Nitrogen Metabolism and
Testing Nitrogen Hypothesis in ME/CFS

To test this hypothesis, we will culture white blood cells in growth media with added nitrogen-labelled amino acids and measure the nitrogen-labelled by-products of energy metabolism. Cells from 30 ME/CFS patients and 20 age and sex-matched controls will be collected and processed at the Stanford Genome Technology Center and cultured for our nitrogen tracking experiment. These experiments will be conducted on blood cells directly from ME/CFS patients and cells cultured from ME/CFS patients.

To capture whether there is an ammonia accumulation in vivo, our best available option is observing the urine concentration of nitrogen-containing metabolites over a short time-course in which varying levels of physical/mental activity have occurred. The 30 ME/CFS patients and 20 sex-match control subjects from the above experiment will be required to provide all urine samples over 4 consecutive days (~20 samples per individual) while their exertion and heartbeat data is collected using wearable technology. Normally, most nitrogen by-products are converted to primary urea along with amino acids, peptides, creatine, creatinine, and polyamines.

In a pathological state, highly reactive, toxic nitrogen by-products are excreted in the urine as nitrate, nitrite, and ammonia. We will measure the ratios of healthy to pathological nitrogen metabolites in the urine and correlate this with the metadata activity.