Inhibition of the orthophosphatase and pyrophosphatase activities of human alkaline-phosphatase preparations

1. Purified human liver and small-intestinal alkaline orthophosphatases release inorganic phosphate at appreciable rates from a variety of organic pyrophosphate substrates. 2. The pyrophosphatase action is inhibited by Mg²⁺ ions at concentrations that activate the hydrolysis of orthophosphate substrates by these enzymes. 3. The results of mixed-substrate experiments, denaturation studies with heat or urea and starch-gel electrophoresis suggest that both orthophosphatase and pyrophosphatase activities are, in each preparation, properties of a single enzyme. 4. Intestinal phosphatase shows greater pyrophosphatase activity relative to orthophosphatase than the liver enzyme.     

Suramin suppresses growth,alkaline-phosphatase and telomerase activity of human osteosarcoma cells in vitro

Neoadjuvant chemotherapy in osteosarcoma improves the survival dramatically, but there is currents drug resistance in about 25% of patients, leading researchers to investigate alternative therapy forms. Suramin has in the last two decades been used as salvage therapy in some cancers. This study was undertaken to investigate suramin as a possible salvage therapy in osteosarcoma. The effect of suramin on three human osteosarcoma cell lines (MG-63, HOS and SaOS-2) and three primary osteosarcoma cell lines isolated from biopsies was investigated. Suramin significantly inhibited cell proliferation, determined by 3H-thymidine incorporation, of osteosarcoma cells at a dose ranging from 250 to 500 microg/ml. Suramin decreased the secretion of alkaline-phosphatase after stimulation by 1,25-dihydroxy-Vitamin D(3) up to 50% and decreased telomerase activity by up to 40%. The data demonstrate that suramin has marked in vitro effects on human osteosarcoma cells supporting further clinical investigation.     

RNase activity in human interferon preparations.

The level of RNase activity in human interferon preparations was examined. Although sequential purification of interferon resulted in nearly a 300-fold increase in specific activity, RNase-specific activity remained more or less constant. The implications of this finding for the analyses of the mode of action of interferon are discussed.     

Properties of alkaline-phosphatase fractions in extracts of human small intestine

1. The chromatographic and electrophoretic heterogeneity of human-intestinal alkaline-phosphatase activity is described. 2. The phosphatase activity has been divided into two main components, of which certain properties have been compared. The components resemble each other in their Michaelis constants for the hydrolysis of phenyl phosphate and in their behaviour towards certain inhibitors, but differ in their stability to heat. 3. The addition of Mg²⁺ or Ca²⁺ ions to the electrophoresis buffers sharpens the electrophoretic zones. 4. The results presented do not support the existence of more than one alkaline phosphatase in small intestine.     

Ribonuclease activity in preparations of human leukocyte interferon]

Preparations of human leukocyte interferon obtained by multi-stage purification procedure exhibited ribonuclease activity with the optimum at pH 7.0--7.5. The enzyme possessed the endonuclease action mechanism. Most substances studied for their effect on the RNA-ase activity in human interferon preparations showed many of them to act on the enzyme in the same way as on other ribonucleases. However, dithioerythritie, a reducing agent for disulfide bounds, activated the ribonuclease in the interferon preparation, as distinct from the pancreatic ribonuclease, which was inhibited by this preparation. Patterns of protein and RNA-ase distribution were obtained by electrophoresis in polyacrylamide gel.     

Association of triiodothyronine binding activity to soluble adenylate cyclase in testicular preparations

Summary Cytosolic adenylate cyclase activity from rat seminiferous tubules was purified by chromatography in DEAE-cellulose, hydroxylapatite and Bio-Gel A-0.5 m as well as by centrifugation in sucrose gradients. In all these purification steps, fractions with adenylate cyclase activity also contained binding activity for L-T₃. Binding studies indicate the existence of two L-T₃ receptor components associated to adenylate cyclase activity. The component exhibiting the highest hormone affinity has the lowest binding capacity.     

Association of centrioles and chromosomes observed in preparations of whole-mounted human chromosomes

The replication of extrachromosomal rDNA molecules from the macronucleus of Tetrahymena was studied by electron microscopy. Replication begins in the center of the palindromic molecule and proceeds by means of bidirectional fork movement toward the free ends of the molecule.     

Association of tyrosine protein kinase activity with mitochondria in human fibroblasts

A tyrosine protein kinase activity has been detected in the mitochondrial fraction purified from human fibroblasts. By enzymatic and sedimentation analysis this activity appeared to be localized in the mitochondrial outer membrane. Mitochondrial tyrosine phosphorylation was strictly dependent on the presence of Mn²⁺ ions. An inverse relationship between cell proliferation and mitochondrial protein phosphorylation on tyrosine residues has been found: a marked increase in the mitochondrial tyrosine kinase activity occurred when a significant reduction in the growth rate followed serum step-down. In mitochondria purified from resting cells, a protein band with apparent molecular weight of 50 kd appeared to be phosphorylated on tyrosine.     

Controlled gamma-irradiation mediated pathogen inactivation of human urokinase preparations with significant recovery of enzymatic activity

Human sources of urokinase have led to the contamination of in-process lots of commercially available material with human pathogens. Effective pathogen inactivation of urokinase preparations can be achieved through the use of gamma-irradiation. Additionally, the presence of a free radical scavenger (ascorbate) and the control of temperature have resulted in maintenance of the enzymatic activity of urokinase without a significant effect on the pathogen inactivation properties of gamma-irradiation. In this study we have optimized the conditions during gamma-irradiation to achieve inactivation of porcine parvovirus by 5 logs and vaccinia virus to levels below the limits of detection, while maintaining 92% of urokinase activity. Product specific optimization of gamma-irradiation has the potential to provide effective pathogen inactivation while maintaining substantial functional activity for many therapeutic proteins.     

Electron-microscopic cytochemistry of alkaline-phosphatase activity in endothelium, pericytes and oligodendrocytes in the rat brain

The fine structural localization of alkaline-phosphatase (ALP) activity was investigated in the endothelial cells and pericytes of blood vessels and in the oligodendrocytes of rat cerebral cortex and corpus callosum by means of electron-microscopic (EM) cytochemistry using the lead-citrate method. ALP activity was associated with both the luminal and abluminal plasma membranes of some endothelial cells, but in other endothelial cells, this activity was found inside the cytoplasm. In some pericytes, ALP activity was associated with the plasma membrane but in others, strong activity was exhibited within both the cytoplasm and nucleus. Light, medium and dark oligodendrocytes showed ALP activity on their plasma membranes; on the other hand, immature oligodendrocytes exhibited activity within the cytoplasm and on the part of their plasma membrane. Within the cytoplasm of these reacted immature cells, the rough endoplasmic reticulum, nuclear membrane and outer membrane of the mitochondria were the main sites of ALP reaction. Endothelial cells, pericytes and oligodendrocytes demonstrated ALP activity along their plasma membrane or within their cytoplasm, and pericytes showed it within their nuclei. In particular, oligodendrocytes retained ALP activity throughout their cell life, and the intracellular distribution of this activity altered as they matured.     

Potential mutagenic activity of some vitamin preparations in the human gut.

Rutin is a nonmutagenic flavonol glycoside, whereas its aglycone quercetin is mutagenic. Cell-free preparations from fecal cultures (fecal preparations) contain a beta-glucosidase that, when incubated with rutin, hydrolyzes it to quercetin. This activity can be further induced when rutin is added to the fecal culture from which the cell-free preparation is made. When vitamin pills that contain rutin are added to the cultures, this induction is equally effective. The vitamin extracts by themselves, like rutin, were nonmutagenic; however, when the vitamin extracts were incubated with fecal preparations containing induced beta-glucosidase, a great increase in mutagenicity was observed.     

Radioprotective activity of hypotensive preparations

In experiments on (CBA X C57B1)F1 mice a study was made of the radioprotective efficiency of some hypotensive drugs. Hemiton administered in different ways, within a wide dose range and at different times before irradiation was shown to possess a pronounced protective action. The substitution of the acid residue in hemiton decreased its activity.