Project Title

Genomic Approach to Find Female-Specific Mechanisms of GWI Pathobiology

Principal Investigator/Project Director

Lubov Nathanson

Colleges / Centers

Dr. Kiran C. Patel College of Osteopathic Medicine

Funder

DOD - U.S. Army Medical Research Acquisition Activity

Start Date

9-15-2017

Abstract

Background: GWI is a debilitating condition that reflects disruption of normal cellular signaling and function, manifesting itself with a complex variety of symptoms affecting multiple organ systems. The exact causes of these dysfunctions are not well understood. Recent research efforts revealed that the disease is likely attributable to a combination of genetic predisposition and environmental influences, potentially due to exposure to toxic chemicals and/or pathogens. Our laboratory has developed a quantitative and reproducible method for the evaluation of physiological changes and activation of relapse symptoms typical of GWI using a measurable stressful trigger such as exercise challenge. Previous results utilizing a microarray-based approach demonstrated that immune defense, regulation of RNA biosynthesis, transcription and some other nucleic pathways are significantly changed in GWI patients. Although men with GWI have been extensively studied, we know much less about women veterans with GWI due to the very limited numbers of female veterans studied so far. The few studies assessing gender differences have revealed that women appear to present with a greater degree of autonomic dysfunction, e.g., heart rate variability, T-cell regulated hypertension, differences in the patterns of genomic DNA methylation. Preliminary data of our group show that female and male GWI patients differ in the cytokine secretion and in the gene expression response to the exercise challenge. Objective: The objective of this study is to combine the latest state-of-the-art genomics and computational biology methods to identify novel, female-specific mechanisms of transcriptional regulation in GWI, which will facilitate better understanding of GWI pathobiology in women and provide further insights in mechanisms of GWI in men. Our studies will provide insight into sex-specific disease onset and progression. This will lead to a more comprehensive understanding of aberrant underlying pathways of disease activation and progression. Our studies will reveal potential therapeutic targets for treatment of GWI and provide insight into sex-specific disease onset and progression, with the goal of curing GWI. Specific Aims: Specific Aim I: Identify metabolic pathways and possible regulatory RNAs affected in women with GWI. Specific Aim Ia. Identification of differentially expressed known transcripts using RNA-seq approach. Specific Aim Ib. Analysis of the functions, affected by differentially expressed known transcripts in women with GWI. Specific Aim Ic: Analysis of the functions, affected by differential splicing of transcripts in women with GWI. Spesific Aim Id: Identification of novel transcripts that are expressed differentially in women with GWI. Specific Aim Ie: Validation of the results by NanoString technology. Specific Aim II. Investigate possible mechanisms of transcriptional regulation in women with GWI. Specific Aim IIa and IIb: Determine gene expression differences between female GWI patients and female HC that can be attributed to CNV. Specific Aim IIc and IId: Identify the genomic DNA methylation patterns and correlation of these patterns to discrepancies in gene expression between female GWI patients and female HC. Specific Aim IIe: Evaluate differences in miRNA expression between women with GWI and female HC using pre-built human miRNA Nanostring panel. Specific Aim IIf: Combine the results of RNA-seq, CNV, genomic DNA methylation assays and miRNAs expression and do a correlation analysis to determine mechanisms of transcriptional regulation that may cause GWI in women. Specific Aim III. Compare affected metabolic pathways and mechanisms of transcriptional regulation in women and men with GWI. Specific Aim IIIa: Calculate differences in gene expression, CNV and DNA methylation changes in men and women with GWI. Specific Aim IIIb: Validate results of Specific Aim IIIa. Study Design: This project will use PBMCs that have already been isolated from patients recruited for the recently VA funded research project. We will select samples from 40 female GWI patients and 40 matched female HCs at three designated time points: (i) T0, before exercise, (ii) T1, at the peak of exercise effort (VO2 max) and (iii) T2, four hours after the exercise challenge. To evaluate changes in gene expression and mechanisms of transcriptional regulation, we will perform RNA-seq, CNV, genomic DNA methylation and miRNA expression assays for 20 GWI patients and 20 HC (“experimental” cohort). We will use for validation of the results our “experimental” cohort and, in addition, samples from the remaining 20 GWI patients and 20 HC (“validation” cohort). We will compare results of this project with the results of the DOD funded ongoing study of the genomic mechanisms of transcriptional regulation in men with GWI. Impact: Conventional GWI treatments have failed to effectively target the underlying dysfunctions associated with GWI, aside from managing symptomatology. The identification of the female-specific genomic mechanisms of GWI will elucidate underlying causes of pathobiology involved in the triggering and progression of GWI symptoms. Furthermore, our approach and collaboration with the DoD funded GWI consortium (W81XWH-13-2-0085) will provide identification of novel gender-specific promising targets to help improve diagnosis, prognosis and specifically designed FDA-approved drug reassignments for treatment of GWI, fully addressing the scope of funding method.

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