Elena Napoli

Serum N-acetyl-beta-glucosaminidase Activity in Diabetic Patients

In a group of seventy-three diabetics, a statistically significant (P < 0.001) increase (+ 41 per cent) of serum N-acetyl-beta-glucosaminidase activity was found. The enzyme level seemed to correlate with both the vascular complications (either microangiopathic or macroangiopathic in nature) and the blood sugar level measured simultaneously. In fact, in patients with moderately elevated glycemia (145 mg./100 ml. ± 41) and without vasculopathies, the enzymatic activity was slightly changed (P > 0.05), while the activity was significantly (P < 0.001) increased (+ 31 per cent), in diabetics with essentially similar gly-cemia (151 mg./100 ml. ± 30) but with vasculopathies, and even more elevated (+ 80 per cent, P < 0.001) in diabetics without vasculopathies but with marked hyper-glycemia (309 mg./100 ml. ± 160). Since lysosomal enzymes, to which N-acetyl-beta-glucosaminidase belongs, are capable of degrading various compounds, these enzymatic changes were regarded as due to an activation of lysosomal enzymes in tissues, occurring in diabetes in response to the metabolic need of degrading either mucopolysaccharides and glycoproteins (as in diabetics with vasculopathies), or various constituents of cells themselves in a context of increased tissue catabolism (as in decompensated diabetics). However, the possibility cannot be excluded that the enzyme is elevated merely because of decreased lysosome stability. Reduction in the destruction rate of the enzyme might also contribute to its elevation.

Enzymes of Glucose Metabolism in Liver of Subjects with Adult-onset Diabetes

An enzyme study was made on needle biopsy specimens of liver from thirty-two subjects with adult-onset diabetes and normal body weight and thirty-two controls. The enzyme pattern in the patients with diabetes was different from that seen with alloxan diabetes. The activities of the two glucose phosphorylating enzymes tested were changed in opposite directions, hexokinase being enhanced and glucokinase moderately decreased. Total glucose phosphotransferase activity remained unchanged. Phosphofructokinase had a reduced activity, which suggested depressed glycolysis, especially if considered together with the enhanced activity of the opposing enzyme, fructose-1, 6-diphosphatase. Normal activity was found for most other glycolytic enzymes, as well as for key gluconeogenic enzymes, including glutamic oxalacetic and glutamic pyruvic transaminases, phosphoenolpyruvate carboxykinase and glucose-6-phosphatase. The finding suggests normal glucose release. Glucose-6-phosphate- and 6-phosphogluconate dehydrogenase activity was elevated. This would indicate an increased metabolism of glucose through the oxidative pathway and, therefore, increased formation of NADPH. This metabolic condition, which is known to favor fatty acid synthesis, might contribute to fatty liver changes. On the other hand, NADP-isocitrate dehydrogenase, which does not provide NADPH for fatty acid synthesis, was little changed.

Enzymes of Glucose Metabolism and of the Citrate Cleavage Pathway in Adipose Tissue of Normal and Diabetic Subjects

Enzyme activities operative in glucose degradation and citrate cleavage pathway were studied in the adipose tissue of twenty-four patients with adult-onset diabetes and normal body weight, aged 59±9 years, and twenty-four matched controls. In normal tissue, type II (heat-inactivated) hexokinase moderately predominated over type I (heat-resistant). 6-Phosphofructokinase had an extremely low activity, which was by far the lowest among the ten glycolytic enzyme activities investigated, and which therefore might greatly limit the glycolytic rate. The level of glucose-6-phosphate dehydrogenase and phosphogluconate dehydrogenase (decarboxylating) was elevated above that occurring in other tissues. This, especially if considered together with the low 6-phosphofructokinase activity, would suggest a major role of pentose cycle in glucose degradation. Of the citrate cleavage pathway enzymes, ATP citrate-lyase, although having a lower activity than malate dehydrogenase and malate dehydrogenase (decarboxylating) (NADP), was readily measurable, which contrasts with previous data by others. This finding is consistent with the occurrence of lipogenetic capacity in human adipose tissue. In diabetic tissue, there was a decreased activity, both on a protein and on a wet-weight basis, of enzymes concerned with the glucose entry into metabolic pathways, namely hexokinase (both type I and, especially, type II) and pentose cycle dehydrogenases, as well as of pyruvate kinase. This could be connected with the defective glucose utilization by adipose tissue in diabetes. Beside the above-mentioned dehydrogenases, malate dehydrogenase (decarboxylating) (NADP) was also diminished. The reduction of these NADPH-forming enzymes, which supply reducing equivalents for fatty acid synthesis, would suggest a depressed lipogenesis.

Enzymes related to lipogenesis in the adipose tissue of obese subjects

In a group of ten adult obese subjects, maintained for 15 days on a normal caloric intake and balanced diet, the activity of hexokinase (EC 2.7.1.1),6-phosphofructokinase (EC 2.7.1.11), and ATP citratelyase (EC 4.1.3.8) in the adipose tissue was significantly increased, both on a protein and on a fat cell number basis, compared to matched normal subjects. The activity of glucose-6-phosphate dehydrogenase (EC 1.1.1.49), malate dehydrogenase (EC 1.1.1.37), and malate dehydrogenase (decarboxylating) (NADP) (EC 1.1.1.40), on the other hand, was unchanged. Since both hexokinase and 6-phosphofructokinase are rate-limiting in glycolysis, their enhanced activity would indicate the occurrence of an increased capacity to metabolize glucose and therefore to generate alpha-glycerophosphate. The elevation of ATP citrate-lyase would suggest increased lipogenesis, owing to the regulatory role that this enzyme plays in fatty acid synthesis. The normal activity of glucose-6-phosphate dehydrogenase and malate dehydrogenase (decarboxylating) (NADP), which supply NADPH for the reduction of acetyl-CoA to fatty acids, would suggest that the change in lipogenesis is of moderate degree, thereb) affecting only the most rate-limiting enzyme, ATP citrate-lyase. These data, on the whole, are consistent with the occurrence of enhanced triglyceride formation. Whether the enzyme changes observed are adaptive or genetic in nature remains to be clarified.

Increased serum activity of beta-n-acetyl-glucoseaminidase in atherosclerosis

In 33 male patients with clinical manifestations of atherosclerosis (angina pectoris, myocardial infarction, or cerebral vascular accidents having occurred no less than six months before study) the serum activity of the lysosomal enzyme beta-N-acetyl-glucosaminidase (EC 3.2.1.30) was found increased by 30 per cent (p<0.001) compared to 76 control male subjects. The activity of enzymes located mainly or exclusively in the cytosol—aspartate aminotransferase (EC 2.6.1.1) and lactate dehydrogenase (EC 1.1.1.27)—was normal. This shows that there was no cell damage that could cause enzyme elevation. Based on the role of lysosomes in the catabolism of macromolecules of ground substance of connective tissue, it is speculated that the enhanced serum beta-N-acetyl-glucosaminidase activity in atherosclerosis might be a manifestation of extracellular secretion of lysosomal enzymes (exocytosis) by the cells of the arterial walls, occurring in response to the accumulation of mucopolysaccharides that takes place in the early stage of disease.

Serum acid phosphatase activity in diabetes mellitus.

In a group of 90 diabetics, an increased serum activity (+137 per cent, p<0.001) of the lysosomal enzyme acid phosphatase was found. The increase was moderate (+55 per cent, p<0.01) in uncomplicated diabetics with slightly elevated glycemia (148 ± 24 mg glucose/100 ml), while it was more accentuated (about twice normal, p<0.001) in diabetics with either vasculopathies (micro- or macroangiopathy) or marked hyperglycemia (343 ± 108 mg glucose/100 ml). Serum aspartate and alanine aminotransferases were normal. Thus, acid phosphatase in diabetes behaves similarly to other lysosomal enzymes: beta-glucuronidase and N-acetyl-beta-glucosaminidase. This is a further evidence of lysosomal enzyme activation in diabetes, apparently linked to both the degree of metabolic decompensation and the presence of vasculopathies. Since acid phosphatase, unlike beta-glucuronidase and N-acetyl-beta-glucosaminidase, is not concerned with degradation of mucopolysaccharides and glycoproteins, the higher activity found in vasculopathic diabetics would indicate that in these patients the activation process is not restricted to the enzymes capable of degrading the aforementioned compounds, which accumulate in the walls of diseased vessels, but involves a variety of lysosomal hydrolases.

Enzymes of citrate-cleavage pathway in liver of subjects with adult-onset diabetes.

In the liver of adult diabetics, the activity of two enzymes of the citrate-cleavage pathway was increased, the change being statistically significant for NADP-malate dehydrogenase (+ 46%, p less than 0.05) but not significant for ATP citrate-lyase (+ 55%, p greater than 0.10). The increased activity of NADP-malate dehydrogenase, together with the previously described elevation of pentose cycle dehydrogenases, suggests enhanced NADPH generation. Considering the recently proposed possibility of extramitochondrial acetyl-CoA formation by routes other than the citrate-cleavage (i.e., via cytoplasmic acetyl-CoA synthetase), our data is consistent with the occurrence of increased lipogenetic capacity.