Discriminação das isoenzimas da adenosina desaminase (ADA) em fluidos corporais humanos

Abstract

Adenosine deaminase (ADA – E.C.3.5.4.4.) is a fundamental enzyme in the catabolism of the purines. It catalyzes the deamination of adenosine or 2’deoxy-adenosine producing ammonium and inosine or 2’-deoxyinosine, respectively. Its activity is expressed by two isoenzymes presented in three isoforms. ADA1 (36 kDa) and ADA1 bound to CD26 (280kDa) are widely distributed in the body tissues. Their action is particularly important because high levels of 2’deoxy-adenosine are toxic for the immune system cells. ADA2 (100kDa) is normally found in serum and is synthesized only in monocyte-macrophage system. The biological importance of ADA2 is not yet fully clear, especially for its kinetics characteristics. The objective of the present work was to discriminate the isoenzymes of human adenosine deaminase using agarose electrophoresis and by mathematical model proposed by Vale and Almeida (1998). In addition, we performed a study of the profile of ADA tests in State of Ceara (Brazil). Samples of of ascites, pleural and pericardial effusion were submitted to electrophoresis in 1% agarose at 80V for 7 hours. The gel was sliced and each slice was incubated in adenosine (22 or 0,55mM) for 20 hours to detect the ammonium released by enzymatic reaction. The results found from electrophoresis were compatible with the model proposed by Vale and Almeida (1998). The pleural fluid is the most frequently requested for the determination of ADA, followed by ascitic fluid, cerebrospinal fluid, pericardial fluid and serum. We observed that the value of enzymatic activity is influenced by corporal fluid type where the enzyme is localized. These data can be associated with the corporal barrier, like brain barrier. We concluded that the proposed mathematical model could be used in clinical laboratories to discriminate ADA isoenzymes to improve the diagnostic method.