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PubMed1984

Multiple forms of alkaline phosphatases in human liver tissue.

Alkaline phosphatases (AP) extracted in the presence of n-butanol from human liver are separated by affinity chromatography on phenylsepharose Cl-4B into two fractions named APII and APIIII. By repeated chromatography, APII was purified to a single enzyme entity with a specific activity of 1,684 kU/g protein. APIIII was purified to a specific activity of 535 kU/g protein. It consisted of only APIIII enzyme activity, but still contained gamma-glutamyltransferase activity. These two forms of AP are different in chromatographic and electrophoretic behaviour, APIIII being a larger molecule than APII. APII and APIIII are very similar in enzyme kinetic behavior, such as substrate activity, thermolability and sensitivity to different inhibitors. It is concluded from these experiments that multiple forms of AP in liver bear identical active centres, the difference is due to a modification of protein residue. It is possible that both are modified forms of one enzyme. Both are different from the AP isoenzyme that appears in serum in cholestatic patients.

Alkaline Phosphatase↗
PubMed1974

Biochemical localization of alkaline phosphatase in the cell wall of a marine pseudomonad.

The various layers of the cell envelope of marine pseudomonad B-16 (ATCC 19855) have been separated from the cells and assayed directly for alkaline phosphatase activity under conditions established previously to be optimum for maintenance of the activity of the enzyme. Under conditions known to lead to the release of the contents of the periplasmic space from the cells, over 90% of the alkaline phosphatase was released into the medium. Neither the loosely bound outer layer nor the outer double-track layer (cell wall membrane) showed significant activity. A small amount of the alkaline phosphatase activity of the cells remained associated with the mureinoplasts when the outer layers of the cell wall were removed. Upon treatment of the mureinoplasts with lysozyme, some alkaline phosphatase was released into the medium and some remained with the protoplasts formed. Cells washed and suspended in 0.5 M NaCl were lysed by treatment with 2% toluene, and 95% of the alkaline phosphatase in the cells was released into the medium. Cells washed and suspended in complete salts solution (0.3 M NaCl, 0.05 M MgSO(4), and 0.01 M KCl) or 0.05 M MgSO(4) appeared intact after treatment with toluene but lost 50 and 10%, respectively, of their alkaline phosphatase. The results suggest that the presence of Mg(2+) in the cell wall is necessary to prevent disruption of the cells by toluene and may also be required to prevent the release of alkaline phosphatase by toluene when disruption of the cells by toluene does not take place.

Alkaline Phosphatase↗
PubMedDate not supplied

Alkaline phosphatase from Nocardia asteroides.

Intracellular alkaline phosphatase (Apase) produced during growth of Nocardia asteroides in dephosphated nutrient broth supplemented with 1% glucose and some properties of the purified enzyme were studied. Filamentous cells of N. asteroides synthetized the Apase in high yields (7.0 x 10(3) units/mg cell protein) during exponential growth phase. The intracellular level dropped drastically and the extracellular amounts increased during stationary phase, probably due to the loss of the enzyme in the medium because of the fragmentation of the filamentous cells into rod-like and spherical forms and/or due to cell lysis. The purified enzyme showed typical Apase activity with optimal pH at 10 and optimal temperature at 37 degrees C. The Km of the enzyme for p-nitrophenyl phosphate was close to the value reported for Apase from other sources. Other properties of the enzyme which resembled those from other species included respectively the uncompetitive, non-competitive and competitive inhibition of L-phenylalanine, urea and inorganic phosphate. The effect of these compounds on the enzyme may be useful in the regulation of Apase activity in vivo and hence may play a vitalrole during morpho genesis of N. asteroides.

Alkaline Phosphatase↗
PubMed1981

Subcellular localization and properties of pyridoxal phosphate phosphatases of human polymorphonuclear leukocytes and their relationship to acid and alkaline phosphatase.

Using a novel fluorimetric assay for pyridoxal phosphate phosphatase, human polymorphonuclear leucocytes were found to exhibit both acid an alkaline activities. The neutrophils were homogenised in isotonic sucrose and subjected to analytical subcellular fractionation by sucrose density gradient centrigfugation. The alkaline pyridoxal phosphate phosphatase showed a very similar distribution to alkaline phosphatase an was located solely to the phosphasome granules. Fractionation experiments on neutrophils treated with isotonic sucrose containing digitonin and inhibitor studies with diazotised sulphanilic acid and levamisole further confirmed that both enzyme activities had similar locations and properties. Acid pyridoxal phosphate phosphatase activity was located primarily to the tertiary granule with a partial azurophil distribution. Fractionation studies on neutrophils homogenised in isotonic sucrose containing digitonin and specific inhibitor studies showed that acid pyridoxal phosphate phosphatase and acid phosphatase were not the result of a single enzyme activity, Neutrophils were isolated from control subjects, patients with chronic granulocytic leukaemia and patients in the third trimester of pregnancy. The specific activities (munits/mg protein) of alkaline pyridoxal phosphate phosphatase an alkaline phosphatase varied widely in the three groups and the alterations occurred in a parallel manner. The specific activities of acid pyridoxal phosphate phosphatase and of acid phosphatase were similar in the three groups. These results, together with the fractionation experiments and inhibition studies strongly suggest that pyridoxal phosphate is a physiological substrate for neutrophil alkaline phosphatase.

Acid Phosphatase↗
PubMed2001

Effect of fasting and refeeding on duodenal alkaline phosphatase activity in monosodium glutamate obese rats.

In the present work the effects of fasting and refeeding on fat pad weight and alkaline phosphatase activity in the brush border of individual duodenal enterocytes have been evaluated in male Wistar rats with obesity induced by monosodium glutamate (MSG) treatment during the early postnatal period. Neonatal rats were treated subcutaneously with MSG (2 mg/g b.w.) or saline (controls) for 4 days after birth. At 4 months of age, two types of experiments were performed. In the first experiment rats, were submitted to 3 or 6 days lasting food deprivation. In the second experiment the rats were refed for 3 or 6 days ad libitum or restrictedly (60% of pre-fasting intake) after a 6 day-fasting period. Fasting and refeeding influenced the body fat and function of the duodenum in MSG-treated rats differently as compared to the controls. However, alkaline phosphatase activity and the weight of epididymal and retroperitoneal fat depots were significantly increased in MSG obese rats (P<0.001) during all the periods examined. While 3 days of food deprivation resulted in both groups in a similar loss of adipose tissue weight and alkaline phosphatase activity, the decrements of these parameters after 6 days of fasting were lower in obese rats suggesting that their capacity to spare body fat stores was enhanced. After 3 days of ad libitum refeeding, a more marked adaptational increase of food consumption and also a significantly increased alkaline phosphatase activity above the pre-fasting level (P<0.01) was observed in the MSG-treated rats. Consequently, a more rapid body fat restoration was demonstrated in these animals. Refeeding of rats at 60% of the pre-fasting intake level resulted in a significant increase of alkaline phosphatase activity in both the MSG and control group; moreover, as food restriction continued, MSG-treated rats tended to further increase the enzyme activity. Our results revealed that MSG treatment of neonatal rats may significantly change the intestinal functions. Permanently increased alkaline phosphatase activity observed in MSG obese rats during all investigated periods suggests that this functional alteration is probably not a consequence of actual nutritional variation but could be a component of regulatory mechanisms maintaining their obesity at critical values.

Adipose Tissue↗
PubMed1986

Purification of placental alkaline phosphatase and its monoclonal antibody.

Monoclonal antibodies (MAbs) to placental alkaline phosphatase (PAlP) were raised by using the hybridoma technique. Spleen cells from immunized mice were fused with the mouse myeloma line NS-1 using 50% polyethylene glycol and cultured in a selection medium. Antibodies were screened by the enzyme immunoassay using immobilized solid-phase antigen and anti-mouse immunogloblin Fab' (rabbit)-beta-D-galactosidase complex. Four double-cloned hybridomas were obtained. The cross-reactivity with different kinds of alkaline phosphatases of the raised MAbs was examined. At first, MAb 11-D-10, with which placenta was stained but liver and small intestine were not stained in indirect immunofluorescence, was selected. Then, the cross-reactivity of MAb 11-D-10 was further investigated by immunoblotting (Western blotting). MAb 11-D-10 reacted with PAlP but did not react with hepatic and intestinal alkaline phosphatase at all. The binding activity of 125I-labeled MAb 11-D-10 with different choriocarcinoma cell lines (SCH, BeWo, NaUCC-1, NaUCC-2, and NaUCC-3) was highest in SCH, then in the order of BeWo, NaUCC-3, NaUCC-1 and NaUCC-2, and correlated to the PAlP content of the cells (SCH greater than BeWo greater than NaUCC-3 greater than NaUCC-1 greater than NaUCC-2). Also the intensity of indirect immunofluorescence of the above cell lines with MAb 11-D-10 correlated with the PAlP content of each cell.

Alkaline Phosphatase↗
PubMed1991

Expression of alkaline phosphatase in murine lymphoma cells.

Alkaline phosphatase (ALP) was secreted and expressed at the cell surface of the lymphoma A/63-2 cell line but not on another clone A/63-1 deriving from a single thymoma (A/63) induced by a wild-type Abelson-Moloney viral complex. The enzyme was heat-sensitive and strongly inhibited by L-p-bromotetramisole and L-homoarginine but not by L-phenylalanine. All these data indicated that this enzyme was most likely identical to the L/B/K ALP isoenzyme. Southern blot analysis showed that neither amplification nor polymorphism were responsible for the high expression of the ALP gene observed in A/63-2 cells. On the opposite, the mRNA transcripts of ALP were only detected in A/63-2 cells indicating that a modulation of the ALP gene transcription occurred which could be due to the insertion of the v-abl gene within or near the 5'-flanking region of the ALP promotor in A/63-2 cells. Butyrate strongly increased both the secretion and the expression of the enzyme on A/63-2 cell surface. This induction was strongly inhibited by cordycepin, an RNA biosynthesis inhibitor, and at a lesser degree by cycloheximide, a translation inhibitor suggesting that butyrate induction occurs both at the transcriptional and the translational level.

Alkaline Phosphatase↗
PubMedDate not supplied

Kinetic determination of granulocyte alkaline phosphatase by the GEMSAEC.

Granulocyte alkaline phosphatase (ALP) activity is measured kinetically using the GEMSAEC centrifugal analyzer and p-nitrophenylphosphate as substrate. Measurements of enzyme activity are made at pH 9.8 utilizing 1.0 M diethanolamine as a buffer containing 0.5 mM magnesium chloride. Granulocytes are separated from heparinized blood using dextran sedimentation, followed with ficoll-hypaque separation. The separated cells are suspended in saline, and the red cells are lysed. Manual counting of stained smears of the cells isolated showed that 95 percent of these cells were granulocytes. After lysing the red cells, the granulocyte suspension is rinsed in 12.5 percent heat inactivated serum in saline and separated by centrifugation. After 200 microliters of the diluent solution (12.5 percent heat inactivated serum in saline) is added to the cells, they are sonicated for seven sec at 0 degrees in an ice bath. The extracts are then assayed kinetically for ALP activity. Granulocyte ALP activity varies from day to day in normal individuals. The daily variation of ALP activity observed correlates closely with and is confirmed by the Sigma Histochemical Procedure based on Kaplow' Scoring Method. The coefficient of variation of the ALP method within assay performed on two consecutive days on a pooled sample is 0.75 percent and 1.38 percent, respectively, and between assays 4.82 percent.

Adult↗
PubMed1991

Evidence for glycosylphosphatidylinositol anchoring of intralumenal alkaline phosphatase of the calf intestine.

1. Considerable amounts of intestinal alkaline phosphatase (AP) were found intralumenally in all animal species investigated, i.e. calf, pig, goat, rat, mouse, guinea pig, hen and carp. The ratios between the total activity of AP found intralumenally and the total intestinal activity vary considerably. Calves and pigs show the highest, i.e. 0.77 and 0.44, respectively, while rodents have much lower ratios. Only 20-34% of the intralumenal alkaline phosphatase (IAP) of the calf and pig is soluble and not within the sediment after centrifugation at 135,000 x g for 60 min. whereas the IAP of rodents is soluble in the range of 60-72% of the total IAP. 2. For the IAP of the mucosa and chyme of calf, all criteria were found which are generally used, indicating a glycosylphosphatidylinositol (GlcPtdIns) anchor as proved by strong hydrophobicity using Triton X-114 phase partitioning, phenyl-Sepharose binding and enzyme aggregation, and the susceptibility to phosphatidylinositol-specific phospholipase C (PtdIns-PLC) and papain digestion. 3. More than 80% of the mucosa alkaline phosphatase (MAP) of the proximal part of the intestine and of the particulate fraction of IAP exhibit these criteria indicating the presence of the GlcPtdIns-anchor structure, whereas the anchor content of the soluble intralumenal enzyme decreases from the pylorus to the ileocecal junction. 4. MAP partially purified to a specific activity of 1747 IU/mg retains the anchor structure. 5. The results presented indicate that the release of large amounts of AP into the chyme is realized without splitting the GlcPtdIns anchor. The possible intralumenal function of this form of AP is discussed.

Alkaline Phosphatase↗
PubMed1997

Identification of human pulmonary alkaline phosphatase isoenzymes.

An increase of alkaline phosphatase (ALP) activity has been observed in the bronchoalveolar lavage fluid (BALF) of patients affected by pulmonary fibrosis in chronic interstitial lung disorders. To characterize the ALP isoenzymes in such cases, we used gel filtration, agarose gel electrophoresis, heat and amino acid inhibition assays, wheat-germ agglutinin (WGA) precipitation, and an immunoassay specific for the bone-isoform of ALP. Only one anodic band representing a high-molecular-weight isoform of ALP (Mr approximately 2,000 kDa) was observed on electrophoresis of BALF. The inhibition assay results were consistent for a tissue-nonspecific isoenzyme sensitive to a temperature of 56 degrees C (71.9 +/- 2.5% inhibition) and to homoarginine (65.7 +/- 1.9%), and resistant to L-phenylalanine and L-leucine. Less than 13% of ALP activity was heat-stable. After incubation of BALF specimens with glycosyl-phosphatidylinositol-phospholipase D plus Nonidet P-40, or with phosphatidylinositol-phospholipase C alone, an electrophoretic cathodic band (Mr approximately 220 kDa) appeared near the bone band of a standard serum. With the WGA assay, 84.4 +/- 3.3% of ALP precipitated and the band disappeared. After immunoassay for the bone isoform, a mean of less than 5% enzyme activity was measured. We conclude that the ALP found in BALF is a pulmonary isoform of a tissue nonspecific isoenzyme.

Adult↗
PubMed1984

Altered isozyme patterns of leucocyte alkaline phosphatase in disease states.

Leucocyte alkaline phosphatase (LAP) is a granulocyte enzyme whose concentration varies in disease states. In order to determine whether the pattern of expression is altered in leukaemic granulocytes, we have analysed the LAP isozyme pattern of a series of normal subjects and patients with various haematological diseases. Electrophoretic patterns of partially purified LAP samples were determined by polyacrylamide gel electrophoresis in the presence of Triton X-100. These patterns were reproducible on repeated samples from the same patient. Presence of the LAPf and LAPs isozymes were determined after staining with the dye Fast Blue BB. Granulocytes were obtained from 15 normal subjects. Thirteen of these samples had only the LAPs isozyme. The other two had LAPs, plus a small amount (less than or equal to 10% of total) of LAPf activity. Eight patients with stable phase chronic myelogenous leukaemia (CML) had only small amounts of the normal LAPs isozyme and no evidence of LAPf . Of 11 patients with CGL who clinically had blast crisis. 10 had both LAPs and LAPf . The eleventh patient who was Ph1 negative had only LAPs. Three of five patients with polycythaemia vera had only the LAPs isozyme while two had both isozymes. Six patients with non-malignant leucocytosis had only LAPs. We interpret this data to indicate that the increased levels of LAP activity in some CGL blast crisis patients are primarily related to synthesis of the LAPf isozyme.

Alkaline Phosphatase↗
PubMed2003

Induction of glomerular alkaline phosphatase after challenge with lipopolysaccharide.

Alkaline phosphatase (AP) can be considered as a host defence molecule since this enzyme is able to detoxify bacterial endotoxin at physiological pH. The question emerged whether this anti-endotoxin principle is inducible in the glomerulus and if so, which glomerular cells might be involved in the expression of ectoAP after stimulation with pro-inflammatory agents. Therefore kidneys of rats treated with either lipopolysaccharide (LPS), E. coli bacteria or non-toxic monophosphoryl lipid A (MPLA) were examined for AP activity 6 or 24 h after challenge. In addition cultures of endothelial cells or mesangial cells were evaluated for AP activity after stimulation with either LPS, TNFalpha or IL-6, and mRNA for AP was studied in TNFalpha-stimulated and control mesangial cells. The results show significant up-regulation of glomerular AP in LPS- or E. coli-injected rats compared to rats injected with MPLA. Endothelial and mesangial cells in vitro showed significant up-regulation of AP activity following stimulation with LPS, TNFalpha or IL-6, whereas increased mRNA for AP was observed in mesangial cells after TNFalpha stimulation compared to non-stimulated control cells. Since it appeared that hydrolysis occurred when endotoxin was used as a substrate in the histochemical staining, we concluded that inducible glomerular ectoAP may reflect a local endotoxin detoxifying principle of the kidney.

Alkaline Phosphatase↗
PubMed1997

Effect of calcium ions on rat osseous plate alkaline phosphatase activity.

Rat osseous plate alkaline phosphatase is a metalloenzyme with two binding sites for Zn2+ (sites I and III) and one for Mg2+ (site II). This enzyme is stimulated synergistically by Zn2+ and Mg2+ (Ciancaglini et al., 1992) and also by Mn2+ (Leone et al., 1995) and Co2+ (Ciancaglini et al., 1995). This study was aimed to investigate the modulation of enzyme activity by Ca2+. In the absence of Zn2+ and Mg2+, Ca2+ had no effects on the activity of Chelex-treated, Polidocanol-solubilized enzyme. However, in the presence of 10 microM MgCl2, increasing concentration of Ca2+ were inhibitory, suggesting the displacement of Mg2+ from the magnesium-reconstituted enzyme. For calcium-reconstituted enzyme, Zn2+ concentrations up to 0.1 microM were stimulatory, increasing specific activity from 130 U/mg to about 240 U/mg with a K0.5 = 8.5 nM. Above 0.1 microM Zn2+ exerted a strong inhibitory effect and concentrations of Ca2+ up to 1 mM were not enough to counteract this inhibition, indicating that Ca2+ was easily displaced by Zn2+. At fixed concentrations of Ca2+, increasing concentrations of Mg2+ increased the enzyme specific activity from 472 U/mg to about 547 U/mg, but K0.5 values were significantly affected (from 4.4 microM to 38.0 microM). The synergistic effects observed for the activity of Ca2+ plus magnesium-reconstituted enzyme, suggested that these two ions bind to the different sites. A model to explain the effect of Ca2+ on the activity of the enzyme is presented.

Alkaline Phosphatase↗
PubMed2003

Dimer asymmetry and the catalytic cycle of alkaline phosphatase from Escherichia coli.

Although alkaline phosphatase (APase) from Escherichia coli crystallizes as a symmetric dimer, it displays deviations from Michaelis-Menten kinetics, supported by a model describing a dimeric enzyme with unequal subunits [Orhanović S., Pavela-Vrancic M. and Flogel-Mrsić M. (1994) Acta. Pharm.44, 87-95]. The possibility, that the observed asymmetry could be attributed to negative cooperativity in Mg2+ binding, has been examined. The influence of the metal ion content on the catalytic properties of APase from E. coli has been examined by kinetic analyses. An activation study has indicated that Mg2+ enhances APase activity by a mechanism that involves interactions between subunits. The observed deviations from Michaelis-Menten kinetics are independent of saturation with Zn2+ or Mg2+ ions, suggesting that asymmetry is an intrinsic property of the dimeric enzyme. In accordance with the experimental data, a model describing the mechanism of substrate hydrolysis by APase has been proposed. The release of the product is enhanced by a conformational change generating a subunit with lower affinity for both the substrate and the product. In the course of the catalytic cycle the conformation of the subunits alternates between two states in order to enable substrate binding and product release. APase displays higher activity in the presence of Mg2+, as binding of Mg2+ increases the rate of conformational change. A conformationally controlled and Mg2+-assisted dissociation of the reaction product (Pi) could serve as a kinetic switch preventing loss of Pi into the environment.

Alkaline Phosphatase↗
PubMed1990

Production of a mouse monoclonal antibody against the alkaline phosphatase of adult Schistosoma mansoni.

This study describes the production and characterization of a monoclonal antibody (mAb) against the alkaline phosphatase of Schistosoma mansoni from splenocytes of chronically infected mice. Convenient selection of the mAb was achieved using the catalytic activity of the antigen in a developed enzyme-antigen immunoassay. The mAb was of the IgG1 subclass and it specifically recognized the alkaline phosphatase in adult worm sections by indirect immunofluorescence. Preincubation of the antibody with partially purified adult alkaline phosphatase did not result in inhibition of the enzyme activity and it did not mediate complement-dependent cytotoxicity against mechanically transformed schistosomula in vitro. The mAb was able to immunoprecipitate under reducing conditions a polypeptide of 65 kDa, similar in size to the monomeric subunit of the schistosome enzyme. The specificity of the mAb was assessed by competitive inhibition with antibodies of infected human sera in an immunoadsorption assay. Periodate treatment of the antigen resulted in altered electrophoretic mobility of alkaline phosphatase, which confirmed the presence of carbohydrate in the molecule, but this did not prevent binding by the mAb. Although the use of the mAb in capture assays for detection of circulating alkaline phosphatase in infected host sera was unsuccessful, the production of this antibody confirmed that the enzyme is exposed by adult worms to the host and that it is immunogenic; additionally, a monoclonal probe is available for further characterization of the structure and function of this important parasite surface molecule.

Alkaline Phosphatase↗
PubMed2005

Production of monoclonal antibody against alkaline phosphatase.

BACKGROUND: Monoclonal antibody against alkaline phosphatase (Alp) has many applications which Alp-anti Alp complex (APAAP) formation is one of the most important ones. This complex is applicable in many immunohistochemical and immunocytochemical techniques such as diagnosis of various kinds of leukemias, lymphomas, skin diseases, kidney dysfunctions, etc. OBJECTIVE: Production of anti-Alp monoclonal antibody for utilization in APAAP complex. METHODS: After several arranged injections of Alp to Balb/c mice and determining the specific antibody titer by ELISA test, the spleen lymphocytes of immunized mice and Sp2/0 myeloma cells were fused using polyethylene glycol as fusing agent and hybridoma cells were cultured in HAT medium. Identification and selection of anti-Alp producing clones were done by performing ELISA test on supernatants of all resulting clones. Limiting dilution method was used to attain monoclones and the effect of obtained antibodies on enzyme activity was investigated by a specific ELISA test. For production of concentrated Ab, the hybridoma cells were injected to peritoneal cavity of mice and the produced ascetic fluids were collected. Finally class and subclass of the obtained antibodies were determined by Isostrip kit. RESULTS: After six rounds of fusion, 104 Hybridoma clones were obtained and two Anti-Alp producing clones (A_1G_8 and A_1G_9) were selected and subcloned. Both antibodies were IgG_1 with kappa (kappa) light chains. These antibodies did not affect the enzyme activity and the electrophoresis of ascetic fluids showed an obvious band in gamma (gamma) position. CONCLUSION: Because these antibodies are from IgG class and don't affect the enzyme activity, it seems that they are suitable for APAAP complex formation.

Alkaline Phosphatase↗
PubMedDate not supplied

Evaluation of osteoblastic activity by morphometric comparison of alkaline phosphatase cytochemistry vs. tetracycline fluorescence.

Alkaline phosphatase (ALP) activity was used as a novel histomorphometric index of osteoblastic surfaces involved in mineralization. The enzyme cytochemical reaction was done on sections of low temperature processed, glycol methacrylate (GMA) embedded bone biopsies from 39 patients with various types of renal osteodystrophy (age 48 +/- 12 yrs; 19 males, 20 females) who had received tetracycline labelling. Sets of three serial sections were obtained from each tissue block: the 1st section (2 microns thick) was stained with Methylene blue Azure 11 for morphology; the 2nd section (2 microns thick) was used for ALP cytochemistry; the 3rd section was left unstained for UV microscopy. ALP positive osteogenic cells on bone surfaces displayed either of two distinct morphologies: a) typical plump, 'active' osteoblasts, and b) flat, elongated cells otherwise indistinguishable from 'bonelining cells'. These ALP+ flat cells were in contact with sites of active osteoid and mineral deposition and also codistributed with tetracycline labels outside of, and in continuity with, osteoid seams. Flat lining cells which were ALP negative were never associated with labels. Therefore, ALP activity also provided an objective criterion for differentiating two different 'phenotypes' among flat bone lining cells (ALP+ and ALP-), associated or not associated with matrix mineralization, respectively. The following histomorphometric variables were measured: Ob.S/BS, OS/BS, MS/BS and ALP.S/BS. Ob.S/BS, OS/BS and MS/BS were different in different types of ROD. However, OS/BS always exceeded MS/BS which, in turn, always exceeded Ob.S/BS. ALP.S/BS exceeded OS/BS in controls, mixed ROD and hyperparathyroidism, whereas the reverse occurred in osteomalacia and aplastic bone, due to the abundance of ALP lining cells over nonmineralizing surfaces.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1990

Expression of a Nagao-type, phosphatidylinositol-glycan anchored alkaline phosphatase in human choriocarcinomas.

The alkaline phosphatase (AP) synthesized by human tumor cells closely resembles human placental AP (PLAP). Little is known about the molecular events that lead to the expression of a placenta-like AP in tumor cells. The complementary DNA encoding the AP expressed by a choriocarcinoma cell line, BeWo, was isolated and characterized. The complementary DNA is the product of the germ cell AP (Nagao isozyme) gene and not of the term PLAP gene. Like placental AP, the tumor AP can be released from the cell membrane by a phosphaditylinositol-specific phospholipase C and has a phosphaditylinositol-glycan (PI-glycan) moiety at the COOH terminus. Immunoprecipitation of phosphaditylinositol-specific phospholipase C-treated AP and analysis by polyacrylamide gel electrophoresis or isoelectric focusing demonstrates that at least 95% of the AP contains PI-glycan. Two-dimensional gel electrophoresis reveals two precursors of the mature AP. One of these does not bind an antibody against the Trypanosoma variable surface glycoprotein cross-reacting determinant and probably does not contain PI-glycan. This precursor had a shorter half-life than the more prominent PI-glycan-containing precursor in pulse-chase experiments, suggesting a precursor-product relationship between the two proteins. These data demonstrate that BeWo AP is the product of a gene normally expressed in testis, thymus, and germ cells, but not in placenta. Thus, the expression of BeWo AP results from the repression of the PLAP gene and derepression of the germ cell AP gene and, as such, the expression is ectopic. The BeWo AP (Nagao isozyme) is modified with PI-glycan that is added soon after translation, not cotranslationally.

Alkaline Phosphatase↗