to CNS. RESULTS/ANTICIPATED RESULTS: MENs were successfully localized to target cortical regions within 30 minutes of magnetic application. After wirelessly applying ~450 G magnetic field between 10-20 Hz, we observed a dramatic increase of calcium signals (i.e. neuronal excitability) both in vitro cultured neurons and in vivo treated animals. Whole brain imaging of awake mice showed a focal increase in calcium signals at the area where MENs localized and the signals spread to regions further away. We also found MENs stimulatory effects lasted up to 24 hours post treatment. MEN stimulation increases c-Fos expression but resulted in no inflammatory changes, up to one week, by assessing microglial or astrocytes activations. DISCUSSION/SIGNIFICANCE OF IMPACT: Our study shows, through controlling the applied magnetic field, MENs can be focally delivered to specific cortical regions with high efficacy and wirelessly activated neurons with high spatial and temporal resolution. This method shows promising potential to be a new non-invasive brain modulation approach disease studies and treatments.

OBJECTIVES/SPECIFIC AIMS: The study aimed to determine the effects of bilateral frontal active transcranial direct current stimulation (tDCS) at 2 mA for 12 minute Versus sham stimulation on functional connectivity of the working memory network during an fMRI N-Back task. METHODS/STUDY POPULATION: Stimulation was delivered over bilateral frontal dorsolateral prefrontal cortex via and MRI-compatible tDCS device during an fMRI working memory task in healthy older adults in a within-subject design. RESULTS/ANTICIPATED RESULTS: Active stimulation compared with sham resulted in significant increases in functional connectivity in working memory related brain regions during the N-Back task. DISCUSSION/SIGNIFICANCE OF IMPACT: Older adults typically have reduced functional connectivity compared with young adults. Our findings demonstrate that a single session of tDCS can increase functional connectivity of the working memory network in older adults. Based on this mechanism of effect, tDCS may serve as an adjunctive method for interventions aiming to enhance cognitive processes in older adults.

OBJECTIVES/SPECIFIC AIMS: Obesity is a rapidly growing epidemic and long-term interventions aimed to reduce body weight are largely unsuccessful due to an increased drive to eat and a reduced metabolic rate established during weight loss. Previously, our lab demonstrated that exercise has beneficial effects on weight loss maintenance by increasing total energy expenditure above and beyond the cost of an exercise bout and reducing the drive to eat when allowed to eat ad libitum (relapse). We hypothesized that exercise’s ability to counter these obesogenic-impetuses are mediated via improvements in skeletal muscle oxidative capacity, and tested this using a mouse model with augmented oxidative capacity in skeletal muscle. METHODS/STUDY POPULATION: We recapitulated the exercise-induced improvements in oxidative capacity using FVB mice that overexpress lipoprotein lipase in skeletal muscle (mLPL). mLPL and wild type (WT) mice were put through a weight-loss-weight-regain paradigm consisting of a high fat diet challenge for 13 weeks, with a subsequent 1-week calorie-restricted medium fat diet to induce a ~15% weight loss. This newly established weight was maintained for 2 weeks and followed with a 24-hour relapse. Metabolic phenotype was characterized by indirect calorimetry during each phase. At the conclusion of the relapse day, mice were sacrificed and tissues were harvested for molecular analysis. RESULTS/ANTICIPATED RESULTS: During weight loss maintenance, mLPL mice had a higher metabolic rate (p=0.0256) that was predominantly evident in the dark cycle (p=0.0015). Furthermore, this increased metabolic rate was not due to differences in activity (p=0.2877) or resting metabolic rate (p=0.4881). During relapse, mLPL mice ingested less calories and were protected from rapid weight regain (p=0.0235), despite WT mice exhibiting higher metabolic rates during the light cycle (p=0.0421). DISCUSSION/SIGNIFICANCE OF IMPACT: These results highlight the importance of muscular oxidative capacity in preventing a depression in total energy expenditure during weight loss maintenance, and in curbing overfeeding and weight regain during a relapse. Moreover, our data suggest that the thermic effect of food is responsible for the differences in metabolic rate, because no differences were found in activity or resting metabolic rate. Additional studies are warranted to determine the molecular mechanisms driving the ability of oxidative capacity to assist with weight loss maintenance.

ABSTRACT IMPACT: My work focuses on understanding the role of an immunomodulatory pathway in Alzheimer’s disease as a potential therapeutic target. OBJECTIVES/GOALS: My long-term goal is to develop effective drugs to halt the progression of Alzheimer’s disease (AD). The overall objective of this translational work is to study the deregulation of the Lanthionine Synthetase C-like 2 (LANCL2) immunomodulatory pathway in AD. I hypothesize that stimulation of the LANCL2 pathway will inhibit progression of AD. METHODS/STUDY POPULATION: These studies use a transgenic rat model of AD, and a LANCL2-binding drug, BT-11, that crosses the BBB. Tg-AD Fisher 344 rats express human APP with the Swedish mutation and human presenilin-1 with the ? exon 9 mutation. Tg-AD rats develop age-dependent AD-like pathology. Rodent chow containing BT-11 (10mg/Kg bw) was administered orally from 5 months of age (pre-pathology) to 11 months of age. Eight groups of rats (8 rats/group) were included: WT and Tg-AD x 2 sexes x 2 conditions (BT-11 treated and untreated). Rats are analyzed for: (1) Spatial learning and memory with an active place avoidance test. (2) AD pathology including amyloid plaques and activated microglia, with hippocampal immunohistochemistry. (3) LANCL2-signaling and other AD relevant pathways, with hippocampaI western blot and RNAseq analyses. RESULTS/ANTICIPATED RESULTS: I predict that BT-11 will prevent and/or mitigate some aspects of AD pathology displayed by the AD transgenic rats (Tg-AD). Tg-AD rats treated with BT-11 should perform better in the spatial learning and memory tasks, than the untreated Tg-AD rats. I also expect that the immunohistochemical analysis will reveal reduced pathological hallmarks, including amyloid plaques and reactive microglia, in hippocampal tissue of BT-11 treated versus untreated Tg-AD rats. Through western blot and RNAseq analyses, I will establish how BT-11 treatment affects the LANCL2 signaling cascade as well as other AD relevant pathways, in treated and untreated rats from both sexes and both genotypes. DISCUSSION/SIGNIFICANCE OF FINDINGS: My proposed translational research will address the potential of targeting the LANCL2 pathway to improve AD treatment. Discovering the effects of stimulating the LANCL2 pathway on AD pathology is a novel approach in AD drug development. I expect that my studies with BT-11 will positively influence therapeutic outcomes of this devastating condition.

OBJECTIVES/SPECIFIC AIMS: To compare methods of ascertaining prevalence for adrenal incidentalomas METHODS/STUDY POPULATION: Retrospective electronic medical record study using Looking Glass Clinical Analytics (Streamline Health, Atlanta, GA, USA) at an urban university medical center. All patients with CT or MR imaging of the abdomen between 1997 and 2014 were identified. Patients with a documented diagnosis (ICD-9 code or problem list) for any history of adrenal disease were excluded. The prevalence of adrenal incidentalomas was ascertained by 2 different detection strategies: (1) documented diagnosis of adrenal incidentaloma or (2) imaging reports containing in the same sentence “adrenal” and “nodule*,” “adenoma*,” or “mass*,” and not containing “no” and “adrenal” in the same sentence. Adrenal pathology surprise was further established in the second approach by excluding patients having previously undergone adrenal lab testing (cortisol, aldosterone, catecholamines, adrenocorticotropic hormone, renin) or having been registered in the cancer registry for any cancer excluding superficial skin cancers. RESULTS/ANTICIPATED RESULTS: In total, 194,624 individuals were identified in our initial search, from which 1056 were excluded for past adrenal disease (Table 1). Detection by the documented diagnosis method yielded 1578 cases (0.8%), compared with 13,697 cases (7.1%) by the imaging report method (Figure 1). Further restricting detection to true “Adrenal Surprise” by excluding those with any past adrenal lab testing and cancer history yielded 10,568 cases (6.1%). Validation studies for the 7.1% prevalence with 100 records revealed an adrenal incidentaloma positive predictive value (PPV) of 98%. When restricted to size ≥1 cm the PPV was 84%. DISCUSSION/SIGNIFICANCE OF IMPACT: Comparing our first strategy using documented diagnoses as criterion for incidentaloma as used in a recent paper by Lopez D (Annals of Internal Medicine 2016; 165: 533–542), we found a prevalence of 0.8% in our population similar to her 0.6%. However, when searching at the level of radiology report text, we found a prevalence ten-fold greater at 7.1%. Therefore, adrenal incidentalomas are more robustly identified by searching radiologic reports.

OBJECTIVES/SPECIFIC AIMS: Although two Ribeye protein isoforms have been identified in zebrafish, information about the identities of their variants is incomplete. This study aims to identify and characterize both of the Ribeye isoforms and their splice variants. METHODS/STUDY POPULATION: Immunohistochemistry was performed on the retina and neuromasts of zebrafish larva and adults.Ribeye expression was analyzed by western blot. Ribeye proteins will be separated, isolated and identified by mass spectrophotometry. RESULTS/ANTICIPATED RESULTS: Immunohistochemistry performed on larval and adult zebrafish retinas revealed the expression of Ribeye A in the inner and outer plexiform layers. Ribeye B was likewise expressed in both plexiform layers in larval zebrafish, but more pronounced expression in the outer plexiform layer in the adult zebrafish retina. Immunohistochemical experiments also demonstrated the co-expression of both Ribeye isoforms in the hair cells of both larval and adult neuromasts. Analysis of Ribeye expression by western blot showed the presence of more than the three previously identified variants. Current experiments are being conducted to characterize the additional Ribeye variants. We expect to identify the residual Ribeye protein as a result of this analysis. DISCUSSION/SIGNIFICANCE OF IMPACT: This study is necessary in order to gain a clear understanding of Ribeye expression in zebrafish tissues. Doing so will enable us to target this protein for gene editing to address outstanding questions about the mechanisms that govern ribbon synapse function. Synapse and synapse-associated proteins are involved in a wide-array of diseases that arise as a result of their dysfunction (e.g. blindness, deafness, bradycardia, autism spectrum disorders, and schizophrenia). Thus, it is important for us to identify the shared and distinct mechanisms that give rise to diseases associated with synaptic dysregulation. Such information could provide the basis for novel therapeutic interventions for synaptic disorders.

OBJECTIVES/SPECIFIC AIMS: Müller cells, radial glial cells of the retina, are the principal repository of xanthophyll pigment (lutein, zeaxanthin, meso-zeaxanthin), which are modifiable by diet and visible clinically by autofluorescence imaging. To understand the structural basis of xanthophyll visualization in vivo, we used 3-dimensional electron microscopic reconstruction of Müller cells surrounding one cone in a healthy human fovea. METHODS/STUDY POPULATION: From a 21-year-old male organ donor, dissected retinas were rejuvenated by oxygenated Ames medium then fixed in 4% glutaraldehyde. A tissue block 3.5 mm2 centered on the fovea was prepared for Automated Tape Ultramicrotomy (Kasthuri et al., Cell 162: 648–661, 2015). From 1462 serial 65 nm horizontal sections, an area ~250×250 μm was imaged at 6 nm xy resolution. Images were stitched and aligned. TrackEM software on a pen display was used to trace, reconstruct, and display cone #5 (of 186) and its contacting Müller cells. RESULTS/ANTICIPATED RESULTS: Cone 5 is ensheathed by 2 types of Müller cells, outer and inner (Dacey, ARVO, 2016). The outer cell is first seen at the external limiting membrane (ELM) between cones 5 and 17. Moving inward from the ELM, it tightly wraps around cone 5’s fiber in a C-shape profile for 78 µm. This Müller cell also intermittently projects to neighboring cones, 2 of which were close to cone 5 at the ELM. As cone 5’s axon approaches the pedicle, it contorts into a corkscrew. The outer cell fluidly molds to this changing shape. At this level, this Müller cell doubles in volume to encompass not only cone 5, but also cone 17 and another Müller cell. In the final 17 µm of the block the Müller cell’s volume quickly dissipates as it sends a small projection towards the internal limiting membrane, eventually encasing an OFF midget bipolar cell also associated with cone 5. In contrast to this outer cell, an inner Müller cell adjoining cone 5 spans only 19 µm, interacting directly with cone 5 and the outer cell for 3.9 µm. DISCUSSION/SIGNIFICANCE OF IMPACT: Neural-glial relationships in a human fovea are visible through 3-dimensional volume EM. The volume of Müller cells in the fovea was impressive, consistent with a pivotal role in the health of cone photoreceptors and xanthophyll homeostasis. It is possible that individual glia also ensheath the post-receptoral neurons in a cone-driven circuit, supporting the concept that xanthophylls contribute to neural efficiency in vision.

OBJECTIVES/SPECIFIC AIMS: Retinitis pigmentosa (RP), also known as night blindness, is an incurable disease which affects ~1 in 4000 individuals globally. Since there are no effective treatment options for RP, the goal of this project is to identify novel drug treatments that can prevent or slow the disease progression. To this end, we optimized a behavioral assay, visual-motor-response (VMR) assay, to investigate rod function (Ganzen et al., ARVO, 2017; Ganzen et al., IJMS, 2017). This was done utilizing a transgenic zebrafish RP model expressing human rhodopsin with the Q344X mutation. In this study, we used this model to perform a proof-of-concept screen for drugs which may improve the vision of the larvae. METHODS/STUDY POPULATION: To screen for beneficial drugs, the SCREEN-WELL® REDOX library was chosen for screening. This library was selected to identify a compound that may alleviate any excessive oxidative stress in the diseased retina. The Q344X zebrafish line suffers from significant rod degeneration by 7 days postfertilization (dpf) and displayed deficits in VMR under scotopic conditions (Ganzen et al., ARVO, 2017). The Q344X larvae were drug treated beginning at 5 dpf at 10 μM. Compounds that were toxic at this concentration were retested at 1 μM. The 5 dpf stage was chosen as most of the rods are intact, and these concentrations were chosen to optimize the drug effect based on similar studies. Hits were identified by assays that provided a robust and reproducible enhancement in the Q344X VMR. The retinae of any drug hits were dissected from larvae crossed with a rod EGFP reporter line and whole-mounted to analyze rod survival via fluorescence. To determine if drug effects were exerted through the retina, eyeless chokh mutant zebrafish were exposed to the drug and tested with the same assay. RESULTS/ANTICIPATED RESULTS: Of the 84 compounds tested, we identified 1 drug that ameliorated the VMR of the Q344X scotopic VMR. Eyeless chokh mutant zebrafish larvae did not exhibit the same VMR when treated with the same drug. Histological analysis suggested increased rod survival in the drug-treated retina of Q344X mutants. DISCUSSION/SIGNIFICANCE OF IMPACT: These results indicate that the vision of the Q344X zebrafish was improved via this beneficial drug treatment. Since eyeless chokh larvae did not respond to the same treatment, the drug likely mediated its positive effects through the Q344X retina, likely by improving rod survival. Together, our results have identified a beneficial drug that may treat RP.

OBJECTIVES/GOALS: Our overarching aim is to examine, in an African American population, cognitive, affective, and neurophysiological processes, as well as risk-taking behavior, in response to racial stigma cues. While accounting for individual differences, we aim to see how these processes and drug-cue reactivity are impacted or altered by exposure to racial cues. METHODS/STUDY POPULATION: Participants will be African Americans between 18 and 25 years of age, equally distributed across genders. We will recruit 75 participants in order to have adequate power to conduct our intended analyses—particularly pertaining to individual differences in risk behavior outcomes. Participants will be asked to complete demographic and self-report questionnaires. Participants will also be asked to complete computerized tasks while their physiological responses (heart rate, skin conductance, and electroencephalographic (EEG) data) are recorded. The tasks are as follow: resting, gambling, go/no-go, picture viewing (positive, negative, and neutral images), and a drug cue image set. These tasks will be repeated after the participant views a racial stigma image set to evaluate the impact of discrimination. RESULTS/ANTICIPATED RESULTS: Data from 18 participants has been collected. Data will be periodically preprocessed and validated (e.g., 1 participant was removed due to data recording errors, so the current valid Nis 17). Generally, we anticipate that behaviors and neural activity will be modulated across all tasks after viewing the racial stigma image set. Specifically, (a) cognitive and affective processing of singular events of racial stigma may indicate a stress response, (b) modulation from chronic experiences of racial stigma render neural systems increasingly sensitive to stigma cues, and thereby less equipped to regulate stress response, (c) the impact of these processes on altering risk behavior (may increase such behaviors), and (d) the impact of these modulations on altering drug-cue reactivity (may amplify reactivity). DISCUSSION/SIGNIFICANCE: The study will identify factors that contribute to stress and risk behavior among African Americans. A substantial gap continues to exist regarding the nature of risk behavior among African Americans, despite the fact that African Americans represent a health disparity population with unique vulnerabilities to health-relevant risk behavior.

OBJECTIVES/GOALS: Chronic kidney disease (CKD) affects nearly 40% of adults with diabetes. Our objective is to assess trends in risk factor control and use of 2022 ADA and KDIGO guideline-recommended medications. METHODS/STUDY POPULATION: Using data from 1988 to 2020 from the National Health and Nutrition Examination Survey, we will examine trends in sociodemographic risk factors and glycemic, blood pressure, and lipid control among adults with CKD and diabetes. Glycemic control will be defined as a hemoglobin A1c (HbA1c)<7%, blood pressure control will be examined at cutoffs of 130/80 and 120/80 mmHg, and lipid control will be defined as a fasting triglyceride level ≥150 mg/dL, a low-density lipoprotein (LDL) <100 mg/dL without atherosclerotic cardiovascular disease (ASCVD), or LDL<70 mg/dL if they have ASCVD. We will assess changes in the use of commonly used antidiabetic, antihypertensive, and lipid-lowering medications. RESULTS/ANTICIPATED RESULTS: We hypothesize that from 1988-2020, blood pressure control has improved while glycemic and lipid control has not improved among adults with diabetes and CKD. We expect decreases in sulfonylurea use and increases in DPP-4 inhibitor, metformin, ACE inhibitor, angiotensin receptor blocker, statin, and insulin use over time among those with diabetes and CKD. In addition, there is likely a significant gap between those who are eligible to use newer medications like SGLT2 inhibitors or GLP-1 receptor agonists and who are currently using them within this subpopulation. DISCUSSION/SIGNIFICANCE: This study will examine adherence to guideline-recommended management and identify gaps in care for adults with CKD and diabetes, which may inform how best to optimize medication use for cardiorenal protection in this high-risk patient population.

OBJECTIVES/GOALS: All IN for Health has initiated social media engagement strategies to create bi-directional communication around improved health and research literacy with the public. In addition, the All IN for Healths community-led advisory board, comprised of diverse individuals and organizations, will provide insight on research design and dissemination. METHODS/STUDY POPULATION: The Indiana Clinical and Translational Sciences Institute (CTSI) has committed to enhancing the publics engagement efforts in the All IN for Health program. This program was established in 2017 to improve the health of Indiana residents through the promotion of health resources and opportunities to participate in research. In 2021, the Indiana CTSI decided to integrate public engagement more formally. An official collaboration with the Indiana CTSIs Community Health Partnerships program was created, a digital engagement specialist was hired, and several new community and population health advisory board members were welcomed for quarterly meetings representing the rural and urban areas of the state, faith-based communities, health departments, and underrepresented groups in research. RESULTS/ANTICIPATED RESULTS: A social media campaign, informed by the All IN for Health advisory board was launched in November 2021, and the advisory board has been meeting on a quarterly basis. All IN for Healths established priority area is currently COVID-19 vaccination uptake, and our prioritized audiences are underserved and under resourced in urban and rural communities, and minoritized populations. All IN for Health has posted approximately 20 times per month on topics including COVID-19, Alzeheimers Disease, Diabetes, fitness, mental health, and research studies with a goal to increase followers and engagements by 20%. The hope is that this engagement will also result in an increase in the monthly e-newsletter subscribers and study volunteer registry participants. DISCUSSION/SIGNIFICANCE: All IN for Health has initiated new public engagement efforts to increase the value of the services offered to Indiana residents. Through consultation and co-design with members of the public and dissemination of research opportunities All IN for Health will be better suited to improve health and research literacy.