Reduced Liver Glucose-6-Phosphatase in Human Leukemia. ∗

Abstract

During experiments concerned with rat liver glucose-6-phosphatase it became of interest to study human liver glucose-6-phos-phatase. Specimens of human liver obtained during post-mortem examinations were homogenized and assayed immediately. It was noted that each of the livers from children with acute lymphatic leukemia was very deficient in this enzyme activity. Some of the mechanisms responsible for this reduction in activity were investigated and form the basis of this report.

Methods. At time of autopsy the patient's age, diagnosis, drug therapy, and hours since death were recorded. A 10 g specimen of liver was placed immediately in 0.12 M KC1 at 0°C, and was kept at 0°C during subsequent steps. A 10% homogenate was prepared using a glass-teflon homogenizer at 1000 rpm for 2 to 3 minutes. The homogenate was centrifuged at 600 × g for 10 minutes and the residue was discarded. The supernatant was centrifuged in a Servall refrigerated centrifuge at 37000 × g for 30 minutes and the residue was suspended in 0.12 M KC1 at a concentration 5 to 10 mg protein per ml. It had been previously demonstrated that rat liver glucose-6-phosphatase activity was purified 3-fold in this residue, when compared to liver homogenate. The activity of the 37000 Xg residue was determined by the method of Swanson(l), at pH 6.5 in malate buffer. Protein was determined by Lowry's(2) method.

The effects of protein synthesis inhibitors on enzyme activity were determined in two ways. First, 6-mercaptopurine, methotrexate (4-amino-N-methylpteroyl glutamic acid), and cortisol were added during the assay of enzyme activity. Second, 6-mercaptopurine (1.0 mg/150 g rat/day), and methotrexate (0.1 mg/150 g rat/day) were fed in 2 groups of rats with their regular Purina Laboratory Chow diet, and other rats Received 100 μg actinomycin D by intraperitoneal injection daily for 4 days.