Platelet-Activating Factor Modulates a Secreted Phosphatase Activity of the Trypanosomatid Parasite Herpetomonas muscarum muscarum

In the present work we characterized the secreted phosphatase activity of the trypanosomatid parasite Herpetomonas muscarum muscarum. This housefly parasite hydrolyzed p-nitrophenylphosphate at a rate of 10.26 nmol Pi/mg protein/min. Classical inhibitors of acid phosphatases, such as sodium orthovanadate (N_aVO₃), sodium fluoride (NaF), and ammonium molybdate promoted a decrease in this phosphatase activity. When the parasites were assayed in the presence of sodium tartrate, an inhibitor of Leishmania spp-secreted acid phosphatases, this activity was drastically diminished. Cytochemical analysis showed the localization of this enzyme on the external surface and in the flagellar pocket of these parasites. Sodium tartrate inhibited this reaction, confirming the biochemical data. Platelet-activating factor (PAF) inhibited the phosphatase activity determined in the supernatant of living H. m. muscarum. Received: 2 February 2001 / Accepted: 5 March 2001     

ENZYME DISTRIBUTION IN THE RAT FETUS AND PLACENTA FOLLOWING THE ADMINSTRATION OF ETHIONINE

Enzyme changes which accompany ethionine-induced resorption of the rat conceptus have been studied by both histochemical and biochemical techniques. Pregnant rats were injected with ethionine over a 3-day period prior to autopsy on day 12 of pregnancy. Sections of the whole conceptus were studied for acid phosphatase with both the Burstone and Gomori methods and for succinoxidase activity with nitro-BT. Biochemical determinations of cathepsins, acid phosphatase, and succinoxidase were performed on homogenates of the fetuses, placentae, and deciduas basalis from ethionine-treated and saline-treated rats. The histochemical study has shown that resorption is accompanied by an increase in the size and number of acid phosphatase granules in the decidual tissues and a concurrent loss of acid phosphatase granules in the fetal tissues. Biochemical methodology indicated that there was no increase in total cathepsin or acid phosphatase activities in the resorbing tissues. No change in succinoxidase activity was found with either histochemical or biochemical techniques. The significance of these results was discussed with reference to the lysosome hypothesis.     

Cytochemical studies on the localization of alkaline phosphatase and HCO-3-activated adenosine triphosphatase in the brush border membrane of rat duodenal enterocytes

Cytochemical techniques were used to demonstrate, with appropriate controls, alkaline phosphatase and HCO-3-activated adenosine triphosphatase (ATPase) in rat duodenal brush border microvillus membranes. Intense activity of ecto-alkaline phosphatase activity was demonstrated with 2-glycerophosphate as substrate. Although biochemical assays suggested that L-phenylalanine inhibited both alkaline phosphatase and HCO-3-activated ATPase, cytochemical studies indicated that there was marked inhibition of alkaline phosphatase revealing a specific HCO-3-activated ATPase on the inner aspect of the microvillus membrane. While it is tempting to suggest that this HCO-3-activated ATPase is implicated in active bicarbonate secretion by the duodenum, decisive identification is not yet possible.     

Localization of acid and alkaline phosphatases in Myxococcus coralloides D

Myxococcus coralloides produces two different phosphatases, one acid and the other alkaline. Both enzymes were localized by physical and biochemical techniques. Spheroplasts from M. coralloides released 20–30% of the phosphatase activities. Osmotic shock or treatment with high MgCl₂ or LiCl concentrations did not produce a greater release. Cytochemical localization situated the phosphatases in the outer membrane and the periplasmic space. Separation of the cytoplasmic membrane and outer membrane of the cells by sucrose gradient centrifugation showed that phosphatases are located primarily in the outer membrane. membrane.     

Cytochemical Localization of Certain Phosphatases in Escherichia coli

Cytochemical studies of Escherichia coli at the light and electron microscopic levels have revealed alkaline phosphatase, hexose monophosphatase, and cyclic phosphodiesterase reaction products in the periplasmic space and at the cell surface. In preparations for both light and electron microscopy, reaction product filled polar caplike enlargements of the periplasmic space, such as those described in plasmolyzed cells, indicating significant terminal concentrations of these enzymes; dense substance was often seen within these polar caps in morphological specimens. Staining of the bacterial surface was commonly encountered, but could represent artifactual accumulation of precipitate along the cell wall. Alkaline phosphatase was demonstrated with several substrates (ethanolamine phosphate, glycerophosphate, p-nitrophenylphosphate, and glucose-6-phosphate) over a wide pH range in a bacterial strain (C-90) known to be constitutive for this enzyme, whereas strains deficient in this enzyme (U-7, repressed K-37), showed no activity with these substrates. Hexose monophosphatase and cyclic phosphodiesterase activities were characterized by reaction-product deposition with specific substrates at acid or neutral, but not at alkaline, pH in strains of E. coli lacking alkaline phosphatase (U-7 and repressed K-37). Fixation in Formalin or the use of calcium as a capture reagent seemed to interfere with periplasmic staining in cells prepared for electron microscopy. Formalin fixation had little effect on biochemical assays of the phosphatase activity of intact cells in suspension, but partially reduced the activity evident in sonically treated extracts or in suspensions of dispersed cryostat sections. Glutaraldehyde treatment impaired enzyme activity more drastically.     

Cytochemical studies on alkaline phosphatase production during sporulation in Bacillus subtilis.

During sporulation of Bacillus subtilis 168 there is an increase in activity of alkaline phosphatase in the presence of Pi. This enzyme was shown by cytochemical techniques to be associated with the cytoplasmic membrane of the mother cell and also with the membranes of the developing prespore. There is a strong correlation between an increasing number of electron-dense deposits due to phosphatase activity and the formation of the spore septum, i.e. stage II of sporulation. Cytochemical and biochemical evidence shows that cells well advanced in spore formation can be derepressed to produce the very much higher amounts of alkaline phosphatase characteristic of phosphate-starved vegetative cells.     

Histochemical studies on reserve substances and enzymes in female gametophyte of Zea mays.

Cytochemical changes during the early development of maize caryopsis are reported. Changes in the localization of different reserve substances (e.g. polysaccharides, proteins, nucleic acids and lipids) and enzymes (acid phosphatase, esterase, lipase, phosphorylase, succinate dehydrogenase, cytochrome oxidase and peroxidase) have been studied in unfertilized and fertilized ovules. Before pollination very feeble enzyme activity (acid phosphatase, succinate dehydrogenase, cytochrome oxidase and peroxidase) was observed. Reserve substances were present in low amounts before pollination. Pollination stimulated the accumulation of several substances and enzymes in the tip of the nucellus, micropylar zone. Just prior to, during and after fertilization, the cells in the micropylar zone had strong reaction for several enzymes indicating temporary enhancement of metabolic activity in the micropylar zone. The role of antipodals in the storage of reserve food products and nutrition of embryo and early stages of endosperm development is discussed. The pattern of enzymatic changes within the embryo sac reflected the biochemical changes operative during quiescent and active stages. The nucellus of Zea mays contains many enzymes required for hydrolysis of reserved food substances. A role of acid phosphatase in autolysis of nucellar cells, after fertilization is suggested. Post-fertilization increase in the activity of enzymes and accumulation of reserve materials is interpreted as reflecting a presumed increase in the metabolic rate relative to growth and differentiation.     

Lipid mobilization and acid phosphatase activity in lytic compartments during conidium dormancy and appressorium formation of Colletotrichum graminicola

Colletotrichum graminicola, a pathogen of sorghum and corn, was investigated prior and during germination as to certain aspects of acid phosphatase activity and lipid mobilization. Ungerminated conidia cytoplasm was filled with lipid deposits, which were mobilized during the germination process. Cytochemical ultrastructural examination showed that conidia vacuoles exhibit acid phosphatase activity, which is suggestive of lytic activity. Lipid bodies, stored in the ungerminated conidia cytoplasm, were internalized by vacuoles in a process analogous to microautophagy and were apparently digested inside them. The lipid bodies disappeared and vacuoles became enlarged in conidial cells during germination. Appressoria also showed acid phosphatase activity in multiple heterogeneous vesicles which were, in most cases, juxtaposed with lipid bodies. These results suggest that the vacuolar system plays an important role during C. graminicola germination and that the initial stages of lipid metabolization are taking place inside the vacuoles.     

THE FATE OF DNA-CONTAINING PARTICLES PHAGOCYTIZED BY MAMMALIAN CELLS

Particulate DNA protein coacervates were digested immediately after being phagocytized by L strain fibroblasts in suspension culture. Enlargement of the phagocytotic vacuoles occurred simultaneously with a loss of the electron opacity of the phagocytized particles. Cytochemical reactions positive for non-specific esterase, acid phosphatase, and nucleoside phosphatase in the phagocytotic vacuoles provided additional evidence for the probability of complete hydrolysis of the phagocytized nucleoprotein.     

Epithelial differentiation in the fetal rat colon: I. Plasma membrane phosphatase activities

During the last week of gestation of the fetal rat, the epithelium of the colon is rapidly remodeled. At 16 days a primitive stratified epithelium surrounds a small central lumen. Over the next 3 days, the main lumen extends narrow clefts down to the basal cell layer and small secondary lumina appear within the stratified epithelium between these clefts. At 19 and 20 days, secondary lumina enlarge but remain discrete; an infusion of cationic ferritin into the main lumen does not enter secondary lumina. During the 2 days prior to birth (21–22), the secondary lumina join the main lumen as superficial cells are sloughed, and the epithelium becomes simple columnar. Freeze-fracture replicas indicate that luminal and nonluminal membrane domains of epithelial cell plasma membranes are separated by continuous tight junctions throughout the conversion process. Cytochemical analysis of tissue slices from 16- to 22-day fetal colon demonstrated the appearance and segregation of two phosphatases on apical and basolateral membrane domains during epithelial conversion. Cysteine-sensitive pH 9.0 (alkaline) phosphatase activity was first detected along the luminal membranes of cells bordering both primary and secondary lumina at 18 days gestation and increased to a maximum at 20–21 days; weaker activity was present on basolateral membranes. Phosphatase activity at pH 8.0 also appeared at 18 days and increased thereafter, but was localized primarily on nonluminal membranes. At pH 8.0, reaction product appeared on both inner and outer sides of the membrane, and was only partially abolished by omission of K⁺ or addition of ouabain; thus the reaction may be only partially due to K⁺-dependent ATPase activity. Biochemical analysis of the cytochemical media confirmed the appearance of phosphatase activities at 18 days. Thus, plasma membrane phosphatase activities appear while the epithelium is still stratified, but are segregated to luminal and nonluminal membrane domains at the onset of activity. Segregation is maintained throughout the process of conversion of a simple columnar epithelium.     

Histochemical studies of developing human fetal small intestine

Summary The histochemical characteristics of absorptive and mucus-producing cells have been described in 16 specimens of small intestine from human fetuses between 7 and 22 weeks of age. In the youngest specimens a slight to moderate reaction was found for all enzymes examined which included those known from biochemical or ultrastructural studies to be located predominantly in Golgi apparatus (thiamine pyrophosphatase), surface membrane (alkaline phosphatase, adenosine triphosphatase, leucine aminopeptidase), lysosomes (acid phosphatase), and mitochondria (succinic dehydrogenase). Mucus granules of somehat unusual location and staining properties were also found at this time. A progressive intensification of a majority of these histochemical reactions was observed between 3 and 6 fetal months.These findings indicate that the histochemical apparatus for degradation and absorption of nutrients and for elaboration of protein and mucus is already well established in the human fetus by the end of this 6 month period. It is not known, however, whether these activities occur in utero or whether they contribute significantly to fetal nutrition.     

百合花粉母细胞间染色质穿壁运动前次生胞间连丝形成的研究

百合花粉母细胞间染色质穿壁运动前(细线期到偶线期)的花药,用一般电镜制片法和铅沉淀法对酸性磷酸酶活性的细胞化学反应产物的定位实验,其结果总结如下:(1)形成次生胞间连丝通道水解作用所需的酶可能是由“类溶酶体”小泡或由内质网腔直接分泌的;(2)次生胞间连丝通道的水解作用,可在细胞壁的两边细胞同时开始,先形成半胞间连丝,然后贯穿??在一起;或从一侧开始,一直穿孔到另一边,最后两者都能形成胞间连丝;(3)用铅沉淀法进行的酸性磷酸酶细胞化学的定位实验表明:在质膜、内质网、类溶酶体小泡中的酶活性反应产物沉积的部位与一般电镜法制备的切片上看到的电子致密度物质的分布情况完全一致,(4)用X-射线微区能谱分析的结果表明:沉淀物中含有铅元素,确实是磷酸铅。因此我们推测所谓“类溶酶体”以及内质网所分泌的水解酶,可能具有果胶酶、纤维素酶和半纤维素酶的性质,它们都能降解、穿孔各自的细胞壁形成胞间连丝。     

Studies on secretory activity in the pars intermedia of Xenopus laevis 2: A biochemical and electron cytochemical investigation of acid hydrolase activity following the stimulation of secretory activity in vivo

In the MSH cell at the onset of secretory activity, acid hydrolase activity increases. This increased activity, shown quantitatively by assaying beta-glycerophosphatase and R-glucuronidase within the stimulated gland, has been shown by electron cytochemical methods for beta-glycerophosphatase (acid phosphatase) and aryl sulphatase to be related to the production of large numbers of dense bodies. Cytochemical evidence also supports the view that these lytic bodies arise from GERL-like cisternal elements since it is shown that in addition to the flattened, parallel Golgi cisternae these elements are also R-glycerophosphatase-positive. The similarities between the dense bodies and those of other cell types are described and discussed.     

Activity of acid and alkaline phosphatase in neutrophils of rats exposed to benzene and treated with selenium.

Cytochemical studies on activity of acid phosphatase (AcP) and alkaline phosphatase (AP) in peripheral blood neutrophils of rats chronically exposed to benzene vapours (1,200 mg/cm3) demonstrated that the exposure results in the increase of AcP and the decrease of the AP activity. The changes noted correlate with the time of exposure and are observed even after 5 months after exposure. The administration of sodium selenate in a dose of 1.0 microgram/kg before exposure prevented the above enzymatic alterations caused by benzene. In contrast, the administration of selenium in a dose of 5.0 microgram/kg only prevented the change of AP activity and caused reactive neutrophilic leukocytosis.     

Phosphatase activity of blood leukocytes and wound exudate during the experimental healing of an aseptic wound

Cytochemical investigations of plain aseptic wounds simulated in 110 Wistar rats revealed a clear-cut dependence between the variations in the activity of alkaline and acid phosphatases in neutrophilic leukocytes, monocytes and lymphocytes of the blood and wound exudate and the stage of the healing process. Elevated activity of the blood alkaline phosphatase correlated with the term of inflammation phase.     

GROWTH AND DIFFERENTIATION OF NEUROSPORA HYPHAE

Zalokar , Marko . (Yale U., New Haven, Conn.) Growth and differentiation of Neurospora hyphae. Amer. Jour. Bot. 46(8): 602–610. Illus. 1959.—Neurospora hyphae grow only at their tips. New protoplasm is formed throughout the hyphae and transported to the tips by active cytoplasmic streaming. This enables the fungus to grow at a prodigious rate. The use of radioactive precursors demonstrated that the rate of protein and ribonucleic acid formation does not decrease substantially in hyphal regions distant from the growing point. Apical growth of hyphae results in a morphological differentiation into young regions filled with cytoplasm and dense with nuclei and old regions, vacuolated and laden with fat droplets. Cytochemical methods were used to study the biochemical differentiation of hyphae and attention was focused on the growing region. It was found that hyphal tips, i.e., the first 50–100 μ of the growing end of the hypha, differed substantially from the rest of the young hyphae. Hyphal tips were slightly richer in proteins and significantly richer in protein-bound sulfhydryl groups and ribonucleic acid. Glycogen was absent at the tips. Peroxidase increased at the tips, while cytochrome oxidase and succinic dehydrogenase decreased. Alkaline phosphatase exhibited strong activity in a short region behind the extreme tip. β-galactosidase, in a few cases, was more active at the tips than in the rest of the hyphae. The results of cytological and cytochemical investigations are discussed in connection with the problem of cellular differentiation.     

Cytochemical studies of the types and localization of acid phosphatases in the various regions of the midgut epithelium of Carausius morosus

Synopsis Cytochemical studies on the localization and substrate specificities of acid phosphatase activities in the epithelial cells of the midgut ofCarausius morosus have revealed the presence of two distinct types of phosphatases. Acid naphthol AS-BI phosphatase activity was present at particulate (lysosomal) sites in all regions of the midgut and its activity was particularly high in the pear-shaped organs. Acid -glycerophosphatase of low activity was present in the mid and posterior midgut regions, but was absent from the pearshaped organs. In the anterior region of the midgut, acid -glycerophosphatase activity could only be found associated with the concentrically laminated vesicles.     

Morphocytochemical and electron microscopic research on murine hybridoma cells producing and not producing monoclonal antibodies

Cytochemical and ultrastructural features of mouse hybridomas and also of the parental cells--myeloma P3-X63-Ag8.653 and spleen cells of the Balb/c mice immunized with cell line RPMI-1788 have been studied. Differences in cytomorphological signs and activity of acid phosphatase, acid nonspecific esterase, nonspecific-alpha-naphthyl acetate esterase were shown in hybrid cell lines secreting and not secreting monoclonal antibodies.     

Cytochemical localization of tartrate-resistant acid phosphatase, alkaline phosphatase, and nonspecific esterase in perivascular cells of cartilage canals in the developing mouse epiphysis

Cytochemical localization of tartrate-resistant acid phosphatase (TRAP), tartrate-sensitive acid phosphatases (TSAP), alkaline phosphatase, and nonspecific esterase was used to characterize perivascular cells within cartilage canals. In the distal femoral epiphyses of 5- to 7-day-old mice, three stages of canal development can be distinguished, and at each developmental stage different perivascular cells were present with morphological characteristics of degradative cells. Vacuolated cells resembling macrophages, fibroblastic cells, and chondroclasts were present adjacent to the matrix in superficial, intermediate, and deep canals, respectively. In order to characterize these perivascular cells cytochemically, nonspecific esterase and TSAP staining was used to identify macrophages, alkaline phosphatase staining was used to identify fibroblastic cells, and TRAP staining was used to identify chondroclasts. There were no cells present in the canals at any developmental stage that were positive for TSAP or strongly positive for nonspecific esterase, placing doubt on the identity of the vacuolated cells as macrophages. Alkaline phosphatase-positive perivascular cells were present in the intermediate and deep canals adjacent to matrix containing alkaline phosphatase-positive chondrocytes. These alkaline phosphatase-positive cells were found in the same location within canals as the fibroblastic cells. Tartrate-resistant acid phosphatase was localized in chondroclasts at the tips of deep canals but was not confined exclusively to chondroclasts. Except for the very early stage of canal development prior to chondrocyte hypertrophy, TRAP-positive cells were present at the tips of superficial and intermediate canals as well as at the tips of the deep canals.(ABSTRACT TRUNCATED AT 250 WORDS)     

Acute lymphoblastic leukemia hand-mirror cells with OKT 8 phenotype

Acute lymphoblastic leukemia with hand-mirror cells (ALL-HMC) was diagnosed in a 20-year old patient. Cytochemical investigations revealed a positive reaction for PAS and acid phosphatase. Lymphoid blast cells were studied with various monoclonal antibodies in order to determine their derivation and differentiation. The data obtained (positivity for Leu 9, OKT 11 and OKT 8) suggest that blast cells were of T lineage with OKT 8 phenotype. This report supports the phenotypic heterogeneity of ALL-HMC.