Evidence has shown that Gulf War Veterans (GWV) with Gulf War illness (GWI) have impaired production of energy, which is generated by the energy-producing parts of cells called "mitochondria." Mitochondria are vulnerable to damage with a range of drug/chemical/toxin exposures, including exposures that have been shown to relate to development of GWI. Additionally, animals exposed to Gulf-relevant exposures have shown altered expression in genes related to mitochondria. However, the exact nature of the mitochondrial (mt) problems in GWI has yet to be characterized. Understanding the details may open important avenues to diagnosis and treatment.

We propose to assess in what way(s) mitochondria are affected. To do so, we will recruit 27 GWV who meet criteria for GWI and 27 controls, matched to the affected Veterans 1-to-1 on age, sex, and ethnicity. Participants will undergo a muscle biopsy under local anesthesia by an experienced investigator. Biopsy samples will be used to examine the structure and function of mitochondria for signs of abnormality and injury. Do they look normal? Are mitochondria normal in number -- or too sparse or numerous? Do the mitochondria break apart and latch together ("fission" and "fusion") with the normal balance? Do they have lots of "cristae" or infoldings to provide high surface area for function, as is normal, or are the cristae sparser or distributed abnormally?

Is the electrical "potential difference" across the mt membrane normal -- or high or low? Is the mt membrane abnormally rigid or fluid? Does the membrane serve as an effective barrier, or fail to defend against entry of substances that should not pass? Is there excessive mt production of free radicals? This can affect mt energy production or trigger cell death. How vulnerable are the mitochondria to swelling caused by calcium?

Production of energy is a pivotal function of mitochondria needed for cell function and survival. How effectively do the different parts of the mt "respiratory chain" -- involved in producing energy -- function? Are there defects in this process? If so, do these predominantly occur at a specific stage? How much "uncoupling" is there -- more uncoupling means more diversion of mt processes away from ATP (cell energy) generation. What is the reserve capacity for energy production?

These aspects of mt status are assessed directly in muscle tissue. Muscle and brain are the most affected domains in mt disorders and in GWI, and muscle is preferred as it is generally not possible to sample brain tissue in people. Statistical tests will compare results in GWV with GWI to results in healthy matched controls.

Additional tests will be done that do not require biopsy, using blood, urine, or breath, focusing on tests that are abnormal in some settings of mt dysfunction, like the respiratory exchange ratio (carbon dioxide produced per oxygen consumed). Clinical tests relevant to mitochondria will assess physical function, fatigue, and GWI symptoms. Mt measures affected in GWI will be correlated with these assessments to see how aspects of mt status predict symptoms and function.

Importance: Mt mechanisms have strong prospects to be fundamental to GWI -- they can account for known findings related to exposure relations, symptom profiles, variable latency to onset, objective abnormalities, and correlated conditions in a fashion that no other explanation put forward to date can equal. Investigation of the nature of the problems affecting energy production by mitochondria in GWI may elucidate core mechanisms of GWI. This may foster approaches to objective diagnosis. It may provide objective markers for GWI that may be tracked with treatments. Inclusion of less invasive lab and bioenergetic assessments provides added information, and also enables determination of whether any sufficiently reflect core defects to provide a possible substitute noninvasive measure that might be tracked with treatment. The mt abnormalities identified can be investigated in animal studies, to see which candidate Gulf-relevant exposures appear most strongly to contribute. This may foster implementation of protections, limiting future exposure for current Veterans with GWI (to protect from further injury) and limiting exposure for current and future military personnel to reduce risk of GWI-like problems. Identifying specific mt mechanisms of GWI will define specific targets for treatment. Treatments that target fundamental mechanisms may reduce symptoms and improve function in affected Veterans and might protect from development of more serious GW-related sequelae in which mt dysfunction is known to play a role, such as neurodegenerative conditions. Importantly, treatments that are designed to target fundamental mechanisms of GWI (mechanisms that this study seeks to elucidate) have the best possible prospects of leading to "cure" for GWI -- rendering identification of these fundamental mechanisms a particular priority.