TLR Antagonism by Sparstolonin B Alters Microbial Signature and Modulates Gastrointestinal and Neuronal Inflammation in Gulf War Illness Preclinical Model

Abstract:

The 1991 Persian Gulf War veterans presented a myriad of symptoms that ranged from chronic pain, fatigue, gastrointestinal disturbances, and cognitive deficits. Currently, no therapeutic regimen exists to treat the plethora of chronic symptoms though newer pharmacological targets such as microbiome have been identified recently. Toll-like receptor 4 (TLR4) antagonism in systemic inflammatory diseases have been tried before with limited success, but strategies with broad-spectrum TLR4 antagonists and their ability to modulate the host-microbiome have been elusive.

Using a mouse model of Gulf War Illness, we show that a nutraceutical, derived from a Chinese herb Sparstolonin B (SsnB) presented a unique microbiome signature with an increased abundance of butyrogenic bacteria. SsnB administration restored a normal tight junction protein profile with an increase in Occludin and a parallel decrease in Claudin 2 and inflammatory mediators high mobility group box 1 (HMGB1), interleukin-1β (IL-1β), and interleukin-6 (IL-6) in the distal intestine. SsnB also decreased neuronal inflammation by decreasing IL-1β and HMGB1, while increasing brain-derived neurotrophic factor (BDNF), with a parallel decrease in astrocyte activation in vitro.

Mechanistically, SsnB inhibited the binding of HMGB1 and myeloid differentiation primary response protein (MyD88) to TLR4 in the intestine, thus attenuating TLR4 downstream signaling. Studies also showed that SsnB was effective in suppressing TLR4-induced nod-like receptor protein 3 (NLRP3) inflammasome activation, a prominent inflammatory disease pathway. SsnB significantly decreased astrocyte activation by decreasing colocalization of glial fibrillary acid protein (GFAP) and S100 calcium-binding protein B (S100B), a crucial event in neuronal inflammation. Inactivation of SsnB by treating the parent molecule by acetate reversed the deactivation of NLRP3 inflammasome and astrocytes in vitro, suggesting that SsnB molecular motifs may be responsible for its anti-inflammatory activity.

Source: Bose D, Mondal A, Saha P, Kimono D, Sarkar S, Seth RK, Janulewicz P, Sullivan K, Horner R, Klimas N, Nagarkatti M, Nagarkatti P, Chatterjee S. TLR Antagonism by Sparstolonin B Alters Microbial Signature and Modulates Gastrointestinal and Neuronal Inflammation in Gulf War Illness Preclinical Model. Brain Sci. 2020 Aug 8;10(8):532. doi: 10.3390/brainsci10080532. PMID: 32784362; PMCID: PMC7463890. https://www.mdpi.com/2076-3425/10/8/532 (Full text)

Neurotoxicant exposures and rates of Chronic Multisymptom Illness and Kansas Gulf War Illness criteria in Gulf War deployed women veterans

Abstract:

Aims: This study analyzed deployment-related exposures and risk of Persian Gulf War Illness (GWI) in women veterans from the Veterans Affairs (VA) Cooperative Studies Program 585 Gulf War Era Cohort and Biorepository (GWECB CSP#585).

Main methods: We examined the associations between GW deployment-related exposures and case definitions for GWI in deployed GW women. Multivariate regression analyses controlling for demographic outcomes were performed.

Key findings: Surveys were obtained from 202 GW deployed women veterans. Self-reported exposure to smoke from oil well fires as well as chemical and biological warfare were the only exposures significantly associated with the Center for Disease Control and Prevention (CDC) GWI criteria. Seventy-nine women were excluded from the rest of the analyses as they met Kansas GW illness exclusion criteria. Eligible women who self-reported deployment-related exposure to smoke from oil wells, pyridostigmine bromide (PB) pills, pesticide cream, pesticide treated uniforms, and insect baits were significantly more likely to meet the Kansas GWI criteria (n = 123) than those unexposed and exposures were related to Kansas symptom subdomain endorsements.

Significance: These results suggest that women GW veterans reporting deployment related exposures of pesticide, oil well fire and PB pills are significantly more likely to meet the Kansas GWI criteria in this national cohort of GW women suggesting its utility in future studies. In addition, based on these results it appears that women exposed to particular toxicants during the war may benefit from more targeted treatment strategies dependent upon the mechanism of exposure of their toxicant induced outcomes.

Source: Krengel M, Sullivan K, Heboyan V, Zundel CG, Wilson CC, Klimas N, Coughlin SS. Neurotoxicant exposures and rates of Chronic Multisymptom Illness and Kansas Gulf War Illness criteria in Gulf War deployed women veterans. Life Sci. 2021 Sep 1;280:119623. doi: 10.1016/j.lfs.2021.119623. Epub 2021 May 15. PMID: 34004246. https://pubmed.ncbi.nlm.nih.gov/34004246/

Dysregulation of cellular energetics in Gulf War Illness

Abstract:

Gulf War Illness (GWI) is estimated to have affected about one third of the Veterans who participated in the first Persian Gulf War. The symptoms of GWI include chronic neurologic impairments, chronic fatigue syndrome, as well as fibromyalgia and immune system disorders, collectively referred to as chronic multi-symptom illness. Thirty years after the war, we still do not have an effective treatment for GWI. It is necessary to understand the molecular basis of the symptoms of GWI in order to develop appropriate therapeutic strategies. Cellular energetics are critical to the maintenance of cellular homeostasis, a process that is highly dependent on intact mitochondrial function and there is significant evidence from both human studies and animal models that mitochondrial impairments may lead to GWI symptoms.

The available clinical and pre-clinical data suggest that agents that improve mitochondrial function have the potential to restore cellular energetics and treat GWI. To date, the experiments conducted in animal models of GWI have mainly focused on neurobehavioral aspects of the illness. Additional studies to address the fundamental biological processes that trigger the dysregulation of cellular energetics in GWI are warranted to better understand the underlying pathology and to develop new treatment methods. This review highlights studies related to mitochondrial dysfunction observed in both GW veterans and in animal models of GWI.

Source: Raju RP, Terry AV. Dysregulation of cellular energetics in Gulf War Illness. Toxicology. 2021 Aug 10:152894. doi: 10.1016/j.tox.2021.152894. Epub ahead of print. PMID: 34389359. https://pubmed.ncbi.nlm.nih.gov/34389359/

The effect of stress on the transcriptomes of circulating immune cells in patients with Gulf War Illness

Abstract:

Aims: In an effort to gain further insight into the underlying mechanisms tied to disease onset and progression of Gulf War Illness (GWI), our team evaluated GWI patient response to stress utilizing RNA-Seq.

Main methods: The protocol included blood collection before exercise challenge (baseline), at maximal exertion, and after exercise challenge (recovery – four hours post-exercise challenge). Peripheral blood mononuclear cell (PBMC) transcriptomics data were analyzed to understand why GWI patients process stressors differently from their healthy counterparts.

Key findings: Our findings validate previously identified dysregulation of immune and inflammatory pathways among GWI patients as well as highlight novel immune and inflammatory markers of disease activity. These results provide a foundation for future research efforts in understanding GWI pathophysiology and creating targeted treatments.

Significance: Gulf War Illness is a complex, chronic, and debilitating multi-system illness impacting 25%-30% of the U.S. troops deployed to the 1990-1991 Gulf War. The condition is characterized by medically unexplained fatigue and affects multiple organ systems. Because the underlying mechanisms are largely unknown, patients receive symptom-based treatment, rather than targeting fundamental biological processes. To the best of our knowledge, this is the first study that applies RNA-Seq to analyze the effect of GWI, and the response to stressors in GWI, on the transcriptomic changes in circulating immune cells.

Source: Van Booven D, Zarnowski O, Perez M, Sarria L, Collado F, Hansotia K, Riegle S, Finger T, Fletcher MA, Klimas NG, Nathanson L. The effect of stress on the transcriptomes of circulating immune cells in patients with Gulf War Illness. Life Sci. 2021 Sep 15;281:119719. doi: 10.1016/j.lfs.2021.119719. Epub 2021 Jun 16. PMID: 34144055. https://pubmed.ncbi.nlm.nih.gov/34144055/

A randomized phase II remote study to assess Bacopa for Gulf War Illness associated cognitive dysfunction: Design and methods of a national study

Abstract:

Aims: Gulf War Illness (GWI) is a chronic, debilitating, multi-symptom condition affecting as many as one-third of the nearly 700,000 U.S. troops deployed to the Middle East during the 1990-1991 Gulf War (GW). The treatment of GWI relies on symptom management. A common challenge in studying the efficacy of interventions for symptom management is participant recruitment related to factors such as the burden of travelling to study sites and the widespread dispersion of Veterans with GWI. The goal of this study is to assess the efficacy of a novel low-risk therapeutic agent, Bacopa monnieri, for cognitive function in Veterans with GWI and to evaluate the utility of a remote patient-centric study design developed to promote recruitment and minimize participant burden.

Main methods: To promote effective participant recruitment, we developed a remote patient-centric study design. Participants will be recruited online through social media and through a web-based research volunteer list of GW Veterans. An online assessment platform will be used, and laboratory blood draws will be performed at clinical laboratory sites that are local to participants. Furthermore, the assigned intervention will be mailed to each participant.

Significance: These study design adaptations will open participation to Veterans nearly nationwide and reduce administrative costs while maintaining methodologic rigor and participant safety in a randomized, placebo-controlled phase II clinical trial.

Source: Cheema AK, Wiener LE, McNeil RB, Abreu MM, Craddock T, Fletcher MA, Helmer DA, Ashford JW, Sullivan K, Klimas NG. A randomized phase II remote study to assess Bacopa for Gulf War Illness associated cognitive dysfunction: Design and methods of a national study. Life Sci. 2021 Oct 1;282:119819. doi: 10.1016/j.lfs.2021.119819. Epub 2021 Jul 10. PMID: 34256038. https://pubmed.ncbi.nlm.nih.gov/34256038/

A common language for Gulf War Illness (GWI) research studies: GWI common data elements

Abstract

Aims: The Gulf War Illness programs (GWI) of the United States Department of Veteran Affairs and the Department of Defense Congressionally Directed Medical Research Program collaborated with experts to develop Common Data Elements (CDEs) to standardize and systematically collect, analyze, and share data across the (GWI) research community.

Main methods: A collective working group of GWI advocates, Veterans, clinicians, and researchers convened to provide consensus on instruments, case report forms, and guidelines for GWI research. A similar initiative, supported by the National Institute of Neurologic Disorders and Stroke (NINDS) was completed for a comparative illness, Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS), and provided the foundation for this undertaking. The GWI working group divided into two sub-groups (symptoms and systems assessment). Both groups reviewed the applicability of instruments and forms recommended by the NINDS ME/CFS CDE to GWI research within specific domains and selected assessments of deployment exposures. The GWI CDE recommendations were finalized in March 2018 after soliciting public comments.

Key findings: GWI CDE recommendations are organized in 12 domains that include instruments, case report forms, and guidelines. Recommendations were categorized as core (essential), supplemental-highly recommended (essential for specified conditions, study types, or designs), supplemental (commonly collected, but not required), and exploratory (reasonable to use, but require further validation). Recommendations will continually be updated as GWI research progresses.

Significance: The GWI CDEs reflect the consensus recommendations of GWI research community stakeholders and will allow studies to standardize data collection, enhance data quality, and facilitate data sharing

Source: Cohen DE, Sullivan KA, McNeil RB, Klimas NG; Gulf War Illness Common Data Elements Working Group; Symptoms Assessment Working Group, McNeil R, Ashford W, Bested A, Bunker J, Cheema A, Cohen D, Cook D, Cournoyer J, Craddock T, Golier J, Hardie A, Helmer D, Lindheimer JB, Lloyd PJ, Kerr K, Krengel M, Nadkarni S, Nugent S, Paris B, Reinhard M, Rumm P, Schneiderman A, Sims KJ, Steele L, Turner M; Systems Assessment Working Group, Sullivan K, Abdullah L, Abreu M, Abu-Donia M, Aenlle K, Arocho J, Balbin E, Baraniuk J, Block K, Block M, DeBeer B, Engdahl B, Filipov N, Fletcher MA, Kalasinsky V, Kokkotou E, Lidie K, Little D, Loging W, Morris M, Nathanson L, Nichols MD, Pasinetti G, Shungu D, Waziry P, VanLeeuwen J, Younger J; GWI CDE Administrative Team, Klimas N. A common language for Gulf War Illness (GWI) research studies: GWI common data elements. Life Sci. 2021 Aug 2:119818. doi: 10.1016/j.lfs.2021.119818. Epub ahead of print. PMID: 34352259. https://pubmed.ncbi.nlm.nih.gov/34352259/

Acetylcholinesterase inhibitor exposures as an initiating factor in the development of Gulf War Illness, a chronic neuroimmune disorder in deployed veterans

Abstract:

Gulf War Illness (GWI) is a chronic multi-symptom disorder, characterized by symptoms such as fatigue, pain, cognitive and memory impairment, respiratory, skin and gastrointestinal problems, that is experienced by approximately one-third of 1991 Gulf War veterans. Over the nearly three decades since the end of the war, investigators have worked to elucidate the initiating factors and underlying causes of GWI. A significant portion of this research has indicated a strong correlation between GWI and exposure to a number of different acetycholinesterase inhibitors (AChEIs) in theater, such as sarin and cyclosarin nerve agents, chlorpyrifos and dichlorvos pesticides, and the anti-nerve agent prophylactic pyridostigmine bromide.

Through studying these exposures and their relationship to the symptoms presented by ill veterans, it has become increasingly apparent that GWI is the likely result of an underlying neuroimmune disorder. While evidence indicates that AChEIs are a key exposure in the development of GWI, particularly organophosphate AChEIs, the mechanism(s) by which these chemicals instigate illness appears to be related to “off-target”, non-cholinergic effects. In this review, we will discuss the role of AChEI exposure in the development and persistence of GWI; in particular, how these chemicals, combined with other exposures, have led to a chronic neuroimmune disorder.

This article is part of the special issue entitled ‘Acetylcholinesterase Inhibitors: From Bench to Bedside to Battlefield’.

Source: Michalovicz LT, Kelly KA, Sullivan K, O’Callaghan JP. Acetylcholinesterase inhibitor exposures as an initiating factor in the development of Gulf War Illness, a chronic neuroimmune disorder in deployed veterans. Neuropharmacology. 2020;171:108073. doi:10.1016/j.neuropharm.2020.108073 https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7398580/ (Full article)

Subcortical brain segment volumes in Gulf War Illness and Myalgic Encephalomyelitis/Chronic Fatigue Syndrome

Abstract:

Aims: There is controversy about brain volumes in Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (CFS) and Gulf War Illness (GWI). Subcortical regions were assessed because of significant differences in blood oxygenation level dependent signals in the midbrain between these diseases.

Materials and method: Magnetization-prepared rapid acquisition with gradient echo (MPRAGE) images from 3 Tesla structural magnetic resonance imaging scans from sedentary control (n = 34), CFS (n = 38) and GWI (n = 90) subjects were segmented in FreeSurfer. Segmented subcortical volumes were regressed against intracranial volume and age, then iteratively analyzed by multivariate general linear modeling with disease status, gender and demographics as independent co-variates.

Key findings: The optimal model for all subjects used disease status and gender as fixed factors with independent variables eliminated after iteration. Volumes of anterior and midanterior corpus callosum were significantly larger in GWI than CFS. Gender was a significant variable for many segment volumes, and so female and male subjects were analyzed separately. CFS females had smaller left putamen, right caudate and left cerebellum white matter than control women. CFS males had larger left hippocampus than GWI males. Orthostatic status and posttraumatic distress syndrome were not significant covariates.

Significance: CFS and GWI were appropriate “illness controls” for each other. The different patterns of adjusted segment volumes suggested that sexual dimorphisms contributed to pathological changes. Previous volumetric studies may need to be reevaluated to account for gender differences. The findings are framed by comparison to the spectrum of magnetic resonance imaging outcomes in the literature.

Source: Addiego FM, Zajur K, Knack S, Jamieson J, Rayhan RU, Baraniuk JN. Subcortical brain segment volumes in Gulf War Illness and Myalgic Encephalomyelitis/Chronic Fatigue Syndrome. Life Sci. 2021 Jun 29:119749. doi: 10.1016/j.lfs.2021.119749. Epub ahead of print. PMID: 34214570. https://pubmed.ncbi.nlm.nih.gov/34214570/

Sex-Based Differences in Plasma Autoantibodies to Central Nervous System Proteins in Gulf War Veterans versus Healthy and Symptomatic Controls

Abstract:

Veterans from the 1991 Gulf War (GW) have suffered from Gulf War illness (GWI) for nearly 30 years. This illness encompasses multiple body systems, including the central nervous system (CNS). Diagnosis and treatment of GWI is difficult because there has not been an objective diagnostic biomarker. Recently, we reported on a newly developed blood biomarker that discriminates GWI from GW healthy controls, and symptomatic controls with irritable bowel syndrome (IBS) and myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS). The present study was designed to compare levels of these biomarkers between men and women with GWI, as well as sex-specific effects in comparison to healthy GW veterans and symptomatic controls (IBS, ME/CFS).

The results showed that men and women with GWI differ in 2 of 10 plasma autoantibodies, with men showing significantly elevated levels. Men and women with GWI showed significantly different levels of autoantibodies in 8 of 10 biomarkers to neuronal and glial proteins in plasma relative to controls. In summary, the present study addressed the utility of the use of plasma autoantibodies for CNS proteins to distinguish among both men and women veterans with GWI and other healthy and symptomatic control groups.

Source: Abou-Donia MB, Krengel MH, Lapadula ES, Zundel CG, LeClair J, Massaro J, Quinn E, Conboy LA, Kokkotou E, Nguyen DD, Abreu M, Klimas NG, Sullivan K. Sex-Based Differences in Plasma Autoantibodies to Central Nervous System Proteins in Gulf War Veterans versus Healthy and Symptomatic Controls. Brain Sci. 2021 Jan 23;11(2):148. doi: 10.3390/brainsci11020148. PMID: 33498629. https://pubmed.ncbi.nlm.nih.gov/33498629/

Exercise modifies glutamate and other metabolic biomarkers in cerebrospinal fluid from Gulf War Illness and Myalgic encephalomyelitis / Chronic Fatigue Syndrome

Abstract:

Myalgic encephalomyelitis / Chronic Fatigue Syndrome (ME/CFS) and Gulf War Illness (GWI) share many symptoms of fatigue, pain, and cognitive dysfunction that are not relieved by rest. Patterns of serum metabolites in ME/CFS and GWI are different from control groups and suggest potential dysfunction of energy and lipid metabolism. The metabolomics of cerebrospinal fluid was contrasted between ME/CFS, GWI and sedentary controls in 2 sets of subjects who had lumbar punctures after either (a) rest or (b) submaximal exercise stress tests. Postexercise GWI and control subjects were subdivided according to acquired transient postexertional postural tachycardia. Banked cerebrospinal fluid specimens were assayed using Biocrates AbsoluteIDQ® p180 kits for quantitative targeted metabolomics studies of amino acids, amines, acylcarnitines, sphingolipids, lysophospholipids, alkyl and ether phosphocholines.

Glutamate was significantly higher in the subgroup of postexercise GWI subjects who did not develop postural tachycardia after exercise compared to nonexercise and other postexercise groups. The only difference between nonexercise groups was higher lysoPC a C28:0 in GWI than ME/CFS suggesting this biochemical or phospholipase activities may have potential as a biomarker to distinguish between the 2 diseases. Exercise effects were suggested by elevation of short chain acylcarnitine C5-OH (C3-DC-M) in postexercise controls compared to nonexercise ME/CFS. Limitations include small subgroup sample sizes and absence of postexercise ME/CFS specimens. Mechanisms of glutamate neuroexcitotoxicity may contribute to neuropathology and “neuroinflammation” in the GWI subset who did not develop postural tachycardia after exercise. Dysfunctional lipid metabolism may distinguish the predominantly female ME/CFS group from predominantly male GWI subjects.

Source: Baraniuk JN, Kern G, Narayan V, Cheema A. Exercise modifies glutamate and other metabolic biomarkers in cerebrospinal fluid from Gulf War Illness and Myalgic encephalomyelitis / Chronic Fatigue Syndrome. PLoS One. 2021 Jan 13;16(1):e0244116. doi: 10.1371/journal.pone.0244116. PMID: 33440400. https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0244116 (Full text)