Determination of Stelar Elements in Roots of Pisum sativum L.

Cytochemical studies of esterase activity in 0.5 mm segmentsfrom root tips of Pisum sativum explanted for up to 9 days inbasal culture medium containing 2 per cent sucrose showed retentionof this activity. During this time, all segments from the secondand third 0.5 mm segments of the root tip developed xylem elementsas did the proximal end of the first segment. No xylem elementswere found in the 12–14 cells behind the quiescent centre.It is concluded that the central group of meristem cells aregenerally programmed to form tissues of the stele immediatelyon leaving the quiescent centre, and that the programming forxylem and phloem elements occurs as a second step. Pisum sativum L., garden pea, determination, histochemistry, esterases, stele, root     

Identification of Shoot Apical Meristem Cells Committed to Form Vascular Elements in Pisum sativum L. and Vicia faba L.

A cytochemical study of naphthol AS-D esterases in vegetativeshoot apices of Pisum sativum and Vicia faba L. has shown thepresence of carboxyl esterases (E.C. 3.1.1.1 [EC] .) in those meristemcells already committed to form vascular elements. These cellsform a sequence linking the morphologically identifiable procambiumto the cells of the tunica layers at a site either already identifiableas the next primordium or which will form the next primordium.The implications of this result are briefly discussed in relationto the control of primordia formation and procambial cell development. Pisum sativum, Vicia faba, determination, vascular tissue, shoot apex, cytochemistry     

An Early Cytochemical Marker of Commitment to Stelar Differentiation in Meristems from Dicotyledonous Plants

A cytochemical study of root apices from Vicia faba and Pisumsativum showed esterase activity to be present in the stele,root cap and rhizodermis, but almost completely absent fromthe developing cortex and quiescent centres. The meristem cellsgiving rise to the cortex were almost negative whilst thosegiving rise to the stele were positive for esterase activity.Cambia from roots, shoots and petioles of a number of dicotyledonousspecies were all positive for esterase activity. It is proposedthat esterase activity may be used as an early marker of commitmentto differentiation into stele in roots of dicotyledonous plants,and that the cambia are fully committed meristems. Pisum sativum L., Vicia fabaL., garden pea, broad bean, meristems, stelar differentiation, esterase activity, xylem differentiation, cytochemistry, cambium     

Nature of human spleen red pulp cells with special reference to sinus lining cells

Summary A histological and cytological as well as enzyme histo- and cytochemical analysis (alpha-naphthyl acetate esterases, naphthol AS acetate esterases, naphthol AS-D chloroacetate esterases, acid and alkaline phosphatases) of human spleen cells in sections and imprints was carried out with special reference to sinus lining cells. These cells show strong naphthol AS esterase activity and no or only little alpha-naphthyl acetate esterase and acid phosphatase activity. Thus they can be distinguished from reticular cells in pulp cords and from other macrophages in cords and sinuses. From the morphological and enzyme histochemical aspect it can be deduced that the sinus lining cells are a distinct cell type of the human spleen. The comparison of these enzyme cytochemical findings with the results of biochemical and electron microscopical investigations suggests that reticular cells of pulp cords and littoral cells of sinuses also have different functions: reticular cells seem to have a high phagocytotic activity while littoral cells seem to be only facultatively phagocytic.This work was supported by the Deutsche Forschungsgemeinschaft.     

Activation of carboxylesterases in root cortical parenchymal cells of Pisum sativum during xylem induction in vitro

A quantitative cytochemical study of naphthol AS-D esterase activity in explants from roots of Pisum sativum grown on basal medium for 3 or 6 days showed similar levels of activity to those seen in sections of cortex and stele from intact roots of similar ages. Explants grown in the presence of auxins or cytokinin alone showed a threefold or twofold increase in cortical parenchymal activity, respectively. On adding both hormones to initiate xylem element formation, there was also a threefold increase in activity in the cortex. In all three cases, the stimulated activity was totally inhibited by either 10(-4) M diisopropylfluorophosphate (DFP) or 10(-4) M diethyl p-nitrophenylphosphate (E600), indicating carboxylesterase activity. The low level of activity normally present in cortical cells was inhibitor resistant, so indicating acetylesterase activity. Thus, auxins and cytokinins appear to activate mainly similar carboxylesterases during the initiation of xylem elements from cortical parenchyma cells.     

Localization of Acid Hydrolase Activity in Zea mays L. Root Tips

Naphthol AS-BI phosphatase, esterase, aryl sulphatase, glucuronidase,and ß-glycerophosphatase have been studied in frozensections of maize root tips. In general these enzymes showedhighest activities at the root surface and at particulate sitesin the cytoplasm although the indigogenic method for esteraseshowed no particulate activity and the naphthol AS-D acetatereaction gave no pronounced surface activity. With electronmicroscopy highest activity for ß-glycerophosphatasewas observed in the cell walls and associated with the vacuoles.The significance of these observations are discussed in relationto the function of surface hydrolytic activity and to the presenceof lysosome-like bodies in higher plant cells.     

Simultaneous Demonstration of Nonspecific Esterase and Chloroacetate Esterase in Human Blood Cells

A new cytochemical method is described for the simultaneous demonstration of nonspecific esterase in monocytes and chloracetate esterase in granulocytes. The procedure uses both alpha-naphthyl butyrate and naphthol AS-D chloroacetate as substrates and hexazotized pararosaniline as the coupler. The enzyme reaction products are highly chromogenic and their localization is precise. This method is potentially useful for the accurate diagnosis of the acute monocytic leukemias. Its advantages and limitations are also discussed.     

Simultaneous demonstration of nonspecific esterase and chloroacetate esterase in human blood cells

A new cytochemical method is described for the simultaneous demonstration of nonspecific esterase in monocytes and chloracetate esterase in granulocytes. The procedure uses both alpha-naphthyl butyrate and naphthol AS-D chloroacetate as substrates and hexazotized pararosaniline as the coupler. The enzyme reaction products are highly chromogenic and their localization is precise. This method is potentially useful for the accurate diagnosis of the acute monocytic leukemias. Its advantages and limitations are also discussed.     

Acid hydrolases in HeLa cells: comparison of methods for light microscopy

To distinguish lysosome populations of HeLa cells, acid phosphatase, beta-glucuronidase, arylsulfatase and esterase were demonstrated using various substrates and couplers with different fixations, pHs and inhibitors. The substrates chosen were for acid phosphatase, naphthol AS-BI phosphate with fast red violet LB at pH 4.6; for beta-glucuronidase, naphthol AS-BI beta-D-glucuronide with fast red violet LB at pH 4.4; for arylsulfatase, p-nitrocatechol sulfate, with lead as the capturing ion, at pH 4.8 and 5.6; and for esterase, naphthol AS-D acetate with fast blue BB at pH 6.5. In the azo-dye methods, the coupling was always simultaneous and results were satisfactory with unfixed cells. For optimal demonstration of arylsulfatase, cells were fixed in glutaraldehyde in 0.1 M cacodylate buffer pH 7.2, 2% for 24 hr or 6.25% for 2 hr, and washed for 1-9 days in 0.1 M veronal acetate buffer pH 7.2, 7.5% with respect to sucrose. Two groups of lysosomes were distinguished. One comprised small bodies, probably primary lysosomes, which lay in a cluster near the nucleus. They had quite stable membranes and were mostly acid phosphatase-positive. They sometimes contained beta-glucuronidase or esterase, but rarely arylsulfatase. The other group included all the acid hydrolase-positive bodies scattered throughout the rest of the cytoplasm. They were mostly larger, with more labile membranes, and contained beta-glucuronidase, esterase or arylsulfatase, but rarely acid phosphatase.     

A biochemical study of non-specific esterases from plant cells, employing the histochemical substrate, naphthol AS-D acetate

Synopsis The reagents routinely employed in the histochemical detection of nonspecific esterases, namely naphthol AS-D acetate and Fast Red Violet LB salt, have been used to study these enzymes biochemically with spectrophotometric procedures. A range of parameters that affect the coupling of the hydrolyzed substrate and diazonium salt were examined and their relevance to future histochemical procedures is noted.TheK _m of broad-bean root-tip esterases was estimated to be approximately 0.07 mM, but other kinetic data suggest that the true value is lower. From an analysis of the kinetics of the hydrolytic reaction it appears that they are of a mixed nature over the time course and substrate concentrations used.The effect of pH on root tip esterase activity has been examined and the recorded optimum of 5.5 is similar to that reported by other workers for plant cells. Non-enzymic hydrolysis of the substrate at alkaline pH levels prevented measurement of enzyme activity beyond pH 7.2.Magnesium ions at a final concentration of 5 mM are important in retaining esterases in their natured state during enzyme extraction, although the addition of increasing amounts of the ion to the assay system caused a corresponding increase in esterase inhibition.     

Cooperative effects of gamma-interferon and 1 alpha, 25-dihydroxyvitamin D3 on in vitro differentiation of the blast cells of RAEB and RAEB-T

We added recombinant human gamma-interferon (gamma-IFN) and 1 alpha, 25-dihydroxyvitamin D3 (1 alpha, 25 (OH)2D3) to the bone marrow cells from six patients with RAEB or RAEB-T in liquid suspension cultures. After cultivation for 7 to 9 days, numerical, morphological and functional changes of the cells were assessed. gamma-IFN and 1 alpha, 25 (OH)2D3 additively suppressed cell growth, especially the number of blast cells decreased. The expression of alpha-naphthylbutyrate esterase (NBE) activity appeared to be promoted but that of naphthol AS-D chloroacetate esterase (NAE) activity was apparently suppressed by the addition of gamma-IFN and/or 1 alpha, 25 (OH)2D3. The percentage of NBT reduction-positive cells and latex-phagocytizing cells was only slightly increased by both agents. These results indicate that gamma-IFN and 1 alpha, 25 (OH)2D3 cooperate to induce monocytoid differentiation of the patients' blast cells. Combination therapy with both agents merits further study.     

Genetically defined lysosomal acetylesterase EC 3.1.1.6 in the cauda epididymidis of mouse,rat, and man

After inhibition by bis-p-nitrophenyl phosphate and subsequent staining for esterase using naphthol AS-D acetate as the substrate, a strong lysosomal esterase was demonstrated in the cauda epididymidis of mouse, rat, and man. Owing to its behaviour towards the classifying inhibitors eserine, diisopropyl fluorophosphate, bis-p-nitrophenyl phosphate, and p-chloromercuriphenylsulphonate, this lysosomal esterase was shown to be an acetylesterase (EC 3.1.1.6). Control experiments involving isoelectric focusing revealed that this acetylesterase was identical with the genetically defined homologues ES-17, ES-6, and ES-A4 in mouse, rat, and man, respectively.     

Acid Hydrolases in Hela Cells: Comparison of Methods for Light Microscopy

TO distinguish lysosome populations of HeLa cells, acid phosphatase, /8-glu-curonidase, arylsulfatase and esterase were demonstrated using various substrates and couplers with different fixations, pHs and inhibitors. The substrates chosen were for acid phosphatase, naphthol AS-BI phosphate with fast red violet LB at pH 4.6; for β-glucuronidase, naphthol AS-BI β-D-glucuronide with fast red violet LB at pH 4.4; for arylsulfatase, p-nitrocatechol sulfate, with lead as the capturing ion, at pH 4.8 and 5.6; and for esterase, naphthol AS-D acetate with fast blue BB at pH 6.5. In the azo-dye methods, the coupling was always simultaneous and results were satisfactory with unfixed cells. For optimal demonstration of arylsulfatase, cells were fixed in glutaraldehyde in 0.1 M cacodylate buffer pH 7.2, 2% for 24 hr or 6.25% for 2 hr, and washed for 1-9 days in 0.1 M veronal acetate buffer pH 7.2, 7.5% with respect to sucrose. Two groups of lysosomes were distinguished. One comprised small bodies, probably primary lysosomes, which lay in a cluster near the nucleus. They had quite stable membranes and were mostly acid phosphatase-positive. They sometimes contained β-glucuronidase or esterase, but rarely arylsulfatase. The other group included all the acid hydrolase-positive bodies scattered throughout the rest of the cytoplasm. They were mostly larger, with more labile membranes, and contained β-glucuronidase, esterase or arylsulfatase, but rarely acid phosphatase.     

Experimental control of the activity of the quiescent centre in excised root tips of Zea mays

Summary Quiescent centres have been demonstrated in cultured excised root tips of both Pisum sativum and Zea mays. Upon addition of sucrose to Zea roots which have been deprived of carbohydrate, the cells of the quiescent zone as well as those of the rest of the meristem undergo DNA synthesis. Following the onset of proliferative activity in the meristem, DNA synthesis in the quiescent-centre cells is again arrested. It is suggested that the dividing cells of the meristem are responsible for the maintenance of the quiescent centre. It has also been shown that DNA-synthesising cells do occur within the quiescent centre and that they appear to be localised in specific regions.     

Effect of Chlorpromazine on Human and Murine Intracellular Carboxylesterases

Clinical use of chlorpromazine (CPZ), an antipsychotic drug, is limited due to its hepatotoxicity. CPZ is found to inhibit in vitro intracellular carboxylesterases (CE), such as alpha-naphthyl acetate esterase, naphthol AS-D chloroacetate esterase, and alpha-naphthyl butyrate esterase in polymorphonuclear neutrophils, hepatocytes, and neuronal brain cells from mice. CPZ inhibits CE of all these cell types, whereby the degree of the inhibition depends on the incubation time and CPZ concentration. The polymorphonuclear neutrophils are most sensitive to CPZ. Comparable results were obtained with polymorphonuclear neutrophils from mice and humans. Since leukocytes are much more available than hepatocytes or neuronal cells in humans, we assume that CE in peripheral blood leukocytes (neutrophils and monocytes) can be used as markers for indication of pending liver damage by CPZ.     

Genetically defined lysosomal acetylesterase EC 3.1.1.6 in the cauda epididymidis of mouse, rat, and man.

After inhibition by bis-p-nitrophenyl phosphate and subsequent staining for esterase using naphthol AS-D acetate as the substrate, a strong lysosomal esterase was demonstrated in the cauda epididymidis of mouse, rat, and man. Owing to its behaviour towards the classifying inhibitors eserine, diisopropyl fluorophosphate, bis-p-nitrophenyl phosphate, and p-chloromercuriphenylsulphonate, this lysosomal esterase was shown to be an acetylesterase (EC 3.1.1.6). Control experiments involving isoelectric focusing revealed that this acetylesterase was identical with the genetically defined homologues ES-17, ES-6, and ES-A4 in mouse, rat, and man, respectively.     

The Response of the Primary Root Meristem of Zea mays L. to Various Periods of Cold

Primary roots of maize (Zea mays L.) grown in nutrient solutionat 5?C elongate at about 1% of the rate found at 20?C. The apicalmeristem becomes shorter and shows little proliferative activityat 5?C, but following transfer to 20?C mitoses increase in frequencyand the meristem regrows to its original length. Both the amountby which the meristem shortens and the time for its completeregrowth are related to the period spent at 5?C. The shorteningof the meristem suggests that at the lower temperature the balancewhich normally exists between cell production and differentiationis altered, the latter continuing at a relatively faster ratethan the former. A new, steady-state balance between the twoprocesses is re-established during the recovery period. Themeristem recovers as a result not only of its own mitotic activitybut also through stimulation of cell division in the quiescentcentre. The degree to which the quiescent centre is activated,as judged by its mitotic index and the number of nuclei labelledby feeding with tritiated thymidine, increases as the durationof the preceding cold treatment increases. The close relationshipbetween proliferative activity in the quiescent centre and theminimum length of the meristem following the cold treatmentsuggests that there is communication between these two zoneswhich co-ordinates their respective rates of cell productionand helps to maintain a normal meristem structure. The resultsemphasize the importance of the quiescent centre as a reservoirof cells that can re-establish a meristem rendered non-functionalthrough the impact of unfavourable environmental conditions. Key words: Meristem, quiescent centre, root, temperature, Zea mays     

The enzyme cytochemistry and composition of elasmobranch granulocytes

Peroxidase, alkaline phosphatase, acid phosphatase, β-glucuronidase, α-naphthyl acetate esterase (ANAE), α-naphthyl butyrate esterase, naphthol AS-D chloroacetate esterase, acetyl-L-tyrosine-α-naphthyl esterase (ATNE), tosyl-L-lysine-α-naphthyl esterase (TLNE) and periodic acid-Schiff (PAS) were studied in 17 species of elasmobranchs in which granulocytes had previously been identified at the ultrastructural level.
Eosinophils, eosinophilic and neutrophilic granulocytes contained variable acid phosphatase, esterases and PAS, but they were strongest in neutrophilic granulocytes; particularly ANAE. Esterases were released into surrounding plasma and therefore probably function as ectoenzymes. In eosinophils and some neutrophilic granulocytes there were indications of weak peroxidase, but this could not be conclusively demonstrated. Alkaline phosphatase was diffuse between granules in some eosinophils of Pavoraja , and (β-glucuronidase was diffuse in neutrophilic granulocytes of Etmopterus baxteri , otherwise granulocytes lacked these enzymes. Neutrophilic granulocytes stained moderately to strongly for ATNE and weakly and inconsistently for TLNE in Squalus acanthias and Dalatias licha . with a similar reaction in granular lymphocytoid and thrombocytoid cells of Galeorhinus ausiralis and Raja nasuta . The enzyme composition of these granulocytes is discussed.     

Regeneration of the Root Cap of Zea mays L. and Pisum sativum L.: A Study with the Scanning Electron Microscope

Removal of the root cap from a root apex initiates regenerationof a new cap. The process has been followed using scanning electronmicroscopy. Quantitative data have been obtained for the growthin area of the exposed acroscopic surface of the quiescent centre(QC) and the increase in volume of the regenerating cap tissue.In Zea the surface of the QC shows an initial rapid increasein area followed by a slower increase. In Pisum the surfacearea increases uniformly, a rapid initial phase being absent.Together with observations on the behaviour of an incision atthe exposed surface, the results indicate that in Zea the capnormally imposes a constraint upon radial growth at the acroscopicsurface of the QC; in Pisum the QC appears not to be so constrained.The different responses may be related to the different arrangementsof cells at the apex of the meristem of these two species. Zea mays, Pisum sativum, maize, peao, scanning electron microscopy, root apex, regeneration     

Acid phosphatase and esterase activity in orchid mycorrhiza

Summary A cytochemical study was made to examine the possibility that acid phosphatase may be specifically involved in the digestion of endophytic hyphae in orchid mycorrhiza. Esterase activity was studied for comparison. Frozen sections of unfixed or glutaraldehyde-fixed protocorms of Dactylorhiza purpurella infected by Thanatephorus cucumeris (Rhizoctonia solani) were reacted for acid naphthol AS BI phosphatase, acid -glycerophosphatase or naphthol AS D esterase.A marked increase in particulate acid naphthol AS BI phosphatase activity was observed during infection of host, central, parenchyma cells shortly before hyphae lysed; a diffuse reaction of high activity was localised on lysed fungus. Acid -glycerophosphatase was present at particulate sites only in fungal cytoplasm and as a diffuse reaction on lysed fungus.Naphthol AS D esterase showed highest activity at hyphal apices. Esterase seems to be associated with growth and differentiation of hyphae in orchid cells, rather than lysis of the fungus.