Disrupting Six/Eya Signalling as New Therapy for Lung Fibrosis

Idiopathic pulmonary fibrosis (IPF) is the most common type of interstitial lung disease, with a median survival of 2-4 years from the time of diagnosis [1]. It is estimated that the prevalence of IPF in the US is approximately 10-60 cases per 100,000 people, with limited pharmacological therapies available [2, 3]. IPF is a chronic, progressive disease characterized by alveolar injury, increased extracellular matrix (ECM) deposition and resultant alveolar destruction. Macroscopically, this leads to poor lung compliance, impaired trans-alveolocapillary membrane gas exchange and ultimately, end-stage respiratory failure, necessitating lung transplantation [2, 4, 5]. Several non-genetic risk factors, such as male sex, older age, and smoking, increase the risk of developing IPF [4, 6]. More recently, several genetic risk factors for IPF have also been discovered, including a single-nucleotide polymorphism (rs35705950) in the promoter region of MUC5B [7-9], which codes for an essential protein for airway clearance and innate immune response, along with genes associated with telomere maintenance, such as telomerase RNA component (TERC) and telomerase reverse transcriptase (TERT) [1, 10].

Effect of Dry Needles on Volume of Subcutaneous Immunotherapy

Allergen Immunotherapy (AIT) is a safe therapy approved for patients with allergic rhinitis, allergic asthma, and atopic dermatitis.

Mammary Tissue Host-Microbiome Interactions in Breast Cancer Development

Through this project we will determine the role of the mammary tissue microbiome in breast cancer development using 16S ribosomal RNA sequencing and dual-transcriptomic sequencing. In the first two years of this project we have selected and received 165 samples from the Susan G. Komen and Indiana University Simon Cancer Center TissueBanks. We completed 16S rRNA sequencing on DNA isolated from all samples in this cohort and note a distinct microbial compositional signature that is associated with breast cancer development. We anticipate submitting this work for publication by February 2021. Due to COVID-19, the RNA isolations from these samples have been delayed until we can return to campus. We anticipate completing RNA isolations in Summer 2021 and will begin our analysis of the RNA sequencing data in Fall 2021. Regardless of these delays, the project is well underway. Results from this work will be key in characterizing host-microbiome cross-talk in the pathogenesis of breast tumor development.

Impact of Operational Sleep Disruption on PTSD-Relevant Fear Learning Processes

This report covers Year 2 of the project. This project examines the impact of disturbances in normal sleep and circadian regulation on mechanisms underlying vulnerability to, and maintenance of, posttraumatic stress disorder (PTSD). The goal of Year 2 was data collection and initial validation of data collection procedures and harmonization with Monash. We proposed to complete 30 participants (45 in total with year 1). Progress was slowed due to COVID19, which prompted our VA facility to shut down all human research the fall. Our study, which requires a hospital stay, was finally approved for collection in October with mitigation procedures in place. We have completed 18 subjects (30 of target). However we do have 6 weeks now booked for collection and a large list of applicants for the study. During the COVID period we examined data for errors and trained and harmonized sleep scoring across the study. We have seen exceptional interest in the study since recruitment opened and hope to make up for lost time this year.

Factors Associated With Outcomes in Patients With Vestibular Symptoms Related to Traumatic Brain Injury

Objective: Dizziness and imbalance are common symptoms following head injury that can continue for months or years in some individuals. Chronic dizziness is a serious health concern that can lead to increased fall risk, emotional and psychological distress, as well as work disability. Multiple causes of post-concussive dizziness can present a challenge to diagnosis, and obstacle to treatment. The Departments of Veterans Affairs and Defense (VA/DoD) developed clinical practice guidelines that outline recommendations for managing individuals with post-concussive dizziness, but it is unclear how these recommendations impact treatment for post-concussion dizziness/imbalance. In addition, some Veterans may seek care outside the VA or use alternative approaches to manage their post-concussive dizziness. The purpose of this study is to determine factors that contribute to successful outcomes of patients with concussion/blast-related dizziness. Methodology: We will use VA and DoD health databases to identify and examine characteristics of Veterans with specific (for example, inner ear balance dysfunction) and non-specific diagnoses of post-concussion dizziness. Then, we will survey Veterans using questionnaires and access the VA electronic medical records to determine factors that impact long-term recovery of post-concussion dizziness. Specifically, we will examine the impact of factors such as type of treatment, the presence or absence of health conditions such as headache and anxiety, the severity of head injury, as well as age and gender. Findings: We have sent survey invitations to 2724 Veterans, and received 674, for a response rate of 25%. Due to COVID, our mailing schedule has been delayed and additional surveys will be mailed in FY21 to achieve our goal of 1050 completed surveys. Chart abstraction has been completed on 486 subjects.

Airborne Pollutants as Triggers of Parkinson's Disease via the Olfactory System

We are interested in determining whether ambient air pollutants impact the development of Parkinson's disease (PD) by increasing -synuclein pathology via inflammation. After completing Specific Aim 2, wherein we found no differences between experimental groups in spread of -syn, nor the expected nPM-induced neuroinflammatory changes. We repeated Specific Aim 2 mouse experiments with a new batch of nPM that showed in vitro activity. We collected tissues during the first week of SARS-CoV-2 quarantine. After restrictions were relaxed, we began analyzing tissues for neuroinflammation. This on-going analysis has yet to show strong inflammatory effects of the in vivo nPM exposure. Due to continued pandemic quarantine and travel restrictions we are discussing option with our collaborators. We will plan and execute novel experiments to explore the effects of LPS-induced olfactory inflammation on spread of -syn by histological (VAI) and biochemical (USC) analyses. We are interested in determining whether ambient air pollutants impact the development of Parkinson's disease (PD) by increasing -synuclein pathology via inflammation. After completing Specific Aim 2, wherein we found no differences between experimental groups in spread of -syn, nor the expected nPM-induced neuroinflammatory changes. We repeated Specific Aim 2 mouse experiments with a new batch of nPM that showed in vitro activity. We collected tissues during the first week of SARS-CoV-2 quarantine. After restrictions were relaxed, we began analyzing tissues for neuroinflammation. This on-going analysis has yet to show strong inflammatory effects of the in vivo nPM exposure. Due to continued pandemic quarantine and travel restrictions we are discussing option with our collaborators. We will plan and execute novel experiments to explore the effects of LPS-induced olfactory inflammation on spread of -syn by histological (VAI) and biochemical (USC) analyses.

Establishing Human and Animal Stool Testing for SARS-CoV-2 to Determine Feasibility of Fecal/Oral Transmission

The Current COVID-19 disease pandemic is caused by the Coronavirus SARS-CoV-2. SARS-related viruses in humans are generally carried by animals and human infections are only due to contact with infected animals. But in the case of SARS-CoV-2, human to human transmission is the most studied route of infection and primarily through fomites or respiratory particulates. The objective of this project was to determine feasibility of methods to test and detect infectious SARS-CoV-2 virus and viral RNA in stool. Once detected, the results were used to support the hypothesis that fecal/oral exposure to SARS-CoV2 virus is a potential route of infection in animal models and humans and an area to address in the control and prevention of COVID-19 disease.

Effect of Mecp2 Mutation on Cortical Projections Revealed by Correlated Single-Cell Transcriptomics and Projectomics

Rett Syndrome is caused by mutations in Mecp2, which result in a constellation of language, cognitive, motor, and autonomic deficits later in life. Although changes in long-range neuronal connectivity likely underlie the behavioral defects in Rett syndrome, it is unclear how long-range axonal projections are disrupted. Here we develop and apply high-throughput single-cell techniques to identify cell type-specific changes in projections in Mecp2 animals. We identified two subtypes of cortical projection neurons with potential changes in long-range projections, including the corticothalamic neurons and L6b neurons. Our results provide candidate cell types for future in depth studies on the long-range circuitry changes associated with Mecp2 mutation. Furthermore, our approach is generally applicable to other brain areas and disease models to reveal cell type-specific changes in projections that are difficult to detect using conventional methods.

Prediction and Evaluation of Strength and Fatigue to Prevent Warfighter Musculoskeletal Injuries

Musculoskeletal disorders are common for people who work on strenuous and demanding tasks, such as nurses, construction workers, and soldiers. The injuries come from overexertion of the individuals related muscle groups that are required to be activated to complete a specific task in various environments. A current solution is the use of robotic-aid devices. These robotic devices are being investigated to treat musculoskeletal injuries and significantly increase a humans capacity in heavy working conditions. However, the design of the human-machine interaction force remains an obstacle to the implementation of effective assistance to people in practical working scenarios due to the environmental variety, task complexity, and human variation and uncertainty. Previous studies showed that the metabolic cost of exercise will be increased instead of reduced if improper forces are provided by the machine. Thus, accurate estimations of the required machine force application to the individual are critical to labor saving and muscle health.

Probiotic (VSL#3) for Gulf War Illness

The overall objective of the study is to determine whether probiotic VisbiomeTM will improve 1) Intestinal symptoms of Irritable Bowel Syndrome and 2) Non-intestinal symptoms (fatigue, joint pain, insomnia, general stiffness and headache) associated with IBS. All these symptoms are part of the Gulf War illness. We screened our first participant in September 2013. Overall, we have screened 101 and enrolled 62 Gulf War Veterans. We have stopped enrolling patients in this study. The stool samples analysis ongoing as well the statistical data analysis.

Neutrophil Elastase Reprograms Macrophage Function in Chronic Obstructive Pulmonary Disease

The central hypothesis of this proposal is that extracellular NE is taken up by macrophages and accumulates in both cytoplasmic organelles and the nucleus. NE activity degrades histone deacetylase 2 (HDAC2) and possibly other HDACS and Sirtuins resulting in increased acetylation of several targets including histone H3, High Mobility Group Box 1 (HMGB1) and nuclear factor kappa B (NFkB) p65, resulting in increased cytokine transcription and release of HMGB1 (AIM 1). Nuclear NE cleaves histone H3 and increases H3 citrulline resulting in chromatin decondensation and release of vital nuclear METs (AIM 2).

Trauma-Informed Guilt Reduction (TrIGR) Intervention

Posttraumatic guilt and shame are common among Veterans and have been implicated in the development and maintenance of posttraumatic distress and a range of adverse outcomes, including posttraumatic stress disorder (PTSD), depression and suicidality, and alcohol/substance use disorders. There is a pressing need for effective treatments targeting transdiagnostic mechanisms such as guilt. We developed Trauma Informed Guilt Reduction (TrIGR) therapy as a therapeutic tool to help Veterans accurately appraise deployment-related guilt and to re-identify and re-engage with their values. The overall objective of this study is to examine the efficacy of TrIGR in reducing deployment-related guilt. The overarching hypothesis is that TrIGR will reduce guilt, shame, and related distress, and these improvements will be significantly greater than in the comparison condition, Supportive Care Therapy (SCT). The study is a Stage 2 randomized, controlled trial of TrIGR compared to SCT. Recruitment of participants takes place at two VA Medical Centers (San Diego, CA and Providence, RI). 150 OEF/OIF Veterans will be randomized to TrIGR or SCT (at least 75 in San Diego). All eligible participants complete an in-person baseline assessment, receive 6 sessions of TrIGR or SCT in individual format, complete brief bi-weekly self-report measures during treatment, and complete follow-up assessments immediately post-treatment, and 3- and 6-months later.

Planning For On Campus K-12 Education During Covid -19

The complex decision-making regarding reopening of schools compels careful thought, planning, and collaboration, withinand across communities. Through this document, the COVID-19 Healthcare Coalition provides clarifications, interpretations,and estimates that can be used as a resource by school leaders to develop and implement plans for returning to on-campuslearning, in the context of the COVID-19 pandemic.

Improving Voluntary Engagement for PTSD Treatment Among Soldiers

Year 2 focused on completing pre-trial activities, including finalizing the clinical and control PTSD interventions, corresponding counselor manuals, research protocols, recruitment advertisements, and recruitment plan. Year 2-4 have primarily focused on randomized controlled trial (RCT) activities (Phase 2), including recruitment, enrollment and data collection that began in January 2018. We obtained necessary approvals for Phase 2 from University of Washington IRB, Madigan IRB, and HRPO.

Investigating Exercise-Induced Neuroplasticity and its Mechanisms in Parkinson's Disease: Targeting Executive Function and Brain Circuitry

Parkinsons disease (PD) is the second most frequent neurodegenerative disorder of old age. A common problem in PD is cognitive impairment. There are no effective treatments. Impairment in executive function (EF) is the most common subtype of cognitive impairment and leads to challenges in daily function, including decision making, multi-tasking, and quality of life. Exercise studies in the aging field and preliminary studies in our PD animal work support the role of high intensity skill practice and high motor fitness in promoting greater EF performance compared to aerobic exercise, and cardiovascular fitness (e.g. V02max). While, a wide range of exercise modalities have shown to improve motor performance in PD patients, investigations of the relationship between exercise and EF in PD and mechanisms of neuroplasticity remain a significant gap in knowledge. This application will address this gap through complementary translational studies in humans and animals. The purpose of this 18-month longitudinal clinical study is to examine the association between EF related cognitive performance and fitness levels, specifically cardiovascular and motor fitness, as well as exercise intensity. We hypothesize that high intensity regular exercise as well as High level of Motor fitness will be associated with greater level of cognitive EF performance over the 18-month period, than High levels of Cardiovascular Fitness or low intensity exercise. We also hypothesize that EF related brain circuity and connectivity will have a greater association with high level motor fitness than cardiovascular fitness and mediate the association between higher level of cognitive EF performance and motor fitness seen at 18 months.

Dysregulation of the PACT-mediated Crosstalk Between Protein Kinases PKR and PERK Contributes to Dystonia 16 (DYT16)

Currently, the available treatment options for dystonia are merely palliative and the drug development has not progressed significantly due to a lack of understanding about the involved molecular pathomechanisms. We investigated if PACT, the gene mutated in dystonia 16 (DYT16), causes a disruption in the normal regulatory crosstalk between PERK and PKR kinases leading to a loss of cell homeostasis after ER stress. Both PERK and PKR kinases phosphorylate eIF2 alpha and activate a downstream signaling pathway that allows recovery and survival after ER stress. The most significant finding during the last funding period was that PACT is a substrate of PERK kinase and PERK phosphorylates PACT in vivo after ER stress. This is a paradigm-shifting finding as it was previously unknown that PACT could participate and regulate both PKR and PERK pathways. The molecular etiology of DYT16 has now been conclusively shown to be a dysregulation of eIF2 alpha signaling. Thus, our research has uncovered a PACT-mediated novel regulatory pathway and laid the foundation for more in depth drug development to target PACT-PERKinteractions in future. In addition, it has added significant new knowledge about how cells respond to ER stress. In the brief period of 8 months we made significant progress included in this report, before the COVID-19 pandemic slowed down our work significantly.

Characterizing Concussion Using Brain Derived Exosomes

Traumatic brain injury (TBI) currently afflicts 357,000 enlisted military men and women in the US Armed Services. For the most common form of TBI, Mild Traumatic Brain Injury (mTBI) most patients recover within a year following the incident, but 10-20 of mild cases result in a long-term disability including seizures and emotional and behavioral issues. Although much has been learned about molecular changes in the brain following injury, access to these biomarkers following mTBI is lacking. The accurate diagnosis and precise individual clinical management of traumatic brain injury (TBI) is limited by the lack of accessible molecular biomarkers that are informative regarding the unique mixture of injury mechanisms in each TBI patient.

Exploiting Inhibitory Siglecs to Combat Food Allergies

During the fourth year of this award, we have continued to generate important data related to targeting of CD22 on B cells and CD33 on mast cells to abrogate food allergies. Unfortunately, the COVID-19 pandemic shut down our labs in March 2020 for several months, causing some delays in our work. With that said, we have still produced new data and are now back in the labs on a regular basis to carry out additional experiments. For the CD22 project, we have now developed a humanized mouse model using NSG mice lacking mouse B and T cells, transfused with human PBMCs. These mice make human IgG against peanut allergens upon exposure to peanut and in pilot experiments, we were successful in stopping this IgG production by use of Ah1 STALs. We have also prepared mouse CD22L Ah1, Ah2, Ah3, and Ah6 for use in our conferred memory model to block IgE production to all major allergens. In terms of targeting CD33 in this past year, we have developed a novel approach by conjugating human CD33L directly to anti-human IgE, without the use of liposomes for scaffolding. This molecule is effective in inducing tolerance in humanized mice. Overall, our results move us closer to translating our STALs platform into human studies by focusing now on the use of humanized mouse models and human CD22 and CD33 ligands in our systems. Finally, we have applied for an Expansion Award for this project.

Examining the Efficacy of the TEACCH School Transition to Employment and Post-Secondary Education Program

The purpose of this project is to conduct a RCT trial examining the efficacy of the TEACCH School Transition to Employment and Postsecondary Education (T-STEP) Program. 60 Community college students (18-21 years of age) with autism spectrum disorder will participate in either the T-STEP Program or manualized counseling services (career, academic, self-counseling) with both proximal (executive function, social communication, and emotion regulation) and distal (employment, postsecondary education success) outcomes measured. The long-term impact of this intervention is to promote a more positive quality of life for young adults with high functioning ASD including increased postsecondary education completion, employment, self-determination, and decreased difficulties with coping and depression. Due to the COVID-19 epidemic preventing in person interactions, we have adapted the protocol to conduct online interventions. This adapted protocol has received approval from all regulatory bodies and the adapted intervention and RCT trial will begin in the second year of funding.

Assessing Rehabilitation Outcomes after Severe Neuromusculoskeletal Injury: Development of Patient Reported Outcomes Assessment Instruments

Currently, there is a paucity of patient reported outcomes (PRO) measures of secondary health effects and complications that result from neuromusculoskeletal injuries, which greatly limits the clinical care and successful rehabilitation, reintegration, and return to duty/work of injured individuals. This study will create valid, standardized, psychometrically robust, and clinically useful PRO measures for traits and symptoms relevant to understanding quality of life and the health and rehabilitation outcomes of Wounded Warriors and civilians with neuromusculoskeletal trauma. Furthermore, this study will develop clinical score reports in an actionable format to improve the clinical workflow and standard of care for individuals with traumatic limb injuries. During year three, we successfully prepared for and launched large-scale field testing. To date, 159 participants have begun baseline and 154 have completed the baseline interviews. Work to plan for data analyses is underway. We have also worked this year amid challenges from the COVID-19 pandemic, which affected our daily operations and caused the pace of recruitment to slow. However, we have established processes for remote recruitment, consenting, and data collection. We have identified several additional sites that have eligible participants and are willing to recruit them, to help increase recruitment throughput.