scholarly journals Alkaline phosphatase protein increases in response to prednisolone in HeLa cells

1981 ◽  
Vol 200 (2) ◽  
pp. 461-464 ◽  
Author(s):  
W C Hanford ◽  
R H Kottel ◽  
W H Fishman
Keyword(s):  
Alkaline Phosphatase ◽  
Hela Cells ◽  
Specific Protein ◽  
Active State ◽  
Placental Alkaline Phosphatase ◽  
Prednisolone Treatment ◽  
Presence And Absence ◽  
Alkaline Phosphatase Isoenzyme

Quantification of term-placental alkaline phosphatase isoenzyme protein in HeLa TCRC-1 cells grown in the presence and absence of prednisolone indicates that there is a net increase in amount of enzyme-specific protein in prednisolone-stimulated cells. In a similar analysis of HeLa D98AH2 cells, prednisolone treatment causes the appearance of term-placental alkaline phosphatase protein and the loss of the intestinal isoenzyme protein. These results support the interpretation that the response of these cells to corticosteroids is the net accumulation of alkaline phosphatase protein rather than the modification of pre-existing enzyme to a more active state.

Use of monoclonal antibodies to detect human placental alkaline phosphatase.

1983 ◽  
Vol 29 (1) ◽  
pp. 115-119 ◽  
Author(s):  
G De Groote ◽  
P De Waele ◽  
A Van de Voorde ◽  
M De Broe ◽  
W Fiers
Keyword(s):  
Monoclonal Antibody ◽  
Alkaline Phosphatase ◽  
Monoclonal Antibodies ◽  
Solid Phase ◽  
Enzymatic Reactions ◽  
Placental Alkaline Phosphatase ◽  
Human Placental Alkaline Phosphatase ◽  
Human Sera ◽  
Starch Gel ◽  
Alkaline Phosphatase Isoenzyme

Abstract Convenient, sensitive, and specific solid-phase immunoassays involving monoclonal antibody are described for the determination of human placental alkaline phosphatase (hPLAP). An endogenous enzyme immunoassay combined the specificity of the immunological and the enzymatic reactions. Alternatively, a solid-phase "sandwich" radioimmunoassay involving immobilized polyclonal rabbit anti-hPLAP in combination with iodinated monoclonal antibody provided some additional advantages. Both tests can be used to detect hPLAP from various sources, e.g., in human sera during pregnancy or as a tumor marker. The radioimmunoassay detected an increase in hPLAP at nine weeks of gestation. We discuss the use of monoclonal antibodies for the differentiation of different alkaline phosphatase isoenzyme types by electrophoresis on starch gel.

scholarly journals The effect of bromodeoxyuridine and dibutyryl cyclic AMP on the induction of placental alkaline phosphate in human choriocarcinoma cells

1981 ◽  
Vol 198 (1) ◽  
pp. 29-35 ◽  
Author(s):  
T A Hamilton ◽  
H H Sussman
Keyword(s):  
Alkaline Phosphatase ◽  
Cyclic Amp ◽  
Polyacrylamide Gel Electrophoresis ◽  
Specific Protein ◽  
Placental Alkaline Phosphatase ◽  
Dibutyryl Cyclic Amp ◽  
Specific Enzyme ◽  
Alkaline Phosphate ◽  
Choriocarcinoma Cells ◽  
Human Choriocarcinoma Cells

The effect of 5-bromo-2′-deoxyuridine (BrdUrd) and dibutyryl cyclic AMP (Bt2cAMP) on the expression of the placental isoenzyme of human alkaline phosphatase was examined in BeWo choriocarcinoma cells. By using a combination of specific immunoprecipitation and polyacrylamide-gel electrophoresis of cells labelled either metabolically with [35S]methionine or cell-surface-labelled with 125I, both BrdUrd (5 micrograms/ml) and 1 mM-Bt2cAMP were shown to result in the enhanced accumulation of a specific protein. This protein has immunochemical identity and co-electrophoreses with placental alkaline phosphatase in two-dimensional gels. These results clearly demonstrate that the induction of placental alkaline phosphatase activity in choriocarcinoma cells treated with these agents is a consequence of the accumulation of specific enzyme protein rather than of altered catalytic activity.

Characterization of placental isoenzyme of alkaline phosphatase in human pregnancy serum

1969 ◽  
Vol 47 (2) ◽  
pp. 147-155 ◽  
Author(s):  
N. K. Ghosh ◽  
W. H. Fishman
Keyword(s):  
Alkaline Phosphatase ◽  
Serum Alkaline Phosphatase ◽  
Heat Stability ◽  
Biochemical Properties ◽  
Heat Inactivation ◽  
Total Serum ◽  
Placental Alkaline Phosphatase ◽  
Human Placental Alkaline Phosphatase ◽  
Alkaline Phosphatase Isoenzyme ◽  
In Pregnancy

Human placental alkaline phosphatase isoenzyme has been characterized in pregnancy serum by several biochemical criteria. The total serum alkaline phosphatase, its L-phenylalanine-sensitive moiety, heat inactivation, and the ratio of enzyme activity at pH 10.7 versus 9.8 (10.7/9.8 R) were measured during parturition and 59 weeks of pre- and post-natal periods. The extent of L-phenylalanine inhibition, heat stability, and 10.7/9.8 R of serum alkaline phosphatase progressively increased during gestation attaining maximum values during the delivery, after which they gradually declined. The electrophoretic behaviors of alkaline phosphatase isoenzymes of pregnancy sera were followed by starch- and Sephadex-gel electrophoreses. Alkaline phosphatase has been purified 300-fold from the placenta of the subject whose serum enzyme was investigated. The biochemical properties, including the electrophoretic behavior and neuraminidase sensitivity of heat-stable alkaline phosphastase in pregnancy sera at term, were comparable to those of purified placental alkaline phosphatase. The values for 10.7/9.8 R of the pregnancy sera were statistically different from those of sera from normal nonpregnant women. The results obtained in this study suggest that the enhanced level of pregnancy serum alkaline phosphatase is due to the enrichment of the circulation with an isoenzyme of placental origin.

L-Phenylalanine inhibition of human alkaline phosphatases with p-nitrophenyl phosphate as substrate.

1982 ◽  
Vol 28 (12) ◽  
pp. 2426-2428 ◽  
Author(s):  
T Komoda ◽  
S Hokari ◽  
M Sonoda ◽  
Y Sakagishi ◽  
T Tamura
Keyword(s):  
Alkaline Phosphatase ◽  
Human Placenta ◽  
Placental Alkaline Phosphatase ◽  
Specific Inhibition ◽  
Human Intestine ◽  
Alkaline Phosphatases ◽  
Substrate Complex ◽  
Nitrophenyl Phosphate ◽  
Organ Specific ◽  
Alkaline Phosphatase Isoenzyme

Abstract With p-nitrophenyl phosphate as the substrate, there reportedly is no organ-specific inhibition of alkaline phosphatase (EC 3.1.3.1) activity by L-phenylalanine. However, we found that at pH 10.0, with p-nitrophenyl phosphate as the substrate, L-phenylalanine obviously inhibits the alkaline phosphatase isoenzyme from human placenta, whereas there is little if any inhibition of the isoenzyme from human intestine. Because of the differing effects of substrates (p-nitrophenyl phosphate and phenyl phosphate) and their enzymic products (p-nitrophenol and phenol) for L-phenylalanine action on the placental alkaline phosphatase isoenzyme, we suggest that the isoenzyme--inhibitor--substrate complex and the effect of released phosphate on L-phenylalanine inhibition of the isoenzyme activity differ from each other.

Micro-scale two-dimensional electrophoresis of alkaline phosphatase from serum.

1988 ◽  
Vol 34 (4) ◽  
pp. 730-735 ◽  
Author(s):  
G J Sanderink ◽  
Y Artur ◽  
F Paille ◽  
M M Galteau ◽  
G Siest
Keyword(s):  
Alkaline Phosphatase ◽  
Liver Microsomes ◽  
Molecular Complexes ◽  
Placental Alkaline Phosphatase ◽  
Two Dimensional ◽  
Two Dimensional Electrophoresis ◽  
Micro Scale ◽  
Interindividual Variations ◽  
Alkaline Phosphatase Isoenzyme ◽  
Dimensional Electrophoresis

Abstract Isoenzymes of alkaline phosphatase (EC 3.1.3.1) were separated by micro-scale two-dimensional electrophoresis, with isoelectric focusing in capillary gels in the first dimension and polyacrylamide gradient-gel electrophoresis in the second. The isoenzymes detected were identified by several treatments--e.g., incubation with sialidase, papain, Triton X-100, and wheat-germ agglutinin--and by comparison with alkaline phosphatase from liver microsomes. Liver and bone isoforms in normal sera showed overlapping isoelectric points but differed in molecular mass, estimated as 172 and 185 kDa, respectively. Sera of patients with liver disease showed several additional groups of alkaline phosphatase isoforms, two of which were found to consist of multi-molecular complexes. Others probably correspond to incompletely glycated enzyme proteins. A further isoform with a mass of about 250 kDa does not seem to correspond to any known isoform of alkaline phosphatase in serum. With this technique, we demonstrated intra- and interindividual variations of the placental alkaline phosphatase isoenzyme in pregnancy sera.

scholarly journals Biosynthesis of glycosylphosphatidylinositol-anchored human placental alkaline phosphatase: evidence for a phospholipase C-sensitive precursor and its post-attachment conversion into a phospholipase C-resistant form

1994 ◽  
Vol 301 (1) ◽  
pp. 205-209 ◽  
Author(s):  
Y W Wong ◽  
M G Low
Keyword(s):  
Alkaline Phosphatase ◽  
Phospholipase C ◽  
Hela Cells ◽  
Placental Alkaline Phosphatase ◽  
Gpi Anchor ◽  
Chase Period ◽  
Resistant Species ◽  
Resistant Form ◽  
Human Placental Alkaline Phosphatase ◽  
Triton X

Previous studies have shown that some cells (e.g. SKG3a) express human placental alkaline phosphatase (AP) in a form which can be released from the membrane by bacterial PtdIns-specific phospholipase C (PI-PLC) while others (e.g. HeLa) are relatively resistant to this enzyme. Chemical and enzymic degradation studies have suggested that the PI-PLC resistance of AP is due to inositol acylation of its glycosylphosphatidylinositol (GPI) anchor. In order to identify the biosynthetic origin of PI-PLC resistance we determined the PI-PLC sensitivity of AP in 35S-labelled cells (10 min pulse; 0-60 min chase) by Triton X-114 phase separation. At the beginning of the chase period, the majority of the AP synthesized was hydrophilic, indicating that it had not acquired a GPI anchor. The concentration of hydrophilic AP species decreased with a t1/2 of 30-60 min but was not processed to an endoglycosidase H-resistant species or secreted into the medium. In both SKG3a and HeLa cells all of the hydrophobic, GPI-anchored AP detectable at the beginning of the chase was PI-PLC sensitive. PI-PLC-resistant species of AP were only observed in HeLa cells and these only appeared after about 30 min. The delayed appearance of PI-PLC resistance was unexpected as previous studies have suggested that candidate GPI-anchor precursors are PI-PLC-resistant as a result of inositol acylation. This work reveals unanticipated complexities in the biosynthesis of AP and its GPI anchor.

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