Investigation on anti-glutamic acid decarboxylase antibodies in type I diabetes mellitus

ABSTRACT

Antibodies directed against the enzyme glutamic acid decarboxylase [GAD] are believed to be the main cause of destruction of pancreatic islet cells in type I [insulin dependent] diabetes mellitus. The enzyme was found both in the brain and pancreatic beta cells. Although similarities in identity of GAD in human and rat brain have been demonstrated, little is known about the interaction between the enzyme and antiserum in type 1 diabetic patients. In the present study GAD was partially purified from rat brain homogenate. The four-step procedure involves, sequentially, an ultracentrifugation, DEAE-cellulose, hydroxyapatite resin, and Sephadex G-200 gel filtration chromatography. The enzyme activity was assayed either manometrically or fluorimetrically. The results showed a positive correlation between the rates of CO[2] production with the changes of fluorescence intensities of the enzyme after addition of glutamate. The collected fraction from the gel filtration chromatography showed approximately 140-fold purification of the enzyme with a 15% yield. The specific activity of the enzyme of brain supernatant and the partially purified enzyme collected from every chromatographic step was measured upon addition of the serum samples from type I diabetes [n= 11]. A marked decrease in the rate of CO[2] production or the change of fluorescence intensities of the enzyme was observed, indicating an interaction between the enzyme and the patients' sera. However, serum samples from healthy control individuals had little effect on the enzyme activity of the partially purified GAD. The results suggested that rat brain GAD might be used as an in vitro reagent for screening of type I diabetes, using an enzyme inhibition assay