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PubMed1983

Expression of multiple isozymes of granulocyte, monocyte, and macrophage esterases in polycythemic Friend erythroleukemia cells.

We examined the expression of cytochemical markers of myeloid and monocyte-macrophage differentiation in conjunction with ultrastructural studies of different malignant erythroleukemic cells isolated from mice infected with the Friend polycythemic virus complex (FLV-P). The amounts of fluoride-sensitive and resistant nonspecific esterase activity increased with the progression of malignancy. Isoelectric focusing resolved this enzyme activity into 13 isozymes in the most malignant Friend cell type tested. These same isozymes were found in the adherent cell population of normal spleens. Two of these isozymes were shown to have chloroacetate esterase activity characteristic of granulocytes. Despite these myeloid and monocyte characteristics, light and electron microscopy showed no morphological evidence of differentiation in either of these lineages. This study demonstrates that the Friend erythroleukemic cell contains markers of three different hemopoietic cell types. The expression of myeloid, monocytic, and erythroid traits in these erythroleukemic cells can be used to monitor their malignant progression.

Animals↗
PubMed1984

[Post therapeutic myeloblastic leukemia in non-Hodgkin's lymphoma].

A 28-year-old man developed AML 18 months after a diagnosis of non-Hodgkin's lymphoma, diffuse small cell type, clinical stage IIA. Induction therapy for the lymphoma consisted 60Co 4000 rads bilaterally to the cervical areas and 2000 rads to the right cervical area. Complete remission was attained. Nineteen courses of combination chemotherapy with Vincristine (VCR), 6-meraptopurine (6 MP), cyclophosphamide (CY) and predonisolone (pred) was added (Total dose: VCR; 26.5 mg, 6 MP; 3320 mg, CY; 3350 mg, pred; 4310 mg). Seven days after the final chemotherapeutic treatment he developed AML with DIC. Leukemic cells were peroxidase and specific esterase (naphthol AS-D chloroacetate) positive. Induction therapy for the AML consisting of DCMP (Daunomycin, Cytosine arabinosid, 6 MP and pred) and VCR (vindesine, CY and pred) was unsuccessful. The patient died of cranial hemorrhage 3 month after the diagnosis of acute leukemia. Autopsy revealed no recurrence of non-Hodgkin's lymphoma in the lymph nodes, bone marrow, spleen and liver. Seven other cases reported in the Japanese literature are reviewed.

Adult↗
PubMed1984

Release of granulocyte-specific colony-stimulating activity by human bone marrow exposed to phorbol esters.

Granulocyte-macrophage colony growth depends on the presence of colony-stimulating activity (CSA). Phorbol esters induce concentration-dependent colony formation in the absence of exogenous CSA. We questioned whether phorbol esters mimicked the action of CSA by directly stimulating colony growth, or whether phorbol esters acted indirectly by inducing marrow cells to release CSA. First, after incubating human bone marrow cells with phorbol 12,13-dibutyrate (PDB) for 3 days, we separated PDB from the protein peak of the conditioned medium by Sephadex G-10 gel filtration and tested this peak for the presence of CSA. When diluted 1:10 in the agar colony assay, this material induced 133 +/- 15 colonies/10(5) bone marrow cells. Second, to determine whether bone marrow cells required the continued presence of PDB in order to release CSA, PDB was removed from bone marrow cells by washing, and these cells were reincubated in fresh medium in the absence of PDB. CSA was found in the medium of these cultures; its release was maximal after preincubation of bone marrow cells with 5 X 10(-8) M PDB for 3 days, followed by incubation for 3 days in the absence of PDB. This CSA stimulated granulopoiesis out of proportion to monocytopoiesis, with 85% +/- 17% of the colonies being granulocytic (as indicated by histochemical staining for chloroacetate esterase), and 12% +/- 3% being monocytic (as indicated by nonspecific esterase). Inhibitors of monocyte colony formation, including PGE1, were not present in the medium that contained this CSA. These studies demonstrate that normal human bone marrow cells exposed to PDB release CSA and that this CSA selectively stimulates granulopoiesis in vitro.

Alprostadil↗
PubMed1984

Development of human connective tissue mast cells from purified blood monocytes.

Highly purified subfractions of human peripheral blood monocytes, when cultured in the presence of 30% L cell supernatant and 30% horse serum, assumed all the characteristics that define human connective tissue mast cells. After three weeks of culture, 75% of the cells developed metachromasia and granular chloroacetate esterase staining, and their intracellular histamine levels increased from 0.0 to 50.5 ng/10(6) cells. On electron microscopy, the cells developed intracytoplasmic granules with all the features typical for mature and immature mast cells. Cultured cells bound 55 pg 125I-IgE/10(6) cells, while labelling was negligible with cells prior to culture and with heat-denatured 125I-IgE. Fluorescent staining with anti-IgE increased slightly as well, while staining with monoclonal anti-monocyte and anti-HLA-Dr markers decreased. Purified lymphocytes did not assume mast cell characteristics, and lymphokines did not induce or enhance in vitro mast cell development or IgE binding. The data therefore further support the concept that connective tissue mast cells arise from the monocytoid lineage.

Binding Sites, Antibody↗
PubMed1984

Effect of phenoxy acids on rat liver regeneration.

Phenoxy-acids incorporated in a basal region on a weight basis, were fed to male rats from which two-thirds of the liver had been removed and the extent of regeneration or the liver increment was ascertained over a period of 10 days p.o. Stimulatory responses occurred with the hypolipidemic drugs, ethyl clofibrate, at levels of 0.25% higher or by gavage over the 1st 8 days p.o. at an overall dosage of 1275 mg/kg, 2-(p-chlorophenoxy)propionic acid (0.50, 1.0%), the corresponding o-chloro-acids (0.20 and 0.50%, respectively) and several other with branching of the acid side-chain. The latter included phenoxyisobutyric acid (0.50%), 2-phenoxypropionic acid (1.0%), 2-phenoxybutyric acid (1.0%), the phenoxydimethyl-pentanoic acid, gemfibrozil (0.40%), the herbicide, 2-(2,4,5-trichlorophenoxy)propionic acid (0.10%) and as screened on a few males, possibly, halofenate (0.20%). The liver increments were in the control range for compounds containing straight chain acid substituents as 4-(p-chlorophenoxy)-butyric acid (0.35%), 3-phenoxypropionic acid (0.75%), 3-(p-chlorophenoxy)-1,2-propanediol (0.50%) and phenoxyacetic acid derivatives, including p-chloroacetic acid and the herbicides 2,4-dichlorophenoxy-, 2-methyl-4-chlorophenoxy- and 2,4,5-trichlorophenoxyacetic acids at levels of up to about 0.50%. Where effects resulted with the phenoxyacetic acids, these were in the direction of inhibition of the regenerative process. The response of the female rat to clofibrate either by diet or gavage was rather poor or equivocal but was more definite with 2-(2,4,5-trichlorophenoxy)propionic acid (0.10%). However, intact females were more consistent in eliciting wet and dry liver enlargement with clofibrate (0.25%) as noted earlier by others and with gemfibrizol (0.040%) and the trichlorophenoxypropionic acid (0.10%) but not with 2,4,5-trichlorophenoxyacetic acid at 0.10%.

Animals↗
PubMed1984

Morphologic, cytochemical, and functional studies of peripheral blood cells of young healthy American alligators (Alligator mississippiensis).

Samples of peripheral blood collected from healthy young American alligators (Alligator mississippiensis) were studied to determine baseline values and characterize cell types. Mean total leukocyte counts were 6.4 +/- 2.9 X 10(3)/mm3, with mean differential values of 54.7 +/- 47.5% heterophils, 10.4 +/- 6.0% eosinophils, 12.7 +/- 16.8% basophils, 23.9 +/- 4.9% lymphocytes, and 0.7 +/- 0.5% monocytes. Mean total thrombocyte counts were 23.0 +/- 7.0 X 10(3)/mm3, and mean total erythrocyte counts were 3.84 +/- 8.7 X 10(5)/mm3. Morphologic descriptions of Wright-Giemsa-stained alligator blood cells generally corresponded to those of their counterparts in other reptiles, birds, and mammals. Cytochemical tests were positive as follows: chloroacetate esterase-lymphocytes and monocytes; nonspecific esterase-heterophils and monocytes; acid phosphatase-heterophils, basophils, and monocytes; and alkaline phosphatase-heterophils, eosinophils, and monocytes. Phagocytic and microbiocidal capacity was tested by adding live Staphylococcus aureus to whole blood. Subsequent staining with acridine orange and observation with fluorescence microscopy showed in vitro phagocytic and microbiocidal activity of monocytes, heterophils, and to a lesser degree, eosinophils.

Acid Phosphatase↗
PubMed1982

[Immuno and enzyme histochemical demonstration of granulocytes--assay of a morphometric evaluation].

Using the naphthol AS-D-chloroacetate esterase method and the indirect immunoperoxidase technique we tried to stain selectively granulocytes in the lung and in the liver. For the immuno technique we used an antibody raised in rabbits against dog granulocytes, which were prepared by a new separation technique. Comparing the two staining methods we found the immunoperoxidase to be superior at least for quantitative evaluation. In a pilot study with polytraumatized dogs versus controls we could detect a more pronounced leucostasis within the trauma group. This was similar to a parallel study which used radioactive labeled granulocytes where a significant increase could be shown. Granulocytes, at least in part, seem to be important for cellular changes within the "lung in shock".

Animals↗
PubMed1982

[Treatment of acute myeloid leukemia with a protocol combining intensive induction chemotherapy, early consolidation treatment, splenectomy and long-term maintenance chemotherapy. Preliminary study].

Twenty-seven patients aged from 10 to 60 years (mean 34.4 +/- 13 years) in the first perceptible phase of acute myeloid leukemia were subjected to intensive induction chemotherapy consisting of adriamycin (ADM), vincristin (VCR) and cytosine arabinoside (ARA-C). Twenty-four patients (89%) attained complete remission (CR) after 1 to 3 cycles and were then given an early consolidation treatment with one of the previous cycles. This was followed by long-term continuous maintenance chemotherapy with 6-mercaptopurine (6-MP) and methotrexate (MTX) alternatively and 3-monthly reinforcement courses of donaurubicin (DNR) and VCR. Twenty of these 24 patients were splenectomized soon after the consolidation treatment. None of the spleens were enlarged, and histological sections of the spleens, liver biopsies and mesenteric lymph-nodes stained with routine dyes and by the naphthol AS-D chloroacetate esterase method revealed mature granulocytes but no demonstrable leukaemic cells. In the group of splenectomized patients, the probabilities of staying in complete remission at 27 and 44 months were 70 +/- 12.6% and 52 +/- 18.5% respectively, and the probabilities of remaining alive at 32 and 55 months were 79 +/- 11% and 57 +/- 19% respectively. Age over 40 and evidence of extramedullary infiltration at presentation appeared to leave little hope of disease-free survival. The rationale for the present therapeutic study is discussed.

Adolescent↗
PubMed1983

Preparation and characterization of acid dehydration products of aflatoxicol.

Treatment of aflatoxicol with acid resulted in the formation of dehydrated derivatives with the 2,3 double bond either hydrated or unaffected. Chromatographic and physicochemical analyses of the purified derivatives revealed that hydration of the 2,3 double bond of aflatoxicol with simultaneous dehydration occurred when sulfuric acid was used. When a weaker acid such as chloroacetic acid was used, no hydration occurred, and 2 isomeric forms of the dehydrated aflatoxicol were obtained. Chicken embryo assay and the Ames test revealed that dehydrated aflatoxicols were much less toxic and mutagenic than aflatoxicol. A mechanism for the dehydration of aflatoxicol is proposed.

Aflatoxins↗
PubMed1983

Activity of nonspecific esterases in the marrow of mice following intraperitoneal administration of concanavalin A and PHA.

The activity of nonspecific esterases was studied in the bone marrow cells of mice after single intraperitoneal injections of Con A and PHA-M in the dose of 30 mg per 1 kg of body weight. Samples were collected 24, 72 and 120 hours after the injection of lectins. A cellular reaction to the activity of nonspecific esterases was tested after the method of Heckner using naphthol chloroacetate AS-D as an esterase substrate. In the control group the cells belonging to the granulocytic system except these being in mature forms, showed an apparent activity. A negative esterase reaction was revealed also in megakaryocytes, monocytes and lymphocytes. The administration of Con A intensified the esterase reaction in the cells belonging to the granulocytic series, except the mature forms. The esterase activity appeared in megakaryocytes, monocytes and lymphocytes. Most numerous changes in the activity were noted 72 hours following the injection. PHA-M was found to exert a reverse effect, weakening the esterase reaction in comparison to both the controls and the Con A--group.

Animals↗
PubMed1980

Granulocytic sarcoma of the skin.

A patient with aleukemic leukemia of the acute granulocytic type, who initially had granulocytic sarcoma of the skin, is described. The skin contained focal infiltrates of pleomorphic mononuclear cells that were identified as granulocytes by demonstration of intracytoplasmic naphthol-ASD-chloroacetate esterase and lysozyme.

Aged↗
PubMed1982

Structure-activity relationships for the induction of differentiation of HL-60 human acute promyelocytic leukemia cells by anthracyclines.

Marcellomycin, a newly developed anthracycline antibiotic with antineoplastic activity, was tested as an inducer of differentiation of the human promyelocytic leukemia cell line HL-60 in vitro. The percentage of cells reducing nitro blue tetrazolium, an indication of a stimulus-induced respiratory burst typical of mature phagocytes, was used as a functional measure of the extent of differentiation. Marcellomycin was a potent inducer of maturation, with 95% of the cells expressing a terminally differentiated state after 10 days of exposure to a concentration of 40 nM anthracycline. Cells exposed to marcellomycin exhibited a 35-fold increase in their total superoxide anion-generating capacity, an 80% increase in acid phosphatase activity, and a loss of myeloperoxidase and chloroacetate esterase activities. In addition, marcellomycin-treated cells stained negatively for alpha-naphthyl acetate esterase. These findings provided evidence for the granulocytic nature of the mature cells. In contrast, marcellomycin was not an effective inducer of differentiation of Friend murine erythroleukemia cells. Studies on the relationship between structure and the ability to induce differentiation of HL-60 leukemia cells demonstrated that removal from the marcellomycin molecule of the terminal 2-deoxyfucose (musettamycin) or its substitution by cinerulose (aclacinomycin A) did not alter differentiation-inducing capacity. However, removal of the carbomethoxy group from the C-10 position of marcellomycin substantially reduced its potency as an initiator of maturation, and removal of the two terminal 2-deoxyfucose moieties (pyrromycin) decreased both potency and the maximal percentage of differentiated cells produced in the population. The monosaccharide anthracyclines Adriamycin and carminomycin were completely inactive as inducers of HL-60 leukemia cell maturation. The results suggest that certain anthracyclines would be reasonable candidate drugs to use in a clinical trial aimed at reducing the leukemic stem cell burden through maturation rather than through cytodestruction.

Anthracyclines↗
PubMed1981

Modification of normal human myelopoiesis by 12-0 tetradecanoylphorbol-13-acetate (TPA).

The tumor promoter, 12-O-tetradecanoylphorbol-13-acetate (TPA), induces macrophage characteristics in the HL-60 promyelocytic leukemia cell line. These same cells can be induced to develop mature myeloid characteristics with a variety of other stimuli. Since normal colony-forming units-culture (CFU C) also have the dual capability of developing colonies with myeloid or monocyte characteristics, the effect of TPA on normal human CFU-C development was studied. To carry out these studies, a method was developed to identify cells histochemically within agar cultures as containing either the myeloid marker, chloroacetate esterase (CAE), or the monocyte marker, nonspecific esterase (NSE). Cells from normal donors were placed into agar cultures with placenta conditioned medium (PCM), TPA in various concentrations, or combinations of PCM and TPA as stimulating materials, and examined after 7-14 days of incubation. TPA alone at concentrations of 5 x 10(-7) M to 10(-9) M stimulated cluster formation. With increasing concentrations of TPA, the percentage of clusters positive for NSE progressively increased, while CAE-positive clusters decreased. I contrast, when TPA at concentrations greater than 10(-9) M was added to PCM, the total number of clusters and colonies decreased. This resulted from a decrease in the number of clusters and colonies that contained the myeloid marker CAE, whereas the number positive for the monocyte marker NSE remained unchanged. These studies demonstrate two effects of TPA on normal CFU-C. In the absence of other sources of colony stimulating activity (CSA), TPA induces clusters. In the presence of PCM, it inhibits the production of myeloid colonies and clusters. Under both conditions, it favors the development of colonies and/or clusters containing predominantly monocytes.

Bone Marrow Cells↗
PubMed1982

Proliferative, morphologic, and cytochemical characteristics of bovine bone marrow macrophage colonies in liquid culture.

Bovine macrophage colonies were grown in liquid medium. Serum collected from a heifer 3 hours after injection of Pseudomonas aeruginosa endotoxin promoted the growth of macrophage colonies. The optimum concentration of postendotoxin serum was 16%. The numbers of colonies increased exponentially between 3 and 6 days of incubation and leveled off at 6 days. Monoblasts, promonocytes, and macrophages were observed in these colonies. A direct linear relationship between the number of nucleated bone marrow cells cultured and the number of colonies formed was found. The macrophage colony cells were positive for nonspecific esterase activity and were negative for chloroacetate esterase activity. Weak peroxidase activity was noticed in monoblasts, promonocytes, and occasionally in macrophages.

Animals↗
PubMedDate not supplied

Granulocyte elastase at the site of cartilage erosion by rheumatoid synovial tissue.

Elastase, an enzyme in the azurophilic granules of polymorphonuclear cells (PMN), is like other granular PMN proteases characterized by its degradative activity at physiological pH towards native macromolecules as shown in a serum free medium. Joint tissue specimen obtained during elective surgery in cases of various rheumatic conditions were examined in order to elucidate the role of this enzyme during joint cartilage destruction. An indirect immunofluorescence microscopic technique utilizing a rabbit immunoglobulin G preparation raised against purified elastase was used for this purpose. Immunoreactive elastase was seen bound to cells which were recognized as PMN by their nuclear characteristics and staining in a histochemical reaction with naphtol AS-D chloroacetate. PMN were encountered more or less often in the pannus but clearly accumulated in a significant amount at the pannus-cartilage junction in one case of rheumatic monarthritis and three out of four cases with rheumatoid arthritis. This finding shows that PMN--contrary to other descriptions--belong to the morphologic characteristics of inflammatory rheumatic conditions and directly supports the hypothesis that PMN enzymes play an active role in rheumatoid cartilage destruction.

Arthritis, Rheumatoid↗
PubMed1981

Enzyme histochemistry and immunohistochemistry on biopsy specimens of pathologic human bone marrow.

We have systematically investigated a variety of fixation and plastic embedding procedures and arrived at a method that allows processing of approximately 2-micron sections of bone marrow biopsies for examination by light microscopy. More importantly, this method permits the use of enzyme histochemical and immunohistochemical procedures that are rapidly becoming mandatory in the diagnosis of hematologic malignancies. Over 200 full-length bone marrow biopsy specimens were fixed in a mixture of paraformaldehyde, glutaraldehyde, and acrolein, dehydrated in acetone, and embedded in a mixture of methyl and glycolmethacrylate. All procedures were carried out at 4 degrees C. Decalcification was unnecessary. Sections 2-micron thick were cut and incubated for peroxidase, naphthol AS-D chloroacetate esterase, alpha-naphthyl butyrate esterase, acid phosphatase (with and without tartrate), or alkaline phosphatase and then examined by light microscopy. Specimens could be prepared for examination within 48 hr. This approach, which provides definitive markers for various hematopoietic cell lines in intact tissues, is invaluable when aspirated material is unavailable. It is also useful in the analysis of focal lesions of bone marrow due to inflammation or neoplasia and shows potential as an investigative tool. For example, we have discovered that early myelofibrosis is accompanied by a marked increase in the number of alkaline-phosphatase-positive reticulum cells.

Bone Marrow↗
PubMed1977

Monocytic differentiation of cluster cells in agar culture of normal human bone marrow. A cytochemical investigation.

A cytochemical investigation was carried out on the cells that make up the clusters forming in agar cultures of normal human bone marrow after 3, 4, and 5 days of incubation. All the clusters were found to comprise young monocytes that were PAS, peroxidase, Sudan Black, and alpha-naphthyl and naphthol-ASD-acetate esterase positive. Lysozyme activity, investigated by the cytobacterial test, was absent in all cases. After 3 days of incubation a large number of pairs of lymphomonocytes or monocytes with the same cytochemical characteristics were observed. When a method of double esterase incubation (alpha-naphthyl or naphthol-ASD-acetate and naphthol-ASD-chloroacetate) was performed on the same slide, no cells of the granulocytic line or intermediate cells between these and the monocytes were observed in any of the clusters. These results indicate that there is direct monocytic differentiation of the colony-forming cells cultured. The origins and stages observed in the maturation of the monocytes are discussed.

Agar↗
PubMed1977

[Studies on the morphology of blood of the European eel (Anguilla agnuilla). III. Studies on monocytic and lymphoid cells].

Investigations concerning the monocytic and lymphocytic cell series were carried out on the peripheral blood and dabbings of the kidney, the spleen and intestines of healthy eels (anguilla anguilla), and eels taken ill with vibrio anguillarum. By means of histological, cytochemical, phaseoptical, and electronmicroscopical examinations independent development lines of the monocytes and lymphocytes were established. The place of origination for the monocytes was found to be the kidney, for the lymphocytes the spleen and the intestines additionally. The evidence of the cytochemical assay of enzymes for the differentiation of these blood cells and their cellular systems is primarily dependent on the distribution of the activity of the unspecific esterase, the naphthol-AS-D-chloroacetate-esterase, the peroxidase and the alkalic phosphatase. Macrophages were - on the basis of enzymocytochemical results - seen as derivatives of the monocytes. Plasma cells and lymphoid cells are transformation forms of the lymphocytes. Our results emphasize the significance especially of cytochemistry for the hematological diagnostics of fish diseases.

Animals↗