- Biomarkers of Gulf War Veterans' Illnesses: Tissue Factor, Chronic Coagulopathy, and Inflammation (An Update)
- Treatment of Memory Impairment and Sensorimotor Deficits in an Animal Model for the Gulf War Veterans' Illnesses
- Biomarkers of Gulf War Veterans' Illnesses: Tissue Factor, Chronic Coagulopathy, and Inflammation
- Identification of Biological Pathways Implicated in Hippocampal Dysfunction and Cognitive Impairment in Gulf War Illness
- Analysis of Paraoxonase Status among U.S. Navy Gulf War Veterans with Increased Postwar Symptoms, Psychological Morbidity and Medical Conditions
- Structural MRI and Cognitive Correlates in Pest-Control Personnel from Gulf War I
- Novel Pharmacological Approaches for Treatment of Neurotoxicity Induced by Exposure to Depleted Uranium
Gulf War Illness (GWI) is an unexplained chronic multi-symptom disorder, resulting from service in the 1990-1991 Gulf War, that afflicts an estimated 200,000 Gulf-deployed U.S. Veterans. Frequently observed symptoms related to GWI include pain, fatigue, cognitive difficulties, and gastrointestinal problems. Dr. Ronald Bach at the Veterans Affairs Medical Center in Minneapolis (VAMC Minneapolis) is using a GWIRP Investigator-Initiated Research Award to further develop findings from VA-funded studies that indicated that ill Gulf War Veterans may be in a hypercoaguable state of unknown etiology.* The earlier work showed strong correlations between the plasma concentrations of inflammation-related proteins and symptoms of GWI. Thus, the hypothesis emerged that chronic inflammation is part of GWI pathophysiology.
In the current study, Dr. Bach studied 85 Minnesota-based Gulf War Veterans, beginning with an in-depth health history of participants followed by a blood sample. During the structured interview each Veteran was evaluated for symptoms of chronic pain, chronic fatigue, and cognitive impairment using the established survey instrument. Platelet-free plasma fractions were analyzed for a panel of 89 inflammation-related biomarkers along with the plasma concentration of thrombin-antithrombin complex and D-dimer. In the course of this analysis it was initially determined that C-reactive protein (CRP) levels, a marker of systemic inflammation, were significantly higher in Gulf War Veterans with three symptoms (as defined in the health surveys) versus asymptomatic Veterans. Dr. Bach subsequently observed statistically significant linear correlations between CRP and a group of 18 plasma proteins. This set of pro-inflammatory potential GWI biomarkers has been labeled “The Gulf War Proteome,” though more in-depth analysis is pending. Dr. Bach states, “To the best of our knowledge this is the first study that brings [these proteins] together in a single proteome. The identities of these biomarkers have much to say about the ill health that many Gulf War Veterans are now experiencing as well as their risks for additional chronic illnesses in the future.” The Gulf War Proteome is an inter-related collection of pro-inflammatory biomarkers whose direct functional relationships are well established. It is comprised of acute phase proteins, cytokines, adipokines, metabolic factors, growth factors, and proteins involved in tissue matrix remodeling.
These findings have focused the search for therapeutic agents to treat GWI. With chronic inflammation emerging as a possible underlying cause of some GWI symptoms, it has become a potential therapeutic target. Thus, anti-inflammatory drugs are being examined for the treatment for GWI. Dr. Bach plans to propose a randomized double-blind placebo controlled clinical trial of an anti-inflammatory drug in veterans with GWI. The goal will be to determine if the intervention improves the health and well-being of veterans with GWI and corrects any of the observed biomarker abnormalities.
* Hannan KL, Berg DE, Baumzweiger W, Harrison HH, Berg LH, Ramirez R, and Nichols D. 2000. Activation of the coagulation system in Gulf War Illness: A potential pathophysiologic link with chronic fatigue syndrome, a laboratory approach to diagnosis. Blood Coagulation and Fibrinolysis 11(7):673-678.
Coenzyme Q10 (Q10) is a vitamin-like antioxidant, naturally produced in human cells and important for cellular energy production. Unfortunately, natural Q10 levels can be inadequate to meet the needs of those with increased "oxidative stress", a build-up of free oxygen radicals, or impaired energy production. This condition could describe thousands of veterans who served in the 1990-91 Persian Gulf War, and are now suffering with Gulf War Illness (GWI).
Dr. Beatrice Golomb and a research team at the University of California at San Diego recently completed a 3-1/2 year study involving 46 veterans with GWI that examined the benefits of daily Q10 administration. Dr. Golomb hypothesized that mitochondrial dysfunction, which is linked to cellular energy production, might contribute to GWI symptoms, and sought to examine whether Q10 supplements conferred benefit to overall health and symptoms in GWI. Two dose levels (100mg/day and 300mg/day) were compared to each other and against placebo treatment, and subjects were treated for 3-month periods in a "cross-over" design (i.e. successive treatment then placebo periods). Self-rated responses for overall health and symptom-based questionnaires were used as study outcome measures.
The presence of carry-over effects confounded analysis of the intended cross-over design, however, analysis of the first 3-month treatment period, which by itself represented a randomized, double-blind, placebo-controlled parallel design study, the 'gold standard' for randomized trials, remained valid. Results demonstrated that Q10 at 100mg led to significant benefit to Gulf War symptoms as well as improved physical function when compared to placebo. Study outcomes were assessed by examining effects from the 20 health symptoms most commonly reported by study participants. These symptoms spanned categories including fatigue, mood, cognition, muscle function, pain, skin, and autonomic symptoms, and with each included symptom present in at least half of the veterans. Not only was benefit to the summed score significant with Q10 at 100mg, but the direction of difference for Q10 at 100mg vs. placebo was favorable for every one of the 20 symptoms - an event that would be highly unlikely by chance (less than one in a million).
These findings provide important preliminary information on effect size and variance relative to self-rated health which could inform a larger trial of Q10 at 100mg better powered to show benefit to global self-rated health - and affirm benefits to Gulf War symptoms.
Roughly one-fourth of the almost 700,000 veterans of the 1990-91 Persian Gulf War report health issues including widespread pain, chronic fatigue, and memory and cognitive deficits, now characterized as Gulf War Illness (GWI). Research has shown that these health issues may have been caused by exposures to pesticides and insect repellents, as well as anti-nerve gas medication, experienced during Gulf War deployment. Dr. Mohamed Abou Donia at Duke University developed a rat model of Gulf War (GW) exposures that uses topical treatment of permethrin (insecticide) and DEET (insect repellent) to simulate chronic GWI symptoms. Dr. Abou Donia is using a Fiscal Year 2008 Gulf War Illness Research Program Investigator-Initiated Research Award to systematically study the potential benefits of the analgesic Flupirtine to treat GWI using this rat model. Flupirtine has been used to treat pain and memory deficits associated with other disorders and has been shown to be safe and non-addictive, though it is not approved by the U.S. Food and Drug Administration (FDA).
In the study, rats in the GW model, either Flupirtine-treated or untreated, are compared with Flupirtine-treated and untreated normal controls for performance in a water maze test. Rats are timed as they try to find a hidden platform in a water tank from different starting points. Reduced swimming times indicate spatial learning and memory. The water maze test also gauges sensorimotor parameters such as reflexes, motor strength, and coordination. In addition, researchers learn about cognition from observing the rats' probing activity during these tests. The study is currently underway, and data will be collected over the next 3 years. Dr. Abou Donia's GW exposures model has effectively reproduced GWI symptoms, as the rats developed alterations in memory and neuropathology. The effects of Flupirtine will be analyzed after all water maze and histopathological tests have been completed. If successful, this research would provide proof of concept that could ultimately lead to FDA approval of Flupirtine for the treatment of GWI.
Many veterans of the 1990-1991 Persian Gulf War suffer from unexplained chronic and often debilitating health issues. It is estimated that 25% to 32% of the nearly 700,000 Gulf War veterans currently experience widespread pain, headaches, fatigue, cognitive deficits, and other symptoms. Preliminary evidence has indicated that a high percentage of ill Gulf War veterans may be in a hypercoagulable state, i.e., an abnormally increased tendency toward blood clotting, the basis of which is unknown. Dr. Ronald Bach of the VA Medical Center in Minneapolis is using funds from an FY08 GWIRP Investigator-Initiated Research Award to further investigate this concept. Dr. Bach will examine the level of tissue factor (TF), the biological initiator of blood coagulation, in blood samples from ill Gulf War veterans and healthy controls. Overexpression of TF in the bloodstream can lead to disseminated intravascular coagulation (DIC) and impaired blood flow in the smallest of blood vessels. This restriction of microcirculation applied chronically can produce symptoms commonly associated with Gulf War Illness (GWI), including fatigue, somatic pain, and cognitive difficulties.1
Dr. Bach will initially try to find more evidence of clotting system abnormalities in ill veterans' blood by measuring an expanded panel of coagulation markers, including D-Dimer, which indicates fibrin clot formation, thrombin-antithrombin III complex (TAT), to indicate ongoing coagulation activity, prothrombin fragment 1.2 (F1.2), a measure of ongoing thrombin generation, and TF-procoagulant activity (TF PCA), which can initiate DIC. Serial samples taken over 24 months will be assayed to establish the chronic nature of the abnormalities. This analysis may also uncover new biomarkers for GWI. Dr. Bach will also assess immune function by measuring levels of inflammatory markers in the blood samples. This may indicate a possible state of chronic inflammation in ill veterans and may support a connection between chronic coagulation and immune function.
This study seeks to contribute to the understanding of GWI etiology. The results generated could identify a novel biomarker for the diagnosis of GWI, lead to clinical trials of therapies, and improve the health of veterans with GWI.
1 Hannan KL, Berg DE, Baumzweiger W, Harrison HH, Berg LH, Ramirez R, and Nichols D. 2000. Activation of the coagulation system in Gulf War Illness: A potential pathophysiologic link with chronic fatigue syndrome, a laboratory approach to diagnosis. Blood Coagulation and Fibrinolysis 11(7):673-678.
Identification of Biological Pathways Implicated in Hippocampal Dysfunction and Cognitive Impairment in Gulf War Illness
Posted February 24, 2011
Fiona Crawford, Ph.D., Roskamp Institute, Sarasota, Florida
Gulf War Illness (GWI), a multisymptom condition associated with service in the 1990-1991 Persian Gulf War, is thought to affect approximately 200,000 to 250,000 veterans. Among the complex and chronic symptoms experienced by these veterans, persistent cognitive impairment has been widely reported and largely attributed to combined exposure to pyridostigmine bromide (PB) and overuse of pesticides and insect repellants (referred to as Gulf War agents). Evidence of structural and metabolic abnormalities of the hippocampus provided from imaging studies, and memory impairment from neuropsychological testing both suggest that hippocampal damage may be the main culprit. Hippocampal damage has also been cited as an important clinical feature associated with neurocognitive disorders such as Alzheimer's disease. With funds from a Fiscal Year 2008 GWI Research Program Investigator-Initiated Research Award, Dr. Crawford is developing a mouse model of cognitive dysfunction following exposure to Gulf War agents; she will characterize the accompanying hippocampal-related cognitive and neuropathological changes and apply state-of-the-art proteomic technology to identify molecular pathways of interest. Pilot studies have generated a mouse model of delayed and persistent cognitive dysfunction and increased astrogliosis presenting at 4 months after combined exposure to PB and permethrin. Proteomic analyses of these mice reveal distinct brain and plasma protein profiles unique to the Gulf War agent-exposed mice compared to controls.
Replication of these findings in additional cohorts of mice is underway. These studies include analysis of APOE-E4-expressing transgenic mice, to determine whether the APOE genotype influences outcome after GW agent exposure; the E4 allele of APOE is a known risk factor for Alzheimer's disease and is also associated with worse outcome after brain injury. Neurobehavioral, neuropathological, and proteomic analyses will all be applied to identify brain molecular profiles corresponding with exposure and response to GW agents. Collectively, these experiments will significantly increase our understanding of the molecular basis of Gulf War agent-induced cognitive dysfunction.
Although the precise cause or causes of Gulf War Illness (GWI) remain unknown, evidence indicates that veterans of the 1990-91Gulf War were exposed to a variety of toxins, such as chemical nerve agents and pesticides of the organophosphate (OP) family. These drugs have been implicated in GWI’s initiation and progression. One OP of interest is sarin, a highly toxic nerve agent whose low-dose, sub-symptomatic exposure has been connected with GWI etiology.
Dr. Mariana Morris and colleagues used a Fiscal Year 2006 GWI Research Program Investigator-Initiated Research Award to develop a mouse model of GWI which examines low-dose exposure to sarin. In the model, a non-symptomatic dose of sarin produced long-lasting changes in central neural function and decrements in cardiac function seen as reductions in heart rate variance.
The global objective of the study was to elucidate the pathophysiological mechanisms of GWI in an animal model, study the autonomic and cardiac biomarkers, and evaluate the effectiveness of some commercially available drugs targeting adrenergic (such as beta blockers in cardiac dysfunction) and cholinergic nervous systems. Dr. Morris also examined complementary cardiac changes focused on the structural and functional effects on the mouse heart using echocardiography, electrocardiography, and histology.
Some of Dr. Morris’ findings on the effects of low-dose sarin exposure in the model include:
- Sarin produced delayed and long-term effects on cardiac function such as reduced ejection fraction.
- Sarin caused left ventricular dilation two months after an asymptomatic dose. This is a marker for dilated cardiomyopathy.
- Levels of atrial and brain natriuretic peptides in the heart were increased, indicating cardiac remodeling possibly due to volume overload.
- Low-dose sarin produced regionally specific changes in brain dopamine.
- A stress test with dobutamine, a selective B adrenergic agonist, showed reduced cardiac function, reduced contractile function, and ischemia, as determined by echocardiographic and electrocardiographic analyses.
These findings have important implications for military or civilian populations since low-dose, non-symptomatic exposure to sarin may result in long-lasting cardiac dysfunction.
After completing cardiovascular studies of pharmacological agents, Dr. Morris is continuing studies to further select appropriate agents capable of ameliorating autonomic symptoms and performing behavioral, anatomical, and endocrine tests. Effective agents will move forward to clinical studies leading to potential treatments for GWI. Since the agents used are already approved drugs for other diseases such as Alzheimer’s or hypertension, the timeframe necessary for clinical implementation could be shortened.
Analysis of Paraoxonase Status among U.S. Navy Gulf War Veterans with Increased Postwar Symptoms, Psychological Morbidity and Medical Conditions Posted December 21, 2009
Lt Col Christopher J. Phillips, MD, MPH, Naval Health Research Center, San Diego, CA
Gulf War Illness (GWI) is a complex of chronic multisymptom health problems that affects 175,000 to 210,000 of 697,000 (at least one fourth) of the U.S. veterans who served in the 1990-1991 Gulf War. The multiple concurrent symptoms typically include a combination of memory and concentration problems, chronic headaches, widespread pain, unexplained fatigue, chronic digestive difficulties, respiratory symptoms, and skin abnormalities not explained by established medical diagnoses or standard laboratory tests. No effective treatments have been identified for Gulf War Illness and few veterans have reportedly recovered. There is increasing evidence suggesting that the higher rate of GWI in Gulf War Veterans (GWV) can be associated with neurotoxic exposures of GWV to organophosphate(OP) and carbamate acetyl cholinesterase inhibitors (AChEis) such as pyridostigmine bromide (PB), pesticides, and nerve agents. PB pills were given to protect deployed troops from the potential effects of nerve agents.
US Navy Seabees, both non-deployed and those deployed in the Gulf War participated in a 1994 study by answering extensive questionnaires about war time exposures, postwar symptoms, and also by providing sera. The study reported a higher rate for 35 out of 41 symptoms among GW deployed Seabees versus non-deployed Seabees. However, neither the 1994 study nor a large (n=11,868) Seabee survey study in 1999 could establish a clear link between environmental exposures and post war symptoms.
Prior studies have reported that differences in PON1 enzyme activity may be associated with Gulf War illnesses, or perhaps more generally with service in the Gulf War. Serum PON1 has been reported to be polymorphic as an Arg192 isoform (PON1R192) and/or a Gln192 isoform (PON1Q192) in human populations. The hydrolysis activity (detoxification activity) of PON1 isoforms varies among different organophosphorous compounds as paraoxon (POX) and diazoxon (DZO).
Lt Col Christopher J. Phillips, MD, MPH, from Naval Health Research Center, received a Department of Defense Fiscal Year 2006 Gulf War Illness Research Program (GWIRP) Exploration - Hypothesis Development Award to investigate PON1 status, defined by the phenotype (enzyme activity) and genotype (PON1-192 Q/Q; Q/R; R/R) and associations with the frequency of self-reported symptoms of the 1994 US Navy Seabees study cohort classified as GWI cases.
Of the 294 US Navy deployed Seabees who participated in the 1994 study, 51 (17%) met the GWI case definition. PON1 status was determined using a validated spectrophotometric assay to study hydrolysis activity of serum or plasma PON1 for substrates such as diazoxon and paraoxon. PON1-192 isoforms of the deployed Seabees who met the case definition of GWI were determined to be QQ (41%), QR (27%), and RR (31%). Adjusted model results indicated that the RR PON1 genotype correlated with the highest odds (OR=4.0) for meeting the GWI case definition. Also, taking PB pills, exposure to burning jet fuel in tent heaters, and exposure to petroleum spraying increased the odds of developing GWI among the deployed Seabees, while self-reported exposure to pesticides did not increase the odds of having GWI.
Pesticides were widely used during the 1991 Gulf War (GW) to protect troops from infectious disease-carrying insects. During their deployment, GW veterans were exposed to pesticides where they worked, slept, and ate. One class of these pesticides, organophosphates, is known to produce chronic neurological symptoms at sufficient exposure levels by inhibiting the enzyme acetylcholinesterase (AChE). Exposure to AchE-inhibiting organophosphate pesticides and anti-nerve gas pills (pyridostigmine bromide [PB]) has long been a suspected cause for the persistent health complaints of GW veterans. Drs. Kimberly Sullivan and Maxine Krengel, of Boston University, used a Fiscal Year 2006 Gulf War Illness Research Program Investigator-Initiated Research Award to follow up on their earlier work (also supported by the CDMRP) with pesticide applicator personnel who also took PB tablets. Drs. Sullivan and Krengel will analyze structural brain MRIs with a highly exposed subgroup of this cohort and record volumetric measurements of key brain areas including total gray matter, white matter, and ventricle volumes. This follow-up neuroimaging study will evaluate the combination of exposures to AChE inhibitors as factors in the expression of GWI veterans' continued health symptoms. In addition, this study could provide a biomarker of brain pathology in these veterans if structural brain differences are identified and can be correlated with functional differences seen on cognitive test measures. Knowledge of these relationships will be useful in identifying objective indicators of pathology that distinguish ill from healthy Gulf War veterans and may help identify potential avenues for treatment.
Depleted Uranium (DU) is a byproduct of natural uranium whose radioactivity is roughly half that of the natural product. According to the World Health Organization, DU is used as counterweights in aircraft, radiation shields in medical devices, and as containers to transport radioactive materials. The military uses DU in its armor plating and in armor piercing rounds to take advantage of its high density. Unfortunately, a number of soldiers who served in the Gulf War have presented with neurologic illnesses for which the unknown long-term implications of DU exposure cannot be ruled out. In an effort to relieve the suggested neurotoxicity caused by exposure to DU, Dr. Stephen Lasley of the University of Illinois at Chicago proposes the long-term administration of a free radical trapping agent and/or an NMDA receptor antagonist to reduce DU neurotoxicity in Gulf War Veterans.
Dr. Lasley received a fiscal year 2006 Investigator-Initiated Research Award through the Gulf War Illness Research Program to study novel pharmacological approaches for the treatment of neurotoxicity induced by chronic exposure to DU. Using an animal model of DU exposure, Dr. Lasley's research aims to determine the ability of drug therapies to reduce the effects DU has on glutamate neurotransmitter function and to assess biochemical markers related to oxidative stress in the hippocampal tissue of rats.
Currently, Dr. Lasley's research is under way as his team continues to monitor the animals and collect raw data for analysis.
Vietti KRN and Lasley SM. 2007. Stimulus-evoked glutamate release is diminished by acute exposure to uranium in vitro. Neurotoxicol Teratol 29:607-612.
Understanding the cause of Gulf War Illness (GWI) has proven elusive since the condition became recognized shortly after the first Gulf War. Research has been devoted to addressing the complex of symptoms observed in GWI-afflicted service members. It is also essential to discover the origins of this disease and methods to detect it, for the protection of the warfighter. Dr. James Baraniuk, of Georgetown University, is researching a potential source for the onset of GWI, examining proteomic disparities between veterans with GWI and healthy control groups in hopes of discovering potential biomarkers and learning more about the mechanism(s) of GWI.
Dr. Baraniuk received a fiscal year 2006 Investigator-Initiated Research Award through the Gulf War Illness Research Program to study carnosine dipeptidase 1 (CNDP1) polymorphisms and carnosine therapy in GWI. The research aims to identify biological markers of disease in GWI-afflicted veterans compared to other veterans from the 1990-91 era. Dr. Baraniuk will compare the treatment effects of carnosine to placebo in GWI subjects, and effects on exercise tolerance, activity, and cognition will be examined. The potential for CNDP1 polymorphisms to alter the outcomes of treatment and the risk of developing GWI will also be investigated. This type of study is required to determine if those with different forms of the CNDP1 enzyme respond to treatment in the same way.
Dr. Baraniuk recently gained Investigational New Drug status from the FDA for the use of carnosine and is recruiting patients for current and future clinical trials. Those interested in more information can contact Dr. Baraniuk at email@example.com.