Changes in the activity of different classes of enzymes in the cerebral cortex of rats in ontogeny and after the transplantation of embryonic nerve tissue

The activity of lactate dehydrogenase (LDH), indophenol oxidase, aspartate aminotransferase (AsAT), alkaline phosphatase, acid phosphatase and aldolase at different stages of rat development was measured. We have also determined changes in the activity of these enzymes resulting from transplantation of embryonic nerve tissue (ENT) into the brain of adult animals. During development from the embryo to the adult animal, LDH and AsAT activities increased, while alkaline phosphatase activity diminished. After ENT transplantation, the most prominent changes were in the alkaline phosphatase activity whereas the activity of LDH, AsAT and acid phosphatase remained unchanged and similar to that in the brain cortex of intact adult animals. Changes in the enzyme activity resulting from ENT transplantation changed in a manner characteristic of the transplant. Local brain damage did not change the activity of the studied enzymes fifty days after surgery.     

Histological and histochemical studies on the distribution of a few enzymes in the neoplastic liver of Uroloncha malabarica (Linnaeus)

The present work describes histological and histochemical observations made on the neoplastic liver of Indian silver bills, Uroloncha malabarica. The histology of neoplastic tissue as well as liver has been discussed. Further, a few enzymes like alkaline phosphatase, acid phosphatase, 5-nucleotides and non-specific esterase have been localized in the diseased liver. The occurrence of lymphocytoma caused a marked change in the localization of the enzymes. Sometimes total inhibition of the enzyme was encountered. Damaged sinusoid cells and bile canaliculi of the neoplasm as well as liver lobules show no reaction for alkaline phosphatase. However, its counterpart, acid phosphatase, exhibits intense activity in both neoplastic tissue and liver cells. Aggregates of neoplastic tissue give moderate 5-nucleotidase reaction while it gives poor activity in hepatic tissue of the diseased liver. Parenchymatous cells are able to give some activity for the non-specific esterase while it is very dull in the neoplastic tissue.     

Acid and alkaline phosphatases activities in vascular smooth muscle: species differences and subcellular distribution

1. Acid and alkaline phosphatase activities were studied in rat and dog aortic muscle using p-nitrophenyl phosphate (p-NPP) as the substrate. Alkaline phosphatase activity was quite comparable to acid phosphatase activity in rat aortic microsomes as well as further purified plasma membranes, but considerably lower than acid phosphatase activity in dog aortic membranes. 2. Subcellular distribution of acid and alkaline phosphatase activities in these vascular muscles indicated that alkaline phosphatases and a large portion of acid phosphatase activities were primarily associated with plasma membranes and the distribution of acid phosphatase showed little resemblance to that of N-acetyl-beta-glucosaminidase, a lysosomal marker enzyme. 3. The rat aortic plasmalemmal acid and alkaline phosphatase activities responded very differently to magnesium, fluoride, vanadate and EDTA. The alkaline phosphatase was more susceptible to heat inactivation than acid phosphatase. 4. These results suggest that these two phosphatases are likely to be two different enzymes in the smooth muscle plasma membranes. The implication of the present findings is discussed in relation to the alteration of these phosphatases in hypertensive vascular diseases.     

Extracellular matrix components and proteolytic enzymes in uterine cervical carcinoma

The important components of mucopolysaccharides and collagen have been analyzed in tissues of control and carcinoma of uterine cervix. Among these components hyaluronic acid and chondroitin sulphate levels were found to be increased, whereas decreased level of collagen was observed in uterine cervical carcinoma. Serum cathepsin B, D and acid and alkaline phosphatases have also been analyzed in controls and carcinoma patients before and after treatments. The activities of these enzymes have been found to increase prominently in advanced stages. Among these enzymes cathepsin B and alkaline phosphatase have exhibited remarkable increase in activity in uterine cervical carcinoma. Different modes of treatment exerted reversion of the elevated activities of these enzymes. However, combined therapy type II (radiation combined with cisplatin and cyclophosphomide) seems to be more effective in reverting the activities of these enzymes to normal levels.     

Specific interaction among some enzymes and sodium dodecyl sulfate

The effect of 1-butanesulfonic acid sodium salt and sodium dodecyl sulfate on the activity of highly purified and crystalline enzymes with marked differences in structure and function has been studied. The enzymes were: alcohol dehydrogenase; lactate dehydrogenase; malate dehydrogenase; isocitrate dehydrogenase; glucose-6-phosphate dehydrogenase; lipase; alkaline phosphatase. While 1-butanesulfonic acid sodium salt, at the studied concentrations, resulted generally inactive, sodium dedecyl sulfate showed a selective inhibitory effect, always under the critical micellar concentration. A kinetic analysis of the inhibitory action was also carried out.     

Production of phosphatates by fungi isolated from desert soils

Twelve fungal cultures isolated from Indian desert soils belonging toAspergillus, Penicillium, Acrophialophora andAlternaria were found to produce both acid and alkaline phosphatases in liquid medium, their amounts varying from culture to culture. Maximum production of these enzymes was observed withA. niger. In general, acid phosphatase activity was much higher as compared to alkaline phosphatase. The optimum incubation period for the production of these enzymes was found to be 14 d and thereafter it started declining. There was a significant and positive correlation between biomass production and acid phosphatase activity but not with alkaline phosphatase.     

Activity of phosphatases and succinic dehydrogenase in the trigeminal ganglion of the corticosteroid-treated mouse.

Immature mice were treated for 21 days with daily doses of triamcinolone diacetate. The trigeminal ganglion was studied enzymatically for the activity of alkaline phosphatase, acid phosphatase, ATPase and succinic dehydrogenase. It became apparent that the prolonged treatment with a potent corticosteroid hormone did not affect significantly the in vivo activity of the above enzymes. Some enzymes even appeared to have increased their intracellular activity. The heterogeneous effect of corticosteroid upon their peripheral target organs is discussed.     

A comparative biochemical research in the Schistocephalus solidus (Cestoda)--three-spined stickleback Gasterosteus aculeatus L. system

The activity of five lysosomal hydrolases (acid phosphatase, DNAase, RNAase, beta-glucosidase, beta-galactosidase), alkaline phosphatase and aldolase have been examined in tissues of the cestode Schistocephalus solidus (Müller, 1776) and the three-spined stickleback Gasterosteus aculeatus (L.) forming a stable parasite-host system. As a rule, the activity of enzymes was higher in a cestode body than in fish tissues. The acid and alkaline phosphatases were the exception. The activity and variation of lysosomal nucleases and aldolase in the parasite differed notably from those in both infested and healthy hosts. The paper discusses the role of lysosomal and cytoplasmic enzymes in a cestode adaptation to parasitism, as well as in the mechanisms of the host's chemical and immunological response to infection.     

两种鼠蚤在新羽化和吸血后不同时间三种酶的组织化学研究

采用组织化学技术,研究了不等单蚤Monopsyllus anisus (Rothschild)和缓慢细蚤Leptopsylla segnis (Schonherr)新羽化和吸血后24 h、48 h和72 h碱性磷酸酶、酸性磷酸酶和三磷酸腺苷酶的分布和活性。显微摄影及定量图像分析结果显示：新羽化蚤碱性和酸性磷酸酶主要存在于中肠、神经细胞核、精子束头部、精巢附腺、射精管、输卵管和受精囊附腺中;三磷酸腺苷酶各组织中均有分布。吸血消化后,两种蚤中肠3种酶活性均有增强;除碱性磷酸酶在消化72 h 后酶活性有所下降外,其余不同消化时间酶活性增强程度上不存在显著差异。两种蚤卵母细胞发育成熟过程中3种酶活性亦见增强。     

Enzyme activity during the growth and aging of human cells in vitro

Acid phosphatase, alkaline phosphatase, and lactic dehydrogenase activities have been compared in normal human diploid cell strains and in SV₄₀-transformed heteroploid cell lines derived from them. A higher level of acid phosphatase activity was observed in diploid cultures derived from adult lung than in cultures derived from fetal lung of similar passage levels. The alkaline phosphatase activity of normal diploid fibroblasts was significantly higher than that of SV₄₀-transformed cell lines derived from them. Generally, the lactic dehydrogenase activities of all these cell cultures were similar. Human diploid cells in culture “age,” in the sense that their ability to proliferate decreases with time during serial subcultivation. Evaluation of the activities of these three enzymes during the “aging” process showed that, although alkaline phosphatase and lactic dehydrogenase activities were similar in “young” and “senescent” cells, acid phosphatase showed a small but significant increase in the senescent cells.     

Acid and alkaline phosphatase activities in the clam Scrobicularia plana: kinetic characteristics and effects of heavy metals

The acid and alkaline phosphatase activities of the clam Scrobicularia plana have been partially characterised in different organs and tissues (digestive gland, gills, foot, siphon and mantle) and the 'in vitro' effect of heavy metals on both types of enzymatic activity have been analysed. The optimal pH ranged between 4.0 and 5.5 for acid phosphatase activity and 8.5 and 9.5 for alkaline phosphatase activity. The apparent optimum temperature was in the 30-60 degrees range for acid phosphatase activity and in the 30-40 degrees C range for alkaline phosphatase activity. The effect of substrate concentration on enzymatic activities in the tissues showed a good fit to the Michaelis-Menten model. For both types of enzymatic activity, the highest values were found in the digestive gland. The effect of heavy metals was dependent on the tissue analysed. Mercury showed the highest inhibition in the organs/tissues and the parameters Km and Vmax were modified when the inhibitor concentration increased, thus indicating a mixed type of inhibition.     

The enzyme histochemistry of developing odontoblasts in cattle, pigs and horses

Synopsis The histochemical distribution of some hydrolytic and oxidative enzymes in developing odontoblasts and subodontoblasts in cattle, pigs and horses has been observed in cryostat sections of teeth that have been decalcified with neutral EDTA.Undifferentiated dental epithelium and immature odontoblasts of the bell stage tooth germ showed lower levels of enzymatic activity as compared with the well-developed tooth germ.When the dentine matrix began to form, the young odontoblasts appeared to have a significantly positive reaction for acid phosphatase, and gradually other enzymes developed an activity at the top of the cusp.Odontoblasts as well as subodontoblastic-rich cells showed strong enzymatic activities for hydrolytic and oxidative enzymes, that is, they were strongly reactive for alkaline and acid phosphatase and lactate and malate dehydrogenases, and moderately reactive for other oxidative enzyme systems.It is suggested that the subodontoblastic layer is concerned with the biosynthesis of dentinal matrix as well as with the odontoblasts themselves.     

Alkaline phosphatases in fixed plant cells

Alkaline phosphatase activity in fixed plant cells has now been demonstrated cytochemically. Presumably cytochemical findings on plant alkaline phosphatases had been lacking because glycerophosphate, which is not hydrolyzed by fixed plant cells, had been used as the substrate.Alkaline phosphatase activity in the onion and corn nuclei has been compared with the activity in rat tissues. In the plant tissues, hydrolysis of phosphates was demonstrated when the substrates guanylic acid, adenosine diphosphate, adenosine triphosphoric acid, diphosphopyridine nucleotide, hexosediphosphates and inorganic pyrophosphate and metaphosphate were used. When the substrates glycerophosphate, adenylic acid and hexosemonophosphates were used, hydrolysis was not found. In the animal tissues however, hydrolysis was demonstrated of all organic phosphoesters employed and of sodium metaphosphate but not the hydrolysis of sodium pyrophosphate.One alkaline phosphatase found in the fixed plant tissues specifically hydrolyzed guanylic acid but no other nucleotide and one specifically hydrolyzed metaphosphate to orthophosphate.The enzymes in both plant and animal cells which hydrolyzed metaphosphates and pyrophosphates were found to require magnesium ions for their activity and to be inhibited by fluoride ions.“Alkaline, phosphatase,” so intimately associated with the chromatin in the nucleus, is postulated to be not just one enzyme but a number of enzymes.     

Resolution, purification and characterization of the orthophosphate releasing activities from fracture callus calcifying cartilage.

Callus calcifying cartilage alkaline phosphatase was resolved by DEAE-cellulose column chromatography into two distinct phsophatase activities. The phosphatase activity which was eluted first from the column, (phosphatase I), was active towards a variety of phosphate esters, sodium pyrophosphatase and several linear polyphosphates, while the second phosphatase activity , (phosphatase II), was active toward simple phosphate esters but not towards sodium pyrophosphate and linear oligo or polyphosphates. All the phosphate esters, sodium pyrophosphate and polyphosphates at higher concentrations were inhibitory for phosphatase I. The modulating effects of magnesium, calcium, zinc and other phosphatase modulators have been investigated. Both phosphatases from callus calcifying cartilage were found to be substrates of neuraminidase with sialic acid as the product. Besides the difference in their specificity, the phosphatases were found to be immunologically different and to have different molecular weights, strong indication that they are different enzymes.     

Alkaline phosphatase and 5'-nucleotide phosphodiesterase from bovine intestine are cross-reactive

Polyclonal antibodies to native alkaline phosphatase and to native 5'-nucleotide phosphodiesterase were found to strongly cross-react with both enzymes. The antibodies also cross-react with both denatured enzymes, with glycopeptides from 5'-nucleotide phosphodiesterase, and with the oligosaccharides remaining after Pronase E digestion of the phosphodiesterase. They do not cross-react with either enzyme after their oligosaccharides have been modified or removed by periodate or trifluoromethanesulfonic acid treatment. Antibodies to denatured 5'-nucleotide phosphodiesterase do not bind to the native phosphodiesterase or alkaline phosphatase but do cross-react with denatured alkaline phosphatase even after removal or modification of the carbohydrate moieties. These results suggest that antibodies to denatured 5'-nucleotide phosphodiesterase may recognize amino acid sequence homology between alkaline phosphatase and 5'-nucleotide phosphodiesterase. However, antibodies to native enzymes apparently recognize cross-reactive determinants of the native enzymes which are carbohydrate in nature. This is the first report of antimammalian alkaline phosphatase antibodies which recognize the carbohydrate moieties of the enzyme.     

Thymidine as synchronizing agent. 3. Persistence of cell cycle patterns of phosphatase activities and elevation of nuclease activity during inhibition of DNA synthesis

Synchronous cultures of HeLa cells were obtained by selective detachment of cells in mitosis and fluctuations in enzyme activity were followed during the subsequent cell cycle. The enzymes measured were alkaline and acid phosphatases and a nuclease active on denatured DNA at alkaline pH (alkaline DNase). Each of these enzymes showed a different pattern of activity in the cell cycle, but a temporal relationship to the DNA synthetic phase was apparent in each case. Treatment of the cultures at the beginning of the cell cycle with 15 mM thymidine did not alter the subsequent pattern of fluctuations in activity of alkaline phosphatase or of acid phosphatase, although DNA synthesis was fully inhibited by this treatment. This indicates that the pattern of activity of some enzymes is not linked to DNA replication. On the other hand, the pattern of fluctuations in the activity of alkaline DNase was abolished by thymidine treatment, and elevation of the activity of this enzyme was observed. These results suggest complex and variable relationships between phases of the cell cycle and enzyme activity, and show that inhibition of DNA synthesis is not a suitable procedure for induction of culture synchrony if enzyme activities are to be studied.     

Histochemical analysis of molluscan stomach and intestinal alkaline phosphatase: A sialoglycoprotein

Summary Though sialoprotein nature of alkaline phosphatase of certain mammalian organs has been suggested by biochemical investigations, no histochemical techniques have yet been applied to elucidate this concept. With this view, the alkaline phosphatase of stomach and intestine of a mollusc—Semperula maculata—was analysed histochemically to elucidate its sialoglycoprotein nature. The localisation of alkaline phosphatase and sialic acid was investigated by employing well known and standard histochemical techniques.Alkaline phosphatase was localised selectively in the brush border of the mucosa of stomach and intestine, it was Mg⁺⁺ nonsensitive but showed a structure-linked sensitivity to phenylalanine. The sialomucins were selectively localised in the brush border, whereas the goblet cells contained both the sialomucins and sulfomucins, and the connective tissue of lamina propria contained sulfomucins. The localisation of alkaline phosphatase and sialomucins in the brush border uniquely coincided with each other. The alkaline phosphatase activity in the brush border was completely lost after neuraminidase treatment at 37.5° C for 16 h. Such effect of neuraminidase on alkaline phosphatase activity was pH dependent and controlled by velocity of reaction. Heat-inactivated neuraminidase showed no effect on alkaline phosphatase activity.These histochemical results have been interpreted as suggesting a sialoglycoprotein nature of alkaline phosphatase in the brush border, and sialic acid somehow seems to be essential for enzyme activity. These results, thus, indicate necessity of visualising some of the sialo-glycoproteins as macromolecules with catalytic activity.     

Enzyme synthesis in the cellular slime mould Dictyostelium discoideum during the differentiation of myxamoebae grown axenically

1. Myxamoebae of the cellular slime mould Dictyostelium discoideum Ax-2 were grown on different media, and were harvested either in the stationary or exponential phases of the growth cycle to yield samples of myxamoebae differing in enzymic composition. 2. Morphogenesis and cell differentiation phenomena in D. discoideum appear to be similar in myxamoebae grown and harvested under different conditions. 3. The specific activity of the enzymes beta-N-acetylglucosaminidase, acid phosphatase, alpha-mannosidase, beta-glucosidase and alkaline phosphatase have been determined during cell differentiation of myxamoebae grown and harvested under different conditions. 4. The pattern of synthesis of these enzymes, all of which have been claimed to be part of the ;developmental programme', either remains unaffected despite the origin of the myxamoebae (alkaline phosphatase) or is qualitatively similar but quantitatively affected (acid phosphatase, beta-glucosidase) or is both qualitatively and quantitatively affected by changes in the myxamoebae (alpha-mannosidase, beta-N-acetylglucosaminidase). 5. The implications of these results for the concept of a ;developmental programme' are discussed.     

Association of Matrix Acid and Alkaline Phosphatases with Mineralization of Cartilage and Endochondral Bone

The activities of acid and alkaline phosphatases were localized by enzyme histochemistry in the chondroepiphyses of 5 week old rabbits. Using paraformaldehyde-lysine-periodate as fixative, the activity of acid phosphatase was particularly well preserved and could be demonstrated not only in osteoclasts, but also in chondrocytes as well as in the cartilage and early endochondral matrices. The acid phosphatase in the chondrocytes and the matrix was tartrate-resistant, but inhibited by 2mM sodium fluoride, whereas for osteoclasts 50–100mM sodium fluoride were required for inhibition. Simultaneous localisation of both acid and alkaline phosphatase activities was possible in tissue that had been fixed in 85% ethanol and processed immediately. In the growth plates of the secondary ossification centre and the physis, there was a sequential localisation of the two phosphatases associated with chondrocyte maturation. The matrix surrounding immature epiphyseal chondrocytes or resting/proliferating growth plate chondrocytes contained weak acid phosphatase activity. Maturing chondrocytes were positive for alkaline phosphatase which spread to the matrix in the pre-mineralising zone, in a pattern that was consistent with the known location of matrix vesicles. The region of strong alkaline phosphatase activity was the precise region where acid phosphatase activity was reduced. With the onset of cartilage calcification, alkaline phosphatase activity disappeared, but strong acid phosphatase activity was found in close association with the early mineral deposition. Acid phosphatase activity was also present in the matrix of the endochondral bone, but was only found in early spicules which had recently mineralised. The results suggest that alkaline phosphatase activity is required in preparation of mineralization, whereas acid phosphatase activity might have a contributory role during the early progression of mineral formation.     

A light- and electron-microscopic study of enzymes in the embryonic shell-forming tissue of the freshwater snail, Biomphalaria glabrata

Abstract. The mode of formation of the molluscan exoskeleton is still poorly understood, but studies on adult snails indicate that enzymes involved in vertebrate bone formation also participate in mollusc shell formation. The enzymes peroxidase, alkaline phosphatase, and acid phosphatase are expressed in a constant pattern and help to identify the different zones of the adult shell-forming tissue. The present study evaluates whether the expression of these enzymes is also a tool for the identification of the developing zones of the embryonic shell-forming tissue. Thus, we analyzed the temporal and spatial activity of the above-mentioned enzymes and of tartrate-resistant acid phosphatase in the shell forming tissues in Biomphalaria glabrata. Embryos of different age groups and adults were studied; alkaline phosphatase activity was seen in very young embryos in the shell field invagination prior to the secretion of any shell material, while peroxidase activity was present from the start of the periostracum production. Acid phosphatase, found in considerable amounts in yolk granules and albumen cells, appeared in the embryonic shell-forming tissue in relatively few Golgi stacks. Tartrate-resistant phosphatase was not present in embryos, but was found in adults in the same zone of the mantle edge as acid phosphatase. Using the enzymes as cell markers, the differentiation of the embryonic shell-forming tissue to the different zones of the adult mantle edge could clearly be followed.