scholarly journals CYTOCHEMICAL LOCALIZATION OF ACID PHOSPHATASE ACTIVITY IN GRANULE FRACTIONS FROM RABBIT POLYMORPHONUCLEAR LEUKOCYTES

1972 ◽  
Vol 54 (1) ◽  
pp. 141-156 ◽  
Author(s):  
Marilyn G. Farquhar ◽  
Dorothy F. Bainton ◽  
Marco Baggiolini ◽  
Christian de Duve
Keyword(s):  
Acid Phosphatase ◽  
Polymorphonuclear Leukocytes ◽  
Mononuclear Cells ◽  
Acid Hydrolases ◽  
Cytochemical Localization ◽  
Dense Bodies ◽  
Pmn Leukocytes ◽  
Secondary Lysosomes ◽  
Isopycnic Centrifugation

When rabbit peritoneal exudates (97% polymorphonuclear [PMN] leukocytes, 2% mononuclear cells) were fractionated by zonal sedimentation or isopycnic centrifugation, four fractions (A, B, C, and D) were obtained, as reported earlier. "A" consisted largely of PMN azurophil granules, "B" of PMN specific granules, and "D" of membranous elements. The source of the more heterogeneous "C" fraction (containing acid hydrolases) was uncertain. To gain further information on the nature of this fraction, cytochemical tests for acid phosphatase (AcPase) were carried out on the starting cells and on the fractions. In intact PMN, lead phosphate reaction product was found in Golgi complexes, perinuclear cisternae, and some azurophil granules (immature forms or disrupted mature forms) of a few cells. The specifics and the intact azurophils were not reactive. Reaction product was also found within Golgi cisternae, secondary lysosomes, and some of the azurophil granules of mononuclear cells. Observations on the A and B fractions confirmed those in situ regarding the localization of reaction product in disrupted PMN azurophils, its absence from specifics, and the latency of the enzyme activity in intact azurophils. In the C fraction, AcPase was found in three structures (a) Golgi cisternae, (b) dense bodies, and (c) small pleomorphic granules Comparison with the starting cells indicates that the Golgi complexes are probably derived from both PMN leukocytes and mononuclear cells, whereas the remaining elements resemble (in size, shape, and density) secondary lysosomes and azurophil granules of mononuclear cells. The results indicate that the bulk of the cytochemically detectable AcPase present in the C fraction is derived from mononuclear cells, rather than from PMN leukocytes

scholarly journals Cytochemical localization of acid phosphatase and trimetaphosphatase activities in exocrine acinar cells.

1980 ◽  
Vol 28 (1) ◽  
pp. 78-81 ◽  
Author(s):  
C Oliver
Keyword(s):  
Acid Phosphatase ◽  
Lysosomal Enzyme ◽  
Secretory Granules ◽  
Acinar Cells ◽  
Pancreatic Acinar Cells ◽  
Metal Chelation ◽  
Acid Hydrolases ◽  
Cytochemical Localization ◽  
Capture Method ◽  
Secondary Lysosomes

Acid phosphatase activity, a lysosomal marker, is commonly demonstrated using the Gomori technique with cytidine 5'-monophosphate or beta-glycerophosphate as substrate. Using this lead capture method on mouse and rat exorbital lacrimal, parotid, and pancreatic acinar cells, reaction product was localized in GERL, forming secretory granules, and secondary lysosomes. However, a different cytochemical localization was observed for inorganic trimetaphosphatase, another lysosomal enzyme. When the technique for trimetaphosphatase activity, a metal chelation method, was applied to exocrine acinar cells, reaction produce was conspicuously absent from GERL and forming secretory granules, but was present in secondary lysosomes, occasionally in Golgi saccules, and in previously unreported basal elongated lysosomes. The differences in the localization of the two enzymatic activities emphasizes the importance of employing more than one substrate where possible, and raises questions concerning the mechanism of delivery of acid hydrolases to secondary lysosomes.

Cytochemical Studies of Protein Uptake by Adrenal Cortical Cells of the Golden Hamster

1973 ◽  
Vol 31 ◽  
pp. 684-685
Author(s):  
J. R. Hillman ◽  
W. G. Seliger ◽  
P. E. Burk
Keyword(s):  
Acid Phosphatase ◽  
Golden Hamster ◽  
Adult Animal ◽  
Zona Glomerulosa ◽  
Cortical Cells ◽  
Cytochemical Localization ◽  
Protein Uptake ◽  
Dense Bodies ◽  
Thiamine Pyrophosphatase ◽  
Adrenal Cortical Cells

In a previous histochemical study, high levels of acid phosphatase were found in the developing adrenal cortex of the golden hamster. The present study describes the cytochemical localization of acid phosphatase and thiamine pyrophosphatase in the adult animal. These localizations are then related to protein uptake by adrenal cortical cells using the exogenous horseradish peroxidase as a tracer protein. Animals were sacrificed at times ranging from 15 minutes to 24 hours after injection of peroxidase. The techniques described by Novikoff et al. were utilized for localization of acid phosphatase and thiamine pyrophosphatase.In acid phosphatase preparations (Fig. 1), dense bodies containing reaction product are found in the cytoplasm of most cortical cells and are often associated with small coated vesicles near the Golgi area. Acid phosphatase activity is present in cells of the zona glomerulosa as well as those of the other two cortical zones.

scholarly journals Human osteoclast and giant cell differentiation: the apparent switch from nonspecific esterase to tartrate resistant acid phosphatase activity coincides with the in situ expression of osteopontin mRNA.

1995 ◽  
Vol 43 (12) ◽  
pp. 1193-1201 ◽  
Author(s):  
J R Connor ◽  
R A Dodds ◽  
I E James ◽  
M Gowen
Keyword(s):  
Acid Phosphatase ◽  
Mononuclear Cells ◽  
Giant Cells ◽  
Mononuclear Phagocyte ◽  
Osteoclast Precursor ◽  
Tartrate Resistant Acid Phosphatase ◽  
Nonspecific Esterase ◽  
Connective Tissues ◽  
Human Osteoclast

Animal model and in vitro cultures suggest that osteoclasts and cells of the mononuclear phagocyte system share a common precursor. However, the human osteoclast precursor has not been positively identified. We attempted to identify the precursor in situ by using a number of osteoclast- and macrophage-selective markers, together with the expression of osteopontin mRNA, previously shown to be abundant in human osteoclasts. Sections of osteophytic bone and a panel of inflammatory connective tissues were processed for in situ hybridization; serial sections were analyzed for tartrate-resistant acid phosphatase (TRAP) and nonspecific esterase (NSE) activity, selective cytochemical markers for the osteoclast and cells of the macrophage/monocyte lineage, respectively. The murine anti-human osteoclast monoclonal antibodies 23C6 (vitronectin receptor) and C35 (osteoclast-selective) were used to further identify the osteoclast phenotype. We compared osteoclasts, giant cells, and their respective putative mononuclear precursors. At resorption sites within osteophytic bone, osteopontin mRNA was expressed in osteoclasts and a distinct population of TRAP+, NSE- mononuclear cells. Adjacent clusters of mononuclear cells were TRAP- and NSE+ or were active for both enzymes; these cells demonstrated variable expression of osteopontin mRNA. In the inflammatory connective tissues, abundant macrophage-like cells (NSE+/TRAP-) did not express osteopontin mRNA. However, TRAP+ mononuclear cells observed among clusters of NSE+ cells did express osteopontin mRNA. At these sites, clusters of putative macrophage polykaryons removing fragments of bone debris were observed. These giant cells and associated mononuclear cells were NSE- and distinctly TRAP+, and expressed osteopontin mRNA, C35, and 23C6 (human osteoclast) reactivity. Therefore, cells involved in the remodeling (resorption) of bone or the removal of bone debris, together with their immediate precursors, switch from being NSE+/TRAP- to NSE-/TRAP+ cells that express osteopontin mRNA. We propose that the clusters of NSE+/TRAP- mononuclear cells represent the immature osteoclast precursor. In support of this, TRAP+/NSE+ cells were occasionally observed in both tissues, representing an intermediate stage in differentiation. These results further suggest that cells of the mononuclear phagocyte lineage within bone and inflammatory connective tissue have the potential to differentiate into osteoclasts.

scholarly journals FINE STRUCTURAL LOCALIZATION OF ACID AND ALKALINE PHOSPHATASES IN CELLS OF RABBIT BLOOD AND BONE MARROW

1967 ◽  
Vol 15 (6) ◽  
pp. 311-334 ◽  
Author(s):  
B. K. WETZEL ◽  
S. S. SPICER ◽  
R. G. HORN
Keyword(s):  
Alkaline Phosphatase ◽  
Acid Phosphatase ◽  
Phosphatase Activity ◽  
Acid Phosphatase Activity ◽  
Mononuclear Cells ◽  
Nuclear Staining ◽  
Alkaline Phosphatases ◽  
Structural Localization ◽  
Dense Granules ◽  
Dense Bodies

In rabbit heterophils, acid phosphatase activity occurs in primary (azurophil) granules which predominate in early cells and persist in mature cells and in tertiary granules which are seen only in mature cells. Alkaline phosphatase activity occurs in secondary granules which appear in intermediate heterophils and later predominate in mature cells. Acid phosphatase activity in heterophil Golgi zones coincides developmentally with the genesis of primary and, later, tertiary granules, whereas alkaline phosphatase in the Golgi complex coincides with secondary granulogenesis. In developing eosinophils, acid phosphatase reaction product occurs in Golgi elements, rims the spherical precursors of angular, mature granules and appears inconsistently within mature granules. Basophil myelocytes show acid phosphatase in Golgi elements but not in specific granules. Additional acid phosphatase reactive structures include: granules of mononuclear cells; phagocytic vacuoles in macrophages; autophagic vacuoles in maturing erythroid cells; small dense granules of platelets; dense bodies in lipocytes; and Golgi elements of mononuclear cells, macrophages, nucleated red cells, megakaryocytes and lipocytes. Localized deposits were absent in control specimens except for enzyme-independent nuclear staining in alkaline phosphatase preparations.

Cell death in the ‘opaque patch’ in the central mesenchyme of the developing chick limb: a cytological, cytochemical and electron microscopic analysis

Development ◽  
1971 ◽  
Vol 26 (3) ◽  
pp. 401-424
Author(s):  
D. S. Dawd ◽  
J. R. Hinchliffe
Keyword(s):  
Cell Death ◽  
Acid Phosphatase ◽  
Phosphatase Activity ◽  
Acid Phosphatase Activity ◽  
Mesenchymal Cells ◽  
Dead Cell ◽  
Acid Hydrolases ◽  
The Dead ◽  
Secondary Lysosomes ◽  
Mesenchyme Cells

Cell death in the ‘opaque patch’ of central mesenchyme of the developing chick forelimb was investigated by a variety of light and electron-microscope cytological and cytochemical techniques. Cell death appears first at stage 23/4 (4 days) and reaches its maximum extent at stages 24 and 25 (4½ and 5 days), at which it separates the ulnar and radial mesenchymal condensations. It then decreases in size to a small area separating the proximal parts of radius and ulna and disappears at stage 28. Cytological studies show the presence of a few isolated dead cells, of mesenchymal cells containing 1–3 ingested dead cells and of macrophages containing up to 18 dead cells in various stages of digestion. These findings are interpreted as showing that isolated dead cells are ingested by neighbouring mesenchymal cells which thus become transformed into macrophages, first ingesting and then digesting further dead cells. Histochemical studies show that isolated dead cells and recently ingested dead cells contain no more acid phosphatase activity, either discrete or diffuse, than either neighbouring living mesenchymal cells, or mesenchymal cells which have ingested 1–3 dead cells. Increased acid phosphatase activity is found within the macrophages, where activity is localized within the digestive vacuoles (‘secondary lysosomes’) containing the dead cells, and within the Golgi apparatus and Golgi vesicles (‘primary lysosomes’) of macrophage cytoplasm. Loss of staining capacity by the dead cell is correlated with high acid phosphatase activity: this is interpreted as indicating the digestion of dead cells within the macrophage by acid hydrolases. There is circumstantial evidence that viable mesenchyme cells in the ‘opaque patch’ area autophagocytose part of their own cytoplasm in secondary lysosomes (1·2–2µm). The role of the ‘opaque patch’ in relation to the pattern of limb chondrogenesis is discussed. It is suggested that cell death may play a role in separation of radius and ulna, and that autophagocytosis may indicate a change in the pathway of differentiation of the mesenchyme cells lying between radius and ulna.

scholarly journals Isolation of pig thyroid lysosomes. Biochemical and morphological characterization

1985 ◽  
Vol 232 (2) ◽  
pp. 529-537 ◽  
Author(s):  
C Alquier ◽  
P Guenin ◽  
Y Munari-Silem ◽  
C Audebet ◽  
B Rousset
Keyword(s):  
Endoplasmic Reticulum ◽  
Polyacrylamide Gel Electrophoresis ◽  
Average Diameter ◽  
Morphological Characterization ◽  
Thyroid Cell ◽  
Acid Hydrolase ◽  
Acid Hydrolases ◽  
Thyroid Cells ◽  
Secondary Lysosomes ◽  
Isopycnic Centrifugation

Open thyroid follicles were prepared by mechanical disruption of pig thyroid fragments through a metal sieve. This procedure allowed preparation of thyroid-cell material depleted of colloid thyroglobulin. Open thyroid follicles were used to prepared a crude particulate fraction, which contained lysosomes, mitochondria and endoplasmic reticulum. These organelles were subfractionated by isopycnic centrifugation on iso-osmotic Percoll gradients. A lysosomal peak was identified by its content of acid hydrolases: acid phosphatase, cathepsin D, β-galactosidase and β-glucuronidase. The lysosomal peak was well separated from mitochondria and endoplasmic reticulum. The lysosomal peak, from which Percoll was removed by centrifugation, was taken as the purified lysosome fraction (L). Lysosomes of fraction L were purified 45-55-fold (as compared with the homogenate) and contained about 5% of the total thyroid acid hydrolase activities. Electron microscopy showed that fraction L was composed of an approx. 90% pure population of lysosomes, with an average diameter of 220 nm. Acid hydrolase activities were almost completely (80-90%) released by an osmotic-pressure-dependent lysis. Thyroglobulin was identified by polyacrylamide-gel electrophoresis as a soluble component of the lysosome fraction. In conclusion, a 50-fold purification of pig thyroid lysosomes was achieved by using a new tissue-disruption procedure and isopycnic centrifugation on Percoll gradient. The presence of thyroglobulin indicates that the lysosome population is probably composed of primary and secondary lysosomes. Isolated thyroid lysosomes should serve as an interesting model to study the reactions whereby thyroid hormones are generated from thyroglobulin and released into the thyroid cells.

scholarly journals Normal rabbit alveolar macrophages. II. Their primary and secondary lysosomes as revealed by electron microscopy and cytochemistry.

1976 ◽  
Vol 144 (4) ◽  
pp. 920-932 ◽  
Author(s):  
B A Nichols
Keyword(s):  
Electron Microscopy ◽  
Endoplasmic Reticulum ◽  
Acid Phosphatase ◽  
Rough Endoplasmic Reticulum ◽  
Alveolar Macrophages ◽  
Digestive Enzymes ◽  
Acid Hydrolases ◽  
Secondary Lysosomes ◽  
Enzymatic Machinery ◽  
Cytochemical Markers

In this investigation, vacuoles containing tubular myelin proved to be digestive compartments with cytochemical reactivity for acid phosphatase and arylsulfatase. These cytochemical markers identify the secondary lysosomes, known to contain enzymes capable of hydrolyzing phospholipids like surfactant. Therefore, it appears that alveolar macrophages possess the enzymatic machinery for the degradation of the tubular myelin found in their digestive vacuoles. Although it thus appears evident that alveolar macrophages participate in the turnover of surfactant, the quantitative significance of this route of disposal is undetermined. This investigation has also established that acid hydrolases, so prominently displayed in the secondary lysosomes, are also localized in the rough endoplasmic reticulum and in Golgi-endoplasmic reticulum-lysosomes (GERL). Moreover, small vesicles which are produced from GERL serve as primary lysosomes in transporting digestive enzymes to the vacuoles.

Demonstration of Secondary Lysosomes in Bovine Megakaryocytes and Platelets Using Acid Phosphatase Cytochemistry with Cerium as a Trapping Agent

1990 ◽  
Vol 63 (01) ◽  
pp. 127-132 ◽  
Author(s):  
Michèle Ménard ◽  
Kenneth M Meyers ◽  
David J Prieur
Keyword(s):  
Acid Phosphatase ◽  
Electron Density ◽  
Golgi Complex ◽  
Initial Report ◽  
Cerium Phosphate ◽  
Virtual Absence ◽  
Phosphatase Cytochemistry ◽  
Secondary Lysosomes ◽  
Trapping Agent

SummaryThe ultrastructure of lysosomes from bovine megakaryocytes (MK) and platelets was characterized using acid phosphatase cytochemistry with beta-glycerophosphate as substrate and cerium as a trapping agent. The technique was easily reproducible; cerium-phosphate precipitates were uniform, readily visualized, and there was a virtual absence of nonspecific reaction product. Acid phosphatase was localized in the trans aspect of the Golgi complex and/or granules of less than 50 nm to 650 nm diameters in MK at all stages of maturation. Forty percent of the MK lysosomes contained inclusions of variable shapes, sizes and electron-density and were classified as secondary lysosomes. Twenty-four percent of the platelet sections contained acid phosphatase-positive granules. Fifty-four percent of these were secondary lysosomes. This is the initial report demonstrating secondary lysosomes in either resting MK or platelets using acid phosphatase cytochemistry. These findings suggest that MK and platelet lysosomes have an intracellular function in resting MK and platelets.

scholarly journals Cytochemical localization of a "basic" ATPase to canine myocardial surface membrane.

1980 ◽  
Vol 28 (12) ◽  
pp. 1286-1294 ◽  
Author(s):  
N N Malouf ◽  
G Meissner
Keyword(s):  
Cardiac Muscle ◽  
Microsomal Fraction ◽  
Surface Membrane ◽  
Buoyant Density ◽  
Membrane Structures ◽  
Intercalated Disc ◽  
Cytochemical Localization ◽  
Fixed Tissue ◽  
T System

Enzymatic properties of a canine cardiac muscle microsomal fraction were determined to localize in situ a "basic," divalent cation dependent adenosine triphosphatase (ATPase) by ultrastructural cytochemistry. The microsomal fraction had a buoyant density of 1.08--1.13 (20--30% [w/w] sucrose) and hydrolyzed adenosine triphosphate in the presence of Mg2+, Ca2+, Mn2+, or Co2+, but not in that of Sr2+ or Ni2+, under conditions that inhibited interfering (Na+ + K+)-ATPase and sarcoplasmic reticulum Ca2+-ATPase activities. "Basic" ATPase was localized in paraformaldehyde-fixed tissue in a medium containing Mg2+ or a high Ca2+ concentration (4 mM). A free Pb2+ concentration of less than 1 microM was used to capture enzymatically released phosphate anions. Electron-dense lead precipitates were present at the plasmalemma, T-system, and intercalated disc membranes with the exception of the nexus. These studies suggest that "basic" ATPase activity is associated with surface membrane structures of canine cardiac muscle.

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