Serum Biopterin as a Marker of Immune Activation in the Spontaneously Diabetic BB Rat

Abstract

Summary Tetrahydrobiopterin (BH4), is a pteridine product which is released by rodent macrophages and fibroblasts upon activation by cytokines. It has previously been reported as a potentially useful marker for monitoring immune activation during the prediabetic prodrome in the spontaneously diabetic BB rat. We have used serial serum BH4 measurements, first to confirm the hypothesis that serum BH4 is elevated as a result of immunostimulation, and second to monitor more closely the serum BH4 levels during the prediabetic prodrome in a cohort of diabetes prone BB rats. A modified biopterin radioimmunoassay protocol was used to establish a reference range of 25.5-80.1 nmolll (95% confidence limits) for 70 Wistar rats of mean age six months. When eight adult Wistar rats were treated with sic BCG there was a significant increase in serum BH4 level (p<O.Ol) which returned to pre-treatment levels by day 16, coincident with the disappearance of visible signs of inflammation. A cohort of 26 diabetes prone BB rats, their diabetes resistant controls (n=20) and a cohort of Wistar controls (n=20) were then used to monitor serum BH4 levels from 30-120 days of age, at frequent intervals. The diabetes incidence in the diabetes prone animals was 100% with a mean age of onset of 80 days. The serum tetrahydrobiopterin level was significantly higher at day 30 in all three groups (p< 0.05) than at any other time point. The serum BH4 level in the diabetes prone group was significantly higher at day 30 (p<O.Ol ) and day 60 (p<0.05) than in either control group. These results show that serum BH4 is a sensitive marker for immune activation following subcubaneous injection of BCG in the rat, and is detectable using RIA. Serum BH4 was maximum in this study at day 30, but the significantly higher levels at days 30 and 60 in the diabetes prone compared with the diabetes resistant animals suggest that the immune activation characteristic of prediabetes can be detected biochemically in blood samples.