Metabolism Clinical and Ewperimental VOL 48, NO 7 JULY 1999 Alterations in the Activity of Several Glycohydrolases in Red Blood Cell Membrane From Type 2 Diabetes Mellitus Patients Giancarlo Goi, Chiara Bairati, Giovanni Segalini, Alberto B. Burlina, Luca Massaccesi, Augusto Lovagnini, and Adriana Lombardo The erythrocyte membrane in 71 patients with type 2 diabetes mellitus was assessed for glycohydrolase activity: N-acetyl-~-D-glucosaminidase, 13-D-glucuronidase, ~- and I~-D-galactosidase, ~- and ~-D-glucosidase, ~-o-mannosidase, and ~-L-fucosidase. Only I~-D-glucuronidase, oL-D-giucosidase, and ~-D-glucosidase showed markedly elevated levels with respect to the controls regardless of the presence of complications. Among the examined patients, those with good metabolic control (not yet submitted to any therapy) showed the same enzyme levels as the reference subjects, while the levels in patients with unsatisfactory metabolic control (treated with oral hypoglycemic and/or insulin) significantly differed from the control levels. For ~-D-glucosidase and I~-D-glucosidase, a correlation with glycemia and the parameters of metabolic control was also evidenced. Alterations of I~-o-glucuronidase, ~-o-glucosidase, and ~-D-glucosidase were also ascertained in the plasma of the same diabetic patients according to the literature; each enzyme correlated with the other, either in plasma or in the erythrocyte membrane. This study shows a correlation between plasma and erythrocyte membrane levels for these three enzymes. The strict parallelism of the glycohydrolases in the two different compartments provides a profile of these enzymes in the pathology of diabetes. Copyright © 1999 by W.B. Saunders Company I NFORMATION on the various degradative enzymes in the erythrocyte plasma membrane, including the glycohydrol- ases 1-6 that were once thought to be exclusively located in lysosomes, 7 has become more readily available. It would seem that erythrocyte plasma membrane glycohydrolases are similar in several properties to glycohydrolases of lysosomal origin. It has been suggested that mature erythrocytes, despite lacking lysosomal particles, need these hydrolases to catabolize por- tions of their own membrane. 3 Attention has been focused on erythrocyte plasma membrane glycohydrolases due to the possibility that these enzymes, given the appropriate conditions of pH and temperature, might become functional not only in degrading plasma glycoproteins but also in altering blood-group substances located at the membrane surface. As a consequence, the conformation of the membrane itself would change, probably with repercussions on the lifespan of the red blood cell) Based on the evidence that in diabetes mellitus, the erythro- cyte plasma membrane undergoes alterations in its physico- chemical, dynamic, conformational, and functional proper- ties, su we performed a systematic study aimed at ascertaining the possible alterations of some of the glycohydrolases present in the membrane: N-acetyl-I3-D-glucosaminidase ([[3-GlcNAc- ase] EC 3.2.1.30), [3-D-glucuronidase ([t3-GlcA-ase] EC 3.2.1.31), [3-D-galactosidase (EC 3.2.1.23), c~-D-galactosidase (EC 3.2.1.22), e~-t)-glucosidase ([oL-Glc-ase] EC 3.2.1.20), a-D-mannosidase (EC 3.2.1.24), ~-L-fucosidase (EC 3.2.1.51), and [3-D-glucosidase ([[3-Glc-ase] EC 3.2.1.21). Since the plasma levels of these enzymes are known to change in both type 1 and type 2 diabetes mellitus, 12-16 we decided to assay these enzymes in the erythrocyte membrane and the plasma of the same patients in an effort to reveal a correlation in the enzymatic activity of the two compartments. SUBJECTS AND METHODS Chemicals and Other Products The commercial chemicals used in the study were the purest available. The water routinely used was freshly redistilled in a glass From the Department of Medical Chemistry and Biochemistry, University of Milan, Milan; Diabetic Unit and Laboratory of Clinical Chemistry and Haematology of Bassini Hospital, Cinisello Balsamo, Milan; and Department of Pediatrics, University of Padua, Padua, ltaly. Submitted June 1, 1998; accepted March 14, 1999. Supported by a grant (to A.L.) from the Italian Ministry of Education (40% contribution for 1995-1996). Address reprint requests to Adriana Lombardo, PhD, Dip. Chimica e Biochimica Medica, Via Saldini, 50-20133 Milano (Italia). Copyright © 1999 by W.B. Saunders Company 0026-0495/99/4808-002351 O. 00/0 Metabolism, Vol 48, No 7 (July), 1999:pp 817-821 817