We have performed lumbar punctures (spinal taps) on two separate groups of subjects with Gulf War Illness (GWI) and chronic fatigue syndrome (CFS), and healthy control subjects. These proteins from the fluid around the brain were compared between these three groups. Ten proteins were found only in the GWI and CFS subjects, but not in the healthy subjects (i.e., GWI/CFS proteome). This is highly remarkable and suggests that these proteins play important roles in GWI. One critical protein was carnosine dipeptidase 1 (CNDP1). It was found in significantly more GWI (4 / 9 subjects) than healthy controls (1 / 12) (p=0.0001). CNDP1 is the enzyme that chops two very important brain antioxidants consisting of two peptides each in half. They are homocarnosine (gamma-aminobutyric acid-histidine, or GABA-his) and carnosine (beta-alanine-histidine). The higher rate of detection of CNDP1 in GWI may be (a) beneficial by cleaving homocarnosine in the brain and releasing GABA that inhibits neuron toxicity and nerve death, or (b) detrimental if CNDP1 degrades these vital antioxidants faster than the dipeptides can be made.

In order to address these issues, it is important to know how CNDP1, carnosine, and homocarnosine function in the body. First, CNDP1 comes as three different genetic forms in the human population. One form (LB5B) leads to a low level of CNDP1 in the blood. Other forms (LB6B and LB7B) have higher levels. It has been shown that the LB6B and LB7B forms of the gene (alleles) are associated with kidney failure in diabetics. Since CNDP1 was increased in the fluid around the brain in GWI and CFS, we wondered if these subjects had the LB6B and LB7B form of the CNDP1 gene. We will test this by doing cheek swabs in 100 GWI and 100 healthy Gulf War veterans and sequencing their DNA. If true, then this genetic test may be a biomarker or indicator of GWI. We will also examine the patterns of different types of proteins in the blood from GWI and healthy veterans. We will determine if the new GWI blood samples contain the 10 proteins that we found in the GWI/CFS cerebrospinal fluid proteome, and if they are absent from the healthy veterans' samples. If so, these proteins may also be indicators of GWI. Additional statistical analysis of the two groups of proteins may identify other biomarkers from GWI that are not found in healthy volunteers. This is biomarker identification. The 100 GWI and 100 healthy veterans will fill out a series of questionnaires and have their pain sensitivity and muscle strength tested. The results of these tests are likely to be very different between GWI and healthy veterans. We may also find important differences if the CNDP1 allele and proteomics data are added. If high levels of the CNDP1 enzyme are breaking down the homocarnosine and carnosine in the brain, skeletal muscle, and other tissues, then replacing some of the carnosine may be beneficial. This will be tested by recruiting the 100 GWI subjects and randomly dividing them into two groups. One group will receive carnosine capsules, while the other group will receive a placebo. The subjects will take their capsules for 6 months to see if there is a significant change in their condition. We anticipate that the GWI subjects, or a segment of the GWI subjects, will experience a big improvement in the level of discomfort and activity if they are taking carnosine. However, we think that the placebo group may have a smaller change or no change. If this is the case, the carnosine may be useful for the treatment of GWI. This would be very interesting, since the conclusion that carnosine may work in some GWI subjects was based on differences in the brain enzyme (CNDP1) that we described earlier. In addition to this testing after 6 months, we will also examine the time course at 0, 1, 3, and 6 months for the carnosine-treated group to determine the time course for improvement of each of the questionnaires and other study tests. We anticipate that problems like sleep, pain, activity, and vitality will improve more than other sensations, like fatigue or blood pressure changes after the hand grip experiment. If this is right, then it may be possible to determine which GWI subjects would respond the most to carnosine, and which ones would not have any benefit. Carnosine may turn out to be a useful new treatment for GWI. Finally, the biology of CNDP1, carnosine, homocarnosine, and nerve damage is very complicated. We will grow nerve cells (neuroblastoma, a tumor of sympathetic nerves) in culture, and use hydrogen peroxide to try to kill the cells. We will then add carnosine and homocarnosine to see if they protect against the peroxide damage. We will also test a brand-new theory in this cell culture system: Blue-green algae in algae blooms in fresh water like the salt marshes of southern Iraq or saltwater like the Persian Gulf can release a neurotoxin. We propose that this chemical may have been inhaled or ingested by military personnel. This toxin can be activated in the body and may lead to atrophic lateral sclerosis (ALS) or a Parkinson's-like disease (PD). Because of the structure of this toxin, it is possible that carnosine or homocarnosine may block the toxin's actions and prevent the development of GWI or ALS-PD syndrome. Carnosine could work in ALS-PD.