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Synthesis of some indole derivatives with potential antiinflammatory activity.

2-Methylindole-3-acethydrazide (1) was reacted with arylisothiocyanate to give the corresponding 4-arylthiosemicarbazides 2a-d. Cyclization of the latter gave the corresponding 3-mercapto-5-[3-(2-methylindolyl)methyl]-1,2,4-triazoles 3a-d. Compounds 3a-c reacted with chloroacetic acid to give the corresponding indolyl-1,2,4-triazolythioglycolic acids 4a-c. The methylmercapto derivative 5 was also obtained from 3a and methyliodide. The hydrazide 1 was also reacted with carbon disulfide to give the corresponding indolymethyl-1,3,4-oxadiazole-2-thione (6) which was condensed with piperidine and formaldehyde to give the corresponding Mannich base 7. Condensation of 1 with aromatic aldehydes gave the corresponding hydrazones 8a-c which were converted into the corresponding oxadiazolines 9a-c.

Anti-Inflammatory Agents↗

Synthesis and antifertility activity of some new fluorine containing 2-([2-(fluoroaryl)-1H-indol-3- yl]methylene)hydrazinecarbothioamides and 2-(fluoroaryl)-([5-(substituted benzylidene)-4-oxo-2-thiazolidinylidene]hydrazone)- 1H-indole-3-carboxaldehydes.

New fluorine containing 2-(fluoroaryl)-1H-indole-3-carboxaldehydes have been synthesized and subjected to reaction with thiosemicarbazide to give corresponding 2-([2- (fluoroaryl)-1H-indol-3-yl]methylene)hydrazinecarbothiamides which were cyclized in the presence of chloroacetic acid, sodium acetate and substituted benzaldehydes to 2-(fluoroaryl)-([5- (substituted benzylidene)-4-oxo-2-thiazolidinylidene]hydrazone)- 1H-indole-3-carboxaldehydes as potential antifertility agents. In preliminary screening, 2-(4'-fluorophenyl)- ([5-(methylene-3,4-dioxyphenyl)-4-oxo-2- thiazolidinylidene]hydrazone)-1H-indole-3-carbocaldehyde exhibited pronounced antifertility activity. All these new compounds have been characterized by analytical and spectral (IR, PMR, MS) studies.

Animals↗

Differential expression of HLA-DR antigens in subsets of human CFU-GM.

Expression of HLA-DR surface antigens by granulocyte/monocyte colony-forming cells (CFU-GM) may be important in the regulation of proliferation of these cells. Using immunological techniques to enrich for progenitor cells, we investigated the expression of HLA-DR in subsets of CFU-GM. "Early" (day 14) CFU-GM express higher levels of HLA-DR than do "late" (day 7) CFU-GM. Among late CFU-GM, cells destined to form monocyte (alpha-naphthyl acetate esterase-positive) colonies express higher levels of HLA-DR than do CFU-GM destined to form granulocyte (chloroacetate esterase-positive) colonies. Because high-level expression of DR antigen was a marker for monocyte differentiation, we examined several lymphokines for their effects on both DR expression and in vitro commitment to monocyte differentiation by myeloid precursor cells. DR antigen density could be increased by more than twofold over 48 hours upon exposure to gamma-interferon (gamma-IFN), whereas colony-stimulating factors had no effect. This was associated with a dose-dependent inhibition of total CFU-GM number, and a relative, but not absolute, increase in the ratio of monocyte colonies to granulocyte colonies. Similarly, in day 7 suspension cultures of purified myeloid precursor cells, gamma-IFN inhibited cell proliferation and increased the ratio of monocytes to granulocytes. Thus, despite the induction of high levels of HLA-DR antigen on precursor cells (a marker of monocyte commitment), the dominant in vitro effect of gamma-IFN was inhibition of granulocyte differentiation.

Antibodies, Monoclonal↗

[Malignant hypereosinophilic syndrome. Report of an exceptional case (author's transl)].

The case of a 43-year-old man with a highly malignant hypereosinophilic syndrome is reported. The condition is classified as such according to Hardy and Anderson's criteria, accepted by many other authors. Other diseases of known etiology which may present high levels of eosinophils in the peripheral blood, such as parasitosis, allergies, neoplasias, collagenosis, etc., were discounted beforehand. The difficulties in distinguishing between these diseases are discussed; they are often accompanied by clinical manifestations which also arise in very different conditions including eosinophilic leukemia, Engfeldt and Zetterström's eosinophilic collagenosis, Löffler's fibroplastic endocarditis, etc. A particularly striking feature of this condition is the formation of large tumor masses of mature eosinophils. They begin in various bones, which they destroy almost completely, and invade the surrounding tissues, destroying them as well. These tumors act similarly to malignant eosinophilic myelocytomas, a fact which has not been reported previously in the literature as far as we know. Although the eosinophils act as though they were neoplastic, they maintain the characteristics of mature cells, both cytomorphologically and ultrastructurally as well as cytochemically (consistently chloroacetate esterase negative). The tendency to diagnose eosinophilic leukemia solely on the basis of the malignancy of the condition and a tissue infiltration of eosinophils without determining the existence of cytologic and/or cytochemical anomalies of the cells showing them to be leukemic is discussed. The authors were unable to find any reports in the literature in which the eosinophils were presented with unmistakeably blastic cellular characteristics. Various nosologic considerations are offered.

Adult↗

[Cytochemical studies in megakaryocytes in hematologic diseases].

Bone marrow smears of 48 patients consisting of 12 normal cases, 36 patients with different haematological diseases-among them 9 cases of idiopathic thrombopenia, 4 cases of polycythaemia, and 9 cases of Hodgkin's disease - were examined cytochemically. Acid phosphatase, unspecific esterases, naphthol-AS-D-chloroacetate esterase, peroxydase, and leucin-aminopeptidase were represented; in addition the PAS reaction, fastgreen staining at pH 1.1, methyl-green pyronin staining and the lipid representation with Sudan black B were carried out. Besides those responses known from literature the different behaviour of acid megacaryocyte phosphatase in different haematological diseases must be particularly emphasized from all reactions.

Acid Phosphatase↗

[Acute mast-cell leukemia. Cytochemical and ultrastructural study, about a particular case (author's transl)].

A case of acute Mast-cell leukemia was studied. A 39 years old female presenting with a brief history of abdominal pain and attacks of flushing; peripheral blood and bone marrow contained up to 60% of poorly differentiated blasts with clumping of deep purpule granules. Peroxydase reaction stains were negative, chloroacetate esterase were strongly positive. Toluidine blue revealed metachromatic stain. Histamine content of the cells was highly greater than normal but nos heparinoid activity could be demonstrated. These abnormal mast-cells have been investigated with the electron microscope; only the dense particular type of granule substructure was found, without any lamellae component. The cells were temptatively classified as "immature" mast-cell. The disease was interpretated as an acute leukemic variety of systemic mastocytosis.

Acute Disease↗

Cytochemistry of blood cells in peripheral smears of some West African reptiles.

The distributions of lipid, glycogen, peroxidase, acid and alkaline phosphatases, beta-glucuronidase and naphthol AS-D chloroacetate esterase have been studied in the cells of peripheral smears from the wall ghecko and the crocodile. The erythrocytes react differently from those of mammalian erythrocytes with regard to peroxidase reactivity. The need to take such species differences into consideration when engaged in histochemical investigations is stressed.

Acid Phosphatase↗

Histochemistry of bone marrow aspirations.

The accuracy of diagnosis of hematological diseases from bone marrow aspirates and peripheral blood smears is considerably improved by use of histochemical stains. As hematopoietic cells differentiate and mature from the pluripotent stem cell and are committed to a particular cell line, the committed cell lines produce different substances, particularly enzymes, which can be identified by various histochemical stains. Histochemical stains are not diagnostic, but are aids in diagnosis, because various cell lines may produce similar substances and because the stains lack absolute specificity. A discussion of the use of histochemical stains for the diagnosis of leukemias is presented. Complete procedures for Prussian blue stain for iron, peroxidase, Sudan black B, naphthol AS-D chloroacetate esterase, nonspecific esterase, periodic acid Schiff (PAS), acid phosphatase, toluidine blue and alkaline phosphatase stains are given in the appendix.

Acid Phosphatase↗

A new approach to the morphological and cytochemical evaluation of human bone marrow CFUC in agar cultures.

A simple procedure is described for the routine morphological examination of different types of colonies and clusters in agar cultures. The following cytochemical methods are used for the identification of cells: naphthol-ASD-chloroacetate esterase, naphthol-ASD-acetate esterase with and without addition of sodium fluoride, Biebrich scarlet and acid phosphatase. Based on their cytochemical staining properties neutrophils, eosinophils, monocytes and macrophages can be readily identified in their different stages of maturation and an assessment made of the cellular composition of colonies and clusters.

Agar↗

Characteristics of granulocyte-macrophage colony formation in patients with chronic myelomonocytic leukemia.

3 patients with chronic myelomonocytic leukemia (CMMoL) were investigated using a culture technique in vitro for granulocyte and monocyte/macrophage precursors (GM-CFC). Time course studies revealed that the growth of colonies and clusters was delayed in CMMoL as compared with that in normal control bone marrow. In 1 of 3 cases of CMMoL, the number of GM colonies increased after removal of phagocytes. Chloroacetate esterase and nonspecific esterase activities in the colonies and clusters were examined by means of our improved cytochemical technique. When we classified colonies and clusters into granulocytic, monocyte/macrophagic, myelomonocytic and mixed type, the incidence of "myelomonocytic' colonies and clusters was significantly higher in two cases of CMMoL than in the normal controls. On the basis of these results, the characteristics of abnormal hemopoietic regulatory mechanisms were discussed.

Aged↗

Freeze-preservation of staining solutions in enzyme histochemistry.

Staining solutions of acid phosphatase and esterase group (naphthol-AS-D-chloroacetate esterase, naphthol-AS-acetate esterase and alpha-naphthyl-acetate esterase) were stored for 6 months using the freezing facilities, and the staining results were compared with those of each set of controls. The results showed that the staining solutions of all enzymes were stable for long-term preservation (at least 6 months) in the deep freezer (-80 degrees C) when they were stored appropriately in separated forms. In such a freezing procedure, the staining results for each enzyme did not show any decrease of positive intensity nor any non-specific staining.

Acid Phosphatase↗

A triple stain technique to evaluate monocyte, neutrophil, and eosinophil proliferation in soft agar cultures.

A useful cytochemical technique has been developed which facilitates the identification of the types of cell aggregates which proliferate in soft agar cultures of hematopoietic stem cells. Staining of fixed agar discs for alpha-naphthyl acetate esterase (ANAE), naphthol AS-D chloroacetate esterase (CAE) and with Luxol fast blue (LFB) in sequence results in permanent preparations in which monocytic, neutrophilic and eosinophilic aggregates can readily be distinguished on the same slide. This represents an improvement over the currently used technique which relies on the removal of a representative sample of aggregates from the agar discs for cell typing. Whole plate staining results in more accurate estimates of eosinophil growth since the frequency of occurrence of these cell aggregates is low under many of the culturing conditions used. The technique provides a useful tool for studying normal and abnormal hematopoiesis.

Bone Marrow Cells↗

Morphological, cytochemical and ultrastructural studies of a case of systemic mastocytosis.

Morphological, cytochemical and ultrastructural studies of mast cells were carried out in a patient affected with systemic mastocytosis. Neoplastic mast cells showed morphological features between classic tissue mast cells and circulating basophils. They showed strong granule metachromasia after toluidine blue, faint positivity to Hotchkiss reaction, strong positivity to chloroacetate esterase, while they were negative to alkaline phosphatase. Ultrastructural observations showed heterogeneity of granules, most of which had homogenous fine dotted contents. The origin and linkage between mast cells and circulating basophils are discussed.

Alkaline Phosphatase↗

The haematopoietic supraneural organ of adult, sexually immature river lampreys (Lampetra fluviatilis [L.] Gray) with particular reference to azurophil leucocytes.

The haematopoietic tissue in the supraneural organ of the freshwater river lamprey (Lampetra fluviatilis L. Gray) was studied in sexually immature animals. Besides erythro- and granulopoietic elements, macrophages, reticular cells, fibroblasts and glycogen-rich fat cells were seen. Developing granulocytes of the lamprey contain one type of azurophil granules originating from small cytoplasmic (Golgi) vesicles. The lamprey's azurophil granulocytes seem to be homologous with those of fishes. However, the granulocytes of fishes, studied thus far, show granules with only one type of inclusion, whereas in lamprey the granulocyte inclusions are variable in size and shape. Thus, lamprey granulocytes are, in this respect, reminiscent of similar cells of higher vertebrates. The PAS and alkaline phosphatase reactions, common markers of vertebrate neutrophil leucocytes, are very weak in the haematopoietic tissue granulocytes of the lamprey, and intense in the blood cells of the same animal. Lamprey granulocytes, similarly to the granulocytes of Chondrostei and Elasmobranchiata, do not stain with peroxidase, naphthol-AS-D-chloroacetate esterase and sudan black B. The haematopoietic tissue contains a relatively high number of degenerated granulocytes.

Animals↗

Establishment of a new peroxidase-positive human myeloid cell line, PL-21.

A myeloid cell line, designated PL-21, was established from the peripheral blood of a patient with acute promyelocytic leukemia. The PL-21 cell line grew in single-cell suspension, with a doubling time of 48-64 hr, and consisted of promyelocytes with fine immature nuclei and prominent azurophilic granules in the cytoplasm. PL-21 cells were positive for peroxidase, naphthol AS-D chloroacetate esterase, and Sudan Black B staining. Under the usual culture conditions, a small proportion of these cells differentiated into mature granulocytes, and this differentiation was enhanced by the addition of dimethyl sulfoxide in the culture medium. PL-21 cells had receptors for the Fc portion of IgG and complement, intracytoplasmic lysozyme and phagocytic activity, but lacked Epstein-Barr virus-associated nuclear antigen. Chromosome analysis of this cell line revealed a human male polyploid karyotype with 13q+ and double minute chromosomes. This new myeloid cell line may provide useful material for the study of proliferation and differentiation of human leukemia cells.

Adult↗

Monoclonal antibody MCS-2 as the marker of phorbol diester-induced myeloid differentiation in acute undifferentiated leukemia.

Leukemic cells from 32 cases of acute leukemia were cultured in vitro with the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) to study their differentiative potential. Three cases of acute undifferentiated leukemia (AUL) were studied intensively. We found that culturing of leukemic cells with TPA can induce changes in cell surface antigens. In particular, MCS-2, a "pan" granulocyte/monocyte marker, was inducible in vitro in AUL and in acute myelogenous leukemia, while it was not inducible in acute lymphoblastic leukemia. BA-2 (recognizing the Mr 24,000 protein) and TA-1 (recognizing the Mr 170,000 and Mr 95,000 proteins) were also inducible in cases of AUL, acute myelocytic leukemia, and acute monoblastic leukemia, although these antigens are not limited only to leukemias of the myelomonocytic lineage. Our studies also indicate that undifferentiated cells could be induced to nonspecific esterase and sometimes to chloroacetate esterase reactivity while losing terminal deoxynucleotidyl transferase. Morphological studies in these cases revealed cytological maturation following TPA treatment. In most cases, these changes were also partially inducible by culturing cells in medium alone or with the addition of dimethyl sulfoxide but not to the extent that was demonstrated by TPA. Our studies showed that MCS-2 is a very good, specific marker of acute nonlymphocytic leukemia. A potential use for TPA to aid in the subclassification of patients with AUL is also suggested.

Acute Disease↗

Eosinophilic leukemia with prominent visceral involvement: histopathological and histochemical observations.

Evidence supporting the view that eosinophilic leukemia exists as a separate entity among myeloproliferative disorders was found in the pathological and histochemical findings in a case of hypereosinophilic disease. Blast cells and eosinophils in all stages of maturity, including unusual atypical eosinophil precursors, were seen in a prominent infiltration of the spleen and the liver. Blasts were far more numerous in the viscera, especially in the liver, than in the marrow and peripheral blood. Enzyme histochemical studies of plastic sections showed that blasts were reactive for cyanide-resistant peroxidase, which is specific for eosinophils, and could therefore be regarded as part of an eosinophilic leukemic proliferation. Some eosinophils showed aberrant reactivity for chloroacetate esterase. The existence of a neoplastic proliferation of eosinophils is consistent with the view that eosinophils represent a distinct marrow line. The possibility that hypereosinophilic patients may harbor a far more prominent blastosis in the viscera than is apparent from marrow and blood picture is also stressed. Finally, a chromosomic abnormality previously described in acute nonlymphocytic leukemia with atypical eosinophils in the marrow is for the first time reported in a patient with hypereosinophilic disease.

Adult↗

Preleukemic granulocytic sarcoma of cervix and vagina: initial manifestation by cytology.

Granulocytic sarcoma is an unusual form of tumefaction caused by acute granulocytic leukemia. On rare occasions, the lesion precedes the leukemic phase and presents as a mass with a normal peripheral white cell count. This report describes the initial manifestation of granulocytic sarcoma by vaginal cytology in a 39-year-old female with Down's syndrome. Six days after admission, the patient died of acute peritonitis following spontaneous perforation of the bowel. Autopsy revealed involvement of cervix, vagina, bowel wall and one pelvic lymph node by granulocytic sarcoma. Bone marrow examination confirmed the preleukemic stage of the disease. Cytologically, the malignant cells occurred singly. No nucleoli were seen. The differential diagnosis between malignant lymphoma and granulocytic sarcoma rests upon a positive naphtol AS-D chloroacetate esterase stain in granulocytic sarcoma. This stain may be performed on paraffin-embedded sections or on smears.

Adult↗